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Malignant rhabdoid tumor (MRT) is an aggressive, highly lethal cancer that strikes young children. Tumors occur in various locations including kidney, brain, and soft tissues. Despite intensive therapy, 80 percent of affected children die, often within 1 year of diagnosis. The vast majority of MRTs have sustained bi-allelic inactivating mutations of the hSNF5/INI1 gene, suggesting that SNF5 may act as a tumor suppressor. Supporting this idea is the fact that 15 percent of children with MRT have somatic mutations in one allele of SNF5 and loss of the remaining allele in their tumors. SNF5 is a core member of the SWI/SNF chromatin remodeling complex, which is required for regulated expression of a subset of genes. In humans, SNF5 binds to HIV integrase and stimulates integration of HIV into human DNA. Studies have also shown that SNF5 binds to EBNA, MLL/ALL- 1, trithorax, and c-MYC and stimulates transcription by these factors. Another member of the SWI/SNF complex, Brg1, directly interacts with pRb and is required for Rb mediated cell cycle arrest. Haploinsufficiency of Brg1 predisposes mice to tumor formation and Brg1 is deficient in several breast cancer cell lines. Together, these data suggest a widespread role for SWI/SNF in tumor suppression. While it is clear that SNF5 and SWI/SNF are involved in tumor suppression, chromatin remodeling and transcriptional activation, their mechanism of action and the relationship between these processes is not clear. We have used gene targeting to inactivate SNF5 in mice. Absence of SNF5 results in embryonic lethality while haploinsufficiency results in 15 percent of mice developing tumors histologically indistinguishable from human MRT. The specific aims of the proposed project are to generate SNF5 deficient cells to determine the function of SNF5 in growth regulation and oncogenic transformation. Second, to improve the MRT model by generating conditionally targeted mice that develop MRT with high penetrance in locations where human tumors occur. Third, to identify genes downstream of SNF5 through use of conditionally targeted cells in DNA micro-array analysis. Lastly, to investigate the function of SNF5 in transcriptional regulation by analysis of SNF5 deficient cells using in vitro transcriptional assays. Since SNF5 is an invariant subunit present in all SWI/SNF complexes, elucidation of its function will provide insight into a newly appreciated mechanism of tumor suppression and may identify new targets for therapeutic intervention against a lethal pediatric cancer. This proposal directly addresses scientific priorities identified by the BT-PRG arm of the NCI.
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{
"pile_set_name": "NIH ExPorter"
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The nuclear pore complex (NPC) is a large assembly 1200-1400 angstroms in diameter and 600-800 angstroms thick which spans the nuclear envelope. The NPC is the communication pathway between the nucleus and the cytoplasm, allowing the passive diffusion of small molecules and ions and actively transporting large macromolecules and ribonucleoprotein particles between these two compartments. Adeno and herpes viruses bind to the NPC prior to release of their DNA into the nucleus. Intact retrovirus capsids are transported through the NPC. Despite the NPC's prominent location in the cell and its importance in controlling nucleoplasmic exchange of (e.g.) hormone receptor complexes, regulatory proteins, mRNPs, ribosomes and virus components, there is relatively little known about this assembly: the NPC cannot be isolated in sufficient quantities for biochemical characterization or analysis by x-ray methods, only very low resolution 3-D information is available on its structure vary few (<10%) NPC proteins have been identified and the molecular mechanisms of the transport processes are unknown. The long term goals of the work are to understand the structure of the NPC ar high resolution and to elucidate the molecular mechanism of active nucleocytoplasmic transport. The accomplishment of the work described herein will represent major progress towards attaining these goals. Cryo-electron microscopy coupled with computer image processing will be used to determine projection maps and a 3-dimensional map at a resolution of about 50 Angstroms from improved preparations of detergent-released NPCs. Subsequently, the 3-D location of a number of important NPC proteins-- including those necessary for nuclear transport -- will be determined by immunoelectron microscopy and image analysis. An examination of freeze-dried, metal-shadowed preparations of manually spread nuclear envelopes will reveal structural differences on the nuclear and cytoplasmic sides of the NPC. These findings will be related to details in the 3-D map. Experiments using synthetic nuclear localization signal peptides have been designed to investigate the identity of the central plug of the NPC and may provide structural information on the transport apparatus.
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{
"pile_set_name": "NIH ExPorter"
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PROJECT SUMMARY/ABSTRACT: This proposal describes a 5-year research and training program for Dr. Maria Nagel to develop an independent academic career studying varicella zoster virus (VZV) infection, latency and reactivation. More than 90% of the population is infected with VZV before adolescence (Seward et al., 2000). As cell-mediated immunity to VZV declines in elderly and immunocompromised individuals (Gershon and Steinberg, 1981; Burke et al., 1982), VZV reactivates and causes multiple serious neurological diseases including zoster (shingles) frequently complicated by chronic pain (postherpetic neuralgia), stroke, blindness and spinal cord disease. VZV replicates in all human cells except neurons where virus becomes latent. The mechanisms that determine productive versus latent infection are unclear, but likely involve cell type-specific interactions of viral proteins and host cell antiviral defense mechanisms. Recent studies on herpesviruses show that viral proteins interact with antiviral promyelocytic leukemia nuclear bodies (PML-NBs) in the host and that the outcome of the interaction contributes to productive infection versus latency (Everett and Chelbi-Alix, 2007); in addition, the structure of PML-NBs is dramatically altered in cells undergoing mitosis compared to PML-NB stability in post-mitotic cells such as neurons (Dellaire et al., 2006a). Based on these observations, we hypothesize that the outcome of VZV infection (lysis versus latency and subsequent reactivation) results from cell type-specific interactions of VZV proteins with the host cell's antiviral PML-NBs. To test this hypothesis, we will: (Aim 1) analyze non-neuronal cells productively-infected with VZV for interactions of PML-NBs with VZV gene 61 protein (VZV 61p), which is homologous to herpesvirus proteins known to interact with PML-NBs, and with VZV gene 63 protein (VZV 63p), the most prevalent and abundant VZV protein expressed in latently-infected human ganglia, using immunochemistry and confocal microscopy. The effects of PML, VZV 61p, and VZV 63p overexpression and underexpression on VZV gene expression and viral titers will also be examined; (Aim 2) analyze latently-infected human ganglionic neurons obtained at autopsy for interactions of PML-NBs with VZV 61p and VZV 63p using techniques in Aim 1 and fluorescent in situ hybridization to detect VZV DNA in ganglionic neurons; and (Aim 3) analyze human ganglion neurons obtained at autopsy during early explant-induced reactivation for interactions of PML-NBs with VZV61p and VZV 63p using techniques from Aim 2. During the award period, Dr. Nagel will be promoted to Assistant Professor in the Department of Neurology, which provides a fruitful environment for physician-scientists. Training will be enhanced with didactic courses in molecular virology, bioethics, and statistical analysis. She will devote 80% effort to research and 20% to patient care/clinical teaching. She will apply for an independent R01 award during her 4th year of training. Her primary scientific mentor will be Dr. Randall Cohrs, Research Professor of Neurology at the University of Colorado Denver, a basic research scientist and one of the world's experts on VZV pathogenesis. Despite the development of a zoster vaccine, even if every American over age 60 received zoster vaccine, more than 500,000 new cases of zoster will occur annually. Thus, an understanding of viral and host cell interactions leading to VZV latency and reactivation will provide the basis for strategies to prevent serious neurological disease caused by VZV.
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{
"pile_set_name": "NIH ExPorter"
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Endometrial carcinoma is the most common gynecological cancer in the United States, with an estimated incidence of over 40,000 cases per year. Endoscopic imaging is an important diagnostic tool for the evaluation of the uterus, but is typically limited to viewing the surface of the endometrium. Therefore, we propose to develop technology for depth resolved imaging of the endometrium. The innovation lies in a unique aperture-based imaging system, which obtains several photon migration paths in a single image, capture. Our Phase I effort is focused on development and validation of the depth-resolved imaging system in a phantom model. First, we will use Monte Carlo modeling to determine crucial design parameters. These results will then be used to create an aperture-based imaging prototype. Finally, a phantom model will be created and used to evaluate the endoscope's ability to probe a turbid medium. If we are successful, we envision a Phase II effort focused on the development and testing of a clinical grade prototype. It is our long-term goal to develop a tool, which, for the first time, would enable the gynecologist to evaluate and visualize submucosal lesions in an office setting.
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{
"pile_set_name": "NIH ExPorter"
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OBJECTIVE: To identify the molecular form of GF-6 immunopositive cells in the fetal forebrain. RESULTS We had previously discovered, using antiserum GF-6, two types of LHRH-like neurons, early and late, in fetal forebrains. The late cells arose from the olfactory placode at E32-E36, migrated into the forebrain at E38-E42, and reached the preoptic area and hypothalamus at E50-70. They are bona fide LHRH neurons based on immunostaining with antisera for mammalian LHRH, but not for any other LHRH forms. The origin of early cells was unclear A small number of early cells were seen in the olfactory placode and the forebrain at E30-E32, and a large number of early cells were present in the forebrain at E32-E34. They settled in the extrahypothalamic area (septum, stria terminalis, amygdala, striatum, and internal capsule) at E50-E70, but not in the preoptic area and the hypothalamus. The molecular form of early cells is unknown, since they were not immunostained with any specific antisera against known forms of LHRH. We studied their form using antisera against LHRH1-5 or metalloendopeptidase E.C.3.4.24.15 (EP24.15), an enzyme that cleaves LHRH at the Tyr5-Gly6 position in 9 fetuses at E36 to E70. We found that the late cells were immunostained with antisera GF-6 and LR-1, but not with antisera for LHRH1-5 and EP24.15, while the early cells were readily immunostained with antisera for LHRH1-5 and EP24.15, as well as antiserum GF-6. Exposure of those antisera, preabsorbed with various peptides, revealed that the early cells (extrahypothalamic cells) contain LHRH1-5 and its cleavage enzyme, EP24.15. A group of neurons containing an LHRH1-5 fragment are present in the basal forebrain before the migration of neurons containing full mature LHRH, and they distribute widely in the brain. FUTURE DIRECTIONS We will examine the function of this LHRH fragment in the brain. KEY WORDS LHRH neurons, LHRH fragments, endopeptidase, olfactory placode, fetal brain
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{
"pile_set_name": "NIH ExPorter"
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Understanding neurogenesis is an essential goal of neuroscience. Naturally occurring sex differences in neurogenesis are of inherent interest but also provide a tractable tool for exploring novel mechanisms not otherwise apparent. Using the laboratory rat we have identified and characterized a profound sex difference in hippocampal neurogenesis restricted to a perinatal sensitive period. Newborn males make up to twice as many new cells as females in the dentate gyrus, CA1 and to a lesser extent CA3 of the hippocampal formation, and 80% of those newborn cells will become neurons while only 40% will do so in females. Thus males have higher rates of both proliferation and neuronal differentiation. Sex differences in the brain are often the byproduct of increased testicular androgen production that is locally converted to estradiol in the brain. We propose the novel hypothesis that the sex difference in hippocampal neurogenesis is the product of two sources of epigenetic repression in females; 1) canonical changes to the DNA and histones via methylation and acetylation that repress pro-proliferation genes, and 2) microRNAs which regulate gene expression by degrading pro-differentiation mRNAs. We further contend that the higher level of microRNAs we detect in neonatal female hippocampus is at least in part determined by chromosome compliment, that is, XX vs XY. A candidate gene, BDNF, has emerged and detailed interrogation of BDNF gene expression and effects will be combined with broader based surveys of the hippocampal methylome and chromatin state to elucidate the source of the sex difference in neurogenesis via the following specific aims: SA1 - Identify the mechanism mediating sex differences in epigenetic control of neurogenesis in the developing hippocampus, SA2 - Determine microRNA profile and mechanism suppressing neurogenesis in developing female hippocampus and SA3 - Establish cross-talk between epigenetics and microRNAs in neonatal hippocampal neurogenesis. Completion of these aims will provide novel insight into the regulation of neurogenesis in the developing healthy brain and illuminate the source of higher male vulnerability to the consequences of neonatal brain injury or insult.
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{
"pile_set_name": "NIH ExPorter"
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The objective of this proposal is to determine how chemokines and newly identified chemokine antagonists, Slits, are regulated, interact with each other, and contribute to inflammation in response to renal injuries. Past research in chernokines has been focused mostly on positive regulation. Thus, in all these situations chemokines are invariably studied as activators or attractants. Recently, we have found that one of the neural axon guidance cues, Slits, was expressed outside of the nervous system, Slit-2 was expressed in the kidney and lung. Leukocytes have been considered as early and important effectors in renal inflammation. Previous studies have shown upregulation of chemokines and downregulation of Slit2 during glomerulonephritis and Slit treatment significantly attenuated nephritic injury. It was reported that Slit/Robe may function through Robe's binding to GTPase activating proteins (GAPs, srGAPs) Recently, we have found that Racl and Cdc42, small GTP binding proteins known for their roles in actin polymerization and cell motility, were shown to be negatively regulated in macrophage-like cells by Slit2. We plan to study the expression and regulation of chemokines, Slits, and Robe in the model of renal ischemia/reperfusion and further determine the role of Slit/Robe in negative regulation of leukocyte accumulation in this model. Another goal of this project is to characterize Slit/Robe mediated signaling of small G proteins in leukocytes. Differences between activation of GTPases between neurons and leukocyte suggests that Slit/Robe in ieukocytes may use other signaling molecules in addition to srGAPs, therefore, we plan to further clone Robe-interacting proteins in mononuclear cells by Two-Hybrid System. The proposed studies will shed new light on how chemokines and Slit interact in leukocyte-dependent renal injury, and uncover the signaling pathway of Slit/Robo, and insights gained from our study will lead to the development of new therapeutic strategies of chemokine-dependent renal injuries in human.
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{
"pile_set_name": "NIH ExPorter"
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Mitochondria are essential, complex organelles of eucaryotic organisms required for a variety of metabolic processes including the generation of energy by oxidative phosphorylation. Biogenesis and maintenance of mitochondria requires the function of molecular chaperones such as Hsp7O. Our long term goal is to understand the mechanism of action of molecular chaperones within mitochondria using S. cerevisiae as a model system. Using genetic and biochemical approaches the analysis of the roles of mitochondrial chaperones of the Hsp7O and Hsp4O classes in the processes of protein translocation, folding and assembly of mitochondrial proteins will be continued. These studies are relevant to issues of human health, as certain human tissues, such as brain, heart, muscle and kidney, are particularly dependent on efficient mitochondrial function. Pathological effects caused by reduced bioenergetic capacity have been found to be caused by mutations in both mitochondrial and human DNA in human populations. In addition, one of the mitochondrial Hsp7Os has been implicated in the maturation of the homologue of human frataxin, which is associated with the neurodegenerative disease Freidrich's ataxia. Ssc1, an Hsp7O of the mitochondrial matrix, is an essential component of the apparatus required for translocation of proteins from the cytosol. Ssc 1 is tethered to the import channel via its interaction with an essential peripheral component of the channel, Tim44. Mge 1, an essential nucleotide release factor for Ssc 1, is also associated with Ssc1 at the import channel. Ssc1, an Hsp4O Mdjl, and the nucleotide exchange factor Mgel are thought to facilitate folding of imported proteins. This proposal is designed to understand the pathway of Ssc1 function in translocation of proteins across the mitochondnal proteins and their subsequent folding in the matrix. Ssq1 and Jaci are additional Hsp7Os and Hsp4Os, respectively, of the mitochondrial matrix. A role for Ssq1 in the regulation of iron metabolism and/or the assembly of Fe-S centers is indicated. Our goal is to understand the function(s) of Ssq1 in mitochondria and to provide insight into the cellular process(es) in which Ssql acts. The primary and secondary effects of the lack of Ssql function, focusing on iron metabolism, including its role in the maturation of Yfh 1, the yeast homologue of frataxin, and assembly of Fe-S clusters will be determined using a combination of genetic and biochemical techniques.
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{
"pile_set_name": "NIH ExPorter"
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Yersinia pestis, Salmonella enterica, and Yersinia enterocolitica use a number of toxins to evade the immune system one of which is the protein tyrosine phosphatase YopH. Following infection, YopH is injected into phagocytic cell resulting in the disruption of focal adhesions, and inhibition of integrin-mediated bacterial phagocytosis, both of which are highly dependent on tyrosine phosphorylation. Hence, targeting YopH against these pathogens represents a sensible yet underexplored strategy for the development of novel anti-bacterial adjuvant to antibiotics. Our hypothesis is that effective small-molecule inhibitors f YopH would render YopH- paralyzed immune cells immediately functional again hence able to initiate both innate and adaptive immune responses. Hence, the molecules that will arise from this pilot study may result very useful not only as potential therapeutics, but also in probing at the cellular level the mechanisms of toxin induced cell death and inhibition of phagocytosis, common to several other pathogens.
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{
"pile_set_name": "NIH ExPorter"
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Interaction of the molecular motor myosin with actin drives muscle contraction and various changes in cell shape and movements that are fundamental to the biology of nonmuscle eukaryotic cells. Essential to the function of myosin in nonmuscle cells is the regulation of its assembly into filaments and higher order structures such as the contractile ring, the assembly of which is spatially and temporally controlled. Little is known about the molecular basis of such spatial and temporal control but phosphorylation of the myosin molecule appears to play a key role. Light chain phosphorylation may regulate myosin filament assembly in some systems, but may primarily control the motor function of myosin. Heavy chain phosphorylation may be primarily involved in regulation of myosin filament assembly. The specific aim of this proposal is to study the roles of myosin phosphorylation in Dictyostelium, an organism that allows the convergence of biochemical, molecular genetic, and physiological approaches. It therefore offers a unique opportunity to investigate the molecular basis of how a cell moves, divides, and changes shape in response to cellular and developmental signals. The experimental plan for the next five year period can be divided into three parts: 1. Biochemical, structural, and molecular genetic studies on the kinases and phosphatases that are responsible for the control of Dictyostelium myosin phosphorylations. 2. Biochemical and structural studies on the effects of these phosphorylations on the functions of purified myosin. 3. Characterization of the effects of phosphorylation on the in vivo behavior and function of the myosin. The latter is made possible by recent applications of sophisticated molecular genetic techniques to Dictyostelium.
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{
"pile_set_name": "NIH ExPorter"
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This study was designed to evaluate the efficacy of training in hypnosis in reducing pain and anxiety in children with acute lymphoblastic leukemia (ALL) undergoing bone marrow aspirations (BMA's). Patients/subjects were selected from the population of children between the ages of 6 and 10 years of age with ALL, receiving outpatient treatment in the Division of Hematology-Oncology at Childrens Hospital of Los Angeles. The design calls for randomization of 40 patients to either hypnosis or attention/placebo groups with stratification of the sample by sex. Baseline measures are collected along four dimensions: Behavioral observation, nurse ratings, structured self-ratings of pain and anxiety. Two training sessions are offered post-baseline and prior to the next outpatient bone marrow aspiration. Further sessions are provided prior to each of three post-baseline bone marrow aspirations and response to training is monitored using the four dependent measures. Seventeen patients accrued to the study in Year 1, 9 in the hypnosis group, 8 in the comparison group. Baseline data was collected on all patients. Fourteen of 17 patients have completed 2 sessions of introductory training (7 each in both hypnosis and comparison groups). Twelve patients have undergone at least one post-training bone marrow aspirations (6 each in both groups). Of these, (three patients in the hypnosis group and one patient in comparison group) have undergone 2 BMA's, and 2 patients (one each, in both groups) have completed the study protocol and have gone off-study (three post-training BMA's). Two methodological modifications were made. Hypnotic susceptibility testing was not done because the major pediatric susceptibility scales were, essentially, hypnotic inductions. Exposing comparison group patients to hypnotic induction would have confounded the controlled nature of the study. A quantified rating of the child's response to either hypnosis or the attention/placebo dimension was added as a factor to be correlated with treatment efficacy. All data were coded prospectively.
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{
"pile_set_name": "NIH ExPorter"
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Funds are requested to develop an NCCU NCMHD Research Center at North Carolina Central University that focuses on minority health disparities. The Center will be housed in the Julius L. Chambers Biomedical/Biotechnology Research Institute and will comprise Administrative, Research, Research Education/Training, and Community Engagement/Outreach Cores. The Administrative Core will have oversight of all of the Center's programs, activities, and budgets. It will manage the selection of core faculty and research/pilot projects, and coordinate the development of all partnership arrangements. A common theme of projects that will be conducted by the research core is that minority health disparities may result [unreadable] from complex gene-environment interactions. The Research Core will support two full research projects and two pilot projects: the first research project is directed at investigating the interaction of psychosocial factors and susceptible genetic polymorphisms in contributing to the physiological blood pressure response to environmental stress and the subsequent development of hypertension in African Americans; and the second will seek to identify and characterize Genetic and Epigenetic Factors associated with Prostate Cancer Disparity. The pilot projects provide unique opportunities for research project expansion and faculty development. Three shared resources facilities will provide technical and logistical support for the research and pilot projects. The research education/training core will oversee laboratory and classroom training for high school, undergraduate, and graduate students, as well as post-doctoral associates and junior faculty. It [unreadable] will also assist the academic units with developing new curricula and degree programs. A Community Engagement/Outreach Core will continue its work on developing methods for effectively communicating health education messages to minority communities. It is anticipated that the NCCU NCMHD Research Center will become a valued, trusted, institution-wide resource for expanding NCCU's capacity to conduct minority health and health disparities research. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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In order to assess the role of the thymus in the acquisition of the T cell self-MHC specific repertoire. The MHC specificity of cytotoxic precursor T cells differentiating within thymuses engrafted into athymic nude mice was determined. The results demonstrated that cytotoxic T lymphocytes from such engrafted thymuses recognized TNP in association with thymic MHC determinants in various haplotype combinations of thymus graft and nude host. In contrast, cytotoxic T lymphocytes from the spleens of these same thymus grafted nude mice recognized TNP on both nude host and thymic MHC determinants. These results are consistent with the thymus being one site which determines MHC restrictions of T cell self recognition, while there exist other factors in this experimental model which have determined the host-restricted MHC specific repertoire observed in the spleens of thymus engrafted nude mice.
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{
"pile_set_name": "NIH ExPorter"
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The hypothalamic-pituitary-adrenal (HPA) axis and its byproduct, cortisol, have been hypothesized to be the mediators that links chronic stress to adverse health outcomes, including increased obesity risk. This mediation is hypothesized to occur via several mechanisms, including stress-related cortisol secretion leading to both increased caloric consumption and a shift in preference to comfort foods (i.e., foods high in fat and/or added sugars). Under conditions of chronic stress, aberrations in the normal diurnal pattern of cortisol secretion occur, and obese, compared to non-obese, adults have been reported to demonstrate these aberrations more frequently. Although it is vital to understand the origins of these associations and the mechanisms through which they are established, few studies have examined links between HPA axis functioning and obesity in childhood. Furthermore, no studies have examined potential links between stress, cortisol, eating behavior, and obesity among low-income preschool children, though this population carries the greatest burden of the childhood obesity epidemic, and disparities in obesity prevalence emerge in this age range. This application, therefore, proposes 3 specific aims: Aim 1: To examine the association between diurnal cortisol patterns and weight status within a population of low-income preschool-aged children. Hypothesis 1: Children with alteration of the normal diurnal pattern of cortisol secretion will demonstrate a higher prevalence of obesity compared to children with the typical diurnal pattern. This alteration of the diurnal pattern will most commonly present as hypercortisolism, but may also present as hypocortisolism. Aim 2: To examine the association between diurnal cortisol patterns and eating behaviors within a population of low-income preschool-aged children. Hypothesis 2: Children with alteration of the normal diurnal pattern of cortisol secretion will demonstrate a higher prevalence of food-related tantrums, emotional eating, and eating in the absence of hunger compared to children with the typical diurnal pattern. Aim 3: To examine the association between diurnal cortisol patterns and dietary composition within a population of low-income preschool-aged children. Hypothesis 3: Children with alteration of the normal diurnal pattern of cortisol secretion will consume more comfort foods (foods high in fat and/or added sugars) compared to children with the typical diurnal pattern. PUBLIC HEALTH RELEVANCE: Childhood obesity is a significant public health problem, particularly for low-income populations. Obesity, once established in childhood, is very likely to track into adulthood. More effective approaches to preventing and treating childhood obesity are needed. A focus on the intersection of stress, eating behavior, and obesity may inform the development of more effective intervention and prevention programs.
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{
"pile_set_name": "NIH ExPorter"
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We have identified a novel pathway for clearing misfolded cytosolic proteins from human cells that directs them to the endoplasmic reticulum (ER). This pathway is used to clear cells of a naturally occurring misfolded NAT1 variant; NAT enzymes acetylate arylamines leading to their detoxification. NAT1 WT by contrast is stable and not present at the ER. We have found other candidates in the literature that may be degraded in this manner and are testing this model further. We are also performing experiments to identify the components involved in this quality control pathway. Importantly, we are looking into exploiting this pathway as a method for clearing cells of misfolded or damaged constituents that persist in human cells and are associated with human diseases. Parkin R42P is an example of a naturally occurring misfolded protein that is trafficked poorly to ER and stable. This Parkin variant is associated with Juvenille Parkinsonism and has a mutation in its N-terminal UBL domain that renders it unstructured. We also find that we can artificially direct Parkin R42P to ER by fusion with the Sec61beta ER-directing transmembrane domain and that this induced ER-localization triggers its clearance. We are expanding on these findings to test further the application of routing misfolded cytosolic proteins to the ER as a strategy for clearance.
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{
"pile_set_name": "NIH ExPorter"
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Olfactory disability represents a danger to the patient resulting from inability to detect hazards such as natural gas and spoiled food and threatens quality of life through the loss of enjoyment of foods and fragrances. Rhinosinusitis is a primary cause of olfactory loss among patients presenting to cliemosensory clinics, and is the most common chronic medical condition in the United States, affecting nearly 33 million people/year. Chronic rhinosinusitis (CR) patients with similar degrees of nasal inflammation may present with normosmia, hyposmia or anosmia, and current therapies are only partially effective. CR may impair olfaction in several ways. Constriction of the airway due to inflammation could alter or impede airflow through the nasal cavity, reducing access of volatile compounds to the sensory receptor cells. Changes in the composition and transport of the watery mucus through which the odorants must diffuse could impair diffusion to or removal from the receptor sites. Proteins secreted by the infiltrating cells of the immune system may directly injure or modulate the function of ORNs or other cells within the neuroepithelium. Fibrosis, gland hyperplasia, keratinization or edema may change the structure of the epithelium or submucosa, impeding axonal outgrowth and preventing successful regeneration of the neuroepithelium. Our limited understanding and the relative importance of these diverse mechanisms and the cellular and molecular processes involved is limiting the development of diagnostic, therapeutic and prognostic tools. This project addresses this need through studies aimed at better understanding the psychophysical (Aim 1), gross anatomical and inflammatory (Aims 1, 2 and collaboration with Project 2) and sensorineural (Aims 3, 4) effects of CR. By comparing data from CR patients with olfactory loss to normosmic CR patients before and after treatment, we will identify features that may be prognostic for olfactory loss/recovery and gain insight into the causes for olfactory loss in CR. We will also develop novel tools for studying airflow dynamics (Aim 1 and collaboration with Project 2) and the effects of CR-associated inflammatory mediators on neurogenesis (Aim 4). Results from this project will enable improved diagnosis and treatment targeting by providing a thorough characterization of the patient population and identifying specific profiles of anatomic, physiologic and sensory characteristics associated with olfactory loss and recovery prospects. Data may also suggest new targets for drug development and will establish a model in vitro system for use in identification and evaluation of novel therapeutic approaches.
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{
"pile_set_name": "NIH ExPorter"
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We propose to test the hypothesis that heterocyclic amine carcinogens (HAs) induce mutations in genes within cells of specific target tissues that are critical to oncogenic progression. Our goals are to identify and characterize these genes using a rat mammary gland carcinogenesis model, and to relate these findings to tumor progression and to the HA which induced the tumors. We propose three specific aims. The goal of the First Aim is to locate regions of gain/loss in DNA from mammary tumors induced by PhIP, MeIQx, IQ, and MeIQ. We will accomplish this by extracting DNA from rat tumors known to be induced by these heterocyclic amine carcinogens, and locate chromosome regions gained, lost or amplified in the tumors using comparative genomic hybridization (CGH). The goal of the Second Aim is to determine which genes are expressed in the chromosomal regions identified in Aim 1 in normal mammary tissue, and then to determine which of those genes are lost, amplified or mutated in the same chromosomal regions in rat mammary tumors. To accomplish this, normal rat mammary cDNA libraries will be hybridized to normal rat metaphase cells. We will microdissect the chromosome regions of these metaphase cells that were shown to be involved in HA-induced tumorigenesis under Aim 1, then sequence the recovered genes and place them on microarrays. mRNA isolated from each tumor will then be hybridized to the microarrays to determine which genes appear over- or underexpressed. Genes that appear to be amplified or lost will be sequenced to identify potential mutations that change DNA consensus sequences as well as protein coding regions that play an important role in HA-induced tumorigenesis. The goal of the Third Aim is to analyze the progression of genetic changes in mammary tumors. This will be accomplished by microdissecting small groups of cells from paraffin-embedded tumor sections, perform CGH on DNA isolated from these cells, and compare these data with CGH results obtained from total tumor DNA. We will also characterize the presence or absence of specific gene deletions and mutations discovered in Aim 2 using PCR and microarray analysis on DNA obtained from tumor sections. By analyzing cells at increasing distances from the center of tumors we will be able to identify and characterize the cellular events that drive tumor progression.
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{
"pile_set_name": "NIH ExPorter"
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Project Summary/Abstract: Cochlear tonotopy describes how different frequencies are represented along the apex to base axis of the organ of Corti. A small number of about 3500 sensory cells constitute each of the four rows of hair cells in the human organ of Corti, which in essence resolves the wide range of frequencies necessary for speech discrimination and communication. For a long time, its limited number of cells and the difficult access hampered investigations on the mechanism determining the fine-tuned molecular architecture of this sensory organ. The overall aim of this study is to overcome limitations caused by the paucity of the target cell population and to establish novel strategies to investigate the highly dynamic process of organ of Corti development using single cell RNA based technology in combination with spatial and temporal transcriptomic analyses. Research proposed in this project will resolve embryonic gene expression gradients along the tonotopic axis using spatial trajectory reconstruction methods from whole transcriptome RNA sequencing data obtained from single murine organ of Corti cells. Gradients depending on sonic hedgehog signaling may potentially confer spatial information consecutively translated in the tonotopic organization of the sensory organ. The aim of this proposal is to test this hypothesis by comparing knock out and over-expression models of sonic hedgehog signaling with respect to changes in previously identified gene expression gradients in the embryo. Embryonic manipulations of sonic hedgehog signaling potentially disturb cochlear tonotopy permanently. Analyzing transcriptional profiles and physiological properties of cochlear hair cells after the onset of hearing will test this hypothesis by comparing loss of function, gain of function, and control specimen.
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{
"pile_set_name": "NIH ExPorter"
}
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IQGAP1 is a 190kD cytosolic protein scaffold that possesses several protein-protein interaction domains. The results of recent research link IQGAP1 to numerous proteins including growth factor receptors, MAP kinase signaling components, and cell junction proteins. Although the details are not well understood, through diverse interactions IQGAP1 has an important role in numerous processes including the definition of cell polarity, directed cell migration, and cell invasion. There is a growing body of circumstantial evidence that IQGAP1 is involved in carcinogenesis, and IQGAP1 is overexpressed in several different neoplasms. Among other intriguing observations, over-expression of IQGAP1 has been shown to destabilize cadherin-based cell-cell junctions and increase cell motility and invasiveness in a Rac1/Cdc42-dependent manner. Currently, there is very little structural information for IQGAP1 and its protein-protein interactions. However, several observations imply that IQGAP1 is not a passive scaffold that merely collects signaling components. First, the oligomerization of IQGAP1 is required for normal function. Second, IQGAP1 is a novel effector of the Rho-family GTPases Rac1 and Cdc42 and binding of activated Cdc42 enhances dimer formation and modulates interactions mediated by the IQGAP1 C-terminus. Third, a mutation that introduces 17 residues within the IQGAP1 GAP-related domain (GRD) confers a constitutively-active phenotype; IQGAP1 molecules that behave as though bound to active Cdc42. The 17 residue insertion is located on the surface of the GRD that would bind GTPases if the mode of binding is similar to the Ras?RasGAP interaction. Lastly, IQGAP1 amino terminal residues inhibit in trans the interaction between the IQGAP1 C-terminus and N-WASP. Based on these and other observations, we believe that through diverse inter- and intramolecular interactions, IQGAP1 exposes and conceals binding determinants necessary for the regulation of its functions. The focus of this proposal is to elucidate the IQGAP1 tertiary structure and the structural requirements for IQGAP1's interactions with binding partners.
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{
"pile_set_name": "NIH ExPorter"
}
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[unreadable] The relevance of the vivarium in regard to public health is to provide the services and resources to assist and support the development and maintenance of animal models used to advance the study of age-related diseases and mechanisms of aging. The vivarium design will be comprised of maintaining current and developing transgenic lines, utilized both internally and externally by collaborating institutions, and of specific services offered to the investigative staff. Specifically, the vivarium will provide an Association for the Assessment and Accreditation of Laboratory Animal Care (AAALAC) accredited facility, exceeding all state and federal regulations, to maintain animal models relating to: changes in gene expression with age, degenerative disease models pertaining to mitotic and post-mitotic tissues, mortality kinetics and experimental drug therapies in conjunction with a variety of transgenic mouse models. The vivarium and these animal models will combine the expertise of investigative staff, technical staff and facility management to facilitate interdisciplinary approaches to cell regulation, differentiation, proliferation, transformation and metastasis. The vivarium will provide husbandry care, and veterinary care of specific-pathogen free and immunodeficient animals, as well as the following services: technical applications (injections, dosing), surgical applications to include micro-injections for induction of specific transgenic lines, ovarian transfer surgeries, breeding colony maintenance, euthanasia training and services, perfusions, necropsy services, routine serology monitoring and testing, quarantine procedures, and staff training for protocol specific needs and will ensure required training policies for animal users. Additional ventilated caging equipment is requested to facilitate housing the additional animals produced on current studies and from the transgenic production services. The ventilated equipment will reduce the husbandry and bedding supply costs. The transgenic and technical service equipment is needed to support approximately 40 new constructs per year and current studies utilizing the vivarium that have anticipated colony expansion within the next year. The Attending Veterinarian will provide expert advice and guidance for all surgical and technical proposals that are submitted to the Institutional Animal Care and Use Committee (IACUC), and will address health issues in the colonies via the vivarium health surveillance program. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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This project (Phase I and Phase II combined) is devoted to the development of a library of homogeneous catalysts, recoverable through liquid/liquid methods, used for asymmetric hydrogenation. Polium Technologies, Inc is seeking SBIR funds for this project because we see a significant demand for such products from industry, but the company cannot afford their development without external funding. The significance of this project is a direct consequence of a huge demand for new chiral technologies fueled by the $160 billion market of single-enantiomer pharmaceutical products. Technology developed in the course of this project would significantly simplify the large-scale separation and recycling of expensive chiral catalysts. This would increase cost-effectiveness of prospective chiral catalytic processes, making them attractive to manufacture a wider range of single-enantiomer compounds. In 2002, the market for such technologies was $7 billion, and it is projected to reach $15 billion by 2009. in Phase I Polium Technologies, Inc will perform a feasibility study. Specifically we will (a) prepare several homogeneous catalysts recoverable by liquid/liquid extraction, starting from several of the most frequently used chiral phosphine ligands; (b) demonstrate activity of these catalysts in model hydrogenation reactions. As of July 29, 2003, there are no publications or patents on catalytic systems for asymmetric synthesis recoverable by liquid/liquid processes or their use in catalysis.
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{
"pile_set_name": "NIH ExPorter"
}
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Research is proposed which will investigate information processing in the caudate nucleus of cats exposed to the dopaminergic toxin N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Chronic extracellular single unit recordings from caudate neurons will be used to assess how these cells respond to specific sensory signals (somatosensory, visual, auditory stimulation, electrical stimulation of cortex) under normal circumstances and at various times after exposure to MPTP. MPTP causes damage to the nigrostriatal system in the cat causing a distinct behavioral syndrome characterized by an acute phase (approximately 90% striatal dopamine depletion) consisting of parkinsonian motor signs (akinesia, bradykinesia, "freezing" during attempted movement) and a reduced responsiveness to external stimulation and subacute and chronic phases in which gross motor behavior approximates normal while sensory responsiveness remains somewhat impaired and substantia nigra pathology is extensive as is the striatal dopamine loss (approximately 70%). The present experiment is designed to examine sensory information processing in the caudate nucleus during the various phases of the MPTP-induced behavioral syndrome to see if disturbances in sensory information processing (during times of sub-maximal and maximal dopamine loss) are apparent irrespective of the appearance of motor symptoms. This is important since many of the cognitive changes associated with basal ganglia disease in humans often appear before motor symptoms (and maximal dopamine loss). These experiments should assess the way in which the caudate nucleus processes afferent signals during varying degrees of dopamine denervation and establish the feasibility of future studies of the behavioral physiology of the MPTP syndrome in the cat.
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{
"pile_set_name": "NIH ExPorter"
}
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Patients with peripheral arterial disease (PAD) experience significant functional limitations due to ischemic symptoms (claudication) and are at high risk for CVD morbidity and mortality resulting from untreated cardiovascular disease (CVD) risk factors and aggressive atherosclerosis. The overall Goal of this randomized controlled clinical trial is to examine the synergistic effect of a multifactor risk reduction program based on components of a chronic care delivery system (i.e. clinical decision support, planned and structured visits, follow-up, application of behavioral change theories, use of a multidisciplinary team approach and self-management) on improving clinical and biological outcomes in 300 patients with PAD. Specifically, we will compare the effects of 24 months of a novel, yet well-tested multiple risk factor reduction program, the Health Education and Risk Reduction Training (HEAR2T) Program for PAD versus enhanced standard care on: 1) symptom limited walking distance as assessed by treadmill exercise testing and walking impairment questionnaire; 2) endothelial function as measured by flow mediated vasodilation (FMVD) via brachial artery ultrasound. We will also explore the association between FMVD and decreased oxidative stress (as measured by oxygen radical absorbance capacity and urinary isoprostanes) and reduced degradation of nitric oxide (NO) and/or increased NO biosynthesis (as measured by urine nitrogen oxide, plasma nitrogen oxide, plasma asymmetric dimethylarginine, plasma, urine and platelet cyclic GMP). Secondary hypotheses examine the association between reducing CVD risk factors, improved endothelial function, increased walking distance, improved quality of life and number of metabolic syndrome abnormalities in PAD patients. This study will contribute to the overall mission of NINR and to nursing science by providing the necessary evidence for the efficacy of a commonly utilized nursing intervention, multiple risk factor reduction, on improving physical function and quality of life in the understudied, elderly PAD patient. This application will also provide preliminary evidence for the biological basis for the efficacy of multifactor risk reduction in restoring vascular homeostasis, critical because of its role in antiatherogenesis and maintaining vasoreactivity, both necessary for slowing the progression of atherosclerosis. The relevance of this study to public health. People with PAD are at increased risk for death and disability because of severe atherosclerosis resulting from untreated CVD risk factors. This maybe due to health care systems that are not equipped to handle the problem. This study will examine the effect of a system that assists providers to reduce a patient's CVD risk factors and explores the biological reasons that this system may be effective in improving health. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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The long-term goal of my laboratory is to understand how Hedgehog (Hh) signal is transduced to control a wild variety of cellular behaviors. Hh family of secreted proteins controls many aspects of animal development. Malfunction of Hh signaling has been linked to numerous human disorders including cancers. Hh family members exert their biological influence through an evolutionarily conserved, yet poorly defined signal transduction cascade. A critical step in Hh signal transduction is the activation of Cubitus interruptus (Ci), which is regulated by large protein complexes containing the kinesin-related protein Costal2 (Cos2). We have developed a novel method for detecting Cos2 interacting proteins in vivo and have identified several novel components of Cos2/Ci complexes. In the proposed study, we will use a combination of genetic, biochemical, cellular, and pharmacological approaches to investigate the mechanisms by which Cos2 and its interacting proteins regulate Ci phosphorylation, activity, and subcellular localization. In particular, we plan to 1) investigate the mechanism by which distinct Cos2 complexes regulate Ci nuclear translocation; 2) study the role and regulation of GSK3/Cos2 interaction in Ci phosphorylation; 3) investigate the role of casein kinase I (CKI) in Hh signaling and the underlying mechanism; 4) determine the role of a novel Cos2 interacting protein, Koro, and identify and characterize additional Cos2 interacting proteins that regulate Ci activity. Knowledge gained from the proposed study shall shed light into how Hh signal is transduced to control cell growth and patterning, and may provide new avenues for diagnosis and therapeutic intervention of cancers caused by mis-regulation of Hh signaling activity.
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{
"pile_set_name": "NIH ExPorter"
}
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DESCRIPTION (provided by investigator): A major public health challenge is to better understand and predict how anthropogenic environmental changes affect vector-borne diseases. In response to rapidly expanding human populations many countries in the Middle East are developing unprecedented large-scale agricultural irrigation systems to increase food production. In Upper Egypt, there are major concerns that projects underway to transform desert environments for irrigated agriculture in the Old Nile Valley (about 70,000 acres) and in the Toshka Valley (nearly 1 million acres) will substantially increase the risks of important mosquito-borne diseases. These projects are creating new suitable habitats for vector populations and facilitating the migration and establishment of large human and animal populations. This project addresses the need to determine how land use and the large-scale transformation of desert environments for irrigated agriculture in Upper Egypt affects the risk and transmission dynamics of the well-established West Nile virus (WNV) and the epidemic potential of Rift Valley fever virus (RVFV) and malaria. There are 3 specific aims: 1) determine how land use and the transformation of desert environments for irrigated agriculture affect the ecology and behavior of mosquito species and their potential for pathogen transmission, 2) evaluate the extent of WNV in birds and RVFV in domestic livestock and assess how geographic heterogeneity in animal host diversity and relative abundance affects the potential for pathogen transmission, and 3) employ eco-epidemiological modeling approaches to assess the impact of environmental changes on the complex dynamics and risks of mosquito-borne diseases. This project represents a unique opportunity to prospectively quantify vector-host-pathogen population dynamics in the context of unprecedented large-scale environmental perturbations in desert environments. The outcomes of this project will contribute to integrated disease surveillance and control programs in Egypt as well as other areas of the Middle East.
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{
"pile_set_name": "NIH ExPorter"
}
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Fibroblast growth factors (FGFs) 19 and 21 are related hormones that have emerged as promising drug candidates for the treatment of metabolic syndrome. Pharmacologically, both FGF19 and FGF21 increase energy expenditure, cause weight loss and improve glucose tolerance, insulin sensitivity and lipid parameters in rodent models of metabolic disease. FGF21 has similar beneficial effects in obese monkeys and in patients with type 2 diabetes. However, the tissues and molecular mechanisms through which FGF19 and FGF21 exert their beneficial effects remain to be determined. In this proposal, we explore the basis for their effects on energy expenditure, weight loss, insulin sensitivity and glycemia. Based on work from our laboratory and others, we hypothesize that FGF19 and FGF21 act directly on the hypothalamus and dorsal-vagal complex in the nervous system to stimulate sympathetic outflow to brown and white adipose tissue depots; and on brown and white adipose tissue to stimulate glucose uptake, fatty acid synthesis and mobilization, and thermogenic gene expression. The net effect of this two-pronged mechanism is increased energy expenditure and weight loss and improved insulin sensitivity. In addition, we hypothesize that FGF19 acts directly on liver to stimulate glycogen synthesis and to suppress gluconeogenesis, which contributes to its beneficial glycemic effects. We propose to test this hypothesis by using diet-induced obese mice in which -Klotho, a cell surface protein that serves as an obligate co-receptor for both FGF19 and FGF21, is selectively eliminated in the nervous system, adipose tissue or liver. Treatment and evaluation of these different tissue-specific -Klotho knockout mouse models in conjunction with complementary in vitro studies with isolated white and brown adipocytes will provide important insights into the tissues and molecular mechanisms whereby FGF19 and FGF21 mediate their pharmacologic effects. We anticipate that these studies will provide important insights into FGF19 and FGF21's mechanism of action and help guide the generation of more efficacious FGF19 and FGF21-based drugs for treating metabolic disease.
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{
"pile_set_name": "NIH ExPorter"
}
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Several promising therapeutic strategies for the treatment of Spinal Muscular Atrophy (SMA) are currently being pursued. As these therapies start to move from the research laboratory to the clinic, it becomes imperative to understand how effective these therapies will be after the onset of disease and if there is a critical window for drug delivery. To address this question of timing in delivering therapies, we have designed a study to evaluate the efficacy of increasing survival motor neuron (SMN) levels throughout development using a new preclinical SMA mouse model, called the Burgheron mouse. In this proposal we describe the Burgheron model, which we generated by breeding a combination of alleles to achieve mice that have intermediate SMN2 levels. Previous attempts to engineer mild SMA models have been relatively unsuccessful, resulting in mice with near normal life spans and little to no measurable phenotype. The Burgheron mice have a noted decrease in body weight, a mean lifespan of 54 days of age and quantifiable abnormalities at the neuromuscular junction, and measure differences in muscle strength. This increased lifespan presents a clear advantage over the currently used severe models of SMA that die early at 17 days of age in that it allows us to begin testing the effects of increasing SMN at later postnatal time points, even well past weaning. By incorporating a conditional inversion allele at the Smn1 locus we can transform the Burgheron model described above into an inducible system that activates SMN production via tamoxifen induction. This allows us to better characterize the effects of introducing SMN into a disease model at various time points in disease progression. In addition, we have chosen a number of different candidate therapeutics to serve the purpose of tool compounds to explore the question of a therapeutic window for treatment.
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{
"pile_set_name": "NIH ExPorter"
}
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This proposed project will involve efforts to determine the control elements for regulation of aminoacyl-tRNA synthetase formation in both Escherichia coli and Salmonella typhimurium, and the elucidation of the exact role(s) of these enzymes in repression control of the corresponding biosynthetic operons. The principal methods of study will involve the use of genetically based and/or physiologically imposed altered repression control responses in cells cultivated under specific growth conditions, along with purification of the cellular components presumed to be critical for control and the performance of appropriate in vitro tests of the capacities of such elements to generate the repression control signals for the aminoacyl-tRNA synthetases and the cognate biosynthetic enzymes.
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{
"pile_set_name": "NIH ExPorter"
}
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The structure and immunologic reactivity of colonic mucins will be studied to determine the potential of these glycoproteins as markers of colonic tissue and colonic carcinoma in particular. Specimens of colonic tissue (normal and neoplastic will be procured and characterized by histologic type. Mucins A and B will be isolated from each tissue specimen and characterized as to amino acid and carbohydrate compostion. The structure of the carbohydrate and protein portions of these mucins will be examined. Oligosaccharides and glycopeptides will be prepared, isolated and characterized as to their composition, primary structure, and immunologic reactivity. An additional portion of each mucin will be stripped of carbohydrate, in order to obtain the protein core. Lower molecular weight peptides will be prepared and examined by peptide mapping. Antisera directed against the mucins will be prepared in rabbits and the specific antibody purified by absorption and affinity procedures. The specificity of the antisera will be analyzed by a retrospective studying of biopsy specimens from individuals with benign and malignant disorders of both colonic and extracolonic tissues. The antisera will be employed in a study of the immunohisto-chemical localization of mucins for staging and prognosis in colon cancer. A sensitive radioimmunoassay (and/or ELISA) will be developed and employed in an examination of the potential role of mucins in detection, diagnosis, staging, prognosis and management of colon cancer.
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{
"pile_set_name": "NIH ExPorter"
}
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Tool to Assess Ongoing Costs of PCMH Project Summary/Abstract The Patient-Centered Medical Home (PCMH) model is expected to help achieve the triple aims of access, quality, and cost effectiveness. Transformation of a practice to a PCMH model requires many changes, including addition of new staff to perform new services, new work flows and systems to implement new avenues for patient access, and new systems to manage the health of a population of patients. Many of these activities and the infrastructure to support them are not covered by current reimbursement models. As practices are transforming to a PCMH model, it is essential that sufficient up-front investments be made available so that practices can create the infrastructure necessary to support high-quality, safe, continuous, coordinated, comprehensive care. It is critical to develop knowledge about the costs of on-going maintenance of the new services required in a PCMH model. With this project, we have an important opportunity to contribute to effective primary care payment reform and to the sustainability of primary care transformation by rigorously defining costs for maintaining PCMH practices. This project will apply a PCMH Cost Dimensions Tool developed to assess the costs to practices associated with activities to achieve PCMH recognition status in 21 practices that differ with regard to their: 1) NCQA PCMH Recognition status (Level III or not), 2) ownership (network of centrally owned practices or independent practices), 3) payers (fee-for-service commercial plans, Medicare, Medicaid, Federally Qualified Health Centers), and 4) patient populations (socio-economic status, health status). Applying the PCMH Cost Dimension Tool in these diverse settings will allow us to examine the ways in which practices and their associated cost structures differ across settings. The findings from this study will be useful to a variety of stakeholders. They will be useful to payers in establishing financial incentives for practices to implement and operate PCMH practices. They will be useful to policy makers in establishing programs that encourage and support the development of PCMH practices. They will be useful to practices in predicting the costs they may experience in maintaining a transformed PCMH practice.
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{
"pile_set_name": "NIH ExPorter"
}
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Biostatistics and Bioinformatics Core SUMMARY/ABSTRACT The Biostatistics and Bioinformatics Core for the Endometrial Cancer SPORE program will support the Projects, and assist the other Cores, in the design and interpretation of clinical and preclinical experiments, and the acquisition and management of data. Core services will be critical for the clinical trials of the Projects, and their use of high-dimensional methodologies of gene expression profiling (GEP), next-generation sequencing (NGS) and Reverse Phase Protein Arrays (RPPAs). The Core will incorporate sound experimental design principles within each Project, specific to the scientific issues being addressed. Each Project will be provided with tailored analyses, accompanied by appropriate and sometimes novel biostatistical or bioinformatics methods. Core leaders will work with Project investigators to identify quantitative measures that can be used to test study hypotheses, including determining sample sizes to ensure sufficient power for the studies. The leaders of the Core will confer regularly with Project investigators to discuss the design and conduct of research projects, evaluate results of analyses, discuss potential new research initiatives and directions within the SPORE, and promote publication of findings. Interactions between the Core and Projects, as well as other integration, will be facilitated by a web-accessible database of relevant information generated by each Project, to be developed and maintained by the Core. The main objectives of the Biostatistics and Bioinformatics Core are to: 1. Provide biostatistics and bioinformatics expertise in the design and conduct of laboratory experiments and clinical trials arising from the research proposed in this application. 2. Provide biostatistics and bioinformatics analysis and interpretation of all data collected under the SPORE Projects, Career Enhancement Program Projects, Developmental Projects, and other Cores. 3. Collaborate and assist all project investigators with the publication of scientific results. 4. Develop and maintain a web-accessible, SPORE-specific database of relevant information for all Projects.
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{
"pile_set_name": "NIH ExPorter"
}
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Functional MRI, diffusion MRI and MR spectroscopy and spectroscopic imaging have great potential for the study and diagnosis of disease and injury, and guiding surgical therapy. All of these methods benefit greatly from the added sensitivity, resolution and contrast from high field strength magnets (3T and above). However, the advantages of higher magnetic fields have not been fully realized due to the increasingly confounding effects of magnetic field inhomogeneity (MFI) caused by magnetic susceptibility differences between air and tissue. MFI leads to signal loss and spatial distortion in MRI and loss in spectral resolution and sensitivity in MRS. The loss of reliability due to these artifacts is a major reason why these techniques have not seen wide use in clinical applications. Current methods of magnetic field homogenization (i.e. shimming) work well on small volumes but are inadequate over larger objects, like the entire human brain. Over the last decade, the MR group at Yale University has developed the technique of dynamic shim updating (DSU) which allows greatly improved magnetic field homogeneity over extended regions. DSU divides a global 3D problem into a number of slices over which adequate magnetic field homogeneity can be achieved. Dynamically updating the pre-determined slice shims in sync with the multi-slice MRI sequence ensures optimal homogeneity for all slices. Following a successful completion of the Phase I STTR grant, the current Phase II application continues the work towards commercialization of DSU. Specifically, DSU hardware will be further improved and combined with professional software to automatically set up the unit. Since DSU is significantly more complicated than regular shimming, additional software will be provided for the every day operation of the unit by standard MR users. Since MFI affects almost all facets of in vivo NMR, the successful commercialization of DSU will have major impacts on the clinical application of MRI and MRS and can thus be labeled as highly significant. PUBLIC HEALTH RELEVANCE: Magnetic resonance imaging (MRI) and spectroscopy (MRS) are the leading techniques to obtain high-quality images and metabolic profiles of intact human tissues in health and disease. Unfortunately, spatial variations in magnetic field strength introduced by the sample can severely degrade the quality of MRI and MRS data, such that distinction of normal from diseased tissues may be compromised. Current technology is not able to completely cancel the spatial magnetic field variations. The current project focuses on the development of novel technology that can significantly reduce spatial magnetic field variations, thus leading to greatly increased MRI and MRS data quality.
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{
"pile_set_name": "NIH ExPorter"
}
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Heart disease, strokes, and limb loss continue to lead mortality and morbidity. Despite advances in surgical and endovascular treatments, long-term success of these interventions remains limited. Researchers have applied a variety of approaches to modify neointimal hyperplasia and vascular remodeling in an effort to improve the outcomes;however, attempts have been largely ineffective due to incomplete understanding of the specific cause/effect links through which hemodynamic factors, biochemical mediators, and cellular effectors lead to an occlusive vascular phenotype after intervention. Prior research has focused almost exclusively on either the static isolated physical or biologic components. For instance, we confirmed that wall shear stress stands as a key regulator of vascular architecture. In parallel, we established that temporally distinct vessel wall inflammatory events (mediated by blood monocytes) predict long-term morphology. However, completely lacking is an understanding of the dynamic interplay between physical forces and cellular inflammatory elements that modulate local variations in wall remodeling, and ultimately success or failure of the intervention. Focusing on clinically relevant vein bypass grafting as a model system, this proposal will fuel the synergy of vascular biologists, engineers, and mathematicians who will employ systems biology approaches to the complex mechanisms of vascular adaptation. This established team will construct and optimize pioneering multi-scale models to understand the interplay of micro-scale physical and biologic forces in vein graft wall behavior. The proposed hypothesis driven model conceptualizes the process of vascular adaptation as two parallel, but interconnected processes. The global remodeling response is mediated through variations in the gene regulatory network (Specific Aim 1), while the focality of lesion development is modulated through the dynamics of monocyte homing to regions of altered flow (Specific Aim 2). The general approach of this proposal is the use of robust but validated computational algorithms for the development of each module. Interfaced with extensive in vivo experimental data, each module contributes a new structural integration level to the overall model. Upon compilation, the final model will utilize an iterative loop, employing a finite element computational technique (hemodynamic module) that drives a Bayesian network (gene regulatory module) and series of partial differential equations (monocyte kinetic module), with resulting outputs interfaced through a cellular automata to predict changes in tissue architecture, which feeds back into the hemodynamic module to initiate a new cycle. Structured to maintain spatial integrity, the underlying power of the predictive model is to examine the regional difference in disease progression in a complex three- dimensional geometry. This work not only provides direct clinical translation toward improved bypass graft durability, but most importantly it stands as a powerful contemporary application of rigorous systems biology approaches to understand and treat complex medical diseases. PUBLIC HEALTH RELEVANCE: Heart attack and stroke continue to be the leading cause of mortality and morbidity in the United States. Available treatments, such as vein bypass surgery or angioplasty, currently provide only short-term improvements, and are prone to failure due to aggressive scar formation. The current proposal uses systems biology approaches to study the complex mechanisms of the vascular response to injury, providing a predictive model with application both as a research tool and in the care of patients.
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{
"pile_set_name": "NIH ExPorter"
}
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The Testoderm C-95-046 Study was a pharmacokinetic (PK) study of a transdermal patch containing the male hormone, testosterone, in HIV- positive and HIV-negative males, who were hypogonadal by measurement of serum testosterone. The PK study was a companion study designed to measure the serum testosterone at baseline, and at the end of a 28 day time period during which participants wear the transdermal patch. A second part of the study measured serum testosterone after another 28 days of medication usage. The drug was given as a transdermal patch worn on the scrotum, and changed daily. The first 2 days of the study (baseline and day one of therapy) were spent as an inpatient in the GCC, during which time a careful physical examination was performed, and 12 blood samples were obtained, as part of the pharmacokinetic data collection. Patients were then instructed in use of the patches and sent home for self-administration. Patients returned to clinic on days 7, 14 and 21 for more blood tests. Patients were again admitted to the GCC on day 27 of the study, for urine and blood testing (16 blood samples). A quality of life questionnaire was filled out at both the first and the second GCC inpatient stay. Clinic evaluations, consisting of a brief evaluation and blood sampling, occurred in the outpatient clinic on days 42 and 56. On day 56, the final day of the study, a physical exam was also performed. The study's potential significance lies in the fact that testosterone, unlike other hormonal manipulations and patient-independent nutrition (e.g., TPN), adds lean body mass to HIV-infected and wasted patients, rather than fat. This is important as we become aware of the need to address the dysfunctional metabolism that studies have now shown to be common in advanced HIV-infection and AIDS. In addition, testosterone appears to contribute to an increased sense of well-being. Results of C-95-012: Nationally, 124 men, with a mean weight loss of 7% of body mass, were enrolled. The Testoderm patches effectively increased serum testosterone levels, with concomitant decrease in FSH and LH levels, as expected. Unfortunately, changes in weight or lean body mass did not reach statistical significance. Quality of life measures and measures of sexual function were also not different between placebo and treatment. There were no differences in HIV viral load between placebo and treatment groups, and no relevant differences in adverse events. Sample size calculations were done to provide 90% power to detect a difference of 2.0 kg in the change in body cell mass from baseline. Co-variate and subgroup analyses were performed for body cell mass using the variables CD4, total and free testosterone, use of protease inhibitor, and history of opportunistic infections. No effects were observed in the co-variate analysis and no differences in the subgroups were observed. Analysis of the continuation, unblinded study (C-95-013) and the PK study are underway.
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{
"pile_set_name": "NIH ExPorter"
}
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CD45 is a transmembrane receptor-type protein tyrosine phosphatase (PTP) that is highly expressed on virtually all cells of the hematopoietic lineage. Expression of CD45 is essential for antigen activation as well as for the differentiation of T and B lymphocytes. A key advantage to the study of CD45 is that the outcome of experiments modifying CD45 can be measured with a relevant functional assay - i.e., antigen activation. The investigator has recently reported the use of novel methods to identify the in vivo, naturally occurring phosphorylation sites of CD45. The hypothesis of this study is that the phosphorylation of CD45 regulates the function and the activity of CD45, and to relate CD45 phosphorylation to its function in antigen mediated activation. The investigator plans to address these goals as follows. The phosphorylation sites of CD45 in antigen activated cells will be identified by radiolabeling cells followed by isolation and analysis of CD45 by immunoprecipitation, peptide mapping, HPLC and MALDI-mass spectrometry. The role of key phosphorylation sties identified above in CD45 PTP activity will be determined by mutagenesis of phosphorylation sites followed by determination of enzyme activity and substrate specificity. Finally, the ability of phosphorylation site-mutant forms of CD45 to reconstitute the signaling responsiveness of CD45 negative cells will be determined. These studies are important to the understanding of phosphorylation events which regulate the receptivity of T cells to antigenic signals.
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{
"pile_set_name": "NIH ExPorter"
}
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There is sufficient evidence indicating that glucocorticoids, thyroid hormone, and insulin are involved in the regulation of fetal lung maturation. The understanding of their precise role in lung growth, cell differentiation and surfactant production and of the factors which modify cell sensitivity to each of these hormones is the major objective of these studies. A primary culture system of fetal lung cell populations highly enriched in type II cells, recently developed in our laboratory, will be further characterized and used to define the role of each hormone in cell growth and differentiation (appearance of lamellar bodies), phospholipid and protein metabolism, and the activity of enzymes involved in lecithin synthesis. The synergistic and antagonistic effects (e.g. cortisol-insulin antagonism) of hormones in these processes will be studied and the possible mechanisms involved will be investigated by measuring receptor levels and their hormonal regulation in vivo and in cell culture. The binding of insulin to both cell surface and intracellular receptor sites as well as the regulation of these sites will be investigated and correlated with the effects of insulin on cell growth and differentiation. Binding of agonists in the absence or presence of antagonists and biological response will be correlated to assess the physiological significance of the binding proteins (receptors). Since the effects of glucocorticoids, and possibly of thyroxine and insulin, on lung maturation are age-dependent, some of the factors determining hormone cell responsiveness will be investigated by examining the ontogenic development of receptors, and by comparing the properties of glucocorticoid receptors and their nuclear acceptor sites, the patterns of chromosomal proteins, and the effects of glucocorticoids on the synthesis or modification (acetylation, phosphorylation) of histones and nonhistones, in lungs from early and late gestation fetuses. The role of glucocorticoids in the development of glucagon receptors will be examined as a possible mechanism for the age-related action of glucocorticoids in glucagen metabolism.
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{
"pile_set_name": "NIH ExPorter"
}
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This contract provides in vitro testing of potential agents to treat parasites and their vectors, maintenance of parasite and vector stocks, and the development of related assays.
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{
"pile_set_name": "NIH ExPorter"
}
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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Primarily working on development of SWIFT, including digital receiver, ringdown compensation, SWIFT compatible multichannel coil testing at 4 T, (magnetization prepared) MP-SWIFT, reconstruction and signal processing. "SWIFT (SWeep Imaging with Fourier Transform) is an MRI sequence capable of rapid and quiet MRI imaging. The SWIFT work in Core 3 is focused on continued development and improvement. We have added the capability for magnetization preparation to give improved T1 contrast, T2 contrast, fat suppression, and water suppression. We continue to improve imaging of bone and tendon, dynamic contrast enhancement (DCE) and general short T2 imaging."
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{
"pile_set_name": "NIH ExPorter"
}
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Cerebral accumulation of amyloid beta (Abeta) protein is a pathological hallmark of Alzheimer disease (AD), and likely plays a fundamental role in its pathogenesis. Research has focused on Aa manipulation as a mechanism for reducing or treating AD, but the clinical utility of findings is limited by lack of a simple way to identify early those at high risk of AD; Abeta levels in the CSF have been measured in some settings, but are impractical for widespread clinical applications. Abeta appears in circulating plasma, and a central aim of this proposal is to investigate the value of plasma Abeta as an early indication of AD risk by exploring its relation to cognitive decline, an early stage in the path of AD development. An additional aim of this proposal is to explore the function of insulin in cognitive decline, and its interaction with Abeta; irecent research has established that insulin modulates levels of Abeta in vitro and in vivo, however, little work has examined the long-term effects of insulin on cognition, or the possibility of different effects in subgroups of the population. This proposal represents a collaboration between basic scientists and epidemologists to test novel hypotheses in men and women regarding plasma Abeta levels, insulin levels, and insulin secretion (as measured by c-peptide) in non-diabetics and their ability to predict development of cognitive decline. The investigation will be conducted within the Nurses' Health Study, including women currently aged 70- 81 years and followed since 1976, and the Physician's Health Study II including men aged 65 years and older and followed since 1982. From each cohort, we will select 2,000 subjects with stored plasma samples collected at two points over 10 years, and with serial tests of cognitive function (3 repeated interviews by telephone, at 1.5-year intervals). The test battery measures general cognitive status, verbal memory, executive function, speed of processing, and working memory; we have conducted extensive demonstrations of the validity and reliability of the telephone method compared to in-person assessments, and have previously detected strong relations with numerous established risk factors for decline. Thus, these two established populations provide a highly cost-efficient setting in which to examine biomarkers of early cognitive decline, and gender and genetic interactions.
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{
"pile_set_name": "NIH ExPorter"
}
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Inflammatory bowel diseases (IBD) are diseases of immune dysregulation. Animal models and human genetic data support a central role for unrestrained Th17 responses in the pathogenesis of at least some forms of IBD. Cytokine reporter mice that we have generated led to the discovery that Th17 cells are flexible in their late program of differentiation, such that cytokine signals received after Th17 commitment impact the stability and phenotype of mature Th17 cells so as to influence their ability to cause colitis. While it has been known for some time that IL-6 is indispensable for initiation of Th17 lineage commitment, its function late in the Th17 pathway is ill-defined. We have recently identified an unanticipated role for IL-6 in the maintenance and pathogenicity of mature Th17 cells. Unlike wildtype Th17 cells, IL-6-deficient Th17 cells are unable to sustain a pathogenic phenotype and do not induce colitis. Although early commitment to the Th17 lineage is mediated via binding of IL-6 to membrane-bound IL-6Ra (mIL-6R), which associates with gp130 for signal transduction ("classical" IL-6 signaling), developing Th17 cells rapidly cleave mIL-6R from their surface and do not re-express detectable mIL-6R. Thus, IL-6 appears to act on mature Th17 cells via trans signaling, in which gp130 is activated by binding of shed, soluble IL-6R (sIL-6R) in complex with IL-6. Accordingly, our data suggest that mature Th17 cells must both produce IL-6 and respond to IL-6 via trans signaling to induce colitis. This identifies an autocrine/paracrine loop by which IL-6 might act on Th17 cells to promote colitis. Here, we will define mechanisms by which IL-6 contributes to late Th17 development and how this impacts IBD initiation and perpetuation. We hypothesize that IL-6 produced by a subset of mature Th17 cells contributes to the maintenance of pathogenic Th17 and Th1-like cells that mediate colitis. Further, we posit that whereas classical IL-6 signaling is essential for Th17 lineage specification, trans IL-6 signaling is essential for the maintenance of mature Th17 and Th1-like effectors that develop from a common Th17 precursor;in the absence of IL-6 trans signaling, pathogenic effector T cells are not sustained and colitis is attenuated. To test this hypothesis, we have generated novel reporter knock-in and knockout models with which to identify and track IL-6 producing cells and dissect specific mechanisms by which IL-6 acts late in the Th17 pathway to regulate its potential for pathogenesis. These transgenic models are complemented by new blocking antibodies and recombinant proteins that we have acquired through collaboration. We anticipate that these studies will elucidate new features of the Th17 pathway that contribute to chronic immune-mediated disease and will facilitate the design of more rational therapeutics that target Th17-mediated disease. PUBLIC HEALTH RELEVANCE: Factors Controlling Effector T Cell Maintenance in the Pathogenesis of Colitis. Inflammatory bowel diseases are diseases of immune dysregulation mediated by CD4 T cells that have poorly restrained reactivity to components of the enteric bacterial flora. This proposal will address mechanisms by which cytokines control the maintenance and pathogenic potential of effector CD4 T that contribute to IBD pathogenesis. We expect that results from these studies will provide a basis for new interventions to curb pathogenic immunity to the intestinal microbiota in the treatment of human IBD.
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{
"pile_set_name": "NIH ExPorter"
}
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The objective of the research is to investigate those factors which regulate the reversible inactivation and reactivation of kidney sodium-potassium adenosine triphosphatase (ATPase). The data supports the hypothesis that renal ATPase is in part regulated by cyclic AMP dependent protein kinase. The current effort is to characterize the factor in the 100,000 xg cell cytosol which mediates the reacivation of ATPase inhibited by protein kinase. Recent evidence by various investigators has indicated that some commercial preparations of ATP are contaminated with vanadium. Vanadium is a potent inhibitor of ATPase. Work is in progress demonstrating that inhibition of ATPase by vanadium is a mechanism separate from the inhibition by protein kinase. The final objective of the research effort is to reconstitute the individually isolated inactivating and reactivating components of ATPase into an in vitro system with ATPase.
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{
"pile_set_name": "NIH ExPorter"
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A comprehensive program of training and research in genomic analysis is proposed for the SERCA candidate. The candidate, currently a staff research associate at Cold Spring Harbor Laboratory, comes to the genome program with a strong research background in theoretical physics. Funds are requested for a combination of training in experimental molecular biology and genetics and for basic research in computational genomic analysis. Development of new methods for detecting genes in genomic sequences and new methods for the analysis of mapping and sequencing strategies will be the focus of the research. Training has already begun with the candidate's attendance of two courses related to genomic analysis, one at Cold Spring Harbor on genomic analysis and one at Life Technologies, Inc., on laboratory methods for recombinant DNA technology. Additional beginning and advanced courses are planned. The candidate has two co-advisors, Drs. David Beach and Dr. Thomas Marr, who are actively engaged in mapping and sequencing the genome of the fission yeast at Cold Spring Harbor Laboratory. Cold Spring Harbor Laboratory is a major research institution in molecular biology with an excellent tract record in training and basic research.
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{
"pile_set_name": "NIH ExPorter"
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The glycoproteins and other envelope proteins of two animal viruses, vesicular stomatitis virus (VSV) and Rauscher murine leukemia virus (RLV), will be studied. The isolated VSV glycoprotein possesses three of the virion-associated biological activities: antigenicity, hemagglutination and the inhibition of cellular macromolecular synthesis. Host-cell modification influences the hemagglutinating ability of the glycoprotein. Biological activities associated with RLV envelope proteins include antigenicity, hemagglutination, and possibly cell fusion. Some of the cell surface antigens of tumors and transformed cells are similar to RLV antigens. In order to do quantitative biological experiments, the VSV glycoprotein will be partially degraded to soluble glycopeptides. The native glycoprotein and the biologically active glycopeptides will be used to study virus-cell interactions and the effects that they have on cellular DNA, RNA and protein synthesis. Chemical analysis of the glycoprotein including amino acid composition, peptide mapping and end group analysis will be performed in order to determine the purity of the glycoprotein which has multiple biological activities. The number, size and sugar composition of the carbohydrate moieties will also be determined. The chemical composition of the non-glycosylated membrane protein will be analyzed in a similar manner. The envelope proteins of RLV will be isolated and purified. Biological and biochemical analyses, similar to those described for the VSV envelope proteins, will be performed on the RLV envelope proteins. Since glycoproteins of viruses and cells are important in the control of intercellular interactions and may play a role in the control of cellular macromolecular synthesis, the biological and biochemical studies on the viral envelope proteins which are described in this proposal should help elucidate the mechanism of action of the glycoproteins of normal and transformed cells. BIBLIOGRAPHIC REFERENCE: McSharry, J.J., R.W. Compans, H. Lackland, and P.W. Choppin. 1975. Isolation and characterization of the non-glycosylated membrane protein and a nucleocapsid complex from the paramyxovirus SV5. Virology 67:365-374.
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{
"pile_set_name": "NIH ExPorter"
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The Office of Education delivers workshops, programs, and individualized opportunities to a population averaging 320 trainees, including postdoctoral, visiting, and research fellows; clinical fellows and medical students; graduate students; and postbaccalaureate and technical fellows, as well as summer trainees. The activities include, typically: public speaking workshops, job interviewing, writing and editorial services, grantsmanship and study section workshops, career presentations and counseling, teaching opportunities through the NICHD Becoming an Effective Scientist course for postbacs and skills workshops and a training program with the University of Maryland, and management programs. An annual retreat for fellows and graduate students is held for 75-100 people each spring to address scientific developments and careers, which includes fellow presentations and a poster presentation by each attendee. The program is developed and run by a fellow-student steering committee. Among the office's accomplishments from the past year: Dr. Yvette Pittman continues in the Office of Education as Associate Director. A former NICHD postdoctoral fellow, she developed our TmT (Three-minute Talks) competition this past year, for which three awards were made to Alex Ritter, Dr. Eva Szarek, and Dr. Monica Gupta for their video clips describing their research, targeting a broad scientific audience. She was instrumental in developing our expanded and validated database of NICHD alumni from 2008 to the present, and earlier. In June 2014, the Division of Intramural Research gave its seventh Mentor of the Year awards to Brant Weinstein, Ph.D., investigator; and Elias Leiva-Salcedo, Ph.D., fellow. Twenty-two FARE awards were made for the 2015 competition. A Fellows Intramural Grants Supplement (FIGS) continues to recognize and stimulate grant applications among fellows, and the Fellows Recruitment Incentive Award (FRIA) continues to support investigators who recruit postdocs from populations traditionally underrepresented in science. The NICHD Scholars program, in its fourth year, has two MD/PhD student members conducting their thesis research in the institute and has recruited two new postbaccalaureate trainees, Gian Rodriguez and Nicole Millan. The Scholars program focuses on developing talent and supporting trainees' academic and career progression. The NICHD Connection monthly newsletter published its 51st issue in August 2014 and reaches all members of the intramural division and our alumni.
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{
"pile_set_name": "NIH ExPorter"
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Manganese (Mn) is an essential metal but elevated levels are cytotoxic. Occupational and environmental exposure to elevated Mn lead to the development of an irreversible parkinsonian syndrome. Despite the clinical significance, mechanisms that regulate Mn homeostasis and detoxification are poorly understood and this has hindered therapeutic progress. An important advance in understanding how homeostatic control of Mn is maintained came from the recent discovery that mutations in the gene coding for SLC30A10 cause a familial form of Mn-induced Parkinsonism. The function of SLC30A10 in higher eukaryotes is ill-defined but, within the last year, we discovered that SLC30A10 was a cell surface localized Mn efflux transporter. Further, we tested several disease-causing mutants and observed that these mutants failed to traffic to the cell surface and also failed to mediate Mn efflux. Our results highlight the significance of SLC30A10 in regulating cellular Mn levels and begin to provide an explanation for the onset of Mn-induced Parkinsonism in individuals carrying mutations in SLC30A10. Based on the above results and the prior genetic studies, we hypothesize that SLC30A10 is the primary efflux transporter responsible for maintaining Mn levels and mediating Mn detoxification at the cellular and organismal level. Our hypothesis is supported by the fact that mutations in the other Mn efflux transporters do not cause Mn toxicity. Our goal is to test the above hypothesis and secure a comprehensive understanding of the regulation of Mn homeostasis and detoxification by SLC30A10 and Mn efflux. The mechanisms by which SLC30A10 mediates Mn transport and Mn regulates SLC30A10 function are unknown. In Aim 1, we will elucidate these mechanisms and, in a set of clinically important studies, also test if chemical chaperones can rescue the trafficking and function of disease-causing SLC30A10 mutants. In Aim 2, we will use a combination of assays in cell culture and genetically-modified mice to directly test our hypothesis that SLC30A10 is the primary transporter that mediates Mn detoxification. The mouse studies include assays to determine the functional consequences of altered SLC30A10 activity and Mn efflux against the neurotoxic effects of Mn in vivo and are translationally significant. Proposed studies will provide novel insights into the biology of SLC30A10 and enable us to better understand the role of this transporter in Mn homeostasis and induced Parkinsonism. Further, these studies will aid efforts to address important disease-related issues such as identifying polymorphisms in SLC30A10 that may alter the risk of Mn neurotoxicity in the general population, and also contribute to the development of innovative new treatments for the management of Mn-induced Parkinsonism.
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{
"pile_set_name": "NIH ExPorter"
}
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Our work on a delivery system for the mucin deficient tear film, and a short-term delivery device using pilocarpine for glaucoma is complete. We are now beginning work on delivery of an antibiotic gentamycin using pseudomonas corneal ulcer in the rabbit as a model. This situation requires a half-time release rate of several hours, ideally, which is quite prolonged in comparison to our other devices. We are studying the kinetics of radioactive gentamycin release from collagen membranes alone, and when complexed with other polymers. A device designed to release tetracycline over several days will then be designed using data from the gentamycin study. The second major area that we will be concerned with this year is delivery of dexamethasone, a corticosteroid in an external ophthalmic inflammative model. Here prolonged delivery is most desirable and a device similar to the gentamycin model will be used. We are clinically testing pilocarpine/collagen membranes and collagen membranes alone in human subjects. BIBLIOGRAPHIC REFERENCE: Bloomfield, S., Dunn, M.W., Rubin, A.L., and Stenzel, K.H. Soluble Artificial Tear Inserts. (Abstract) A.S.A.I.O., April 1976.
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{
"pile_set_name": "NIH ExPorter"
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During the current year, 42 temporal bones have been obtained from 22 interesting cases. During the same period, 42 previously obtained temporal bones (29 cases) have been stained. Already prepared temporal bones - cases of sudden deafness, crista neglecta, Reissner's membrane with blood vessels, and leukemia with sudden hearing loss - have been thoroughly investigated and the reports on these cases are either in print or in press in the Annals of Otology, Rhinology and Laryngology. During the current year, additional cases have also been investigated in depth, and among these the significant findings were as follows: 1. Congenital heart anomaly (20 bones): Significant anomalies such as dehiscence of the facial canal, remnant of mesenchymal tissue, high jugular bulb, persistence of stapedial artery, poor development of horizontal canal, and absence of cochlear aqueduct were observed. 2. Leukemia with low-frequency sensorineural hearing loss (1 bone): The case showed hemorrhage and degeneration of the cochlear nerve in the osseous spiral lamina of the upper middle turn of the cochlea. 3. Branch of the singular nerve (223 bones): The branch was observed near the internal auditory canal in 194 cases (87 percent), and near the posterior canal ampulla in 58 bones (26 percent). BIBLIOGRAPHIC REFERENCE: Sando, I., Harada, T., Loehr, A., and Sobel, J.H.: Sudden Deafness with Pathological Correlations. Temporal Bone Histopathology Report. Ann. Otol. Rhinol. Laryngol. 86:269-279, 1977.
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{
"pile_set_name": "NIH ExPorter"
}
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Obesity and diabetes lead to high circulating levels of insulin and high blood pressure (BP). However, mechanisms underlying these associations are in dire need of clarification. For example, in the kidney, insulin, through its own receptor, can lead to sodium retention, and activate the epithelial sodium channel (ENaC). Nonetheless, insulin can also increase the production of nitric oxide (NO), which has been shown to reduce the activity of ENaC. Thus, these two actions of insulin would be expected to have opposing effects on BP. To better elucidate the role of the insulin receptor in the kidney, we have developed an transgenic mouse with "knockout" (KO) or deletion of the insulin receptor (IR) in the distal tubule of the kidney (IRKO). These mice survive and grow normally, but have significantly higher basal BP. They are also impaired in their ability to rapidly excrete a NaCl load, as well as, have blunted rise in urinary nitrates plus nitrites (a urinary form of NO) excretion in the basal state and after insulin treatment. Thus our hypothesis is that IR in the kidney may have a previously unappreciated role in facilitating volume excretion and in the maintenance of normal BP. We further suggest these deficiencies arise directly as the result of impaired NO production with subsequent over-activity of ENaC. Three main aims are outlined below. Aim 1 is to determine whether reduced renal NOS activity in the distal tubule is a mechanistic determinant of the sodium-excretory defect and elevated BP in the IRKO mice. Renal nitric oxide synthase (NOS) activity will be measured in inner medulla, outer medulla, and cortex, as well as, microdissected proximal tubule, thick ascending limb, and collecting duct from fed and fasted IRKO and WT mice. We will also test the direct effects of insulin on NOS activity and NO levels in inner medullary collecting duct (IMCD) cultures. Finally, we will test whether molsidomine, an NO donor, restores NO levels, as well as normalizes BP and sodium excretion in IRKO mice. Aim 2 is to determine whether increased activity of the epithelial sodium channel (ENaC) is a mechanistic determinant of the sodium-excretory defect and elevated BP in the IRKO mice. For this aim, we will test whether benzamil, an antagonist of ENaC, abolishes differences in BP and natriuresis. ENaC regulation will be examined in native tissue and in primary IMCD cells. Finally, in Aim 3 we determine whether distinct IR signaling relating to ENaC activation and NO generation is altered in the IRKO mouse IMCD. Phosphorylation of critical proteins involved in IR signal transmission from the receptor to the activation of ENaC and nitric oxide synthase will be evaluated. These proteins include, but are not limited to, the insulin receptor substrate (IRS), phosphoinositide-3-kinase (PI-3K), and the serum and glucocorticoid-regulated kinase (SGK1). Overall, these studies will highly elucidate the role of insulin in the the distal tubule with regard to its role in blood control and sodium regulation. Moreover, they may provide insight into the pathology underlying hypertension associated with the metabolic syndrome. PUBLIC HEALTH RELEVANCE: Insulin resistance is associated with high circulating plasma levels of insulin, as well as, increased blood pressure;however, cellular mechanisms relating the two are not well understood. The kidney, a major regulator of blood pressure, expresses insulin receptors along the renal tubule;however, their role there is undefined. These studies are aimed at elucidating insulin's role in the distal tubule, which will hopefully allow for earlier and better targeted treatments in obesity and diabetes.
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{
"pile_set_name": "NIH ExPorter"
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Ninety-seven per cent of right-handed (RH) individuals develop speech and language processing in the left (dominant) hemisphere of the brain, while the "minor" hemisphere controls emotional behavior. We have speculated that a hypothetical gene, <I>RGHT</i>, functions to specify left-brain dominance resulting in developing the right-handed preference. We have been collecting blood samples and cheek swabs from specific families with the aim to genetically map the hypothesized <I>RGHT</i> gene. We have collected enough samples and the next stage is to use these samples for mapping by the sib-pair method. We have published our random recessive model in which individuals with the nonfunctional recessive allele on both homologs have a 50:50 chance of either being RH or LH. We also found interesting association of hand use preference with the clockwise vs. counterclockwise scalp hair-whorl rotation. We suggest that individuals with the right gene are RH and develop clockwise hair whorls, but individuals with the recessive allele are 50:50 in hair whorl orientation. We have previously proposed that human brain and visceral organs laterality might be controlled by the Somatic Strand-specific Imprinting and selective chromatid/strand Segregation (SSIS) model. It predicted a chromosome-specific, nonrandom WW:CC segregation phenomenon where both older "Watson" strand-containing chromatids form a homologous pair of chromosomes are delivered in mitosis to one daughter cell and the older "Crick" strand-containing chromatids are delivered to the other daughter cell. This was proposed as a mechanism for cellular differentiation to produce non-equivalent daughter cells in mitosis. We have now established the existance of such biased strand segregation phenomenon in mice cells concerning chromosome 7 segregation. Interestingly th esegregation pattern is cell type regulated so that there is directed segregatio i soem cell types but random in others. In future exaperiments we want to find the mechanism of biased strand segregation of this chromosome and also we will determine whether other chromosomes are likewise subject to cell type regulated segregation process.
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{
"pile_set_name": "NIH ExPorter"
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We propose to assess biochemical markers of dyslipidemia and endothelial dysfunction, and omega-3 fatty acids in relation to risk of CVD among men and women diagnosed with type 2 diabetes in two large ongoing cohort studies, the Nurses? Health Study (NHS) and Health Professionals? Follow-up Study (HPFS). By 1998, 12,600 confirmed type 2 diabetic cases have already accumulated in the two cohorts. By the year 2002, 5,507 blood samples prospectively collected from persons with previously or newly diagnosed type 2 diabetes will be available for analyses. Using this unparalleled resource, we will specifically evaluate (1) The relationship between plasma levels of cell adhesion molecules (i.e. sICAM-1, sVCAM-1, E-selectin), diabetic dyslipidemia, and risk of CVD among diabetics; (2) the association between Lp(a) concentrations and risk of CVD among diabetics, independent of high triglycerides and low HDL; (3) the association between long-term intakes of omega-3 fatty acids and CVD risk in diabetes. The main NHS and HPFS grants will provide follow-up and documentation of CVD in addition to covariate information. Overall, the large size of these cohorts, the prospective design, the high follow-up rates, and the availability of archived blood specimens provide a unique opportunity to study the relationship between diabetic dyslipidemia and risk of CVD in an extremely cost-efficient and timely manner. In addition, these two cohorts provide an unusual opportunity to compare lipid profiles and endothelial markers of CVD between diabetic men and women. We believe that this study will provide new insight into the understanding of the atherosclerotic process in diabetes and may suggest new nutritional and pharmacological means to prevent and treat CVD complications among diabetics.
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{
"pile_set_name": "NIH ExPorter"
}
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The Nevada Cancer Research Foundation (NCRF) was established in 1983 to conduct cancer research under the National Cancer Institute's (NCI), Community Clinical Oncology Program (CCOP) grant. The NCRF main office is located in central Las Vegas with satellite offices in Reno and Carson City, Nevada. It is Our Mission to provide high quality, state-of-the-art, cancer treatment research and cancer control and prevention research in Nevada through the participation of the local physicians and health care providers at their facilities.' It is also our intent to provide high quality, timely data to the NCI in a research environment that sets the highest standards for the safety and welfare of our participants. The long term NCRF CCOP grant objectives have been: 1) to accrue a minimum of 50 participants annually to CCOP approved cancer treatment clinical trials in Nevada; 2) to accrue a minimum of 50 participants to CCOP approved cancer control and prevention studies in Nevada; 3) to provide to the Research Bases timely data of high quality. 4) to promote quality and state-of-the-art treatment in the community through participation in clinical trials by: (a) involving CCOP physicians and staff in scientific and educational activities of the Research Bases; (b) expanding the knowledge, awareness, and involvement of the primary healthcare providers and specialists in the Research Base activities for the development and use of cancer prevention and control research; 5) to support other professional education programs and cancer prevention and control outreach services to under-served rural areas; 6) to identify women and minority groups that may be underserved and to involve them in clinical trials; and 7) to continue to work with the NCI to develop a community cancer network in order to facilitate the growth, development, and implementation of important new cancer initiatives. To accomplish these goals, the NCRF CCOP makes the program available to all interested professionals from varied medical specialties for state-wide participation thereby providing access for participation to the citizens of Nevada. Essential to our mission is the service we provide to our participants. We encourage involvement by closely following all aspects of their health care, protecting their welfare, and by providing the latest information and resources available. We are here specifically to provide a service to Nevada residents and generally to provide a service to all who are afflicted with this disease. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Human-associated microorganisms (the ?microbiome?) are present in numbers exceeding the quantities of human cells by at least 10-fold, and the collective genome (the ?metagenome?) exceeds our human genome in terms of gene content by more than 150-fold. We and others have recently demonstrated that bacteria are detected in the placenta using a variety of culturable and non-culturable approaches. For several years we have developed and employed metagenomics to characterize the placental microbiome, and observed variation in its community membership and their function by virtue of gestational age at delivery. However, it remains unknown what the different maternal source(s) are of the placental microbiome, and whether it sinks to the fetus via the intra-amniotic cavity and thus is measureable in amniotic fluid. In response to the Human Placental Project, we propose to identify the sources and sinks for and of the placental microbiome employing two large, robust and well characterized existing datasets. We will generate unparalleled metagenomics and metabolomics data, in order to test our central hypothesis that the placenta is populated by commensal microbiota which largely arise from the maternal oral and GI communities, with a lesser contribution from the vagina. We further hypothesize that placental microbes populate the fetus and the intrauterine environment, and are detectable in mid-trimester amniotic fluid. Moreover, their metabolites serve as lasting signatures of the microbiotas functional presence. In order to prove this hypothesis, we will execute three essential aims in a total of over 1230 subjects samples from two existing data sets. The net result of the completion of these aims will be to first identify the maternal source of the placental microbiota, and validate these findings using state of the art single molecule fluorescent in situ hybridization and culturation. We will thereafter recapitulate these findings in early and mid- second trimester amniotic fluid and thus identify the early evidence of the placental microbiota sink. In a final aim, we will use LC/MS full spectral metabolomics on these same subjects samples to identify the stable and lasting metabolic footprint of the microbiome. We present our proof of concept work on intrahepatic cholestasis of pregnancy as evidence for the feasibility, significance and ready translational application of our approach. As a proven team of perinatal physician scientists with an emphasis and history of being at the forefront of big data (and notably metagenomics) science, we are uniquely poised to now undertake complex integration of these unique data sets in studies which are feasible, justifiable, and of likely long-term significance and high impact.
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{
"pile_set_name": "NIH ExPorter"
}
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Excessive stimulation of glutamate receptors of the NMDA sub-type result in the activation of nitric oxide synthase (NOS), the generation of nitric oxide (NO), and neuronal cell death. The apoptotic signaling pathway by which NO exerts its neurotoxic effects remains poorly understood. Events such as protein nitrosylation, mitochondrial dysfunction and activation of stress-activated p38 mitogen activated protein (MAP) kinase have been proposed to act as downstream effectors of NO-induced neurotoxicity. Affected neurons are thought to die by apoptosis, a form of cell death that involves the activation of cell death proteases, known as caspases. However, caspase inhibition often only delays neuronal cell death. Thus cell death determining events, upstream of casr ase activation, are likely to contribute to the commitment to cell death. Cell shrinkage is a universal event of all apoptotic cell death and involves the efflux of intracellular K+ ions. The molecular mechanism that drives K+ efflux during apoptosis is unclear. The purpose of this project will be to explore the possibility whether activation of outward voltage-gated K+ channels and subsequent cell shrinkage and mitochondrial injury via a pathway mediated by free Zn+ may constitute early events that commit neurons irreversibly to NO-induced neurotoxicity. To pursue these goals, primary cerebrocortical neurons will be analyzed using approaches such as time-lapse deconvolution microscopy, whole cell patch-clamp recording, transient transfections, biochemistry, and cell-free systems of apoptosis with isolated mitochondria. Among the specific questions that will be addressed in this project are: (1) Does NO provoke K+ efflux, enhancement of voltage-gated K+ channels, and apoptotic cell shrinkage? (2) Does stress-activated p38 MAP kinase regulate the activity of voltage-gated potassium channels and cell shrinkage? (3) Does NO provoke Zn2+ release from metallothionein (MT) which in turn results in mitochondrial damage, generation of reactive oxygen species, and p38 MAP kinase phosphorylation? [unreadable] [unreadable] Because NO plays an important role in a wide range of neurodegenerative diseases including stroke, Parkinson's disease, Alzheimer's disease, multiple sclerosis, epilepsy, and AIDS dementia, results obtained from this project could provide broad implications for the development of new therapeutic drugs to mitigate or even prevent neuronal cell loss during neurodegeneration.
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{
"pile_set_name": "NIH ExPorter"
}
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Von Willebrand Disease (vWD) type 1 is the most common form of vWD, accounting for as much as 80% of reported cases. With prevalence estimates as high as 1.6% in the general population, upwards of 4.5 million Americans may be affected. Unfortunately vWD type 1 is also the most difficult type to diagnose, leading to greater patient distress and increased cost. The goal of the proposed study is to identify and characterize common polymorphisms within the von Willebrand factor (vWF) gene that account for a sufficient portion of variation in vWF levels to be clinically useful in the diagnosis and management of von Willebrand disease (VWD) type I. Successful execution of this project will advance the understanding of vWD by providing a strong foundation to direct future studies, and an experimental paradigm that can be used to accelerate the dissection of complex traits. The project is designed also to advance this young investigator's knowledge and skills as a nurse scientist and genetics researcher. Moreover, the methodology can be applied to any gene locus in the search for genetic causes of vWD. This research is relevant to public health, vWD type 1 is the most common bleeding disorder. In addition there has been a continued call for a more accurate method of diagnosis, as conventional methods have not proved very useful in the assessment of this particular bleeding disorder. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Functional Photosystem II reaction centers were isolated from the thermophilic cyanobacterium Phormidium laminosum and reconstituted in dimyristoyl-phosphatidyl-choline (')MPC) vesicles. Samples with the maximum protein/lipid ratio were examined in the STEM, both as freeze-dried and negatively stained specimens. The freeze-dried samples showed a number of folded structures (perhaps due to the vesicles breaking up) and because of this, there was little order in the stained samples. Further experiments to preserve the structures are planned.
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{
"pile_set_name": "NIH ExPorter"
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Although a thesis topic has yet to be selected, some areas of interest for the completion of a Ph.D. include: 1) Elucidation of the signaling pathways that lead to lymphocyte activation 2) Understanding of the roles and functions of dendritic cells in antigen presentation, induction of tolerance and immunological memory. 3) Characterization of the human anti-porcine immune response with the goal of aiding the development of xenotransplantation. The selection of these areas of research is based both on my particular interest and on the possible contribution of these studies on the treatment and prevention of disease.
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{
"pile_set_name": "NIH ExPorter"
}
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The long term goal is to understand the molecular mechanism of hepatocarcinogenesis so that novel strategies for the effective prevention and treatment of liver cancer can be developed. Epigenetic abnormity is a major hallmark in hepatocellular carcinogenesis. The histone methyltransferase enhancer of zeste homolog 2 (EZH2) plays a key role in embryonic development via catalyzing trimethylation of histone H3 at lysine-27 and facilitating DNA methylation. EZH2 is overexpressed in human liver cancer, and knockdown of EZH2 causes apoptosis of liver cancer cells and tumor regression in nude mice. However, little is known about the in vivo importance of EZH2 in liver pathophysiology and carcinogenesis, largely due to the embryonic lethality of EZH2-null mice. Additionally, activation of the oncogene c-Myc causes liver cancer in mice, and c-Myc overexpression is associated with malignant transformation of hepatocytes in humans;however, the underlying mechanism of c-Myc-induced hepatocarcinogenesis remains unclear. The objective of this proposal is to elucidate the pathophysiological importance of EZH2 in liver utilizing novel models of mice with liver-specific knockout (LKO) of EZH2 and/or overexpression of c-Myc. We found that EZH2-LKO mice appeared normal, suggesting that loss of EZH2 in liver is well-tolerated. EZH2 is essential for the regeneration of pancreatic beta cells. We found that EZH2 was markedly induced during liver regeneration, suggesting its importance for the regeneration of liver. The central hypothesis is that EZH2 is important for liver regeneration and is required for c-Myc-induced hepatocarcinogenesis through inhibiting c-Myc-induced cell senescence/apoptosis and promoting c-Myc-induced cell proliferation. Knockout of EZH2 will cause a significant delay of liver regeneration and blockage of c-Myc-induced hepatocarcinogenesis. This central hypothesis will be tested in two specific aims. Aim 1 will determine the importance of EZH2 in liver regeneration using models of 2/3 partial hepatectomy in EZH2-LKO mice. The working hypothesis is that EZH2-LKO mice will have considerable delay in liver regeneration but will eventually regenerate the liver due to compensatory mechanism(s). Aim 2 will determine the importance of EZH2 in c-Myc-induced liver carcinogenesis using mice with liver-specific knockout of EZH2 but overexpression of c-Myc. The working hypothesis is that EZH2 deficiency will prevent c-Myc-induced liver cancer via inhibiting cell proliferation but enhancing cell apoptosis/senescence. Results from this study will provide crucial novel knowledge about the importance of EZH2 in liver regeneration and liver carcinogenesis, which may ultimately help to identify EZH2 as a novel target for the prevention and treatment of liver cancer, a deadly disease with increasing incidence in the USA. Moreover, understanding the role of EZH2 in liver regeneration will help to determine how EZH2 inhibitors may influence liver injury-repair and the safety of chemotherapy when EZH2 inhibitors are combined with other anticancer drugs to treat diverse malignancies. PUBLIC HEALTH RELEVANCE: The United States has increasing incidence of liver cancer, a deadly disease that lacks effective treatment. The proposed studies are of importance in the under-investigated area of epigenetic control of liver regeneration and liver carcinogenesis by a novel histone methyltransferase EZH2. Thus, the findings are expected to enhance our understanding of liver diseases and liver cancer, and this knowledge can be used to develop new drug target to prevent and treat liver cancer.
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{
"pile_set_name": "NIH ExPorter"
}
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We propose to create a "Center for GEI Association Studies" in support of the NIH's Genes by Environment Initiative (GEI) at the Broad Institute, within the Genetic Analysis Platform. Our aim is to enable whole genome association scans to succeed by supplying high quality large-scale genotyping in an environment that has proven ability in industrial process, commitment to scientific rigor and a depth of analytic expertise. Specifically, according to the stated goals of the RFA, we will genotype at least 7200 samples in year one, and at least 12,000 in total over the first three years of the award. We will receive and qualify DNA samples from external investigators. Once qualified we will use the Affymetrix whole genome genotyping platform to carry out genotyping of 1M SNPs on a single array. We will also make available three other platforms for genotyping (Illumina Infinium, Illumina Golden Gate and Sequenom MassArray) as options for whole genome studies, or critically, to be used in rapid validation and follow up of putative associations. We will apply rigorous QC metrics and filters to supply "clean" data sets to users. Finally, we will work with the NCBI to deposit data according to the NIH's guidelines on data sharing in a timely and efficient manner. Given the fast pace of development of genetic analysis tools, we will continue to improve upon existing products in terms of content and quality and will drive forward the adoption of new technologies as they become available.
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{
"pile_set_name": "NIH ExPorter"
}
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The University Group Diabetes Program (UGDP) was undertaken for the purpose of evaluating the effect of hypoglycemic treatment on the rate of development of vascular complications in mild adult-onset diabetics. It is a multi-clinic study involving 12 Clinical Centers and a Coordinating Center. Patient recruitment in this study started in February 1961 and was completed in early 1966 after a total of 1,027 patients had been enrolled. Patients were randomly assigned to one of the following treatment groups; insulin given in fixed dose, insulin given in variable dose to maintain blood glucose levels within a normal range, tolbutamide, phenformin or placebo. All patients were placed on diets designed to achieve or to maintain normal body weight. Information obtained from the study is used to evaluate the efficacy of treatments under study in preventing or delaying the onset and development of vascular complications in mild diabetics and to provide detailed information on the natural history of diabetes. This application contains the justification for support for the 12 participating Clinical Centers for one additional year for the purpose of completing data collection under the original study protocol, providing for mortality follow-up and enabling the participation of the clinical investigators in the preparation of first reports.
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{
"pile_set_name": "NIH ExPorter"
}
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Project Abstract Degenerative cervical spondylotic myelopathy (CSM) is the most common cause of spinal cord injury (SCI) representing a significant public health problem. A major shortcoming limiting efforts to improve the treatment of patients with CSM is the lack of quantifiable metrics on which to base clinical decisions. Advanced MRI techniques, such as diffusion tensor imaging (DTI) have shown promise in this area. DTI measures the magnitude, anisotropy, and directionality of water displacement in tissue and provides quantifiable measures of directional diffusivity along white matter tracts. While DTI provides a valuable tool to assess white matter integrity, unfortunately, we have found current DTI techniques are flawed because diffusion properties derived using DTI lose specificity and sensitivity with increasing pathological and anatomical complexity. Thus the prediction of long-term outcome using DTI remains uncertain. To overcome factors confounding DTI analysis, we developed diffusion basis spectrum imaging (DBSI), to more accurately delineate white matter injury, allowing differentiation and quantification of axonal injury/loss, demyelination, and inflammation in the setting of spinal cord compression. DBSI quantifies edema/tissue loss in addition to axon/myelin injury, providing improved imaging biomarkers that more accurately predict a patient's clinical course, response to therapy, and long-term prognosis. The long-term objective of this proposal is to establish and validate non-invasive imaging biomarkers that are predictors of clinical course and therapeutic response to surgical decompression in patients with CSM. The first aim will assess whether spinal cord DBSI pathological metrics reflect neurological impairments and predict long-term neurologic outcomes following decompressive spinal surgery in patients with CSM. This aim will test the hypothesis that clinical manifestations of spinal cord compression in mild CSM are predominantly a reflection of edema and inflammation, with a lower incidence of true axonal loss; In contrast the high variability of functional recovery observed in moderate CSM patients is attributable to a greater risk for permanent axonal loss caused by spinal cord compression. The second aim of this proposal, will refine DBSI modeling for assessing effects of blood flow deficits on spinal cord pathology and improving the accuracy of axonal loss quantification in CSM. This aim will test the hypothesis that the effect of spinal cord blood flow on CSM pathology may be assessed by including Intra-Voxel-Incoherent-Motion (IVIM) in DBSI modeling; the accuracy of DBSI-derived axon volume may be improved by including intra-axonal diffusion component in DBSI modeling. The identification and validation of such non-invasive DBSI biomarkers will provide guidance on clinical management, long-term prognosis, and family counseling. The validation of a non-invasive biomarker for predicting functional recovery in the surgical management of cervical myelopathy would represent a new and substantial advance in the treatment of cervical myelopathy.
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{
"pile_set_name": "NIH ExPorter"
}
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A DMH-induced rat colon cancer is being explored as a model of human colorectal cancer. Studies are designed to determine the immunogenicity of the tumor and examine tumor-host manipulations which lead to enhanced resistance to this neoplasm.
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{
"pile_set_name": "NIH ExPorter"
}
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Varicella zoster virus (VZV) causes chickenpox and shingles. Virus infection of cells is known to trigger several signaling pathways that are important for cell proliferation and prevention of programmed cell death (also known as apoptosis). We have used a virus genome wide approach to identify functions of VZV proteins. Nearly all VZV genes were individually expressed in tissue culture cells along with a reporter system that responds to specific signaling proteins. Screening the VZV genome, we found that VZV ORF12 protein activated two protein kinases (ERK1/2 and p38). VZV deleted for ORF12 was impaired for activation of the kinases. ORF12 protein was shown to be located in the virus tegument (between the virus envelope and the nucleocapsid). We also found that ORF12 inhibits programmed cell death (apoptosis) induced by chemical treatment of the cells. These experiments indicate that ORF12 protein can activate signaling pathways and inhibit programmed cell death.
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{
"pile_set_name": "NIH ExPorter"
}
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The importance of post-translational modifications (PTMs) in regulating the global proteome of the eukaryotic cell cannot be underestimated. Ubiquitin (Ub) and Ub-like proteins (Nedd8, SUMO, ISG15, etc.) play a particularly key role for the host. Depending on the topology of the conjugation, these proteins stabilize members of signaling pathways as well as target proteins for destruction by the proteasome. Removal of Ub by host deubiquitinating enzymes (DUBs) are used by eukaryotes to ensure proper regulation of signaling cascades. Modification of these regulatory pathways on the protein-protein level by bacteria represents a key pathogenic target for promoting infection. ChlaDUB1 and ChlaDUB2 are two virulence factors produced by Chlamydia tra- chomatis which act as deubiquitinating and deneddylating enzymes. As Chlamydia does not possess the cellular machinery for ubiquitination of their proteins, it is anticipated that ChlaDUB1 and ChlaDUB2 target the host proteome directly. Indeed, the presence of functional DUBs in the C. trachomatis genome suggest an important role in virulence. Therefore, it is predicted that ChlaDUB1 and ChlaDUB2 directly disrupt the regulation of host immune pathways at the protein-protein level. In order to test this hypothesis, two simultaneous approaches will be used. First, a series of biochemical in vitro assays will be used to dissect the nature of the ChlaDUB1 and ChlaDUB2 substrates and to identify the precise host targets of ChlaDUB1 and ChlaDUB2. Both enzymes are known to sharply attenuate the NF-kB pathway. Experiments are designed to identify the precise Ub-tagged NF-kB pathway members that are targeted by ChlaDUB1 and ChlaDUB2. Second, recognizing that ChlaDUB1 and ChlaDUB2 may target host proteins beyond those represented in the NF-kB pathway, a global screen for targets is required. As a transcriptional analysis is unable to identify changes on the critical level of post-translational modifications, an innovative global protein stability (GPS) assay will be used. An additional advantage of this approach is that can applied back into eukaryotic systems to advance the ubiquitin field as a whole. ! !
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{
"pile_set_name": "NIH ExPorter"
}
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Naive CD8+ T lymphocytes lack effector function and undergo a differentiation process to exit the naive state and to acquire a primed effector phenotype. Preliminary results indicate that this transition is a complex and highly regulated process, involving receptor-mediated signal transduction, cell cycle progression, chromatin remodeling, and changes in gene expression. Evidence suggests that the naive state may be actively maintained, raising the possibility of a genetic program to place a brake on differentiation. In contrast, primed T-cells gain effector function, as well as altered signal transduction pathways and a more dynamic actin cytoskeleton, suggesting that new genes become expressed to mediate these processes. The overall goal of this application is to identify the molecular phenotype of naive and primed effector CD8+ T-cells, and to study the functional role of identified molecules in normal T-cell populations. The principal model will utilize 2C TCR transgenic mice crossed to a Rag2 background, to extinguish endogenous TCR gene rearrangement and ensure a naive phenotype. Mechanistic experiments in normal, post-thymic, T-cells will be enabled by a novel Coxsackie and adenovirus receptor (CAR) transgenic mouse and adenoviral transduction. The first specific aim is to identify genes and proteins that are differentially expressed between naive and primed effector CD8+ T-cells, by: I) examination of candidates by Western blotting, ii) analysis of expression of defined genes using Affymetrix microarrays, and iii) identification of unknown genes by subtractive library cloning. The second specific aim is to optimize the CAR transgenic mouse system for transduction of normal, post-thymic T-cells. Transgenic mice expressing CAR have been constructed using an LckICD2 promoter/enhancer cassette. Optimization of adenoviral transduction will be performed, and interbreeding with 2C/Rag2-/- mice will be carried out for studies of naive and primed CD8+ T-cells. The third specific aim is to analyze the function of differentially expressed signaling molecules and transcription factors linked to MAP kinase pathways in normal T-cells using CAR transgenic mice. Preliminary results have shown that several proteins involved in MAP kinase signaling become markedly up-regulated upon T-cell priming. Wildtype and mutant Ras, FosB, MKK7, JNK1, JNK2, and other candidates will be introduced into naive and primed CAR transgenic T-cells. Effects on early T-cell differentiation, as well as on apoptosis, TCR sensitivity, CD28 independence, and magnitude of cytokine production, will be examined. The fourth specific aim is to investigate the functional requirement for modulators of actin cytoskeleton dynamics in normal T-cells. Preliminary results have revealed that expression of several molecules associated with cytoskeletal dynamics is increased tremendously upon differentiation from the naive to the primed effector state. Wildtype and mutant a-Pak, Raci, and other candidates will be introduced into primed effector T-cells, and potential effects on TCR capping, conjugate formation, cell spreading, cytolysis, and cytokine production will be investigated. In addition, gelsolin4 -/- mice will be examined for functional defects in the T-cell compartment. The results of these experiments will provide invaluable information regarding the molecular and biochemical changes that accompany T-cell differentiation, pointing the way to new potential targets for manipulating immune responses in vivo.
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{
"pile_set_name": "NIH ExPorter"
}
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Adenoviruses are being used to study regulation of the sequential reactions in viral replication and the effects on their host cells. These viruses are particularly useful for this objective since they can establish productive or abortive infections, depending upon the host cell, leading to either cessation of cell division or transformation of the cells to unlimited growth. Conditionally lethal, temperature-sensitive (ts) mutants are proving to be of particular value for investigating these control mechanisms. Special use will be made of mutants unable to initiate DNA replication at the non-permissive temperature owing to a defective DNA-binding protein (e.g., H5ts125) for studying control of DNA replication, transcription, and transformation. New techniques will be employed to isolate additional mutants in early gene functions, and ts mutants of type 7 adenovirus will be isolated and characterized.
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{
"pile_set_name": "NIH ExPorter"
}
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The purpose of this research is to develop an efficient day-to-day resource allocation system for regional referral networks of special care centers and referring hospitals. This will be accomplished by: (1) development of alternative special care center assignment and transport dispatch procedures to yield more efficient utilization of special care resources and better availability of modern clinical technology to all patients, especially those in rural areas; 2. Evaluation of the dispatch procedures in a simulation model; 3. Incorporation of the successful procedures in an interactive computer decision support system and its application to an existing referral network; and 4. Evaluation of the cost effectiveness of the interactive system as compared to manual systems currently in use. It is expected that the results of this research will serve as a model for the development of special care referral networks as well as for the improvement of existing daily resource allocation decisions in established networks. Results can also serve as inputs to long-term planning for regional networks.
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{
"pile_set_name": "NIH ExPorter"
}
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Research has consistently identified parental monitoring, the extent to which a parent tracks the activities and location of their youth, as an important risk factor for early substance use, delinquency and risky sexual behavior (Crouter & Head, 2002). The literature suggests that parental monitoring may be part of a transactional process between parents and youth and may reflect parent actions such as supervision (e.g., an adult is present to observe youth activities), parental solicitation (e.g., parent attempts to ask youth for information) and child disclosure (e.g., youth decisions to share information with their parents). Measurement issues have made it difficult to identify which specific parent and youth actions are protective, how parent and youth monitoring behaviors work together to influence youth risk, how monitoring strategies change over adolescence, and how parental monitoring and youth disclosure are related to other protective parenting practices (Stattin, Kerr, & Tilton-Weaver, in press). The proposed study aims to further our understanding of the protective process that underlies the association between parental monitoring and risky behavior during middle school. Using latent class analysis, this project will identify unique profiles of monitoring and explore how these profiles are associated with the prevention of early adolescent substance use, delinquency, and deviant peer associations. Specific aims are to (1) Identify latent classes of families who have unique monitoring profiles, explore if these classes are the same across gender of the child, and investigate how youth risky behavior and other aspects of parenting predict membership in these latent classes at the entrance to middle school and (2) Model the stability and change of membership in monitoring latent classes over the middle school period and how youth risky behavior and the affective quality of the parent-child relationship may predict these transitions. The data for the research are from the PROSPER project, a NIDA funded large-scale effectiveness trial of substance use prevention in small towns and rural communities in two states. Data will include measures of monitoring activities reported by mothers and youth from Grade 6 to Grade 9. The monitoring process during middle school may be particularly important for prevention, as early initiation of risky behavior has been linked to later severe antisocial behavior and adult substance abuse (Grant & Dawson, 1997; Dewit et al., 2000; Nagin & Tremblay, 2001). Identifying the specific combinations of monitoring activities that prevent early adolescent substance use may help the field refine the targets of change in family based prevention programs, thus enhancing the efficacy of such programs.
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{
"pile_set_name": "NIH ExPorter"
}
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Infection with an enteric parasite can act as an adjuvant to prime for a Th2 biased response to a typically tolerogenic form of dietary antigen. However, in agreement with recent clinical and epidemiological studies, we have found that helminth infection protects against the anaphylactic symptoms and antigen specific IgE induced in a model of food allergy. Helminth dependent protection against allergy was abrogated when helminth infected, allergen sensitized mice were treated with neutralizing antibodies to IL-10. The unexpected identification of Th2 responses without atopy has led to the suggestion that helminth infection induces a specialized subset of dendritic cells that drive the generation of immunoregulatory T cells. Our data provide the basis for an ideal experimental model in which to test this hypothesis, hi Aim 1 we will examine whether helminth-induced protection against allergy is attributable to IL-10 secreting T cells. Helminth infection may also influence allergen presentation directly, via its effects on antigen presenting cells. This possibility is explored in Aim 2. The impaired responsiveness to oral vaccines observed in helminth-infected individuals may also be a consequence of helminth-induced immunosuppression. Characterization of the mechanisms by which infection influences the response to oral vaccines, as proposed in Aim 3 has practical implications for maximizing vaccine efficacy in the developing world. The induction of immunoregulatory mediators by chronic enteric infection is not likely to be restricted to helminth infection but may be a feature common to all types of infection that serves to maximize protective immunity while minimizing pathology.
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{
"pile_set_name": "NIH ExPorter"
}
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The Gordon Research Conference on Cilia, Mucus, and Mucociliary Interactions, held from February 27 to March 4, 2005, at the Rancho Santa Barbara Marriott in Buellton, California, will provide a unique opportunity for scientists involved in ciliary, mucous and mucociliary research to present their latest progress and to exchange ideas, thus bringing together a very diverse group of scientists who usually do not meet. Abnormal cilia and mucus contribute to a variety of human diseases including diseases of the airways, abnormal left-right asymmetry, and polycystic kidney disease to name a few. Many of these diseases have no effective or only limited treatment options. Thus, there is a clear need for a small meeting focusing on these topics. As a Gordon Research Conference, the meeting will follow the highly successful model of such conference layouts and will offer 9 plenary sessions and additional poster sessions. Three of the plenary sessions will be devoted to cilia, one to primary ciliary dyskinesia, three to mucus and mucins, and two to mucociliary interactions. The organizing committee has invited an international and diverse group of many young but also established investigators to present their work during plenary sessions. Discussions have been recognized to be vital to the success of the conference and ample time for these will be allowed between speakers during plenary sessions, facilitated by discussion leaders. Finally, poster sessions will complement the plenary sessions and free afternoons will further enable idea exchanges between scientists.
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{
"pile_set_name": "NIH ExPorter"
}
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This proposal is aimed at fully characterizing the extent to which retrograde transport and/or endocytosis are involved in a myogenic model of Spinal Bulbar Muscular Atrophy (SBMA). SBMA is a X-linked lower motoneuron disease characterized by adult onset muscle weakness, atrophy, and significant depletion of motneurons. It has been widely accepted that muscle weakness and atrophy result from loss of innervation from motoneurons. Our group has unexpectedly engineered a transgenic mouse model for SBMA which shows disease symptoms in androgen-dependent fashion by selectively over-expressing the wild-type AR gene exclusively in muscle tissue, challenging two tenets of the current dogma about this disease: 1) that SBMA originates in motoneurons, and 2) that an expansion of CAG repeats exceeding 40 within the AR protein is required to trigger SBMA. Our model of SBMA shows many of the hallmark pathological features of this disease, including androgen-dependent loss of motor function accompanied by marked muscle atrophy and motoneuron death. Most recently we have discovered that our transgenic mice showing SBMA disease symptoms also show marked deficits in retrograde labeling of motoneurons using cholera toxin horseradish peroxidase (CT-HRP). The experiments proposed within this fellowship are designed with the intent of identifying the extent to which a deficit in retrograde transport is responsible for the pathogenesis in our muscle model of SBMA. We have designed ours experiments to evaluate the extent to which potential changes in axonal transport and synaptic morphology correlate with changes in motor function. This information should tell us whether changes in retrograde transport represent a potential cause or is simply a byproduct of the disease, these results will be important for guiding future hypotheses regarding the disease. Our model, which over-expresses wt AR protein only in skeletal muscle fibers provides an unique method to investigate the contribution of muscle to the SBMA phenotype. Hence, we have also proposed to investigate the usefulness of known SBMA therapeutics on our muscle model of SBMA. Successful therapeutic intervention for our model of SBMA would provide compelling evidence that targeting skeletal muscle fibers is likely to yield effective therapeutic agents to treat SBMA. This proposal carries profound implications for other polyglutamine and neuromuscluar diseases in that it suggests a muscular component to these diseases. Such implications ask, are these diseases more muscular or neurological in origin? Answering such questions could have profound impact on how neuromuscular disease should be treated in the future.
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{
"pile_set_name": "NIH ExPorter"
}
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The users assembled in this application have a common interest in using cryo transmission electron microscopy (cryoTEM) of vitreous ice- embedded samples as a quantitative tool to investigate the assembly, regulation and function of biologically important macromolecular complexes. Despite continuous progress in x-ray crystallographic and NMR methods, it is still difficult using these techniques alone to obtain atomic level structural information about the many complex macromolecular assemblies that govern fundamental cell biological processes. It is now clear that a powerful approach is to combine structural information obtained at different levels. The structural information obtained from cryoTEM and computer based 3-D reconstruction spans the gap between our ability to obtain structural information about large macromolecular assemblies using conventional light and electron microscopy, and high-resolution data obtained from NMR or X-ray crystallography. This data can be combined to build up a detailed picture of the overall architecture of the complexes and the interactions between the components. This grant is based on projects in which such a complimentary approach is used to analyze complex biological structure. The major users in this application will use the requested cryo-TEM to probe file mechanisms of actin binding protein interactions (Almo, Condeelis), bacterial flagellar motor switching (Khan) and virus entry/exit (Kielian) at the molecular level. At the level of ultrastructure, the developmental program of C. elegans will be elucidated by examination of its fine morphology as revealed in cryo-sections (Hall).
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{
"pile_set_name": "NIH ExPorter"
}
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The purpose of this study is to determine whether Ethyol can protect against the myelosuppression and neuropathy associated with Taxol therapy. This is a phase I dose escalation study of Taxol with Ethyol and G-SF in patients with advanced cancer. Patients will undergo extensive neurologic testing before and after Taxol therapy.
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{
"pile_set_name": "NIH ExPorter"
}
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Peripheral nerve injuries represent a major public health problem, seen frequently in patients admitted to level I trauma centers and in as many as 30% of individuals with traumatic brain injuries. A primary goal of intervention in this arena is t hasten recovery so as to restore function as rapidly as possible. Progress towards this goal, however, has been limited. Interestingly, we recently discovered that systemic erythropoietin (EPO) administration speeds functional recovery after experimental crush injury to the sciatic nerve, even when EPO is administered one week after injury. These findings have already led to clinical studies being initiated overseas. Understanding how EPO induces this beneficial effect would enable us to design conditions and dosing strategies that can optimize its therapeutic effects; this is the goal of th3e studies proposed in this application. In the first Specific Aim, we will address the question of whether local delivery of EPO, rather than systemic delivery, will enable us to further enhance recovery and thus set the stage for the development of innovative local delivery methods to enhance/induce repair. The pleiotropic multi-organ effects of EPO, coupled with the surgical and medical focus for local treatment for local traumatic injury argues for a local application of this potent pharmacologic bio-modulator. This, in effect, focuses the effect of EPO directly at the site of injury, eliminating secondary effects and enabling the optimization of treatment in a clinically relevant and targeted manner. In the second Specific Aim, we will test the hypothesis that at least part of the mechanism of EPO action derives from beneficial effects on Schwann cells, the cells that support the function of peripheral nerves by myelinating the axons that run through these nerves. Overall, the identification of target cell(s) of EPO action will better enable us to determine how benefit occurs. In this Specific Aim, we will also begin teasing apart the underlying signaling that supports the impact of EPO, providing a basis for envisioning adjuvant therapies that focus on manipulating the key signaling players in the EPO receptor pathway. The above Specific Aims will represent the research training component of the proposed program to support the transition of Dr. John Elfar MD, a clinician/surgeon who is expanding his academic role to the clinician scientist track. The proposed research training plan is positioned on the backdrop of a robust mentoring and didactic training program that collectively represents a multi- faceted training environment aimed at ensuring successful career transition for Dr. Elfar.
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{
"pile_set_name": "NIH ExPorter"
}
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The B cell response to antigen is regulated by a variety of co-receptors that convey information to the B cell about the quality of the antigen and the status of the ongoing immune response. Our progress using high resolution live cell imaging to delineate the very early antigen driven events in B cell activation has provided a new context in which the impact of coreceptors can be evaluated. Using high resolution fluorescence resonance energy transfer (FRET) coupled with total internal reflection microscopy (TIRFM) and single molecule tracking we provided evidence for an ordered process that occurs within seconds to minutes of the BCR binding antigen. Antigen bound BCRs form immobile clusters that then grow in size by molecular trapping. The clusters perturb the local lipid environment causing lipid rafts to coalesce around the BCR clusters. As a consequence of the membrane perturbation the first kinase in the pathway, Lyn, that is tethered to the membrane by raft lipids is brought into close molecular proximity to the BCR clusters. Simultaneously, Lyn phosphorylates the Ig alpha beta cytoplasmic domain of the BCR and the Ig alpha beta chains undergo a conformational change from a closed to an open form. Syk is recruited to the phosphorylated BCR and the signaling cascades are triggered. We learned that the FcgammaRIIB is excluded from the growing BCR cluster and that CD19 is recruited to the cluster in a ligand independent fashion. During the coming year we will pursue a new approach to better understand how the CD19/CD21 complex and the FcgammaRIIB regulate B cell responses. We will use super-resolution imaging to describe, at the 10-50 nm level, the spatial relationship between the BCR, CD19 and FcgammaRIIB in resting human B cells and in B cells in which the BCR is ligated alone by Ag or FcgammaRIIB is ligated by immune complexes. We are also characterizing the spatial distribution of these receptors on naive human IgM expressing naive B cells and in IgG-expressing memory B cells. These studies should provide a detailed spatial map of the distribution of these receptors on B cell surfaces and insight into the mechanisms underlying their function.
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{
"pile_set_name": "NIH ExPorter"
}
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The complex bacterial masses which develop on and adhere firmly to teeth, collectively known as dental plaque, provide most of the causes of dental caries and periodontal diseases. In the adherence of plaque, bacterial polysaccharides are important but we are only beginning to understand the variety of molecules, their nature and their interactions which are responsible for this phenomenon. A thorough understanding of these molecular interactions should lead to innovations for the removal of plaque or the suppression of its development, and thereby should contribute importantly to the control of dental caries and periodontal diseases. Among the important interactions are numerous examples of specific coaggregation between bacteria of different species, e.g., Streptococcus sanguis and Actinomyces viscosus. S. sanguis is a very common, and often the most abundant bacterium identified in the early colonization of freshly cleaned teeth in man. Actinomyces, which are implicated in root caries and periodontal diseases, increase in number rapidly during the first several days of plaque formation. The coaggregation between these two may be important in the build-up of the Actinomyces population. The objective of this proposal is to provide an understanding of the molecular basis of coaggregation between S. sanguis 34 and A. viscosus T-14. Toward this goal we have: (1) Established conditions under which the coaggregation is reproducible and can be studied quantitatively, e.g., pH, specific ionic requirements, bacterial culture conditions, etc.; (2) Determined that the coaggregation requires the interaction between a protein on A. viscosus T14V with probably a carbohydrate on S. sanguis 34 and that this interaction is inhibited by lactose and certain other beta-galactosides specifically; (3) Obtained a crude, water-soluble, preparation from S. sanguis 34 which inhibits the coaggregation and, in this regard, is more potent than lactose. Specific aims of this proposal now are to purify and characterize the molecules from S. sanguis 34 which inhibit the coaggregation and to determine specific structural features which are essential for the inhibition.
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{
"pile_set_name": "NIH ExPorter"
}
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The overall goal of the Center of Biomedical Research Excellence (COBRE) in Neuroscience at the University of Vermont (UVM), established in 2001, has been to integrate and expand neuroscience research and training at UVM by building a collaborative intellectual infrastructure, developing cutting-edge shared core facilities, supporting recruitment of new junior faculty, and by providing research project and pilot project funding for neuroscience faculty in multiple colleges. The Neuroscience COBRE expanded interdisciplinary neuroscience research and promoted translational research by increasing the interaction between basic and clinical neuroscientists. COBRE funding has been instrumental in developing an excellent research infrastructure that has allowed the neuroscience community to compete successfully for new institutional funds and extramural support. As the neuroscience faculty enterprise has grown, neuroscience has been designated an area of research and training emphasis at UVM. The primary objective of the next grant cycle is to build upon our success and continue to support the infrastructure of neuroscience research and training at UVM. The goals are: Goal 1: To support the intellectual environment and research opportunities of neuroscientists at UVM by (1) maintaining a vigorous mentoring program to promote success of pilot project grant directors and newly recruited junior neuroscience investigators, (2) supporting a University-wide Neuroscience Seminar Series and Annual Retreat, (3) supporting Pilot Project grants, (4) facilitating the growth and productivity of the university-wide Neuroscience Graduate Program and (5) facilitating dialogue between basic and clinical scientists that promotes development of collaborative and translational research. Goal 2: To support the infrastructure for neuroscience research and training at UVM by fostering the continued growth of two Neuroscience COBRE multi-user research cores: an Imaging/Physiology Core and Cell/Molecular Biology Core. In particular, the Neuroscience COBRE grant will ensure continued growth in sophistication of equipment and research training offered through the Imaging/Physiology Core and Cell/Molecular Biology Core.
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{
"pile_set_name": "NIH ExPorter"
}
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Prevalence of smoking among opioid-dependent individuals is four-fold that of the general US adult population and is associated with increased risk for smoking-related morbidity and mortality. The 2009 passage of the Family Smoking Prevention and Tobacco Control Act gave the FDA regulatory junsdiction over tobacco products, and a public policy mandating a reduction in cigarette nicotine content is currently being considered. Such a policy could dramatically reduce smoking rates and smoking-related adverse health effects in the general population. Unfortunately, little is known scientifically about the effects of reduced-nicotine cigarettes in populations that are especially vulnerable to smoking and adverse health outcomes, including smokers with comorbid other drug dependence. Whether this more dependent group might respond with compensatory increases in smoking rate or inhalation patterns, potentially increasing exposure and adverse health effects, is unknown. The overarching objective of this project is to conduct a thorough experimental evaluation ofthe abuse liability and health effects of very low nicotine content (VLNC) cigarettes in opioid-dependent smokers. We will compare cigarettes varying in nicotine content across a range of doses starting from levels approximating those in usual brand cigarettes to very low nicotine levels (0.83, 0.28, 0.10, 0.03mg) using brief- and extended-exposure protocols. This project will represent the first investigation of VLNC cigarettes in smokers with opioid dependence or other substance use disorders and stands to contribute new scientific knowledge with the potential to inform FDA policy decisions. As Primary Aims, we will evaluate whether VLNC substitute for usual-nicotine cigarettes during bnef exposure, without producing compensatory increases in smoking, and we will compare VLNC and usual-nicotine cigarettes, under conditions of extended (12-week) exposure, on smoking rates, toxin exposure levels, and nicotine dependence. As Secondary Aims, we will assess adherence, withdrawal, cigarette demand, biomarkers of exposure to carcinogens, markers of pulmonary, cardiovascular and neurocognitive function and use of other non-prescribed drugs. Understanding how opioid-dependent smokers respond to reduced-nicotine cigarettes is essential for evaluating the potential impact of a nicotine reduction policy. We propose to provide the first experimental analysis of the effects of VLNC cigarettes in opioid-dependent smokers using measures that are relevant to the abuse liability and potential health impact of these products. Taken together, these data have the potential to directly inform FDA policv decisions regarding reduced-nicotine tobacco products. RELEVANCE (See instructions): Prevalence of smoking in opioid-dependent individuals is four-fold that of the U.S. adult population and associated with significant smoking-related morbidity and mortality. This project will investigate the acute and extended effects of reduced-nicotine cigarettes on smoking rate, toxicants, health effects, cognitive functioning and other measures in smokers receiving methadone or buprenorphine treatment for opioid dependence. This information will help to determine whether a public health policy of reducing the nicotine content of cigarettes would be effective and safe forthis challenging population of vulnerable smokers.
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{
"pile_set_name": "NIH ExPorter"
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An investigation of the relationship between dietary antioxidants, lipid peroxidation and free radical events associated with the metabolism of carcinogenic compounds which promote lipid peroxidation in isolated liver cells is proposed. The tumorigenic activity of such carcinogens is inhibited by antioxidants and a study of the type described in this application may reveal an antioxidant-inhibited, free radical reaction which is obligatory for tumor formation. Current information indicates that the antioxidant content of diets relative to the polyunsaturated fatty acid content may have an important influence on tumor incidence. These two dietary components are antagonistic with respect to the occurrence of lipid peroxidation in tissues. Our current studies have shown that lipid peroxidation results in the formation of singlet oxygen (King, M.M., Lai, E.K., and McCay, P.B., J. Biol. Chem. 250: 6496, 1975) which could contribute to the activation of carcinogenic compounds which promote lipid peroxidation. We will employ spin-trapping compounds in these studies to observe free radical events during lipid peroxidation in isolated liver cells and subcellular fractions thereof. Spectra obtained from electron spin resonance (ESR) signals should permit analysis of the nature of the radical producing the signals. Preliminary studies have already indicated the feasibility of this approach. The function of the selenoenzyme, glutathione peroxidase, with respect to its influence on lipid peroxidation in membranes, is also included. Our current findings indicate that the enzyme does not operate to reduce lipid peroxidases in membranes as currently accepted. In view of the correlation between the selenium status of animals and tumor incidence, a study of the mechanism by which this enzyme inhibits lipid peroxidation in biological membranes is also proposed.
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{
"pile_set_name": "NIH ExPorter"
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PROJECT SUMMARY/ABSTRACT Transcriptional regulation of gene expression plays a critical role in numerous cellular processes. Epigenomics refers to the study of global patterns and dynamic changes of protein molecules and biochemical factors that interact with genomic DNA to affect the chromatin architecture and to regulate gene expression. Epigenomics bridges the mechanistic gaps between genetic variations and cellular phenotypes. Identification of functional epigenomics and transcriptional regulatory relations is essential for understanding fundamental gene regulatory mechanisms. High-throughput genomic approaches have been increasingly applied in the field and a large amount of multi-level genomics data have been generated to characterize molecular profiles of different cell types in various systems. One major challenge in such genomics studies is unbiased model-based computational analysis and integration of these high-dimensional multi-omics data from different platforms to retrieve functional insights. The research program of my lab focuses on developing quantitative models and computational methods for functional multi-omics data analysis. We have developed several computational models and bioinformatics methods for ChIP-seq data analysis and predictive models for functional transcriptional regulation by integrating publicly available multi-omics data. Our long-term vision is that by using novel computational methodologies with adapted cross-disciplinary approaches from statistics, physics, mathematics and computer science, we will be able to understand fundamental mechanisms of gene regulation in human cells and their role in many diseases. Specifically, in the next five years, my lab will mainly focus on the following objectives: (1) Developing accurate predictive models for functional transcriptional regulatory relations and networks with smart integration of multi-omics data. (2) Developing statistical models for unbiased quantification and analysis of chromatin accessibility sequencing (ATAC-seq and DNase-seq) data. (3) Developing computational methods for joint analysis for integrating cross-scale bulk and single-cell multi-omics data to study functional regulatory dynamics in a single-cell level. In the meantime, we collaborate with a few experimental labs and apply our developed computational methods for studying functional epigenomics and transcriptional regulation in a variety of mammalian cell systems. We commit to make all methods and algorithms that we develop into open-source bioinformatics software tools, APIs, and web-based resources that are accessible and useful to the biomedical research community.
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{
"pile_set_name": "NIH ExPorter"
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For the families of nursing homes residents, remaining constructively involved in the care of the resident is a challenge. Family access to clinical and administrative information is essential to effectively participate with care, yet it is usually not available, or is inadequate. Channels of communication between families and facility caregivers are poorly defined. The proposed Phase Il study will complete a curriculum of clinical, administrative, and legal education for families of nursing home residents in an interactive video format accessible through the Web. It will include a "family orientation" for newly admitted residents, and in addition, a facility-specific "Family Communication Room" will enable families and staff to communicate directly on an ongoing basis. A 12-month randomized trial with four facilities will assess family satisfaction, involvement and complaints as well as resident quality of care and quality of life. The Phase I study successfully completed all stated objectives. Approximately 45-minutes of interactive-video family education on dementia was created in conjunction with family advisors. Eighteen family members who completed the program showed enhanced knowledge on pre-post tests (t = 5.9, p<0.0001) and reported high levels of satisfaction. The Phase I family advisors were essential to the development of this Phase II proposal. Building on the successful commercialization of a previous product for nursing home staff education, there is a well-defined commercialization plan. Discussions have begun to market this product with a major insurance underwriter servicing the Iong-term care industry.
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{
"pile_set_name": "NIH ExPorter"
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The Tissue Core will collect tissue from subjects in Project 1 and distribute it to Projects 2 and 3. Specifically: Brains of Project 1 rats (Dr. Roberts) that self-administer cocaine under the DT4 schedule, with and without exposure to candidate medications, will be hemisected and distributed to Project 2 for analysis of immediate early gene expression (Dr. Porrino) and tissue neurotransmitter content (Dr. Jones). Brains of Project 1 rats (Dr. Roberts) that self-administer cocaine under the progressive-ratio schedule, with and without exposure to candidate medications, will be hemisected and distributed to Project 3 for analysis of receptor binding and G-protein receptor function (Dr. Childers) and signal transduction systems (Dr. Howlett). Brains of monkeys that self-administer cocaine under the progressive-ratio schedule in Project 1 (Dr. Nader) will be hemisected and distributed as follows: [unreadable] one hemisphere will be distributed to Project 2 for in vitro receptor autoradiography (Dr. Porrino) and tissue neurotransmitter content (Dr. Jones). [unreadable] the other hemisphere will be distributed to Project 3 for identification of candidate genes and proteins via laser capture dissection (Dr. Hemby). In later years of the Center, these experiments will include synaptosomal preparations. In addition to distributing tissue from Project 1, the Tissue Core will treat separate groups of rats with drugs found to demonstrate efficacy in reducing self-administration rats in Project 1. Brains from these rats will be distributed to Project 3 for assessment of G-protein receptor function (Dr. Childers) and changes in signal transduction systems (Dr. Howlett). In addition to brain tissue, cerebrospinal fluid and venous blood will be periodically collected from nonhuman primates in Project 1 and stored for future use by investigators within and outside the Center. Several additional tissues will be collected at necropsy, including the pituitary gland, heart, myocardium, pancreatic tail, Kver lobe and adrenal glands. This list can be expanded based on future research questions and collaborations within and outside the Center.
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{
"pile_set_name": "NIH ExPorter"
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75% of all breast tumors in women express estrogen receptor alpha (ER?+). Although anti-estrogens have markedly reduced mortality for many women with this luminal tumor subtype, one quarter of these patients eventually succumb to anti-estrogen-resistant cancer. Tumor initiating cells, or cancer stem cells have been implicated in resistance to conventional therapies, recurrence and metastasis. However, the origin of luminal tumors and the characteristics of the cells that defy treatment remain unknown. Mouse models of ER?+ breast cancer that would facilitate these studies are very rare. Multiple studies point to a close link between prolactin (PRL), aand the pathogenesis and therapeutic responsiveness of ER?+ breast cancer. Our in vivo model, the NRL-PRL transgenic mouse, permits dissection of the dynamic processes that lead to diverse carcinomas with transcript profiles that resemble the luminal tumor subtype in women, including ER? expression. In this new application, we will employ this model to investigate the hypothesis that PRL cooperates with ovarian steroids to modulate mammary epithelial progenitor and tumor cell subpopulations, promoting the development of luminal carcinomas. Further, these tumors contain tumor initiating cells, which display hormonal dependencies and therapeutic susceptibilities distinct from the bulk of tumor cells. In Aim 1, we will establish the effect of PRL and crosstalk with ovarian steroids on mammary epithelial subpopulations in nonparous females prior to lesions, as defined by surface markers and functional progenitor assays, and identifies key mediators of these processes. In Aim 2, we will determine characteristics of tumor initiating cells in PRL-induced ER?+ carcinomas, and ascertain their dependence on PRL and ovarian steroids and susceptibility to adjuvant treatment. In Aim 3, we will elucidate interplay between PRL and a well-characterized oncogenic signal, ?-catenin, in regulation of normal mammary subpopulations and tumor subtype by examining the net effect of crosstalk on epithelial subpopulations prior to lesions, in early lesions, and carcinomas. These studies will illuminate the mechanism(s) whereby PRL interacts with endogenous steroids and oncogenes to promote the luminal breast cancer subtype, and its role in treatment sensitivity. They will test the cancer stem cell hypothesis in PRL-induced ER?+ carcinomas, generating a new and biologically relevant paradigm for understanding this prevalent tumor subtype, enabling the design of novel therapeutic approaches.
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{
"pile_set_name": "NIH ExPorter"
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RNA interference (RNAi) is an evolutionarily conserved process in eukaryotic organisms that serves as a regulatory mechanism to target specific RNAs for degradation. A hallmark of RNAi is the incorporation of short interfering RNAs (siRNAs) into RISC (RNA-induced silencing complex). RISC or related complexes are also thought to be involved as defense systems against virus infections, but direct biochemical proof remains to be provided for any organism. In the Preliminary Studies section, evidence is presented for the isolation of an in vitro active Tomato bushy stunt virus (TBSV)-activated siRNA-protein complex with bona fide RISC properties and with novel features not previously reported that provide an explanation for several biological observations associated with infection. The newly developed system and the findings provide a unique opportunity to study the properties of antiviral RISC in detail to reveal principles that may apply to RISC of other systems and thus can potentially yield information to design improved strategies for RNAi-based therapies. This fits the mission of the agency to pursue "fundamental knowledge about the nature and behavior of living systems and the application of that knowledge." The long-term goal is to apply TBSV as a model system for studying the biochemical properties of RISC for future comparison of antiviral RISC-like complexes in other organisms. Within this scope, the objective of this RO3 application is to characterize the anti-TBSV RISC in detail using techniques specifically adapted for this purpose in the laboratory, and to compare the properties with RISC induced by other plant viruses. The underlying hypothesis of the application is that antiviral RISCs not only share many properties but also have biochemical attributes that are virus- specific. To test this, the specific aims of this application are to: 1) Characterize RISC from TBSV infected plants, and 2) Characterize RISC isolated from plants infected with viruses other than TBSV. Relevance for Public Health. First, a better understanding of how organisms defend themselves against certain viruses will be beneficial towards our understanding and implementation of antiviral therapies. Second, since virus infections produce high amounts of small RNAs (i.e. siRNAs); they represent useful model systems towards improving new strategies for therapeutic administration of high dosages of siRNAs. [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Endochondral ossification is a pivotal event in skeletal development and in adult skeletal regeneration. In both, osteoclasts and chondroclasts erode hypertrophic cartilage, hypertrophic chondrocyte die, blood vessels grow in followed by recruitment of osteoprogenitor cells and bone and bone marrow cavity formation. These events require extracellular proteolysis and are essential for normal bone formation and repair. The goal of this proposal is to elucidate the molecular and cellular mechanisms, controlled by matrix metalloproteinases (MMPs), that coordinate the events in endochondral bone formation and regeneration. In the previous funding period we showed that proteolysis by MMP-9 is a critical determinant of the events in endochondral bone formation. MMP-9 and vascular endothelial growth factor (VEGF) act in overlapping pathways to regulate the program of hypertrophic cartilage remodeling, angiogenesis and bone formation in growth plate development and skeletal regeneration. We now propose to use genetic, molecular and cell biological approaches to further characterize how these two MMPs, each separately and together, regulate the program of endochondral bone development. We will use targeted and conditional mutations in mice to evaluate the relative roles various cell types expressing MMP-9 and MMP-13 (collagenase-3) to cartilage remodeling, osteoblast and osteoclast function, vascular recruitment and skeletal remodeling, using morphometric and biochemical approaches coupled with cell and organotypic cultures. From these studies we will be able to determine the steps in skeletal growth plate morphogenesis. We will focus on terminal differentiation of hypertrophic chondrocytes, extracellular matrix remodeling, regulation of vascular recruitment and bone remodeling. We will then use the MMP mutant mice and unstable and stabilized fractures as tools to evaluate how the mechanical environment determines the regenerative response during in skeletal regeneration in wild type and mutant mice. We will then build on these analyses to determine the physiologic substrates of the MMPs during development and skeletal repair. These studies will give us insights into novel and important functions for the MMPs, extracellular matrix proteins and angiogenic regulators in endochondral bone formation. The endochondral ossification processes are misregulated in osteoarthritis and recalcitrant hard tissue lesions. A fuller understanding of the central role of angiogenesis and its regulation by proteolysis in these processes may have implications for therapeutic interventions and the development of therapy for certain types of non-healing fractures. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Our goal is to better understand the biologic basis for lung cancer metastasis, which could potentially have a tremendous public health impact because lung cancer is the primary cause of cancer-related death in Western countries, and metastasis is the primary cause of death from lung cancer. On the basis of our previous reports, we believe that the extracellular signals that repress microRNA-200 (miR-200) levels in lung cancer cells are key drivers of metastasis and are targets for metastasis prevention. Here we show that metastasis-prone tumor cells from mice that develop lung adenocarcinomas owing to expression of mutant K-ras and p53 express both Notch and Notch ligands, and the Notch ligand jagged2 promotes tumor sphere formation, EMT, invasion, and metastasis. Jagged2 increases the expression of all six GATA transcription factor family members, and GATA3 promotes tumor cell EMT and metastasis by suppressing miR-200 levels. We posit that each of the GATA factors regulated by jagged2 plays a distinct transcriptional role and collectively mediate the diverse biologic effects of jagged2 in this tumor model. This hypothesis is innovative because it links the Notch axis to a microRNA that is central to EMT and metastasis, and it opens a new area of investigation into the role of GATA factors as promoters, rather than repressors, of metastasis. The experimental approaches we have proposed are innovative; the mouse model is based on one we created that closely recapitulates biologic and transcriptional features of human lung adenocarcinoma, and the cell micropatterning techniques we have developed are at the forefront of the interface between bioengineering and cell biology. We propose two Specific Aims. The first Aim is to determine whether inactivation of Jagged2 abrogates metastasis in a mouse model of lung adenocarcinoma. For this Aim, we will create mice that conditionally inactivate jagged2 and express oncogenic K-ras and p53 in the lung; we will examine whether jagged2/Notch-dependent interactions between tumor cells are required for polarized sphere formation and EMT using cell micro-patterning techniques we have developed; and we will determine whether high jagged2 levels correlate with low microRNA-200 family member expression in human lung adenocarcinoma biopsy samples and whether this expression pattern correlates with disease recurrence and short duration of survival. The second Aim is to examine whether the GATA transcription factor family members up-regulated by jagged2 are required for tumor cell polarization, sensitivity to TGF2-induced EMT, and metastasis. For this Aim, we will use genetic approaches to deplete GATA factors individually from lung adenocarcinoma cells; we will study the biologic and transcriptional properties of those cells in culture; and we will examine their tumorigenicity and metastatic potential in an orthotopic lung tumor model we have developed. Our long-term goal is to develop novel approaches to identify those patients who are at high risk for recurrence following treatment of early-stage disease and to use pharmacologic approaches to target key mediators of metastasis in those patients.
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{
"pile_set_name": "NIH ExPorter"
}
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This renewal application requests support to continue a long-term study of the genetics of hypertension in populations of West African origin. Defining the genetic architecture of hypertension remains a central challenge for cardiovascular (CV) epidemiology. To date, however, no widely replicated loci have been identified for hypertension. The genome-wide association design has proven to be effective for other complex traits. We propose to build on our population samples in West Africa, the Caribbean and the US and to create a large collaborative network that will allow a comprehensive search for genetic underpinnings of this disorder. A multi-stage design will allow discovery and replication in West African populations, and further replication in Western Hemisphere groups in contrasting environments. We will use the Affymetrix 6.0 array to conduct a genome-wide association study of blood pressure/hypertension in 2,000 Nigerians of Yoruban ethnicity to identify markers/regions of interest (data on 1,250 samples are being collected in the current grant; support is requested for 750 additional samples). Informative markers will then be typed in replication populations from West Africa (N = 4,000), as well as Brazil and the Caribbean (N = 7,500). A sub-set of the replication samples will be used to assess gene-environment interactions by comparing low vs. high risk factor status for BMI and salt intake. We will then examine consistency of the findings in large US studies of African Americans with similar data. The data will be made available for public access. Our study will also benefit from the extensive genetic data already available on the Yoruba population through the HapMap project. This project builds on many years of experience studying hypertension in Afro- origin populations. These populations have a highly informative genetic structure and genetic effects are likely to be strongest in West Africa where risk factor exposure is low. DNA samples and extensive phenotype information have already been collected on the 18,000 study participants. The design makes it possible to conduct the initial search in a population with high levels of internal genetic diversity without recent admixture and a wide range of measured BP. Access to a broad range of replication samples further makes it possible to determine the generalizability of these findings and potential gene- environment effects.
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{
"pile_set_name": "NIH ExPorter"
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New genetic-based methods are being developed to map neuronal interconnectivity. These approaches promise to be extremely powerful since they combine the specificity of cell type-specific promoters with the ability of neurotropic viruses to spread trans-synaptically. However, in their current configuration, these methods rely on transgenic mice to selectively drive the expression of either the tracer or the activator. Several laboratories, including ours, have accumulated experience with the rat or with other mammals for which transgenic technologies are currently cumbersome or unavailable. For this reason, in Specific Aim 1 of the present R21 application we propose a novel method for demonstrating retrograde trans-synaptic labeling of pathways in the mammalian brain, which can be freely applied to different species without the need to generate transgenic animals. Neurons will be directly visualized with a virus vector conditionally expressing green fluorescent protein (GFP), thus providing an extremely high signal to noise ratio and facilitating detection of multisynaptic pathways projecting upon the target. The proposed method utilizes two separate virus vectors delivered in two interconnected regions. The anatomical selectivity with the proposed method is based on the specific connections of the neurons of interest. Subsequent developments could also include the use of cell type-specific promoters for added specificity. In Specific Aim 2, we propose to explore the use of recombinant cholera toxin B subunit (CTb) as a viral vector-encoded tracer to expand the toolbox of genetic tracers. CTb is a reliable, sensitive and robust tracer for both anterograde and retrograde studies. Of particular note is the ability of CTb to trace collateral axons derived from the parent retrogradely labeled somata. Very exciting recent studies have also demonstrated the use of wheat germ agglutinin (WGA) as a genetically-encoded tracer. Recombinant CTb could also be used concomitantly with WGA to simultaneously label two pathways. The results of the present R21 will contribute to the development and refinement of strategies for the regulated delivery of recombinant tracers and, in follow-up studies, other reporters such as functional biosensors.
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{
"pile_set_name": "NIH ExPorter"
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This competitive renewal of the comprehensive Research Training Program in Bone Biology and Disease at DAB provides the rigorous interdisciplinary education required for the development of independent research scientists capable of innovative bone-related research. Since its establishment four years ago, one predoctoral trainee, who is now a postdoctoral fellow in implant biology and materials research, and five post-doctoral students, including an academic clinician, have completed the program;four of whom are pursuing bone biology research, three in academics and one in industry. The Program leverages the resources and research base of in the UAB-Center for Metabolic Bone Disease (Director: J. McDonald, MD), one of five P30-funded bone programs in the nation. The 33 preceptors of the Training Program have been selected based on their complementary research expertise in clinical and basic research, dynamic research programs, and exemplary training records. Available basic research projects include the use of novel models combined with cutting-edge technologies in the analysis of the basic biology of bone loss;osteoporosis, other metabolic bone diseases, and glucocorticoid-induced bone loss, dental disease, metastatic bone disease and osteosarcoma;biomaterials/implant research;and gene therapy transfer technology. The clinical research projects (epidemiology, prevention, outcomes, and effectiveness studies;development of diagnostics;and clinical trials) exploit the participation of clinical faculty in long-term, multi-institutional studies. The Training Program is administered by Dr. McDonald (Chair: Pathology) with the assistance of two Associate Directors, and an Advisory Committee. Trainees in basic research are required to take all or part of the Integrative Biomedical Sciences (IBS) Program, which provides organ- and cell-based basic science training necessary for transition from methods-driven molecular biology and genetics analyses into clinically-relevant, problem-based studies. Predoctoral trainees are enrolled in the IBS, Biomedical Engineering, Cellular and Molecular Biology, or MD/PhD Graduate Programs at UAB. In the past four years, the bone-related curriculum has been expanded and the faculty strengthened through strategic recruitment. The ongoing institutional commitment to bone research has resulted in a major expansion of state-of-the-art facilities and the development of new interdisciplinary programs at UAB that further strengthen the research base and curriculum. Funding is requested for three predoctoral and three postdoctoral trainees. Relevance: This program produces scientists who apply cutting edge molecular and cellular approaches to the discovery, development, and testing of therapeutics for the prevention and treatment of bone loss associated with osteoporosis, rheumatoid arthritis, and other diseases, and the promotion of bone wound healing, as well as the development of orthopedic and dental implants with greater biocompatibility and longevity.
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{
"pile_set_name": "NIH ExPorter"
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During the current funding period, Dr. Wucherpfennig's lab has developed novel approaches for the quantification of autoantibodies and CD4 T cells in autoimmune diseases. This work was conducted as integral component of research projects, and the technologies have now matured to a point where these reagents can be generated most efficiently by a core supervised by Dr. Wucherpfennig. Collaborative work between the Wucherpfennig and Hafler labs resulted in the creation of autoantigen tetramers that enable sensitive detection of autoantibodies in human demyelinating diseases. Tetramers of myelin oligodendrocyte glycoprotein (MOG) were shown to permit sensitive detection of MOG autoantibodies in patients with acute demyelinating encephalomyelitis (ADEM) and pediatric MS. The first major aim for this core will be to generate such antigen tetramers for quantification of autoantibodies against other selfantigens in MS and type 1 diabetes. The core will generate both radiolabeled versions for autoantibody quantification as well as fluorescent versions for characterization and isolation of autoantigen-specific B cells in patients with MS and type 1 diabetes, and in relevant animal models. During the present funding period, the Wucherpfennig lab has also developed a novel approach for the creation of MHC class II tetramers. MHC class II molecules are expressed with a covalently bound CLIP peptide that protects the binding site during biosynthesis and purification. The low affinity CLIP peptide is rapidly released following linker cleavage, and these MHC class II/CLIP precursors can therefore be used for the generation of a variety of different MHC class II tetramers from a single protein preparation. This approach enabled quantification of autoantigen-specific CD4 T cells in animal models of MS and type 1 diabetes. In particular, it allowed simultaneous visualization of regulatory and effector T cells in Foxp3-GFP knock-in mice during the course of experimental autoimmune encephalomyelitis (EAE). The second aim for this core will therefore be to generate MHC class II tetramers for all projects of this PPG and to further develop this technology by generation of multimers with higher valency that can be used to detect lower affinity CD4 T cell populations.
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{
"pile_set_name": "NIH ExPorter"
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The specific purpose of this cooperative agreement is to serve as a research base for the St. Mary's Hospital Community Clinical Oncology Program (CCOP). The University of Rochester Cancer Center will provide: -\approved, non-competing, multi-disciplinary clinical research protocols: - scientific guidance on the implementation of these protocols; -\administrative direction and training for the data management of these protocols. Investigational drugs supplied by NCI and/or pharmaceutical companies for these clinical trials will be procured, distributed, monitored and controlled by the University of Rochester Cancer Center pharmacist.
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{
"pile_set_name": "NIH ExPorter"
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This project aims to elucidate the mechanisms by which the cell exerts control over the metabolism of fatty acids, complex lipids and the overall lipid composition of the cell membrane and outer layers. Regulatory mechanisms governing biosynthesis of the lipid components of the envelopes of procaryotic organisms as model systems will be emphasized. Several different approaches will be employed. The first is the study of regulatory mutants of lipid metabolism in Escherichia coli and other unicellular true bacteria. The lesions of regulatory mutants of E. coli now in hand will be determined by study of the enzymes presumably involved as the sites of genetic lesion, and by mapping the mutations by reference to known genetic markers. The second approach is a continuation of a comparative study of microorganisms to uncover and characterize any repression and/or feedback inhibition of fatty acid and/or lipid biosynthesis by fatty acids or complex lipids that occurs. A third approach involves study of the occurrence and synthesis of lipids in the blue-green algae. This group of procaryotic organisms has been so far only poorly studied, and exhibits a number of unique features (e.g., a high and varied cellular content of polyunsaturated fatty acids) which suggest singular value as models for study of regulation.
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{
"pile_set_name": "NIH ExPorter"
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Chest pain is a frequent symptom of patients with hypertrophic cardiomyopathy (HCM), commonly occurring in the setting of angiographically normal epicardial coronary arteries. In contrast to the generally reproducible effort angina of patients with coronary artery disease, HCM patients ofter report pain with variable threshold of onset, often prolonged in duration even after cessation of activity. Twenty patients with HCM and chest pain despite angiographically normal coronary arteries underwent a pacing study with measurement of great cardiac vein flow, lactate and oxygen content in addition to left ventricular filling pressure. A cohort of 28 patients without HCM who underwent the same pacing study without experiencing chest pain served as controls. During pacing coronary flow rose in both groups although coronary and myocardial hemodynamics differed greatly. In contrast to the linear increase in flow in control up to a heart rate of 150, HCM patients demonstrated an initial rise in flow at an intermediate heart rate, at which point 12 of 20 HCM patients described their typical chest pain. Continued pacing to a heart rate of 150 resulted in a fall in coronary flow in 14 of 20 HCM patients, associated with a substantial rise in left ventricular filling pressures and metabolic evidence of ischemia. A paradoxical narrowing of the arterial-venous oxygen difference was also noted, which may contribute to ischemia. Thus, most HCM patients achieve maximum coronary vasodilatation at modest increases in heart rate. Myocardial ischemia occurs at this point, resulting in elevation in left ventricular filling pressures, probably because of ischemia-related changes in ventricular compliance, with a subsequent adverse effect on coronary blood flow.
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{
"pile_set_name": "NIH ExPorter"
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The overall objective of this project is to determine the composition and role of specific glycolipids in immune effector cell function. Two major aspects will deal with: (1) Testing whether certain glycolipids can function as tumor-specific cell surface markers. We have defined the glycolipid gangliotriosyl ceramide (asialo GM2) as a potential tumor associated marker in two mouse cell lines and have demonstrated both in vivo and in vitro that anti-asialo GM2 specific antibody-dependent immune lytic functions can be directed to the tumor cells. Future experiments will be designed to develop antibody-independent cellular cytotoxicity specific for asialo GM2. (2) Defining glycolipid markers on immune system effector cells. We have recently described the glycolipid ganglio-N-tetraosylceramide (asialo GM1) as a marker for murine natural killer (NK) cells. In addition we are determining the chemical structures of glycolipids on NK-susceptible target cells to define the "target antigen" to which NK cells are directed. The next step will be to determine what role effector cell glycolipids play in lytic phenomena. A vital class of reagents for these studies are monoclonal anti-glycolipid antibodies produced by the hybridoma technique.
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{
"pile_set_name": "NIH ExPorter"
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The heart of the Urologic Oncology Branch is our clinical program- we study the human model of kidney cancer, prostate cancer and bladder cancer. The Urologic Oncology Clinical Core is comprised of a genetic counselor / protocol manager, clinical research nurses, patient care coordinators (PCC), a senior data manager and data managers. RESEARCH NURSESClinical trialsResearch nurse specialists coordinate / manage the operations of our clinical trials in kidney cancer, prostate cancer and bladder cancer. Their responsibilities include: kidney cancer, prostate cancer and bladder cancer patient recruitment, participate in the informed consent process; perform patient education; create patient-specific protocol schedule; schedule blood and urine studies, CT scan, MRI; PET CT, internal medicine, neurosurgical, ENT consults; register patient with CRO; collect / review diaries with patients; assess patient compliance; rapid and accurate data capture; Q/A data entries in c3D; report AEs to NCI IRB and sponsor according to protocol requirements; submit annual Continuing Reviews to IRB; maintain regulatory binder; participate in clinical trial meetings. Familial Kidney CancerA key component of the UOB Branch is the study of families who have rare inherited conditions, such as von Hippel-Lindau, Hereditary Papillary Renal Cell Cancer, Birt-Hogg-Dub Syndrome, and Hereditary Leiomyomatosis and Renal Cell Carcinoma, which predispose to development of kidney cancer. Research nurse specialists and a genetic counselor provide protocol oversight for our natural history protocols in kidney cancer, prostate cancer and bladder cancer. Responsibilities include: patient recruitment; telephone intake covering pedigree; initiation of informed consent process to include purpose of study; provide information about NIH, the study objectives, assess patients expectations; initial medical history; determine appropriate clinical studies; communicate with Patient Care Coordinators (PCC) re: appointment and indicated studies; assist PCC in obtaining pertinent medical records; tissue slides, films; obtain informed consent; assure patient has adequate understanding of information received during visit; prepare / submit annual Continuing Review to NCI IRB.Prostate Cancer and Bladder CancerThe prostate cancer and bladder cancer program is supported by research nurse specialists who have overall oversight of patients with localized prostate or bladder cancer. Responsibilities include: telephone intake to include; purpose of study; information about NIH, the study objectives, assess patients expectations; initial medical history; initiate informed consent process; determine appropriate clinical studies; communicate with Patient Care Coordinators (PCC) re: appointment and indicated studies; obtain informed consent in person ; patient teaching; follow-up visit; assure Quality of Life questionnaires are submitted /received by patients at multiple time points; work with data managers to assure data is captured and submitted. GENETIC COUNSELING A Genetic Counselor actively participates in pursuit of the research questions about known genetic syndromes or unknown familial renal cancer. Responsibilities include: family recruitment through professional genetic societies; develop a 3-generation pedigree; complete a personal medical history; determine the genetic test to be performed; perform literature searches, contribute to the identification of novel clinical manifestations of rare heritable malignant disorders. Patients with or at risk for an inherited kidney cancer cancer susceptibility disorder undergo an extensive counseling / education session. Topics generally covered during these sessions include: Clinical aspects and natural history of the suspected disorder; if the disorder is unknown, the differential, clinical and genetic evaluation plan is presented; psychosocial risks of genetic testing; implications of potential genetic test results; clinical management of condition and /or multiple, bilateral renal tumors; health promotion strategies; provide supportive counseling at-risk or affected patients; provide written educational material and medical papers if appropriate; provide information about support groups; discuss familial communication. Genetic test results are provided per patient preferred modality; a counseling letter is sent with DNA result and recommended surveillance strategies.PROTOCOL MANAGEROverall protocol management is the responsibility of our Genetic Counselor. Protocol manager responsibilities include: draft natural history protocols; prepare and submit protocol amendments; submit Continuing Reviews for natural history and tissue procurement protocols; remain informed about the IRB process as well as the ethical / legal aspects of clinical research; serve as a resource/ mentor to the UOB about protocols and the IRB processPHYSICIANS ASSISTANTThe in-patient and Urology consult service is managed by our Physician Assistant (P.A.). Responsibilities include: manage surgical patients upon decision for surgery; determine with surgical Fellows, the appropriate pre-operative work-up; arrange in-patient admission; schedule / enter orders for operative admission including imaging studies, consultations; refer patients to NIH social service for potential Family Lodge availability; perform H & P upon admission; conduct regular and frequent patient rounds; handle patient concerns while surgeons in operating room; schedule post-operative follow up visit; collaborate with nursing /social service re: discharge plan; see consult patient during post-op visit. The Urology consult service, offered to the entire NIH community is managed by our P.A. Responsibilities include: initially see / evaluate all consult patients; assess patient and, in concert with UOB Fellows, identify probable diagnosis; recommendations to referring service; see all consults at follow-up visits; submit consult note into CRIS system. PATIENT CARE COORDINATORSThe complex task of scheduling multiple appointments and clinical studies is the responsibility of our patient care coordinators (PCC). PCC responsibilities include: schedule new and follow- up patient appointments; communicate with patients regarding reimbursement / travel policy; admission procedures; enter patient information into ATV system; schedule all studies such as CT, MRI. bone scan, PET CT; arrange coordinate consultations with internal medicine, neurology, endocrine, dermatology, cardiology consults, etc.; develop patient schedule; send letter of invitation; copy of protocol consent, appointment schedule; obtain relevant tissue slides, imaging studies and medical records; track / oversee recommendation by UOB physicians for follow-up visits and recommended studies for follow-up visits. DATA MANAGEMENTSenior Data ManagerOverall management of UOB data is the responsibility of our senior Data Manager. His responsibilities include: adapt LabMatrix to UOB data needs; maintain extensive UOB database; responsible for data integrity; query requested reports; Q/A entered data; capture and track: demographics; tumor measurements; clinical manifestations; genetic test results / genotype; provide mentorship to contract data personnel.Data management staff:One data management staff is responsible for tracking/ identifying tumor size and location; growth rates of tumors; captures images for UOB archives. Two contract data staff members are responsible for entering / QA data collected from natural history and clinical trials; tracking enrollment; works closely with research nurses and CRO to assure data accuracy. All data from the clinical trials in entered into the LabMatrix database of the Urologic Oncology Branch.
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{
"pile_set_name": "NIH ExPorter"
}
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The goals will be achieved by a) evaluating obesity, energy balance, and physcial activity in relation to cancer incidence and survival;b) Improving assessment of energy intake, output, and body phenotype;c) Evaluating biomarkers to determine mechanisms including inflammatory cytokines (CRP, IL-6, TNF-alpha), growth factors and their major binding proteins (IGF-1 and IGFBP-3), steroid hormones (testosterone, estrogen, SHBG) in relation to cancer and intermediate outs comes such as weight loss and weight gain d) evaluate novel such as metabolomics in assessing obesity, energy balance, and physical activity and body phenotype and in relation to cancer risk. Although this project is for all cancer, its particular cancer includes breast, colorectal, and pancreatic cancer.
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{
"pile_set_name": "NIH ExPorter"
}
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The research proposed is a three year cross-sequential study of the cognitive and motor skills and social and affective behaviors of 120 children with pervasive developmental disroders (PDD), who have been diagnosed by a child psychiatrist as meeting the DSM-III diagnostic criteria for either infantile autism (IA) or childhood onset pervasive developmental disorder (COPDD). Children diagnosed as having IA or COPDD will be recruited into six age groups of 20 children each. At the time of initial testing the sample will consist of 20 4 year olds, 20 6 year olds, 20 8 year olds, 20 10 year olds, 20 12 year olds, 20 14 years old. Each of the children in the PDD sample will have two normal control matches--a verbal mental age match, and a non-verbal mental age match. Children in the PDD sample will be tested once a year for three years on a neuropsychological test battery: normal controls will be tested once, in year two of the study. In years one and three, children in the PDD group will be videotaped in 2 20 minute structured sessions, and parents and teachers will be asked to complete a behavior checklist. The long-term objectives of this study are fourfold: to determine the pattern of development of cognitive skills in IA and COPDD diagnosed children; to separate normal-but-delayed from deviant aspects of language development in the PDD sample; to construct, through empirical methods, cognitive skill profiles and behavioral profiles for the PDD children; and to explore the relationship between cognitive skill profiles and behavioral profiles and to examine their joint relationship with the DSM-III diagnoses of IA and COPDD. The six specific aims of the study are : (1) to determine whether a previous finding of a post-pubertal decline in cognitive skills is replicated in this sample; (2) to construct and cross-validate cognitive profiles; (3) to analyze the stability of cognitive profiles cross-sequentially; 4) to construct and validate behavioral profiles; (5) to determine joint distributions of cognitive and behavioral profiles and DSM-III diagnostic categories; and (6) to combine present data with previously collected data on the language development of 177 children with PDD in order to generate a large-scale contrastive developmental profile of linguistic skills. It is hoped that the empirical construction of valid subgroups, and clearer understanding of cognitive development will help to better define classification criteria, and aid study of biological markers.
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{
"pile_set_name": "NIH ExPorter"
}
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The long-term objective of the proposed work is to reduce foodborne illness associated with commercial food establishments in Minnesota and the entire U.S. Specific aims are to: 1) increase and improve environmental assessments conducted during foodborne illness outbreak investigations; 2) identify contributing factors and antecedents factors associated with foodborne illness outbreaks; 3) identify and understand environmental factors associated with sporadic foodborne illness through special studies; 4) evaluate food safety programs in Minnesota; 5) synthesize and disseminate the findings from EHS-Net research projects to the environmental health community and the food service industry; and, 6) develop, implement, and evaluate interventions at commercial food establishments that will reduce the incidence of foodborne illness. The Minnesota Department of Health (MDH) has been a leader and active participant in EHS-Net since its inception and proposes to continue EHS-Net involvement in the same fashion going forward. Environmental health specialists from the MDH Division of Environmental Health and epidemiologists from the MDH Foodborne, Waterborne, Vector borne, and Zoonotic Diseases Section will continue to comprise the MDH EHS- Net team and will work closely together to conduct EHS-Net activities. Active population-based surveillance for reportable bacterial and protozoal pathogens will be conducted, along with state-of the art outbreak detection and investigation. Environmental assessments will be conducted on all foodborne outbreaks in commercial food establishments in Minnesota using standard tools to allow collection of high quality data for the National Voluntary Environmental Assessment Information System [NVEAIS]. Special studies to identify practices, behaviors, or other risk factors that lead to sporadic foodborne illness will be conducted at random samples of restaurants in previously selected sampling frameworks. Another proposed project is to synthesize all previous EHS-Network and disseminate it to target audiences in an effective format. Finally, a major part of our proposed ongoing work will be to use data and lessons learned from past EHS-Net activities to develop, implement, and evaluate interventions at commercial food establishments. MDH will continue to build communication and collaborations with EHS-Net partners (other states, CDC, FDA, and EPA) and will actively participate in multi-site studies decided on by the EHS-Net Steering Committee. Education of environmental health specialists and industry groups throughout Minnesota, based on lessons learned from EHS-Net, will continue to be actively pursued.
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{
"pile_set_name": "NIH ExPorter"
}
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The Cancer Epidemiology Program (CEP) is an interdisciplinary research program organized into three scientific areas, each related to a type of cancer risk factor: (1) Viral Risk Factors, (2) Hormonal, Obesity, and Inflammation- Related Risk Factors, and (3) Genetic and Epigenetic Risk Factors. Viral Risk Factor research in the CEP focuses extensively on human papillomavirus (HPV) and its role in anogenital and oral cancers. The CEP has long been a major contributor to HPV research. The goal of this research is to contribute new information important to the development of cancer screening practices, HPV vaccine strategies, and other new prevention and treatment methods. This includes studies of the viral and host factors associated with HPV persistence/progression; biomarkers of cervical and anal pre-cancer/cancer; the effectiveness of HPV vaccines in high risk populations; and the impact of microbicides on risk of HPV infection. In addition. Viral Risk Factor research in CEP addresses the effects of HIV/AIDS on cancer, including studies of the immunologic deficits that drive the relationship of HIV with HPV-related tumorigenesis and risk of other AIDS-associated cancers. Hormonal/Obesity/inflammation research in CEP focuses extensively on the role of the insulin/IGF-axis, sex hormones, adipocytokines, and related pathways in obesity-associated cancers (e.g., colon, breast, prostate, etc.). This includes prospective studies of tumor incidence/recurrence/progression and their relation with circulating and local tissue levels of proteins in these pathways and the expression of their receptors. Given the US obesity epidemic these studies are timely, and will contribute to ongoing efforts to identify biomarkers in these pathways that can be used for patient risk stratification, and/or as targets for chemoprevention and treatment. Genetic and Epigenetic research in CEP focuses extensively on germline and somatic mutations, genetic polymorphisms, DNA methylation, and microRNAs. These studies examine the signaling pathways related to oncogenesis, tumor biomarkers important for selecting and developing targeted therapies, and genetic/epigenetic risk factors that can be used for patient risk stratification. CEP investigators are also conducting methodologic studies to improve the laboratory and statistical tools available for conducting genetic and epigenetic research. The CEP currently has 26 members from 11 departments, of whom 11 are new members, supported by 15 NCI grants ($3.7M Direct), and 11 other peer-reviewed cancer-relevant grants ($2.7M Direct). Since the last CCSG review there have been 414 cancer-relevant research papers in the CEP of which 29% represent intraprogrammatic and 18% represent interprogrammatic publications.
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{
"pile_set_name": "NIH ExPorter"
}
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Knowledge of the molecular structure of trimeric Env on intact viruses and delineating the mechanisms of cell-cell transmission are central to the design of effective immunogens and therapeutic agents to combat HIV/AIDS. We have taken several important steps over the last year towards these goals. Cell-cell interactions that play a critical role in normal immune system function are exploited by HIV to facilitate its transmission from antigen-presenting cells such as dendritic cells to susceptible target CD4+ T-cells via a specialized structure designated a virological synapse. Using ion abrasion scanning electron microscopy, electron tomography, and super-resolution light microscopy, we analyzed the spatial architecture of cell-cell contacts and distribution of HIV virions at virological synapses formed between T-cells and both mature and immature dendritic cells. We demonstrated the striking envelopment of T-cells by sheet-like membrane extensions derived from mature dendritic cells, resulting in a shielded region for formation of virological synapses. Within the synapse, filopodial extensions emanating from CD4+ T-cells make contact with HIV virions sequestered deep within a 3D network of surface-accessible compartments in the dendritic cell. We show that HIV-1 induces membrane extensions in immature dendritic cells through activation of Cdc42. We demonstrated that these extensions are induced following engagement of DC-SIGN by HIV-1 Env via a cascade that involves Src kinases, Cdc42, Pak1 and Wasp. Silencing of Cdc42 or treatment with a specific Cdc42 inhibitor, Secramine A, dramatically reduced the number of membrane protrusions visualized on the cell surface and decreased HIV-1 transfer via infectious synapses. Ion abrasion scanning electron microscopy of cell-cell contact regions showed that cellular extensions from immature dendritic cells that have the appearance of thin filopodia in thin section images are in fact extended membranous sheets with a narrow cross-section. Our results demonstrate that HIV-1 binding on immature DC enhances the formation of membrane extensions that facilitate HIV-1 transfer to CD4+ T lymphocytes. While there are striking differences in the synapses formed by immature dendritic cells, the common theme of the burial of the site of HIV transfer and the receptor-dependent initiation of virion transfer by T-cells are new discoveries that highlight novel aspects of cell-cell HIV transmission. An immunogen that can stimulate the production of broadly neutralizing antibodies to the HIV-1 envelope glycoproteins (Env) gp120 and gp41 is a major goal in the quest for a HIV/AIDS vaccine. Using cryo-electron tomography combined with subvolume averaging, we have analyzed the structure of SOSIP gp140 trimers, which are cleaved, solubilized versions of the ectodomain of trimeric HIV-1 Env. We show that unliganded gp140 trimers adopt a quaternary arrangement similar to that displayed by native unliganded trimers on the surface of intact HIV-1 virions. When complexed with soluble CD4 (sCD4), Fab 17b, which binds to gp120 at its chemokine co-receptor binding site or both sCD4 and 17b Fab, gp140 trimers display an open conformation in which there is an outward rotation and displacement of each gp120 protomer. We demonstrate that the molecular arrangements of gp120 trimers in the closed and open conformations of the soluble trimer are the same as those observed for the closed and open states, respectively of trimeric gp120 on intact HIV-1 BaL virions, establishing that soluble gp140 trimers can be designed to mimic the quaternary structural transitions displayed by native trimeric Env. The discovery that trimeric gp140 immunogens can mimic functionally relevant Env quaternary structural changes on infectious virions is an important step towards development of an Env-based HIV-1 vaccine, and also represents an important technical advance in the use of tomographic methods to determine the structures of protein complexes. Our previous studies on cryo-electron tomography of HIV-1 established several basic features of the function of trimeric Env and the conformational changes associated with CD4 binding. We have extended these investigations to a variety of SIV and HIV-1 strains, including some that were part of earlier controversies in the field about spike structure. Our results showed unequivocally that the molecular architectures of trimeric Env from SIVmneE11S and SIVmac239 strains are closely comparable to that previously determined for HIV-1 BaL, with the V1 and V2 variable loops located at the apex of the spike, close to the contact zone between virus and cell. The location of the V1/V2 loops was definitively confirmed by structural analysis of trimeric Env in an HIV-1 strain with deletions of these loops. Strikingly, in SIV CP-MAC, a CD4-independent strain, trimeric Env is in a constitutively open conformation with gp120 trimers splayed out in a conformation similar to that seen for HIV-1 BaL Env when it is complexed with sCD4 and the CD4i antibody 17b. Our findings suggest for the first time a structural explanation for the molecular mechanism of CD4-independent viral entry and further establish that cryo-electron tomography can be used to discover distinct, functionally relevant quaternary structures of Env displayed on intact viruses.
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{
"pile_set_name": "NIH ExPorter"
}
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The principle function of this laboratory is to provide mass spectrometry analysis for the investigators[unreadable] participating in the program project. Specifically this Core will perform qualitative and quantitative analysis of[unreadable] phospholipid subclasses and molecular species of these subclasses using state-of-the-art electrospray[unreadable] tandem mass spectrometry (ESI/MS/MS). In addition, the facility will quantify 5-HETE, PGE2, and other[unreadable] oxidized polyunsaturated fatty acids generated by tissue culture samples. The source of the samples will be[unreadable] from tissue culture and tissue collected from animal models. The sensitivity and dynamic range of the[unreadable] methodology can measure from pmole to fmole amounts of lipid. As required, the Core will implement and[unreadable] develop advanced methodologies to facilitate the analysis of lipids for the program project.
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{
"pile_set_name": "NIH ExPorter"
}
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