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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. In the first study of poly(IC) and rectal SIV transmission, we asked whether TLR ligation by poly(IC) could prevent SIV transmission or modify the course of infection in animals that were not protected. Poly(IC) was applied to the rectal epithelium of macaques either coincident with (n=7) or 24 hours prior to (n=7) SIVwt challenge. As controls, we challenged 4 animals with SIVwt without receiving poly(IC) treatment and included the 4 animals used to test the infectivity of the virus stock (n=8 total). It does not appear that either the coincident or the 24 h pre-treatment with poly(IC) affected the infection rate (coincident: 4 of 7 infected, 57%;24 h pre: 4 of 7 infected, 57%, control: 6 of 8 infected, 75%). However, we are continuing to evaluate differences in plasma virus RNA over time (at peak and set point) as well as innate and adaptive immune parameters (memory and na[unreadable]ve T cell subsets, regulatory T cells, antibodies, DCs and the ratio of myeloid and plasmacytoid DCs, and cytokine and chemokine production) in these 18 animals.
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Summary of Work: The stability of genomes, particularly humans, is influenced by opportunities for recombination between long (0.3 -10 kb) repeats of diverged DNAs such as ALUs and LINES, rearrangements between very short (4-10 bp) random repeats and replication slippage within the stretches of microsatellites. All these repeats are abundant in the genome. Homonucleotide runs and other microsatellites are a source of frequent replication slippage leading to frameshift mutations inactivating genes. We have demonstrated that unpaired loops resulting from replication slippage in long homonucleotide runs are not subject to proofreading by the DNA polymerase 3' to 5' exonuclease, because they are distant to the 3'-terminus of the nascent strand. As a result, the prevention of mutations would rely primarily on postreplication mismatch repair. This has now been demonstrated by the hypermutability (as much as 10,000-fold) of homonucleotide runs and other microsatellites in mismatch repair deficient cells. We have determined that a mismatch repair defect renders genes with a long homonucleotide run 100-fold more mutable than genes lacking homonucleotide runs in the coding sequence. Thus, homonucleotide runs represent important at-risk sequences in the genome and could be an important source of human disease.
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{
"pile_set_name": "NIH ExPorter"
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The MESA study is on-going. MESA study visit five began in April 2010 and will be completed in late 2011. Analyses of various ocular phenotypes, including retinal microvascular disease, have resulted in multiple published manuscripts with more in press, or under preparation. Genotype and ocular phenotype data began to be released on dbGaP in August 2010. Genetic analyses under the auspices of MESA SHARe are on-going. The MESA Study has several ancillary studies, including a lung component (funded by NHLBI) and an air pollution component (funded by the EPA). Discussions to integrate and analyze data across various study components have resulted in peer-reviewed papers and numerous multi-disciplinary collaborations. More information about the MESA Study and its ancillary studies, including a full list of participating investigators and institutions and a complete list of publications to-date, can be found at http://www.mesa-nhlbi.org.
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{
"pile_set_name": "NIH ExPorter"
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This is a proposal to continue the longitudinal study of adjustment and disorder among three large, population-based cohorts of adolescents from predominantly African-American, low-income, urban families. Each cohort was initially assessed for peer social status, academiC achievement, and social behavior, particularly aggression and prosocial behavior in third grade, and those assessments were repeated in fourth and fifth grades. Adjustment to the first year of middle school, delinquency, and teen parenthood was assessed for each total cohort (n = 1749) at the end of sixth grade, and intensiVe assessments of conduct problems, psychological disorder, and drug or alcohol abuse were made on representatiVe, random, stratified subsamples of each cohort (n =648). These assessments will have been repeated at the end of the eighth and tenth grade year for all three cohorts by the end of this project year. Data on family structure and functioning, as well as stressful life events for the subject have been collected on all subsamples concurrently with the psychological assessment. Police records on delinquency, school records on truancy and drop out, and public health records of teen mother live births have been collected for the full cohorts during this period. Social network data, including information on deviant peer group membership was obtained for one cohort at sixth, eighth, and tenth grades and at eighth and tenth grades for one older cohort. The goal of the project for the next five years is to continue the same schedule of data collection on all three cohorts until their class finishes high school. Retainees will be assessed at the time of their cohort and until they graduate. When the majority of each cohort completes twelfth grade, the representative subsamples will be assessed each year for three years, assuring that all subjects will have reached the age of twenty-one. The samples in each cohort are over-sampled for a group of children who fit both Patterson's and Moffitt's theories of early starting delinquents. The additional years of data collection will allow us to identify the late starter groups postulated by these theories. The validity of propositions about social development and peer network involvement for these two groups will be tested on this inner-city African-American population. A second data analytic goal is to analyze patterns of individual change within each of the three major developmental periods covered by this project - preadolescence, adolescence, and early adulthood - and to identify major patterns of development across the three periods. Within this framework, we will establish predictors of risk for delinquency and disorder both in terms of point-to-point predictions and dynamic growth trends. A final goal is to identify patterns of resilience among this sample and to determine protective processes operating within the family and the community. A subsidiary goal will be to track the longterm implications of two small sample interventions that were successful at one year of follow-up prior to middle school.
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{
"pile_set_name": "NIH ExPorter"
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Delirium is a major challenge facing anesthesiologists and surgeons due to its prevalence, complex etiology, and potential severe impact on patients. Postoperative delirium is associated with longer hospital stays, poor functional outcomes, higher healthcare costs, and increased mortality. A related phenomenon, postoperative cognitive decline (POCD) is also common and associated with worse functioning after hospital discharge. There is recent, but limited evidence to suggest that anesthetic depth is associated with poor postoperative cognitive outcomes. Specifically, several studies suggest that deeper level of anesthesia is associated with increased incidence of postoperative delirium and longer-term cognitive decline. In fact, some suggest that older patients undergoing surgery should routinely be monitored with an anesthetic depth monitor to allow the titration of anesthetics or sedation medications to lighter levels to promote better postoperative cognitive outcomes. However, such a practice has never been validated or proven to be feasible or safe. In this grant proposal, we will undertake a randomized control trial in a cohort of older patients undergoing major spine surgery to receive either a light vs. deep level of anesthesia as determined by a processed electroencephalogram. We will use a validated battery of tests to measure cognitive function before and after surgery. Postoperative delirium will be measured using the Confusion Assessment Method. In contrast to most previous studies that were cross-sectional, we will perform preoperative and serial follow-up assessments to measure the trajectory of cognitive function, and 6-week outcomes. In this study, our aims include determining the feasibility and safety of randomizing patients undergoing major spine surgery to receive deep vs. light anesthetic levels during surgery. Specifically we aim to determine whether two different anesthetic depths as measured by an anesthetic depth monitor can be practiced uniformly by a number of anesthesia providers across a large group of older surgical patients. Second, we will determine an effect size for designing a future larger trial to determine whether anesthetic depth contributes to an increased incidence of adverse postoperative cognitive outcomes as measured by delirium or cognitive decline. Third, we will determine whether preoperative level of cognitive impairment moderates the effect of deep vs. light levels of anesthesia on incident delirium or POCD. Our proposed exploratory clinical trial will be the first to randomize patients undergoing general anesthesia to receive either light vs. deep levels of anesthesia with simultaneous measurements of postoperative delirium and cognitive status. Our study will determine not only the feasibility of anesthetizing patients using two different levels of anesthetc depth, but also the safety of such an approach. In addition to measuring postoperative delirium and POCD, we will also be monitoring possible adverse consequences of light anesthesia. Our study results will provide critically important information to improve perioperative care to the older patients undergoing major surgery.
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{
"pile_set_name": "NIH ExPorter"
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A new class of Ig which has recently been identified on murine lymphocytes has been tentatively described as IgD because of similarities with its human counterpart. Preliminary studies indicate that the expression of this Ig is thymic independent and also perhaps antigen independent. Although it appears after IgM during ontogeny, it may be the major cell surface Ig on lymphocytes from adult mice. For this reason the study of IgD is considered essential in understanding B cell differentiation and function. The objectives of this proposal are to (1) definitively identify this Ig as IgD by the production of a cross- reacting antiserum (2) determine if development of IgD is antigen independent by using antigen-free mice, (3) show that IgD receptor has the same antigenic specificity as IgM by using idiotypic antibody to precipitate both classes from lysates of iodinated immune cells, (4) study the relationship of the presence of IgD to the homing properties of lymphoid cells by using radioiodinated cells in transfer experiments, (5) establish the role of the spleen as a site for differentiation of cells from IgM to IgD by using splenectomized and asplenic mice, (6) study IgD-biosynthesis and shedding in resting and stimulated cells using previously described techniques, and (7) establish whether a switch from IgM to IgD occurs in differentiating B lymphocytes by using irradiated bone-marrow reconstituted animals.
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{
"pile_set_name": "NIH ExPorter"
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During the past decade a surge of experimental evidence has shown that astrocytes release chemical transmitters which modulate synaptic transmission and neuronal excitability. In addition to taking up synaptically released neurotransmitters, astrocytes release glutamate, ATP and D-serine. Though it is now clear that astrocytes listen and talk to synapses, the functional implications of this bi-directional signaling pathway are not yet clear. During the past funding period we have generated astrocyte-specific inducible transgenic mice in which we can block the release of the chemical transmitter ATP from astrocytes in adult animals. With this mouse we have discovered that astrocytes coordinate synaptic networks by regulating the extracellular concentration of adenosine: astrocyte-released ATP is hydrolyzed to adenosine which suppresses the strength of excitatory synaptic transmission. The availability of this transgenic animal puts us in the unique position to test the following hypothesis. "Activity dependent release of ATP from astrocytes leads to the accumulation of extracellular adenosine that suppresses synaptic transmission neuronal excitability. The astrocyte-dependent accumulation of adenosine acts as a spatial filter, enhancing the contrast between neighboring synaptic pathways and limits the spread of excitation thereby suppressing the development of seizures." Using molecular genetics together with brain slice electrophysiology we will perform four specific aims of investigation: Aim I: We will test the hypothesis that the diacylglycerol arm of the phospholipase C (PLC) pathway is required to stimulate ATP release from astrocytes. Aim II: Synaptically stimulated release of purines from astrocytes causes a contrast-enhancement between neighboring active and inactive synapses. Aim III: Astrocyte-derived purines block the induction of LTP. Aim IV: By integrating the overall level of neuronal activity astrocytes have anti-convulsant actions through their control of extracellular adenosine.
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{
"pile_set_name": "NIH ExPorter"
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Elevation of catecholamines are known to occur in stressful situations like hypoglycemia or diabetic ketoacidosis. This study is designed to measure the relative contributions of hepatic glycogenolysis and gluconeogenesis to glucose production when plasma epinephrine in increased. 13C-nuclear magnetic resonance spectroscopy (natural-abundance 13C) will be used to measure rates of whole body glucose production. This study will be performed in healthy subjects and poorly controlled type I diabetic subjects.
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{
"pile_set_name": "NIH ExPorter"
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Many patients who suffer from neurally-mediated syncope eat a low salt diet. This diet may, in fact, increase their risk of developing syncope. At this point in time, there are no established methods to quantify salt intake. This is a pilot project, using a simplified dietary questionnaire and two 24-hour urine collections, to categorize patients according to their salt intake. Thirty normal subjects will be studied, and each will fill out the pilot questionnaire, undergo four detailed dietary interviews, and provide two twenty-hour collections of their urinary sodium excretion.
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{
"pile_set_name": "NIH ExPorter"
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The objective of this research is to define the mechanism action of vitamin A at the molecular level in organs other than eye. Emphasis is given to investigation of possible alterations in the transcriptional mechanism during vitamin A deficiency. Efforts are made to characterize the properties and function of two newly discovered intracellular binding proteins for retinol (vitamin A) and retinoic acid. The levels of these binding proteins are examined during perinatal development of the rat.
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{
"pile_set_name": "NIH ExPorter"
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The goal of this Mentored Patient-Oriented Research Career Award (K23) is to prepare Dr. Heather Niemeier to become an independent scientist studying innovative behavioral approaches for the treatment of obesity. Through a combination of didactic and direct mentored training experiences, this training plan will provide her with skills in the following areas: the etiology and pathophysiology of obesity and the behavioral approach to weight regulation;methodology, statistics, and ethics of clinical research in obesity;and strategies being used to address affective and cognitive factors in eating disorders and addictive disorders and their translation into obesity treatment. Dr. Niemeier will work closely with her mentor, Dr. Rena Wing, and her co-mentors, Drs. Kittichai Promrat, Joseph Fava, Richard Brown, and Carlos Grilo throughout the course of the award. The resources at The Miriam Hospital/Brown Medical School combined with the mentoring by an expert advisory team will provide Dr. Niemeier with the ideal environment to achieve her long-term goals. Obesity is a major public health problem. Behavioral weight loss, the current treatment of choice for moderate obesity, achieves impressive short term results, however, weight regain following treatment is a major problem. Over 50% of participants in behavioral weight loss programs report difficulty with internal antecedents to unhealthy weight control behaviors and this difficulty is associated with weight regain following treatment. Current treatment approaches do not adequately address these antecedents. The aim of this application is to develop, implement, and evaluate a behavioral weight loss program modified to provide participants with skills to deal effectively with affective and cognitive difficulties. Two studies are proposed, both targeting men and women with BMI of 27-40 who self-report difficulty with emotional and/or cognitive antecedents to unhealthy weight control behaviors. Study I is an uncontrolled pilot study in 20 participants to initially assess acceptability and efficacy of the innovative treatment. In study II, 120 participants will be randomly assigned to receive 12 months of standard behavioral therapy for weight loss or standard behavioral treatment plus affective and cognitive skills training. The long-term goal of this research is to improve the weight loss maintenance outcomes of behavioral weight loss programs by addressing affective and cognitive antecedents to unhealthy weight control behaviors.
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{
"pile_set_name": "NIH ExPorter"
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This Master Agreement Order (MAO) is to determine the vital status and current address of persons under study by the National Cancer Institute (NCI). The cohort to be traced are mothers and their offspring born between the 1940s and 1960s. The subjects originate from two geographic areas, Chicago and Boston. There are approximately 400 mothers from the Chicago area who participated in a randomized clinical trial between 1950 and 1952. Approximately 85 offspring from these pregnancies also need to be traced. There are about 200 mothers, originally from the Boston area, who gave birth between 1945 and 1965. There are also about 200 daughters (not offspring of these mothers) born in the Boston area during the same time period. Both of these groups have been followed before but were lost during the 1980s. There are also approximately 700 offspring (males and females) born at the Boston Lying-In Hospital who have never been included in a study and will need to be traced.
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{
"pile_set_name": "NIH ExPorter"
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In order to understand the development and functioning of the thymus, both in terms of T cell differentiation and stromal cell environmental support, we have undertaken a molecular approach to identify genes that are uniquely expressed in this organ. We have created an anchored RT- PCR based cDNA library from 14 day fetal thymuses after culturing them in deoxyguanosine and treating them with an anti-CD45 antibody to deplete the cell population of hematopoietic cells. The cDNA library was subtracted with poly A+ RNA prepared first from a fibroblast cell line and then from whole spleen. Three prime sequencing of 250 random cDNAs revealed 137 which were not in the databases of known sequences. The novel cDNAs were then screened by Northern blotting for expression in various tissues and in a set of SV40 transformed thymic epithelial cell lines. Four genes were selected for further analysis because they were limited in their expression to the thymus or to one of the stromal cell lines. All four were completely sequenced by obtaining full length cDNA clones from a SCID thymus library.During the past year the lab has made progress in understanding the structure and function of two of these genes. 1) The 1C12 gene, encoding a 12 transmembrane spanning co- transporter glycoprotein was disrupted by insertional mutagenesis using a DNA segment encoding both a beta galactosidase and a neomycin resistance gene. The insert was placed in the middle of the first exon. Heterozygous mice had very poor expression of the beta galactosidase gene and proved not be useful for identifying the cell type(s) expressing the 1C12 gene. Furthermore, homozygous gene-targeted mice showed normal thymic development as well as normal repopulation of the thymus in adult mice following sublethal irradiation. Thus, despite the exclusive expression of this protein in the subcapsular region of the thymus, no functional abnormalities were found by disrupting the gene. 2) The 1C10 gene has weak homology to transcription factors containing a POU domain and possesses several nuclear localization signals. To demonstrate that in fact the gene product goes to the nucleus, we engineered a 1C10 construct in which the gene for Green Fluorescent Protein was attached to the 3? end. GFP normally distributes in the cytoplasm, but when attached to 1C10 it accumulated in the nucleus. In a separate set of experiments we examined the expression of the 1C10 gene in thymus and lymph node in the absence of various lymphocyte cell populations, using a series of gene-targeted mutations. We found that expression in the lymph node required B cells, gamma delta T cells, and to a lesser extent alpha beta T cells, while expression in the thymus was independent of the presence of these cells and was even seen in RAG-/- mice which lack all but the earliest lymphoid precursors. Reconstitution of RAG-/- mice with normal bone marrow did not result in 1C10 expression. These observations suggest that 1C10 requires an inductive signal early in development in order to be expressed in the lymph node. - Thymus, genes, mice, knock-out, RT-PCR
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{
"pile_set_name": "NIH ExPorter"
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ABSTRACT The Rush Alzheimer?s Disease Core Center (Rush ADCC; P30AG10161) supports cutting edge and innovative research on the etiology, pathogenesis, diagnosis, treatment, and prevention of mild cognitive impairment (MCI), Alzheimer's disease (AD), and other common conditions of aging, by providing researchers with a stimulating environment, highly valued clinical and post-mortem data, and ante- and post-mortem biologic specimens. The Rush ADCC supports a variety of timely and important areas of research including risk factors for the transition from normal aging to MCI to AD, the neurobiology of normal aging and MCI, and studies that incorporate contemporary omics technologies and neuroimaging to identify novel therapeutic targets. These research areas are supported through the careful design and integration of seven highly successful Cores: an Administrative Core; Clinical Core; Data Management and Statistics Core; Neuropathology Core; Outreach, Recruitment and Education Core; Religious Orders Study Core, and Latino Core, as well as the Research Education Component. The Rush ADCC and its ancillary studies including the Rush Memory and Aging Project and the Minority Aging Research Study are building a vibrant neuroimaging portfolio of studies, some of which include participants of the Clinical and Religious Orders Study Cores for in-vivo imaging, and in collaboration with the Neuropathology Core for ex-vivo imaging. We are now generating autopsies on persons without dementia who have undergone in-vivo imaging. These are mostly non-Latino whites. Now that we have a relatively large number of African Americans and Latinos in the Clinical and Latino Cores, with a growing number of autopsies, we are in a position to expand this work in the minority communities. The overall goal of the proposed Neuroimaging Core is to generate a resource of neuroimaging data from the Rush ADCC African Americans and Latinos, and affiliated studies, and provide neuroimaging expertise to facilitate high quality, cutting edge, externally-funded research focusing on the transition from normal aging to mild cognitive impairment (MCI) to the earliest stages of Alzheimer?s disease (AD) and other dementias. The proposed Neuroimaging Core will markedly extend the ability of the Rush ADCC to interrogate neuroimaging as part of its clinical-imaging-multi-level omics-pathology pipeline supporting studies of the transition from normal aging to MCI to the earliest stages of dementia, and studies of ante- and post-mortem biospecimens, and extend this work into the African American and Latino communities which are vastly under- represented.
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{
"pile_set_name": "NIH ExPorter"
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Our two laboratories have significant experience with the design of biomaterials for tissue engineering and the use of genetic engineering to enhance vascular cell survival and blood vessel formation in vivo. For example, in the Saltzman laboratory, biodegradable cell-adhesive polymer microparticles have been used for assembly of brain cells into neotissues;controlled release of nerve growth factor by the microspheres enhanced brain cell survival and function after transplantation. In the Pober laboratory, conditions have been developed for isolation, culture and retroviral transduction of vascular cells and Bcl-2-transduced human umbilical vein endothelial cells (EC) suspended in gels of natural biopolymers have been shown to form a microvascular network capable of anastomosis with host vessels and to induce remodeling in the host so as to increase local tissue perfusion. The Pober laboratory has also extensively characterized the responses of ECs to tumor necrosis factor (TNF), most recently showing that in human organ culture, TNF can act through a pathway involving TNF receptor 2 (TNF-R2) and the downstream kinase endothelial/epithelial tyrosine kinase (Etk) to stimulate cell growth and tissue repair. Here, these techniques will be combined and optimized to produce engineered systems that are capable of rapid, robust, and reliable revascularization of ischemic tissue. These systems will be tested in animal models that permit dissection of the cellular and molecular features that lead to revascularization of limbs after ischemia. Our working hypothesis is that optimization of cell/polymer transplantable systems with respect to composition of the polymer scaffold, addition of controlled-release functions, and appropriate selection of cells will lead to improved therapeutic recoveries in blood flow and clinical outcomes in ischemic models. To test this hypothesis, we propose to: 1. compare the effect of introduction into human EC of wild type and mutant forms of Bcl-2, TNF-R2 and Etk on revascularization within scaffolds;2. optimize the conditions for transplantation of transduced EC by incorporation of wild type or modified vascular smooth muscle cells (VSMC) or by modifications in the composition of the natural protein polymers or of the scaffold composition;and 3. introduce controlled release of agents into the scaffold design such as a TNF mutein that signals via TNF-R2 but not TNF-R1. Since these experimental systems are flexible with respect to transduced genes, protein release, polymer surface modification, and cell source, they are ideal constructs for testing additional lypotheses once proof of concept is established. Our approaches rely on materials that are already acceptable to the FDA in clinical settings;therefore, our results in animal models will be ready for translation into clinical practice.
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{
"pile_set_name": "NIH ExPorter"
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Atherosclerosis, the underlying cause of most human heart disease, results from a focal imbalance of the normal equilibria of the arterial wall. While metabolic cooperation between vascular cells is essential for the maintenance of normal vascular homeostasis, little is known about the nature of these interactions or whether disturbance of their equilibria will precipitate irreversible pathological changes in arterial tissue. We propose to investigate mechanisms of cellular interactions between vascular endothelium, smooth muscle cells (SMC) and monocyte-derived macrophages in vitro and in intact vascular tissue. Two separate general mechanisms of cell cooperativity will be studied: a) cell contact-mediated communication via gap junctional channels connecting the cytoplasm of adjacent cells, and b) humoral communication in which diffusible substances secreted by one cell type pass via the interstitial fluid to specific receptors on the surface of the other cell population. The effects of cell biological pertubations associated with hypercholesterolemia (cellular cholesterol, lipoprotein metabolism), a major risk factor for atherogenesis, will be investigated. New techniques to probe cellular communication in intact normal and atherosclerotic (fibrofatty lesion) vascular tissue will be tested and developed to integrate the in vitro findings with vessel wall biology and pathology. Gap junctional-mediated cell interactions will be investigated biochemically in vitro (transfer of 3H-nucleotides, fluorescent dyes and putative second messengers such as cyclic nucleotides and Ca++ and electrophysiological measurements) using cocultures of endothelial cells, SMC and macrophages. The effects of various mediators of gap junctional transfer, particularly cellular cholesterol composition, will be evaluated. The effects of heterocellular communication upon receptor-mediated lipoprotein metabolism in endothelial cells will be measured. Gap junctional communication will also be investigated in normal and atherosclerotic intact arterial tissues both at the ultrastructural level and functionally using new methods to deliver tracers to the vascular tissue. The regulation of platelet-derived growth factor-like mitogens (c-sis related) synthesized and secreted by endothelial cells will be investigated in the context of the interactions of these cells with SMC and macrophages using molecular biology techniques.
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{
"pile_set_name": "NIH ExPorter"
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The focus of this application is to define how enamelysin (MMP-20) processes enamel proteins in vitro and in vivo, and to determine how such processing allows for proper enamel development. The proposed research takes advantage of a unique development in the enamel field: the generation of an enamelysin knockout mouse. The enamelysin (-/-) mouse displays a severe enamel phenotype consisting of hypoplastic enamel and an obliterated rod pattern with no other detectable tissue malformations. The knockout mouse proves that proteolysis of enamel matrix proteins is a critical part of Dental enamel formation and allows us to test specific hypotheses concerning the functional effects of enamelysin cleavages in vivo. These hypotheses are: 1) enamelysin is the only protease that cleaves enamel proteins during the secretory stage of mouse amelogenesis, 2) enamel matrix proteins characterized from the enamelysin knockout mouse will be uncleaved, and therefore still in their secreted forms, 3) amelogenin and/or ameloblastin cleavage products are necessary for enamel rod organization, and 4) assemblies of full-length enamel proteins catalyze an increment of crystal elongation that, once completed, is followed by proteolysis to disassemble the structure and make way for another round of crystal extension. By performing a proteomic characterization of the enamel matrix of developing teeth obtained from the enamelysin knockout and from wild-type mice (Aim 1), important new information will be gained on the relative abundance of various enamel protein cleavage and alternative splice products. By determining the substrate specificity of enamelysin (Aim 2) we will understand the relative contributions of enamelysin cleavage site preferences and the accessibility of preferred cleavage sites (i.e., does protein folding play a role?) in determining the make-up of the enamel matrix. To test the validity of our hypotheses, we propose to utilize the knockout mice for transgenic experiments that determine if amelogenin and/or ameloblastin cleavage products are necessary for enamel rod organization (Aim 3) and to determine if proteolysis of full-length enamel proteins is necessary for enamel crystallite elongation (Aim 4). This application utilizes a unique tool (enamelysin knockout mouse) and recent, highly advanced technologies including, the ProteoSep system for protein isolation and mixture based oriented peptide libraries for substrate specificity analysis, to help determine how protein processing allows for proper enamel development. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Investigations of the biosynthesis of two novel microbial metabolites will be continued. The first is the antitumor agent valanimycin, an azoxy compound produced by Streptomyces viridifaciens. As a naturally occurring azoxy compound, valanimycin is a member of a growing family of natural products that includes substances with antitumor, antifungal, and carcinogenic activity. Previous investigations have led to the elucidation of the early steps in the valanimycin biosynthetic pathway and, more recently, to the cloning and analysis of the valanimycin biosynthetic gene cluster. The long term objective of future studies of valanimycin is to identify and investigate the proteins encoded by the gene cluster that are responsible for the formation of the azoxy group of valanimycin, and thereby to elucidate the biochemical mechanism of azoxy group formation. These studies are significant because they should help illuminate the chemistry of N-N bond formation associated with the biosynthesis of other known bioactive natural products that contain N-N bonds. Future studies of valanimycin have several specific goals. The first goal is to investigate the role played by VImL, an apparent seryl-tRNA synthetase, in valanimycin biosynthesis, including an investigation of the possibility that a seryl-tRNA is an intermediate in the valanimycin biosynthetic pathway. The second goal will be to overproduce and investigate the function of several other proteins in the gene cluster that may be involved in the processing of the known intermediate isobutylhydroxylamine and in the processing of a serine derivative such as seryl adenylate or a seryl-tRNA. The third goal will be to create non-polar disruptions in genes of unknown function in the gene cluster and analyze the chemical phenotype of the resulting mutants to detect new intermediates in the pathway. The second metabolite to be investigated is the antitumor agent sparsomycin, a potent inhibitor of protein biosynthesis produced by S. sparsogenes. Sparsomycin is an important target for biosynthetic investigation because it is known to inhibit the peptidyltransferase step in protein biosynthesis and because previous studies have shown that sparsomycin biosynthesis involves novel biochemistry. Future investigations of sparsomycin will focus on cloning the sparsomycin resistance gene from S. sparsogenes to understand the mechanism of self-resistance in this organism and to gain access to the biosynthetic genes.
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{
"pile_set_name": "NIH ExPorter"
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This is a randomized, active-controlled parallel group trial in patients with chronic hepatitis C who have not previously been treated. Intron A 3 MU three times a week (TIW) for 48 weeks will be compared to three doses of PEG-IFN (0.5, 1.0, 1.5 mcg/kg) administered once a week for 48 weeks. Follow up will be for an additional 24 weeks.
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{
"pile_set_name": "NIH ExPorter"
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Our long-term goal is to understand the mechanistic details that underlie the formation of nascent synapses after repeated cocaine exposure. The objective of this application is to use cocaine self- administration, viral-mediated gene transfer and electrophysiological assays to begin to elucidate the formation of these synapses. Within the limbic system, three major glutamatergic afferents that synapse onto NAc neurons originate from the prefrontal cortex (PFC), hippocampus, and the amygdala. Given that these three afferents play distinct roles in addictive behaviors, identifying the specific afferents that undergo cocaine-induced generation of nascent synapses and thus circuitry reorganization will provide a circuitry- based understanding of different aspects of the addictive state. The central hypothesis of the application is that self-administration of cocaine increases the number of silent synapses in NAc neurons among the PFC afferent and that this increase is correlated with an enhancement in the ability to learn about drug reward;thus, forming an apparent pathological memory. We plan to test our hypotheses by pursuing the following two aims: 1) Determine the effect of the contingency of cocaine administration on silent synapses development;and, 2) Establish the glutamatergic afferent to the NAc that express cocaine-generated silent synapses. The proposed work is innovative because it will further our understanding of the cellular and molecular changes that accompany cocaine exposure. Silent synapses have been shown to be crucial in the development of synapses and circuitry in the developing brain. If we can understand the development of addiction from this novel angle, more accurate pharmacological manipulations can be designed for future clinical interventions to block and/or reverse pro-addiction cellular adaptations.
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{
"pile_set_name": "NIH ExPorter"
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Aedes aegypti mosquitoes from several locations in Asia, Africa, southern USA, and Caribbean will be tested for vector competence with yellow fever virus. The rate of yellow fever transovarial transmission after the first ovarian cycle will be determined. An attempt will be made to develop the RIA detection technique for yellow fever antigens in mosquito expectorate. The pathogenesis of Mokola virus in Aedes mosquitoes will be studied using virus titration and FA methods. Emphasis will be on establishing a passage line in reproductive tissues. The development of antibody for Congo-Crimean hemorrhagic virus in laboratory animals will be followed by FA, AGDP, CF, and HI tests. The plaque reduction neutralization test and in vitro immunologic methods in CER cells, will be used to compare virus strains from different geographic areas. Comparison of strains will also be attempted using glycoproteins and biochemical techniques.
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{
"pile_set_name": "NIH ExPorter"
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Health problems in our nation?s communities are not uniformly distributed. While the national infant mortality average is 7.2 deaths per 1,000 live births, the highest prevalence when categorized by race is among African American infants. The African American infant mortality rate is 13.8 nearly three times higher than the racial group with the lowest infant mortality rate. Medical advances alone have not eliminated the health disparity gap implying that social and psychological constructs also play important roles. The goal of this research proposal is to study the effects of stress and coping on birth outcomes. The specific aim is to gain a better understanding of how perceived racism and stressful life events affect the birth outcomes of African American women and to describe how coping, including spirituality and social support, are related to the cumulative stressors that these women experience. Data will be collected from interviews during pregnancy using various tools to measure stressful experiences that women may encounter and coping strategies used in their lives. Each study participant will be tracked to identify birth outcome. Racial health disparities are a complex issue. This proposal will add a dimension to existing knowledge by examining perceived experiences of racism in context with other variables and their effects on birth outcomes among African American women.
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{
"pile_set_name": "NIH ExPorter"
}
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The technology to characterize microvascular network dynamics and assess its impact on tissue oxygen delivery is only now emerging. We have recently demonstrated the unprecedented ability to map oxygen in the microvasculature and cerebral tissue with two-photon microscopy (TPM) [Sakadzic2010] and are proposing to complement this with our novel statistical intensity variation optical coherence tomography (OCT), enabling quantitative dynamic maps of microvessel flow, red blood cell flux, and vessel diameter. Combined, these technologies will provide unprecedented spatio-temporal resolution imaging of oxygen delivery to brain tissue. These technologies will have a broad impact in health science in preclinical models of neuro-degenerative diseases and cancer, and, as we elaborate in Aim 4, help guide the identification of clinically relevant imaging biomarkers. To demonstrate the technologies' utility, we will address fundamental questions about cerebral vascular physiology: How do microvascular flow properties impact tissue oxygenation, and how do age-related vascular alterations compromise tissue oxygenation? We will then relate these findings to clinically relevant imaging biomarkers.
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{
"pile_set_name": "NIH ExPorter"
}
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B cell differentiation centers on changes in immunoglobulin (Ig) gene expression--the production of Ig with high antigen-binding avidity and heavy chain isotype suitable to elicit various physiologic responses to foreign antigen. In the proposed project, the structural features of Ig gene expression will be analyzed in unique, long-term B lymphocyte cell lines (B-Lyl cells) and antigen-specific subclones. In respone to conven- tional stimuli (anti-IgM and lymphokines), these cells initiate high-rate IgM secretion, production of "mature" heavy chain isotypes (class switching), and heavy chain class switching. Consequently, the molecular events related to Ig gene expression can be analyzed in these cells as an inducible, ongoing process. Using recombinant DNA techniques, the molecular basis for the changes in Ig gene expression will be determined: sequential DNA rearrangements, and structural and quantitative changes in mRNA species. Screening methods for somatic mutation will also be evaluated. These molecular events may then be used to more precisely characterize the regulatory influence of various T lymphocyte subclasses, macrophages, and their soluble product. B-Lyl cells are also notable for their similarity by surface phenotye to a minor B cell population believed to account for the polyclonal Ig production in genetic models of murine SLE. In the future, comparison of B-Lyl cell lines derived from "normal" and "lupus" strains may clarify the mechanism of abnormal Ig production in this disease model.
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{
"pile_set_name": "NIH ExPorter"
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Worldwide, head and neck cancer is a major health problem. The incidence rate of head and neck squamous carcinoma (SCCHN) is higher among Puerto Rican Hispanics than among Hispanics living in the US. Advances in the locoregional control of SCCHN have not increased patients' survival due to recurrence, metastases or second primary tumors (SPTs). Development of SCCHN involves the interplay between genetic events and environmental factors. Tobacco and alcohol are the main risk factors. Tobacco and alcohol are the main risk factors. Recently, human papillomavirus type 16 (HPV16) has been implicated in the etiology of 20-30% SCCHNs. Gene-environment interactions play a significant role in determining susceptibility to SCCHNs. Gene-environment interactions play a significant role in determining susceptibility to SCCHN. Variants in DNA repair genes, and metabolic genes associated with tobacco and alcohol metabolism could be associated with the individual's susceptibility to SHHN. Variants in DNA repair genes, and metabolic genes associated with tobacco and alcohol metabolism could be associated with the individual's susceptibility to develop cancer. The underlying hypothesis is that individuals with susceptible genotypes and behavioral risk factors are at greater risk of disease progression and SPTs. In HPV16-positive SCCHN patients, HPV16 DNA serum levels may be a potential biomarker for metastasis. To test our hypothesis we plan a follow-up study of 250 Puerto Rican patients with the following specific aims: 1) Gather baseline and follow-up epidemiological and clinical data on Puerto Rican SCCHN patients (200 new patients and 50 already accrued); 2) Determine HPV16 positive tumors and specific variants; 3) Measure the serum levels of HPV16 DNA in HPV16 DNA in HPV16-positive patients; 4) Correlate HPV16 infection and p53 mutations with clinical data of SCCHN Puerto Rican patients; 5) Analyze selected constitutional polymorphic genes associated with tobacco and alcohol metabolisms, and DNA repair (e.g. GSTs, ADH3, XRCC1, XPD, and p53); and 6) analyze epidemiological and biomarker data as predictors of recurrence and development of SPTs. These patients will be followed for the development of recurrences, metastasis and SPTs. We will be able to obtain preliminary data and to compare lifestyle characteristics and molecular markers in this Hispanic population. The long-term goal of this line of research is to identify markers for predicting the risk of recurrence of STP in these patients could be used to develop specific intervention strategies to target high-risk patients. This project will provide the opportunity to develop the infrastructure for an epidemiological study at UPRCC. These preliminary data will be basis for an R01 proposal.
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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. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The fibrous collagens are the fundamental constituents of the Extracellular Matrix (ECM) of animals, forming the structural basis of all known mammalian connective tissue: bones, skin, cornea, teeth, tendons, ligature, blood and lymphatic vessels and all organ systems including the heart and lungs. Therefore, the aims of this proposal are to: 1) To structurally characterize the fibrillar collagens types II and I by fiber crystallography. 2) To structurally characterize the molecular organization of extracellular matrix molecules commonly found complexed with the collagens via fiber crystallography and bioinformatics / computational methods. 3) To develop fiber crystallography and Micro X-ray Diffraction ([unreadable]XD) techniques that enhances the chance of successful completion of aims 1 and 2.
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{
"pile_set_name": "NIH ExPorter"
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Project Summary The objective of this proposal is to investigate the epigenetic mechanism of genomic imprinting of the imprinted gene Growth Factor Receptor Bound Protein-10 (GRB10) in neuronal versus non-neuronal development. Imprinted genes are unique to mammals and are expressed in a monoallelic, parent-of-origin-specific manner. Dysregulation of imprinted gene expression is associated with neurological, behavioral and growth disorders. GRB10, implicated in growth disorder Silver-Russell syndrome, is uniquely expressed from the paternal allele in neuronal cells and from the maternal allele in non-neuronal cells. Strikingly, the maternal and paternal transcripts initiate from different promoter regions and the nature of this novel switch in neurons is unclear. This proposal aims to investigate how the allelic and isoform switch occurs at the Grb10 locus to enable unique parent-of-origin expression in neurons. We and others have identified binding sites for the critical DNA architectural protein CTCF within the differentially methylated Grb10 imprinting control region (ICR). Our laboratory also described the importance of CTCF in controlling monoallelic Grb10 expression in mouse embryonic stem cells (mESCs). Aim 1 will test the hypothesis that CTCF is required for coordinating the imprinted and molecular switch in Grb10 expression. The ability of CTCF to catalyze neuronal-specific expression will be assessed by mutating binding sites within the ICR in an in vitro neuronal differentiation model. Aim 2 will test the hypothesis that tissue-specific enhancers regulate the Grb10 switch in neurons. A putative enhancer will be removed by CRISPR editing in mESCs, and tissue-specific Grb10 expression will be measured in derived neurons. The studies in this proposal use the imprinted gene Grb10 as a model to study the epigenetic and molecular mechanisms that govern tissue-specific gene expression. Given that many genes in the central nervous system are also tissue and allele-specific, discerning the epigenetic factors coordinating Grb10 expression in neurons will elucidate how other complex genes are differentially regulated in neurodevelopment.
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{
"pile_set_name": "NIH ExPorter"
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In our previous study we have demonstrated the direct relationship between the level of free and bound testosterone in plasma and HDLc. The association persisted after adjustment for age, cigarette smoking, alcohol consumption and obesity. Physical activity did not appear to be related to HDLc in this population because of the relatively sedentary status. We propose to re-evaluate the same participants two years later and again determine the relationship between hormones, the risk factors, and lipoprotein cholesterol. Measurement of both testosterone and estrogen will be done. A replication of our previous findings are necessary prior to doing more detailed lipoprotein and hormone studies. The men participating in this study are part of the Multiple Risk Factor Intervention Trial (MRFIT) cohort, and will be re-examined at their sixth annual examination. The cost of the study will be modest since most of the information is collected as part of the MRFIT study.
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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. This project focuses upon Research in Computational Epidemiology with an emphasis on a health disparity disease of major concern, that of HIV/AIDS. SPECIFIC AIMS: 1) Development of a novel micro-epidemiologic computational model for HIV/AIDS at the intracellular level: The model was developed to provide insights in how an intracellular/molecular level model can be incorporated within a macro-epidemiologic integrative systems dynamics model for examining a variety of computational experimentations. 2) Development of a systems dynamics model and modelling of the multifactorial epidemiology of HIV/AIDS. An epidemiologic model that incorporates cellular/molecular (micro level) and macro level epidemiologic dynamics for examining preventive and therapeutic strategies of HIV/AIDS in African American sub populations was developed. 3) Modelling of the multifactorial epidemiology of HIV/AIDS: Multifactorial and quantitative epidemiologic studies that interrelate (biomedical, behavioral, and socioeconomic factors) determinants have been developed. Using these models as our experimental medium, we evaluate various intervention scenarios to recommend effective strategies for the community to minimize the risk of new infections prevent future infections and manage existing infections. 4) Static modeling to examine if HAART Therapies for HIV/AIDS. To answer the question of which combination of HAART is most effective we collaborated with the University of Alabama at Birmingham (UAB) 1917 HIV/AIDS clinic. Although the results of this study appeared promising in the beginning, they did not provide clear and reliable outcomes. STUDIES AND RESULTS: Manuscripts are being prepared for submission to journals. PLANS: To continue analyzing data, gathering more data and information for the estimation of the parameters and variables of the model for HIV/AIDS with a focus on the macro-epidemiology of AIDS in African Americans.
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{
"pile_set_name": "NIH ExPorter"
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Human TLR2 differs from many of the TLRs in that 1) it forms hetero dimeric [unreadable] complexes with other TLRs (like TLR1 and TLR6); 2) it recognize pathogens with the widest variety. We have optimized the expression of ectodomain using baculovirus expression system, which produce enough quantity samples suitable for structure studies.[unreadable] We have obtained crystals that diffract beyond 3.8 A. Molecular replacement has not been successful so far. We are continuing the study in order to obtain better diffracting crystals and solve the phase.
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{
"pile_set_name": "NIH ExPorter"
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The detection and alignment of locally conserved regions (motifs) in multiple sequences can provide insight into protein structure, function and evolution. A new Gibbs sampling algorithm is described that detects motif-encoding in sequences and optimally partitions them into distinct motif models; this is illustrated using a set of immunoglobulin fold proteins. This algorithm extends the previous work in this area (Lawrence, et. al. Science, 262:208-214, 1993) in three ways: 1) The requirement for the specification of the number of motifs in each sequence has been relaxed. 2) The length of the motif is now automatically determined by the algorithm. 3) A non parametric test for the significance of the alignment has been developed. When applied to sequences sharing a single motif, the sampler can be used to classify regions into related submodels, as is illustrated using helix-turn-helix DNA-binding proteins. This feature permits the algorithm to simultaneously align the sequences and classify segments into submodels. Other statistically-based procedures are described for searching a database for sequences matching motifs found by the sampler. When applied to a set of thirty-two very distantly related bacterial integral outer membrane proteins, the sampler revealed that they share a subtle, repetitive motif. Although BLAST (altschul et. al., 1990) fails to detect significant pairwise similarity between any of the sequences, the repeats present in these outer membrane proteins, taken as a whole, are highly significant (based on a generally applicable statistical test for motifs described here). Analysis of bacterial porins with known trimeric beta-barrel structure and related proteins reveals a similar repetitive motif corresponding to alternating membrane spanning beta-stands. These beta-strands occur on the membrane interface (as opposed to the trimeric interface) of the beta-barrel. The broad conservation and structural location of these repeats suggests that they play important functional roles.
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{
"pile_set_name": "NIH ExPorter"
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The Administrative Core will be responsible for integrating all scientific and infrastructure activities of the Conte Center. Drs. Robert Schwarcz (the Center PI and co-PI of the Core) and William Carpenter (co-PI of the Core), together with the Steering Committee, will have overall responsibility for all administrative functions of the Center. The Steering Committee will include the key scientific leadership for this application: Dr. L. Tonelli (PI, Project 2); Dr. L.E. Hong (PI, Project 3); Dr. R.W. Buchanan (PI, Project 4); Dr. R. McMahon (Biostatistician); and Dr. J. Gold (Neuropsychologist), and will meet monthly to review the progress of each of the projects. They will also review the Biostatistics and Data Management Module, which provides statistical and data base support for the projects. In consultation with NIMH program staff, the Core will establish an External Advisory Committee and consult/meet with the advisors regulariy to review plans. The External Advisory Committee will include experts in relevant translational neuroscience, clinical research and drug discovery academic and industry labs. The Steering Committee will also be directly involved in: review of proposals for Pilot and Feasibility projects; facilitating the involvement of post- doctoral research fellows and junior faculty in Center activities; involving all Center scientific staff in Center progress through an annual retreat; overseeing training in responsible conduct of research and HIPAA regulations, diversity recruitment; overseeing compliance with UMB and Federal Human Subjects and Vertebrate Animals regulations; implementation of an Optional Summer Research Experience for undergraduates; dissemination of research results and sharing of research data; and coordinating the provision of staff support [funded by the host institution (MPRC) rather than this grant] for administrative functions of the Center; data management and data entry; and IT staff, as well as IT hardware and software resources.
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{
"pile_set_name": "NIH ExPorter"
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Collegiate living arrangements represent a microcosm of the undergraduate experience and act as one of the ways in which the culture of the campus influences the individual student. Despite the overall campus norms on drinking, collegiate living arrangements vary greatly and can either encourage or discourage alcohol misuse. Grant funds are requested to examine the potential protective influence of pioneering residential learning communities (RLCs) in reducing alcohol misuse among first-year undergraduate students using a longitudinal panel design. RLCs were developed to enhance the co-curricular connections between formal learning opportunities and students' living environment; however, these RLCs also may provide a milieu that discourages alcohol misuse. The proposed target sample will include a total of 5,200 first-year undergraduate students living in either RLCs (n= 1,200) or traditional residence halls (n=4,000). The specific aims of this study are: (1) To assess whether first-year students in RLCs exhibit less alcohol misuse and fewer alcohol-related consequences than first-year students in traditional residence halls; (2) To longitudinally examine the mechanisms by which RLCs may affect alcohol misuse and alcohol-related consequences. In particular, the study will investigate whether a relationship between RLCs and reduced alcohol misuse and alcohol-related problems operates via environmental factors (e.g. alcohol use by roommates) and/or individual characteristics (e.g. involvement with co-curricular activities); and (3) To determine subgroups of students who benefit most from RLCs. Characteristics of RLCs (e.g. single sex) will be examined by considering appropriate groupings and comparing the degree to which students within these subgroups exhibit decreased alcohol misuse and consequences. Additionally, individual characteristics (e.g. gender, ethnicity) will be scrutinized as possible moderators of RLC effects, by studying interactions between these potential moderators and RLC status. In partnership with a research awardee under RFA AA-03-008 and NIAAA staff, student participants will self-administer a survey at three points in time: (1) baseline during freshmen orientation prior to the first academic semester; (2) during the second academic semester at 6-month follow-up; (3) during the second year at 18-month follow-up. At each measurement point, participants will complete a set of measures that assess substance use behaviors and constructs addressed by RLCs.
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{
"pile_set_name": "NIH ExPorter"
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In the United States, there are 3.9 million adults disabled as the result of traumatic brain injury (TBI). These people live with significant and permanent alterations in social, behavioral, physical, and cognitive functioning. If they are to obtain the services that are necessary to address the effects of TBI, families need to be effective advocates. However, the significant impact of brain injury on families leaves them poorly equipped to advocate for services, especially in this era of shrinking availability of rehabilitation options. This Phase I project successfully developed an interactive multimedia CD-ROM for families affected by TBI, focusing on effective communication skills used to advocate for services to address social behavior in adults with TBI. Phase II development will adapt the CD-ROM materials for Internet use and expand content to include: (a) four additional advocacy skills, (b) additional areas of concern (e.g., physical/medical issues, cognitive functioning, psychosocial concerns, vocational planning, community integration, and substance abuse/misuse), (c) the ability to tailor information by degree of support provided by the family member to the individual with brain injury, and (d) a repeat-visit function. In addition, a web-based chat room will provide social support. This interactive multimedia (IMM) program will provide family members training in key skills needed to advocate for TBI-related services. The benefits of the IMM format include: proven effectiveness; low cost; individualized, self-paced mastery learning; objective tracking of the users' progress; and availability of training materials regardless of busy family schedules. The market for the CD-ROM product includes all families impacted by the TBI of their adult relative, non-family caregivers, and rehabilitation professionals providing TBI- related services. [unreadable] [unreadable] [unreadable]
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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. We continue our collaborative research with Bayer Corp. in areas of in thrombosis and hemostasis. We are engaged in a research project to generate definitive information whether PAR1 antagonists provided by Bayer have therapeutic potential for treatment of thromboembolic diseases. First, 5 compounds will be evaluated ex vivo, and 1 compound will be tested in vivo. Subsequently, 3 compounds will be evaluated ex vivo, and two compounds will be evaluated in vivo.
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{
"pile_set_name": "NIH ExPorter"
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The ability of polarized epithelia to alter their cellular organization to accommodate the formation of complex structures is essential for the proper formation of almost all tissues and organs in the vertebrate embryo. The research put worth in this proposal will investigate the mechanistic basis for how cells control morphology and architecture during development, by elucidating the genetic, cell biological, and biochemical properties of Shroom family proteins during embryogenesis and adult life. The Shroom family of cytoskeletal-associated proteins has been linked to controlling cellular morphogenesis during vertebrate embryogenesis in humans, mice, and frogs, as defects in these genes cause neural tube defects and X-linked mental retardation. The specific aims of this proposal are as follows: Aim I: Utilize genetic approaches in mice to define the in vivo functions of Shroom2 and Shroom3. Aim II: Define the mechanism by which each of the Shroom family proteins are regulated. Aim III: identify the cellular factors that link Shroom3 to myosin-dependent changes in cell shape and architecture. Aim IV: Determine the cellular basis for the genetic interactions between Shroom3 and the PCP pathway in neural morphogenesis. These aims address the hypothesis that Shroom family proteins function through a defined set of proteinacious networks and pathways to specify epithelial behaviors and characteristics, such as cell morphology and organization, that culminate in establishment of the correct vertebrate body plan and tissue architecture.The research in my lab utilizes a combination of approaches to understand the relationship between specific cellular behaviors and proper embryonic development and how errors in these processes can result in birth defects or embryonic lethality. We will continue to use in vivo studies in mice and in vitro studies in cell culture to understand the molecular mechanisms underlying human birth defects.
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{
"pile_set_name": "NIH ExPorter"
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Current clinical immunosuppressive regimens for allograft recipients affect the entire immune response and result in many unwanted side effects. The first aim of this research is to determine which T lymphocyte subsets are essential for rejection of skin and heart allografts in the mouse. In particular, we will try to separate the roles of class I antigen-reactive T cells (cytotoxic/suppressor cells) from class II antigen-reactive T cells (helper cells). We will then use monoclonal antibodies against the appropriate T cell subsets as therapeutic reagents for normal mouse recipients of skin and heart allografts. Another specific aim is to determine whether some monoclonal antibody isotypes are more effective than others and whether treating with more than one isotype at a time might be helpful. We will assess the extent to which monoclonal antibody induces modulation of T cell surface antigens; we will try to compensate for modulation and loss of immunosuppressive effect by conjugating the monoclonal antibodies with the plant toxin ricin. Finally, we will determine the conditions under which infusion of donor strain lymphocytes (B cell and T cell subsets) into the recipient, after treatment with monoclonal antibody and allografting, affects establishment of microchimerism (in spleen and lymph nodes) and long term allograft survival.
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{
"pile_set_name": "NIH ExPorter"
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Prostate cancer is currently the second leading cancer death in American men. Prostate cancer is dependent on androgen acting through the androgen receptor (AR) for growth and survival, and androgen ablation has been used for treatment of prostate cancer. Despite an initially favorable response, hormone ablation therapy eventually fails and the cancer progresses to an incurable metastatic disease. Epidemiological data regarding prostate cancer genetics indicate roles of AR mutation, as well as AR poly-glutamine (poly-Q) polymorphism, in susceptibility to prostate cancer development and response to therapy. The overall objective of this proposal is to create mouse prostate cancer models for studying the role of the AR in prostatic cancer development and metastasis. Using the Cre-loxP conditional gene knockout technology, several derivatives of TRAMP or PTEN-deficient mouse prostate cancer models with cell type-specific AR knockout (ARKO) (post-puberty epithelial, fibroblast-, and smooth muscle cell-specific ARKO), replacement of prostatic epithelial AR with AR(T857A) [murine equivalent of human AR(T877A)] mutant will be generated for in vivo study of the roles of the AR in prostate stromal cells or epithelial cells, the AR(T877A) mutation in prostate cancer initiation, progression, or metastasis. Prostate tumor initiation, progression, and/or metastasis in these models will be examined and compared with their littermates expressing wild type AR through histological analyses, immunohistochemical determination of various cellular markers, biochemical determination of the expression levels of tumor metastasis related genes and proteins, and analysis of invasive properties of their metastatic tumor. Through the differences in these parameters between the various ARKO or AR (T857A) mice and their wild type littermates, the role of the AR in each specific cell type and the AR(T877A) mutation in prostate carcinogenesis will be delineated. Information obtained from these studies not only will provide better understanding of the roles of the AR in prostate carcinogenesis, but also will be useful for designing new treatment regimen for prostate cancer in hormone refractory state. In addition, these models may be useful for testing new drugs for prostate cancer therapy. Project Narrative: New Mice Models for Studies of Androgen Receptor in Prostate Cancer [unreadable] [unreadable] We will generate various mouse models that lack the androgen receptor in individual cells of the prostate and study their influence on prostate cancer progression. Information obtained from our studies of the differences between these mice models will lead to our understanding some roles of the androgen receptor in the development of prostate cancer and also will be useful for designing new treatments for prostate cancer. In addition, these mice models may be useful for testing new drugs and treatments for prostate cancer therapy, which might eventually be used in humans. [unreadable] [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Patients with deep vein thrombosis of the lower extremities andor pelvis are eligible for this protocol if their thrombosis has been symptomatic for no more than 14 days and if their bleeding risk is very low. They are hospitalized for 3-7 days for up to 4 daily injections of rtPA directly into their thrombi. Simultaneously they receive anticoagulation, which is continued for 6 months. Since the last report we have enrolled 13 patients, for a total of 24. Extensive thrombolysis had been achieved with low doses of alteplase in every patient, and there have been no serious adverse events. The protocol is approved for 30 patients.
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{
"pile_set_name": "NIH ExPorter"
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The project is a cross-sectional and longitudinal analysis of alveolar bone loss (ABL) in aging men. Study subjects will be 700 adult male participants in the VA Dental Longitudinal Study (VADLS). Bone loss will be determined using existing radiographs, including (1) intraoral periapical, (2) cephalometric, and (3) hand-wrist radiographs. The VADLS has the most extensive longitudinal radiographic data base available with which to examine the factors associated with progressive alveolar bone loss. The existence of sequential radiographs permits a longitudinal analysis of the actual ABL experienced over a twenty year period at three year intervals. The study will be able to determine actual rates of ABL per site and per subject, while also controlling for any pre-existing ABL found at the baseline examination. Extensive dental and general health information is available for the subjects so relations between ABL and systemic conditions can be observed. Unique opportunities exist to examine the relationship of ABL to changes in bone at non-periodontal sites in the same subjects and the contribution of extra-oral factors to ABL. Relationships of ABL to age-related changes in bone at non-periodontal skeletal sites can be investigated, allowing study of the contribution of systemic changes in bone density to the intraoral changes in alveolar bone loss. The project will yield much improved estimates of the rate of ABL conditional on putatively important explanatory variables.
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{
"pile_set_name": "NIH ExPorter"
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Summary Odor perception and hedonics are highly intermeshed at both the behavioral and neural circuit level in humans and non-human animals. This association may in part reflect the fact that the piriform cortex (PCX) is particularly tightly linked with the amygdala. Amygdala projections from the lateral, basal, accessory basal, and posterior cortical nuclei as well as the periamygdaloid cortex of the amygdala target both the posterior PCX (and to a lesser extent the anterior PCX), with the PCX sending reciprocal connections back to the amygdala. The basolateral amygdala (BLA) is required for odor fear learning, and work from our lab has shown that PCX odor responses are shaped by both fear learning and by BLA input. For example, discriminative odor fear conditioning involving both a CS+ and CS- results in odor-specific learned fear responses, as well as narrowing of PCX single-unit odor receptive fields (i.e., enhanced PCX odor acuity). This modification of PCX odor coding may be due to input from the BLA since optogenetic activation of BLA fibers within the PCX can modify single-unit and single-unit ensemble odor responses in anesthetized rodents. The BLA may exert this modulation of the PCX through its glutamatergic projections which target both excitatory pyramidal cells and inhibitory interneurons in the PCX. Finally, odor fear memory is shaped by post- conditioning sleep-dependent consolidation. Interestingly, manipulations of PCX activity during post- conditioning slow-wave sleep can influence both the strength and the accuracy of learned fear responses. However, how the BLA and PCX work in tandem to shape PCX odor coding and hedonics is unknown. Here, using high precision spatiotemporal manipulations and single-unit ensemble recordings in freely moving animals, I will dissect the BLA-PCX circuit before, during, and after animals learn hedonic associations with an odor. I will assess the BLA?s role in shaping both the strength and acuity of learned odor responses at both the behavioral and PCX single-unit ensemble levels. The proposed work is comprised of three specific aims. Aim 1 will test the hypothesis that BLA input to the PCX is required for acquisition of an odor-specific learned fear response and associated changes in PCX odor coding. Aim 2 will test the hypothesis that BLA input to the PCX is required during post-conditioning slow-wave sleep for consolidation of odor-specific learned fear response and associated changes in PCX odor coding. Aim 3 will test the hypothesis that BLA input to the PCX is required for expression of an odor-specific learned fear response. The techniques required to successfully complete these proposed experiments will also represent a significant advancement of my technical and analytical skill set to use in my future research career. Beyond mere technical skills, the proposed work also allows me to expand my knowledge of the field and literature into both new brain systems (i.e. piriform cortex and amygdala) as well as a new means of investigating the role of memory in perception.
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{
"pile_set_name": "NIH ExPorter"
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The goals of this project are to 1) purify and immunologically characterize the proteins coded for by mammalian retroviruses and 2) utilize radioimmunoassays for these polypeptides to search for retrovirus expression in mammalian cells and study the molecular biology of RNA tumor viruses.
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{
"pile_set_name": "NIH ExPorter"
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Cystic fibrosis is the most common autosomal recessive genetic disease in the United States. With the recent recommendation for multi-mutation CF screening by the ACMG/ACOG, more labs will be implementing multiplexed genotyping assays. Presently, DNA diagnostic tests for CF are expensive, complicated to run and limited to certain ethnic backgrounds. Using a powerful new technology called AEGIS we propose developing a fast, inexpensive, reliable and easy to run a multiplexed mutation detection system for clinical screening of mutations in the CFTR gene that cause CF in the majority of Americans. In Phase I, proof of concept studies will take place to show that the system is feasible for testing CF mutations on human genomic samples. In Phase Il, we intend to develop a clinical diagnostic assay multi-mutation carrier and newborn CF screening. PROPOSED COMMERCIAL APPLICATIONS: This project could result in an easy to run, low-cost, high-throughput, and ultrasensitive method for Cystic Fibrosis DNA mutation screening that minimizes PCR amplicon handling and is run on an inexpensive validated instrument platform. In addition, validation from this project will open the doors to other test kit development such as population carrier screening for other genetic diseases and research based chromosome mapping studies.
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{
"pile_set_name": "NIH ExPorter"
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Bacteria are highly varied, and have developed mechanisms to diversify to enable survival in a dynamic world. Bacteria that are obligate colonizers of specific hosts have additional benefits and challenges compared with those with broader host ranges or those that are free-living. Helicobacter pylori is a gram negative bacterium that colonizes the human stomach. Once acquired, H. pylori persists in its host essentially for life (in the absence of antimicrobial therapy), is intimately related to human gastric tissues, including injection of H pylori constituents into epithelial cells, and when present, is the single dominant microbe in the human stomach. In addition to its major medical importance, H. pylori also is a model system for understanding microbial persistence in a host and the enabling mechanisms. Our hypothesis is that H. pylori evolved specific genetic mechanisms to create and control variation that maximizes its persistence in the gastric niche. We will address this hypothesis, through several Specific Aims: In Aim 1, we will assess how H. pylori controls intragenomic variation involving short sequence repeats (SSRs), using DNA repair and recombination genes. We plan to examine this question under steady state and fluctuating environments in vitro, and employ mathematical models to understand the underlying principles of the dynamics. In Aim 2, we will assess how H. pylori controls susceptibility to transforming DNA, and then determine its costs and benefits in vivo, in a murine model. Finally, in Aim 3, we plan to conduct experiments to understand the spatial localization (biogeography) of H. pylori colonization; to determine whether there is heterogeneity of sectoring of H. pylori strains in the gastric environment, under fixed or oscillating conditions. For each of the experiments to be performed, we will develop mathematical analyses to find the general properties under which the microbial populations diversify and are selected. Through such analyses, we hope to create the basis for a deeper understanding of how microbes are able to persist for long periods in their human hosts.
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{
"pile_set_name": "NIH ExPorter"
}
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The fifth component of human complement (C5) is an important participant in inflammatory and cell killing processes. The activation peptide (C5a) possesses potent spasmogenic and chemotactic activity. The macromolecular cleavage product (C5b) expresses a transient binding site for C6. The C5b, 6 complex is the foundation upon which the membrane attack complex (C5b-9) is assembled. Elucidation of the molecular features germane to these biological properties will require knowledge of the complete primary structure of C5. Available protein sequence data for C5a will be used to synthesize a mixed sequence oligodeoxyribonucleotide hybridization probe complementary to C5 mRNA which will be used to screen a human liver cDNA clone bank. Confirmed cDNA clones greater than 1kb will be subjected to sonication, thereby producing random overlapping fragments of 400 to 500 bp, shotgun cloned in the bacteriophage M13 vector and sequenced using the Sanger dideoxy termination method. The complete nucleotide coding sequence for C5 will be derived in this way. Concurrently, random tryptic peptides generated from purified C5 will be sequenced to assist in generating an unambiguous complete primary sequence. Poly (A)+ mRNA specifying C5 will be prepared from multiple tissues and cell lines and analyzed using C5 cDNA probes by Northern transfer analysis. The molecular basis for C5 deficiency states will be studied. The chromosomal locus for the C5 gene will be deduced by Southern analysis of DNA samples prepared from interspecies somatic cell hybrids. C5 cDNA clones will also be used as hybridization probes to screen two human genomic libraries. The gene for C5 will be isolated and characterized. Correlation of the intron/exon arrangement of the C5 gene with structural and/or functional domains of the protein molecule will be made. Recent data has indicated that the complement proteins C3, C4, and C5 and the protease inhibitor Alpha 2M are structurally homologous. Available Alpha 2M protein sequences will be used to synthesize oligonucleotide probes for identification and isolation of Alpha 2M cDNA clones. Cloned Alpha 2M cDNA fragments will then be used to identify genomic clones. Information derived from these studies of C5 and Alpha 2M gene structures will be used to analyze the pattern of protein structural domain segregation.
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{
"pile_set_name": "NIH ExPorter"
}
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Normal brain development requires appropriate levels of nutrients, hormones, and other signaling molecules presented to the brain at precise developmental timepoints. For example, iodine deficiency results in reduced thyroid hormone (TH) production and severe developmental abnormalities. Similarly, copper (Cu) and iron (Fe) deficiencies during late brain development result in strikingly similar derangements in brain development. The long-term goal of the proposed research program is to understand the molecular basis of Cu, Fe, and TH action in the developing brain. To this end, the investigators have developed a model that seeks to identify a shared molecular mechanism causative of the aberrancies in brain development associated with these three deficiencies. Recent data have revealed that TH synthesis is reliant on adequate Fe levels. Fe is likely required for normal function of the TH synthesizing enzyme thyroid peroxidase. Interestingly, Cu deficiency is also associated with reduced TH levels. The investigators'preliminary data may provide an explanation for this latter finding. They have observed that Cu deficient rodents become Fe deficient. The Fe deficiency in these animals likely results in TH deficiency. Thus, they have formulated the following hypothesis: that Cu and Fe deficiencies lead to reductions in brain TH levels. The associated reduction in TH levels deleteriously affects brain development and therefore, contributes to the derangements in brain development and function observed in Cu and Fe deficient animals. Three specific aims are proposed to test these hypotheses: Aim 1 is to assess the effects of Cu and Fe deficiency during late brain development on circulating and brain TH levels. Aim 2 is to compare the molecular abnormalities associated with Cu, Fe, and TH deficiencies during late brain development. Aim 3 is to assess the effects of TH repletion on molecular abnormalities associated with Cu and Fe deficiencies during late brain development. These data will reveal whether reduced TH levels mediate some of the pathophysiological effects of Cu and Fe deficiency during neonatal brain development. These studies will further provide the preliminary data necessary to conduct mechanistic studies designed to reveal the precise contributions of each constituent towards brain development and function. If the hypotheses prove correct in model animal studies, it will be important to assess the TH status of Cu and Fe deficient infants to ensure that adequate TH status is obtained during neonatal development. Such findings may have a direct and immediate clinical impact, as even transient TH deprivation during late brain development is associated with reduced cognitive abilities later in life.
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{
"pile_set_name": "NIH ExPorter"
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The purpose of this study is: 1) to determine the effects of Pramlintide(Amylin) on glucose control in Type I diabetes through the assessment of hemoglobin A1C, and 2) to determine the safety of two doses of Pramlintide compared to placebo when administered by subcutaneous injection to Type I diabetes mellitus. The hypothesis to be tested is if Pramlintide improves the control of Type I diabetes, particularly reducing blood glucose in the postprandial hyperglycemic state.
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{
"pile_set_name": "NIH ExPorter"
}
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The HDAC6 gene that we propose to study in this project is a novel therapeutic candidate emerged from the burgeoning field of epigenetics. Histone deacetylases (HDACs) are the therapeutic targets of a novel class of anticancer therapies called HDAC inhibitors. Non-selective inhibitors of histone deacetylases, which promote protein hyperacetylation in the brain, have demonstrated consistent behavioral activity across various rodent models, including models of affective disorders and antidepressant response. Although most studies, so far, have interpreted this antidepressant-like activity in the context of chromatin-remodeling mechanisms (ie the canonical role of HDACs) it is now well established that the influence of protein acetylation on cellular processes extends far beyond transcriptional regulation. In fact, certain deacetylases, of which HDAC6 is a prominent example, are involved primarily in non-histone functions. These functions comprise, for instance, the regulation of HSP90's chaperone activity, an important component of steroid receptors signaling. Through this mechanism, HDAC inhibitors have recently been shown to blunt Glucocorticoid Receptor (GR) function, one of the main receptors for stress steroid hormones. Our preliminary data indicate that HDAC6 is enriched in the serotonin system, the primary target of most antidepressants currently used in the clinic. Here, we propose to test the hypothesis that inhibition of HDAC6 constitutes a candidate therapeutic mechanism for antidepressant and pro-resilience interventions through focused inhibition of GR signaling within the serotonergic system. To test this hypothesis we propose the following aims. Our Aim I is to conduct a comprehensive regional and cellular mapping of HDAC6 mRNA and protein in the mouse brain, and to start evaluating the pattern of HDAC6 expression in the human brain. Our Aim II is to evaluate the behavioral consequences of HDAC6 manipulations in 5-HT system. Here we will combine pharmacological approaches with serotonin-specific gain and loss of function of HDAC6 in the live animal. Consequences of these manipulations will be examined in a battery of tests measuring resilience to chronic stress as well as sensitivity to acute and chronic effects of antidepressants. Our Aim III is to examine HDAC6's influence on GR signaling in 5-HT neurons. Here, we will use a combination of in vivo and cell culture-based assays to characterize the effect of HDAC6 KO and HDAC6 pharmacological inhibition on HSP90 acetylation, GR translocation and GR-mediated gene regulation. The goals of this project connect in 2 major ways with areas of high priority defined by the NIH in the field of Neuroscience and Basic Behavioral Science. First, this proposal will increase our understanding of the fundamental mechanisms of complex social behavior relevant to the socio-affective component of depressive symptomatology. Second, we hope to validate HDAC6 as a new molecular target relevant to the treatment of mental disorders. PUBLIC HEALTH RELEVANCE: This project is integral to our long term endeavor to elucidate the molecular and cellular mechanisms underlying antidepressants therapeutic activity. A greater understanding of these mechanisms is a critical prerequisite to the discovery of new antidepressant therapies, with faster onset and enhanced efficacy, which are greatly needed in Psychiatric clinics. In the present project we propose to evaluate, using animal models, the potential of histone deacetylase 6 (HDAC6) as a new therapeutic target for antidepressant treatments.
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{
"pile_set_name": "NIH ExPorter"
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DESCRIPTION: Adapted from the investigator s abstract: Sickle cell disease arises from the polymerization of sickle hemoglobin by a double nucleation mechanism involving homogenous nucleation in solution and heterogeneous nucleation into sickle hemoglobin polymer surfaces. While the conceptual features of that mechanism have been validated, its predictive power is limited by the lack of a structural foundation and a flawed description of heterogeneous nucleation in the presence of non-polymerizing molecules. The experimental core of this proposal is to assemble a data base of solubility (csat), homogeneous and heterogeneous nucleation rates and elongation rates, for sickle hemoglobin assembly as a function of solution conditions (T, pH, non-ideality) and for various amino acid substitutions. Sickle hemoglobin mutants with additional amino mutations will primarily be provided by Dr. James Manning of Rockefeller University, and the hemoglobin of the SAD mouse will be provided by Dr. Frank Costantini of Columbia University. These data will be used to test and refine a new structural model for heterogeneous nucleation which are proposed herein. The data will also allow the description of heterogeneous nucleation to be extended to describe gelation in the presence of oxygen, fetal Hb, or other non-polymerizing species. This work will ultimately provide an energetic map of the interactions which create the rigid, vasoocclusive gel.
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{
"pile_set_name": "NIH ExPorter"
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The principal objective of this study is to increase our understanding of the primary causes of cataract formation in the human. Animal lenses will be used initially; when and where appropriate, clear and cataractous human lenses removed at the time of cataract extraction, enucleation, or autopsy, will be used. Attention will be focused on the factors regulating anaerobic glycolysis, the principal source of biological energy in the lens. Hexokinase, the pacemaker of this metabolic pathway and phosphofructokinase, an equally important point of metabolic control, will be studied in detail. Isolation and characterization of these enzymes will be attempted. Changes in overall glycolytic activity will be correlated with changes in hexokinase and phosphofructokinase activity. A well developed lens-culture system will be used to study an experimental analogue of the "hypoglycemic" cataract; it is in this type of cataract that a link between decreased hexokinase activity, disordered glycolysis and cataract formation may be substantiated. Well established biochemical and physiological techniques will be employed in all these studies. A continuing effort will be made to extend and compare results with the animal lens to the human lens. BIBLIOGRAPHIC REFERENCES: Chylack, L.T., Jr., Bienfang, D.C., Bellows, A.R. and Stillman, J.S.: Ocular manifestations of juvenile rheumatoid arthritis. Amer. J. Ophthalmol. 79:1026, 1975. Chylack, L.T., Jr., Mechanism of "hypoglycemic" cataract formation in the rat lens. I. Role of hexokinase instability. Invest. Ophthalmol. 14:746, 1975.
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{
"pile_set_name": "NIH ExPorter"
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Obsessive-compulsive disorder (OCD) affects 1-2% of children and adolescents, causing significant distress and impairments from the unrelenting obsessional thoughts and compulsive behaviors. Approximately 1/4 of children with OCD report an abrupt, dramatic onset of their symptoms. When these symptoms are accompanied by other behavior problems, cognitive changes and somatic symptoms, the child may qualify for a diagnosis of Pediatric Acute-onset Neuropsychiatric Syndrome (PANS). PANS is characterized by the sudden onset of OCD and/or eating disorder, accompanied by at least two of the following seven comorbidities: 1) Anxiety; 2) Emotional lability and/or depression; 3) Irritability, aggression and/or severely oppositional behaviors; 4) Behavioral (developmental) regression; 5) Deterioration in school performance; 6) Sensory or motor abnormalities; 7) Somatic signs and symptoms, including sleep disturbances, enuresis or urinary frequency and others. (Swedo, Leckman, Rose Pediatr Therapeutics 2:1-8, 2012) For some children with PANS, symptoms appear to arise as a consequence of common childhood infections, including Group A streptococcal (GAS) infections (strep throat and Scarlet fever). Children whose symptoms begin or exacerbate following GAS infections may belong to a subgroup of neuropsychiatric disorders identified by the acronym PANDAS (for Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcal infections) (Swedo et al, AJP 1998). Three decades of research has revealed that PANDAS represents a post-streptococcal neuroinflammatory disorder, similar to Sydenham chorea (the neurologic manifestation of acute rheumatic fever). Five lines of evidence support an etiologic role for GAS infections in PANDAS: 1) Case-control, population-based studies, and school-based observational investigations have shown increased rates of OCD and tic disorders among children with recent GAS infections. 2) Prospective, longitudinal evaluations of children with acute-onset OCD reveal symptom exacerbations following GAS infection or exposure 3) Prompt treatment of GAS infections can ameliorate OCD and other neuropsychiatric symptoms among children newly ill with PANDAS 4) Prevention of GAS infections reduces neuropsychiatric symptom exacerbations (similar to results for rheumatic heart disease or Sydenham chorea) 5) Repeated GAS infections in the nasal-associated lymphoid tissue of mice produce neuroimmune activation via GAS-specific T-cells which enter the brain via the olfactory nerve In PANDAS, symptoms appear to arise when certain rheumatogenic GAS infect genetically vulnerable individuals and provoke the production of antibodies recognizing antigens not only on the GAS cell wall, but also on cells within the human central nervous system. These cross-reactive antibodies produce neuroinflammation of the basal ganglia, resulting in the abrupt onset of neuropsychiatric symptoms which characterize PANDAS. An emerging literature supports the classification of PANDAS as a post-infectious (autoantibody-mediated) autoimmune encephalitis; supportive lines of research include: 1) Prospective, longitudinal clinical observations documenting multiple symptom domains, including abnormalities of emotional, behavioral, motor, sensory, somatic and cognitive functions 2) Abnormalities on clinical laboratory assays, including quantitative immunoglobulins, as well as on paraclinical tests, such as polysomnography or electroencephalography 3) Demonstration of cross-reactive antibodies that recognize both components on the GAS cell wall and human neuronal tissue; titers of these cross-reactive antibodies are higher during acute illness than during convalescence in the cerebrospinal fluid (CSF) as well as blood of affected individuals 4) In animal models, passive transfer of the cross-reactive antibodies produces behavioral abnormalities and restricted food intake 5) Positron emission tomography (PET) studies reveal activated microglia (a sign of neuroinflammation) in the caudate and lentiform nucleus of acutely ill PANDAS patients; these abnormalities resolve with successful immunotherapy 6) Immune therapies, including steroids, intravenous immunoglobulin (IVIG) and therapeutic plasmapheresis, have been reported to improve neuropsychiatric symptoms (although a 2016 NIMH/Yale trial of IVIG failed to show superiority of IVIG over placebo for 35 children with PANDAS) To replicate the findings in PANDAS and extend investigations to the larger group of children with PANS, Dr. Swedo is currently leading a nationwide consortium of clinicians and researchers with interest in PANS/PANDAS. The goals of the collaborative clinical research network are to: (1) improve recognition, diagnosis, and treatment of acute onset neuropsychiatric syndromes, (2) conduct research which improves understanding of etiologic factors, host vulnerability, and disease mechanisms, and c) increase awareness and recognition of PANDAS, PANS, and related disorders in order to improve access to care. To date, the PANS/PANDAS Clinical Research Consortium has published a guide to the medical evaluation and diagnosis of acute-onset neuropsychiatric disorders (Chang et al, J Child Adol Psychopharmacology, Feb 2015) as well as a set of four papers which provide a comprehensive approach to clinical management (Swedo et al) and include recommendations for use of psychiatric medications and cognitive-behavior therapy (Thienemann et al), as well as treatment with antibiotics (Cooperstock et al) and immunomodulatory therapies (Frankovich et al) (J Child Adol Psychopharmacology Sep 2017, Vol 7, Issue 7, pages 562-606) Members of the Consortium are collaborating on a number of research projects, including efforts to identify specific antigen-antibody interactions; document neuroinflammation through the use of PET or magnetic resonance imaging (MRI); evaluate the impact of the cross-reactive antibodies on the blood-brain barrier and evaluate the efficacy of novel therapeutic interventions. In parallel to these clinical-laboratory investigations, Dr. Swedo has launched a nationwide effort to obtain prospective, longitudinal clinical information, samples and genetic material from children with PANS and their families. The archived material will be a rich resource for future investigations of disease mechanism, treatment and prevention of acute-onset neuropsychiatric disorders in children. Work was conducted under clinical protocols NCT01281969, NCT03507218, NCT01778504.
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{
"pile_set_name": "NIH ExPorter"
}
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Most of the axial growth of the eye occurs during the first two years of life. While the growth of the eye is usually precisely regulated by a process known as emmetropization, this process can become deranged in certain conditions resulting in excessive or retarded axial growth and large refractive errors. This project seeks to elucidate the molecular mechanisms responsible for the physiological processes that cause aberrant ocular growth with aphakia. Specifically, we will assay infantile eyes for changes in certain neurotransmitters and growth factors after removing the crystalline lens which we have previously shown retards ocular growth. Understanding the mechanisms whereby aphakia alters ocular growth is of vital importance to accurately select the most appropriate intraocular lens power to implant into infantile eyes after cataract surgery. In addition, by understanding the mechanisms retarding ocular growth, it may be possible to modulate the excessive ocular gro wth which occurs in certain pathological conditions such as retinopathy of prematurity. FUNDING NIH EY08544 $156,904 9/30/98 - 1/31/02 PUBLICATIONS None P51RR00165-38 1/1/98 - 12/31/98 Yerkes Regional Primate Research Center
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{
"pile_set_name": "NIH ExPorter"
}
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Current practice among manufacturers of pulse oximeters is to calibrate an instrument prior to shipment according to a calibration curve based on measurements obtained from a group of human subjects. No means for checking the calibration of pulse oximeters in the clinic now exists. We are designing a device for calibrating pulse oximeters that simulates the optical properties of a pulsating capillary bed containing various fractions of oxygenated and deoxygenated hemoglobin. The design of the calibrator is based on liquid-crystal light modulators which are used to set the dc and modulated (ac) light intensities passing through the "capillary bed. " By adjusting the ac/dc intensity ratios in wavelength bands in the red (630-690nm) and infrared (800-1000nm) the display of the oximeter can be set to the desired oxygen saturation (SO2) value.
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{
"pile_set_name": "NIH ExPorter"
}
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In the intestinal tract, the first line of defense against pathogenic microorganisms is the layer of epithelial cells that line its extensive mucosal surface. These cells provide a physical barrier to infection and play critical roles at mucosal immune sites by detecting and eliminating enteric pathogens, including the human immunodeficiency virus (HIV) and Salmonella enterica. The intestinal mucosa contains, in addition to epithelial cells, an extensive nervous system capable of pre-programmed behavior, and it houses the largest lymphocyte population in the body;these three cell types communicate with each other through molecular signals which modulate inflammation and coordinate mucosal defense responses to infection. Opioid abuse is an important co-factor in host susceptibility to HIV and other mucosal pathogens, but its impact on mucosal host defense is relatively unknown. Opioid drugs may impair neuro-immuno-epithelial interactions at immune inductive and effector sites in the intestine. The proposed experiments will test the general hypotheses that opioids impair the immune responses of epithelial cells and lymphoid cells in the intestinal mucosa through actions mediated by neuronal and extraneuronal opioid receptors, and that opioid neuroimmune signaling is enhanced by mucosal inflammation. In Specific Aim 1, we will determine and compare the effects of opioids on proinflammatory cytokine release and changes in opioid receptor expression before and after inflammation in porcine intestinal epithelial cells in culture and from explants of Peyer's patch and absorptive mucosae through measurements of secreted cytokines and their mRNA expression. We will also assess opioid actions on intestinal epithelial cell wound healing by electric cell-substrate impedance sensing. In Specific Aim 2, we will compare the morphological interrelationships between opioid receptors and opioid peptides in epithelial cells, neurons and leukocytes of Peyer's patch and absorptive mucosae by immunocytochemical and in situ hybridization methods. We will also examine opioid effects on an epithelial-neuronal co-culture from porcine intestine through measurements of cytokine release and changes in neuronal excitability evoked by Salmonella infection. Finally, we will characterize changes in the ligand affinity and G protein coupling coupling of specific opioid binding sites in neural membranes from inflamed and uninflamed Peyer's patches by radioligand binding techniques. In Specific Aim 3, we will characterize delta-opioid receptors mediating chemokine-induced chemotaxis of intestinal T lymphocytes, and determine the effects of opioids on neurally-mediated secretory immunoglobulin-A release from intestinal mucosa explants. The results from this multi-faceted transdisciplinary project will provide a fundamental understanding of how opioid drugs act to alter mucosal defense function. Moreover, they will unveil new drug targets for the modulation of mucosal immune responses to HIV and other gut pathogens as well as oral vaccines offering protection from these microbes. PUBLIC HEALTH RELEVANCE The goal of the proposed transdisciplinary investigation is to elucidate the mechanisms by which opioid drugs of abuse disrupt the interplay between enteric neurons, epithelial cells and immunocytes at sites of mucosal immunity, which are key targets for infection by enteropathogens like S. typhimurium and the human immunodeficiency virus (HIV). Our experiments will provide new and significant information on the ability of opioids to influence mucosal responses evoked by Salmonella typhimurium, an enteroinvasive zoonotic pathogen and potential HIV vaccine vector. The results that we obtain in this multi-faceted project will advance our understanding of how opioid drugs act to alter mucosal defense function. In addition, they will reveal new drug targets for the modulation of mucosal immune responses to enteric pathogens including HIV, and provide new information contributing to the development of oral vaccines based on bacterial vectors that are designed to confer protection against HIV infection in gut-associated lymphoid tissue, the major body reservoir for this virus.
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{
"pile_set_name": "NIH ExPorter"
}
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Separation of reduced and permethylated oligosaccharides by gas chromatography can be facilitated by the use of a fused silica capillary column 100 meters long coated with methyl silicon. The presence of N-acetylhexosamines in oligosaccharides increases retention time and decreases efficiency of separation. Transamidation of hexosamines by trifluoroacetolysis followed by reduction, removal of O-trifluoroacetyl groups and permethylation dramatically reduces the retention time of hexosamine-containing oligosaccharides and permits useful separation of oligosaccharides containing up to six monosaccharide units, regardless of how many of these are hexosamines. The mass spectra of permethylated oligosaccharides with N-trifluoroacetylated amino sugars show unexpectedly high abundances of mass ions containing the N-trifluoroacetyl group. As many of these ions are large, they provide useful information regarding oligosaccharide structure.
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{
"pile_set_name": "NIH ExPorter"
}
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The goal of this work is to understand how bone adapts to its mechanical environment, and to further the validation and development of predictive computational models for strain-induced cortical bone remodeling. We hypothesize that bone formation on the cortical-endosteal surface can be reactivated when mechanical strains are elevated, causing bone formation within the marrow cavity; and that alterations of the mechanical environment act on the differentiation of cells rather than on individual cell vigor. These hypotheses will be tested both experimentally and analytically. The proposed research uses the ulnar osteotomy dog model we have developed. Eighteen skeletally mature (1-2 year old) beagle dogs will have 1-2 cm segment of the distal ulna removed to overload the intact radius. A separate group of eighteen dogs will have ulnar osteotomies, but the osteotomy gap will be spanned with a 5-hole metal fixation plate. This will test the effect of osteotomy without overload of the radius. A third group (n-18) will have sham operations. Following surgery, fluorochrome markers will be given to monitor bone formation. Force plate data will be collected before and after surgery. Two days prior to sacrifice, bone strains will be measured on the radius using three sets of rosette strain gages. Six animals in each group will be sacrificed at one, six, or 12- months. An additional six dogs per group will be used to measure acute strain changes after either an ulnar osteotomy alone, or after a plated osteotomy. Following sacrifice, undecalcified sections of bone from the radius and ulna will be prepared and histomorphometric measurements will quantify the tissue-level and cell-level adaptation of the bone to a change in strain. These data will then be used to relate Hart's computational model to bone cell activity by using cellular and tissue dynamics to define rules governing bone remodeling.
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{
"pile_set_name": "NIH ExPorter"
}
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DESCRIPTION: (Applicant's Description) Research on fibronectin and integrins has expanded greatly in the last 10-20 years. These molecules control many basic biological functions such as cell adhesion, determination of cell shape and differentiation, organ and tissue development, hematopoiesis, immune cell function, tumorigenicity and metastasis, cell migration, cytoskeletal organization, and organization of the extracellular matrix. More recently, it has become obvious that integrins collaborate with growth factors to regulate cell proliferation and apoptosis, and thus have a major involvement in cell signaling. In addition, several of the integrins, including Alpha-2b-beta-3 on platelets, alpha-5-beta-3 on tumor blood vessels, and alpha-4 integrins on immune cells, have become widely recognized as targets for therapeutic intervention. The overall importance of fibronectin and integrins is further emphasized by the results from several knockout mouse experiments. In many cases, null/null mutations have produced embryonic lethal or perinatal lethal phenotypes. The challenge for the cell adhesion field over the next several years will be to comprehend and organize the large amount of new structural information and design experiments based upon it; learn the relative biological importance of the several seemingly redundant adhesion systems; and apply the new insights to the treatment of human diseases. The 1997 Gordon Research Conference on "Fibronectin, Integrins, and Related Macromolecules" will serve as a valuable forum to present and discuss new information, identify problem areas, and plan new approaches.
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{
"pile_set_name": "NIH ExPorter"
}
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There is a current need in the clinical sciences for new technologies to rapidly diagnose cancers based on the detection of dysregulated molecular signatures. Abnormal expression of small metabolites involved in key steps of glucose transport, glutaminolysis, aerobic glycolysis and Krebs cycle, and larger metabolites such as fatty acids and complex lipids has been observed in various types of cancers. Moreover, abnormal metabolic patterns have been associated with specific genes linked to cancer prognosis. The MYC oncogene, for example, which is amplified in 55% of human hepatocellular carcinomas (HCC), is known to play a key role in the regulation of metabolic pathways and has also been associated with poor prognosis. Hence, the ability to rapidly and easily measure metabolites could provide a powerful approach for the clinical diagnosis and prognosis of cancers. New ambient ionization mass spectrometry imaging (MSI) techniques can perform direct analysis of tissue samples for in situ, near real time assessment of their molecular signatures. The goal of this proposal is to develop an ambient ionization MSI technique, desorption electrospray ionization (DESI-MSI), in conjunction with biostatistical tools to measure, define and validate metabolic signatures that are diagnostic and prognostic of human solid cancers, and to test this technology as a clinical tool for intrasurgical diagnosis of cancers. Application of DESI-MSI to analyze human cancerous tissue is a recent line of research developed in the last 6 years, in which I played a leading role during my PhD with Prof. R. Graham Cooks at Purdue University, and that I continue to develop in my postdoctoral research with Prof. Richard N. Zare at Stanford University. DESI-MSI allows hundreds of metabolites to be measured, imaged and accurately identified from an unmodified tissue sample in less than a second per pixel, in the open air, ambient environment. Although powerful, the DESI-MSI experiment is fairly simple: a spray of charged droplets extract metabolites from a sample surface, and are captured by a mass spectrometer for chemical analysis and identification. I believe this technology has the potential to transform the way cancer is diagnosed and treated in the clinical setting. The specific aims of my K99/R00 proposal are: 1. Develop DESI-MSI and refined statistical tools to identify and validate metabolic signatures diagnostic of a solid tumor, human HCC, 2. Investigate if certain metabolic patterns are related with a specific gene, the MYC oncogene, using the refined transgenic mouse models of MYC-induced HCC, 3. Evaluate DESI-MSI as a clinical tool for intrasurgical diagnosis and prognosis of HCC and other solid human tumors. While the initial aims of this proposal are focused on HCC, the developed methods will be applicable to study other human solid cancer, as it will be pursued in my independent phase, and thus have broad significance in human cancer diagnosis, prognosis and treatment. I have strong expertise in analytical chemistry and mass spectrometry, and a track-record of success in developing novel mass spectrometry tools for biological sample analysis. I have published 36 peer-reviewed manuscripts and have been honored to receive few awards for my research achievements, including the Nobel Laureate Signature Award of the American Chemical Society in recognition as 2012's best doctoral dissertation amongst all branches of Chemistry in the USA. However, while my prior research and training experiences in MS and translational research have enabled me to conduct the MS and clinical portions of collaborative research projects, through my interdisciplinary interactions as a postdoctoral researcher at Stanford University I recognized that my chemical training is not sufficient to conduct significant biomedical research as an independent researcher. Prior to engaging in a career as an independent investigator in cancer/biomedical research, I would greatly benefit from training in high-dimensional statistics methods to properly analyze and interpret mass spectral data of clinical samples, and in basic molecular biology methods for understanding cancer biology processes. Stanford University provides a spectacular environment to pursue the interdisciplinary project I propose and to receive training from the most outstanding researchers in these areas. As part of my career development, my mentorship and training will be provided by Prof. Richard N. Zare (Department of Chemistry), an innovator in methods of chemical analysis, Prof. Robert Tibshirani (Departments of Health Research & Policy, and Statistics), famous for the development of biostatistical methods for high-dimensional data analysis, and Prof. Dean Felsher (Department of Medicine, Division of Oncology), a pioneer in the development of MYC-induced transgenic mouse models of cancers. The training will be achieved through experimental work and also formal course work. My long-term career goal as an independent researcher is to develop novel MS technology for clinical diagnosis and prognosis of human cancers. As an independent researcher, I will apply my expertise in MS, and the training in biostatistics and molecular biology that I will receive through the K99 period to develop new, automated MS tools for clinical and intrasurgical diagnosis and prognosis of various human cancers, and to translate this technology to the clinics. I have a particular interest in using MS for assessing cancer margins during surgical resection, procedure for which new and rapid diagnostic methods are greatly needed. Aims 1 and 2 will be performed during the K99 mentored phase, and aim 3 will be pilot for HCC and much further explored in the R00 independent phase for other solid human cancers. The K99/R00 award will support my development into an independent investigator who develops relevant novel mass spectrometry tool in combination with biostatistical methods for clinical diagnosis and prognosis of human cancers.
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{
"pile_set_name": "NIH ExPorter"
}
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Evaluate the efficacy of a number of retinoids of differing chemical structures to prevent the development of bladder cancer during its preneoplastic period using the FANFT model for bladder cancer induction. Determine effects of retinoids on the bladder and on tumor development by using the following parameters: bladder weight; number of tumors per bladder detected grossly; percent of bladder having hyperplasia, papilloma, and carcinomas (staging of urinary bladder pathology will be based on the World Health Organization); and recording of other pathological lesions, including other tumors. Monitor FANFT in the feed or ANFT in the urine by quantitation of Salmonella typhimurium revertants produced by urine of experimental animals. Monitor blood and tissue levels of retinoids in experimental animals.
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{
"pile_set_name": "NIH ExPorter"
}
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The nematode C. elegans has powerful genetics, a well-described nervous system, and a complete genome sequence;thus, it is well suited to analysis of behavior and development at the molecular and cellular levels. In particular, the ability to functionally map the influence of particular genes to specific behavioral consequences makes it possible to use genetic analysis to functionally dissect the molecular mechanisms underlying poorly understood aspects of nervous system function. However, many genes with critical roles in neuronal function have effects on behavior that to a casual observer appear very subtle or difficult to describe precisely. Therefore, to fully realize the potential of C. elegans for the genetic analysis of nervous system function, it is necessary to develop sophisticated methods for the rapid and consistent quantitation of mutant phenotypes, especially those related to behavior. The goal of this proposed work is to develop computer vision tools for quantitatively characterizing the phenotypic patterns caused by mutations or pharmacological treatments in C. elegans. By making it possible to precisely characterize the behavioral phenotypes of mutants with abnormal locomotion or egglaying, these tools will be particularly useful for correlating specific neurotransmission defects with characteristic behavioral patterns. These analytical tools will also be used to generate a comprehensive database containing complex behavioral data on a large set of mutant strains. This database will make it possible to identify groups of mutants and pharmacological treatments that have similar effects on behavior or development. With the accumulation of increasing phenotypic data on known mutants, it should ultimately be possible to record from unknown mutant or drug-treated animals and make informed initial hypotheses about the functions of uncharacterized genes and the targets of uncharacterized drugs.
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{
"pile_set_name": "NIH ExPorter"
}
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Staphylococcus aureus is the second leading cause of hospital-acquired bloodstream infections. In 2003, nearly 300,000 patients in US hospitals acquired an S. aureus infection, which accounted for nearly three million days of hospitalization, $9.5 billion in excess costs, and at least 12,000 inpatient deaths. Additionally, approximately 50% of the US population either persistently or intermittently carries a biofilm of S. aureus in their nose without experiencing any symptoms. Despite the lack of symptoms, nasal carriage of S. aureus is a well-known risk factor for developing a hospital-associated staphylococcal infection. Nasal carriers of S. aureus are three- to six-times more likely to acquire an S. aureus infection in the hospital, and, approximately 80% of the time, that infection is caused by the strain of S. aureus that lives in their nose. Based on infection rates and the prevalence of S. aureus nasal carriers, it has been estimated that eradicating S. aureus from the noses of surgical patients could save $4.5 billion a year in US hospital costs. The only FDA-approved treatment for nasal decolonization of S. aureus is 2% mupirocin. Clinical studies have shown that nasal decolonization decreases the number of infections contracted by S. aureus carriers, but resistance to mupirocin is becoming a concern. In vitro, the concentration of mupirocin required to inhibit growth of S. aureus increases significantly after only one exposure, and there is a growing body of clinical evidence linking mupirocin resistance to unsuccessful nasal decolonization. Nitric oxide, a free radical gas naturally produced by the human body to help fight infection, is a promising alternative to mupirocin for nasal decolonization of S. aureus. In recent years, the use of nitric oxide as an antimicrobial agent has been investigated due to its broad-spectrum antimicrobial activity, ability to diffuse directly through cell membranes, and the belief that it would be very difficult for nitric oxide-resistant strains of bacteria to develop. Additionally, nitric oxide has been shown to be effective against biofilm embedded bacteria. The goal of this SBIR project is to use Novan's proprietary nitric oxide-releasing technology to develop a topical nitric oxide-releasing nasal product capable of eradicating S. aureus biofilms from the noses of carriers without breeding bacterial resistance. In this proposal, Novan aims to: 1) Establish the minimum concentration of nitric oxide-releasing silica (Nitricil(tm)) required for bactericidal activity against 90% of S. aureus clinical isolates under planktonic and biofilm growth conditions in vitro;2) Determine whether S. aureus can develop stable resistance to nitric oxide;and 3) Evaluate the safety of Nitricil(tm) particles for use in the nose. Upon achieving these aims, Phase II efforts will focus on preclinical evaluation of intranasal Nitricil(tm) formulations, including: 1) testing the lead formulations against a mouse model of S. aureus nasal colonization;2) full preclinical toxicology;and 3) the quality scale-up manufacturing of both drug substance and drug product required to submit an Investigational New Drug application to the FDA. PUBLIC HEALTH RELEVANCE: Staphylococcus aureus infections cost US hospitals approximately $9.5 billion per year. Nasal carriage of S. aureus is a major risk factor for contracting such an infection, and estimates project that treatment to remove these bacteria from the noses of carriers could save $4.5 billion annually. The only FDA-approved treatment for nasal decolonization of S. aureus is mupirocin, but the prevalence of mupirocin-resistant S. aureus is increasing and is associated with treatment failure. The goal of this SBIR project is to use Novan's proprietary nitric oxide-releasing technology to develop a topical nasal product capable of eradicating S. aureus biofilms from the noses of carriers without breeding bacterial resistance.
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{
"pile_set_name": "NIH ExPorter"
}
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We have been developing assays to detect oxidative lesions in specific genes and thus to quantitate their formation and repair. We generate oxidative DNA damage is generated by several different approaches including hydrogen peroxide, X-irradiation, irradiation with methylene blue, and treatment with 4NQO which forms at least one adduct with oxidative characteristics. Our main approach is to treat cells in culture with acridine orange, which after activation with light forms oxidative lesions in DNA. The main lesion is 8-OH guanosine which can be detected by use of the FaPy glycosylase. This enzyme creates strand breaks in DNA at sites of the lesions, and the single stranded DNA can then be resolved on alkaline gels. We find that 8-OH guanosine is rapidly repaired in active genes in hamster and human cells. While it has been a general notion that there is no DNA repair in mitochondria, we now find that these organelles do have repair capacity. They can not, however, repair all lesions. They are capable of repairing DNA lesions created by monofunctional alkylating agents, but not UV induced pyrimidine dimers. We find fast repair of oxidative damage in mitochondrial DNA, and the mechanism is under investigation. One question is whether the repair in mitochondrial DNA is transcription coupled. We are investigating whether the common deletions in mitochondrial DNA seen in senescence and other conditions could be due to a localized deficiency in DNA repair.
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{
"pile_set_name": "NIH ExPorter"
}
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Iron absorption is being studied in animal models to ascertain transport mechanisms. Studies are designed to integrate kinetic data with techniques aimed at isolating and studying components in the transport system. Guinea pigs treated in ways that will alter iron absorption constitute the basic animal model. Components of the purported transport system are separated using standard chromatographic and electrophoretic techniques.
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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. Structural information of HIV-1 env spike will significantly enhance our ability to design efficacious immunogens, which might be vaccine candidates for AIDS.
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{
"pile_set_name": "NIH ExPorter"
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The purpose of this Core is to provide the infrastructure for the computational efforts of the research- in this program project grant. The development of NMR spectroscopy is intimately linked to high-performance computation. Recent developments make data acquisition faster, and multidimensional NMR experiments can be acquired at higher resolution. Thus, more and larger data sets are to be handled. This causes problems with data transfer, processing and analysis.This Core will establish the requisite high-performance infrastructure, working towards the following specific aims. Aim 1: Establish and maintain a 128 cpu cluster for data processing and modeling Aim 2: Implement and maintain a core library of parallel applications Aim 3: Maintain and further develop efficient processing and data analysis software Aim 4: Installation and maintenance of additional third party software relevant for NMR-related research Aim 5: Training and dissemination
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{
"pile_set_name": "NIH ExPorter"
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The objective of this project is to uncover the molecular mechanisms of genetic rearrangements. The transposition reaction of bacteriophage Mu is studied as a model system. Critical steps in Mu transposition are a pair of DNA cleavages and strand transfers involving the ends of Mu DNA sequence and a target DNA; these reactions generate a branched DNA intermediate. The two chemical reaction steps take place within higher order protein-DNA complexes called transpososomes, the core of which is composed of two Mu-end DNA segments synapsed by a tetramer of MuA transposase protein. Transpososome assembly is controlled by a number of cofactors: an enhancer type DNA sequence element called IAS that overlaps the Mu operator sequence and the Mu repressor that binds to it, the MuB protein, the E. coli-encoded HU and IHF proteins, ATP, and Mg++. By making use of a simplified transpososome assembly reaction system, we have shown that both the Mu end DNA cleavage and the subsequent strand transfer at one Mu DNA end are catalyzed by the MuA monomer that is bound to the partner Mu DNA end within a transpososome. For Tn10 and Mu transposition, two transposase monomers within the transpososome have been shown to catalyze all the chemical steps at the two transposon ends. By using chiral phosphorothioate containing DNA substrates, we compared the orientation of the substrate engagement at the transposase active site for the successive reaction steps in a number of transposition reactions. MuB ATPase controls each of the early steps of Mu DNA transposition: it assists transpososome assembly, is involved in the target DNA site selection, activates the MuA transposase for strand transfer reaction, and protects transpososome from premature disassembly by ClpX chaperon protein until strand transfer is completed and the transposition intermediate is ready for DNA replication by the host replication proteins. In turn, the functional state of MuB is controlled by the ATPase cycle and by its interaction with MuA. Structural and functional aspects of MuB-DNA complex are currently under investigation by using a variety of physical and biochemical techniques. The molecular interactions involved in the transposition complex of phage Mu were studied by using fluorescence labeled proteins and DNA. Tools have been developed for the assay of transposase-DNA binding, Mu-end pairing, stable synaptic complex formation, and Mu-end DNA deformation. The assembly and disassembly of protein-DNA complexes, and also the conformational changes within the complex during the reaction are studied in real time. Techniques and instruments are currently under development to study these reactions at the single molecule level by using a powerful fluorescence microscope. Efforts are continued toward solving the high-resolution structure of the higher order protein-DNA complexes involved in transposition reactions, in collaboration with scientists at the University of Chicago.
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{
"pile_set_name": "NIH ExPorter"
}
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The objectives of this program are to increase the reproductive-endocrine-genetic data base of nondomesticated, wildlife species. Program strategies emphasize investigation of basic reproductive-endocrine-genetic factors which appear as the most critical prerequisites to the application of either artificial insemination or embryo transfer. The program employes a multidisciplinary approach targeted toward female and male reproduction genetics. The use of suitable, domestic animal models permits testing the efficacy of research concepts before adaptation to rarer, nondomestic species. Areas of effort in the female include: hormonal evaluation for correlation of endocrine profiles of pituitary-ovarian functions and as an index of the influence of manipulative stress; ovulation induction through the administration of exogenous gonadotropins to optimize the timing of the ovulatory event; and embryo collection, culture, freezing and transfer as techniques for cryobanking genetic stock, improving reproductive potential and eventually allowing microinjection of molecularly cloned genes which participate in transformation and inborn errors. Particular emphasis has been applied to the colection, in vitro cultivation, viable freezing, and micromanipulation of embryos of mouse, cat and miniature swine, which hold much promise as animal models. Areas of effort in the male include: semen collection and evaluation to characterize ejaculate norms and correlate these findings to the level of genetic polymorphism in wildlife populations; semen handling and cryopreservation to increase spermatozoal viability and establish optimal methods for chronic storage of genetic material; and hormonal evaluation to improve the understanding of pituitary-gonadal-adrenal relationships with particular emphasis on the marked differences in stress responses among taxonomically related wildlife species. Together, these generalized research concepts not only allow rapid expansion of physiological-genetic norms for any given rare species, but also improve methods of assessing fertility potential or genetic status to optimize managemet efforts for selective natural propagation.
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{
"pile_set_name": "NIH ExPorter"
}
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The focus of this proposed project is to develop the novel technologies necessary for exploiting RNA-binding proteins (RBPs) to comprehensively identify regulatory elements present in expressed human RNA. Eukaryotic organisms depend on the actions of RBPs for successful gene expression, providing not only a link between transcriptional and translational regulation, but also playing essential roles in established regulatory processes including transcription, splicing, export, stability and translation. Understanding posttranscriptional gene regulation and identifying the RNA regulatory elements responsible for this regulation is an important aspect of fulfilling the ENCODE project objectives. We previously developed methods for purifying endogenous mRNA-protein complexes (mRNPs) and identifying the associated mRNA targets using genomic array technologies, a method we termed ribonomics. Our studies have demonstrated that, analogous to transcriptional regulation, groups of mRNAs are coordinately regulated in a combinatorial manner by distinct classes of RBPs at the post-transcriptional level and that RBPs can coordinate the regulation of groups of mRNAs by targeting related regulatory elements located in the transcripts. Presently, ribonomic technology only identifies mRNA targets of RBPs. New high-throughput technologies will need to be developed for the exhaustive identification and mapping of RBP binding sites that function as regulatory elements. Newly developed tiling array technology represents a tremendous breakthrough for the potential identification of regulatory elements in expressed RNA. This new array platform represents a significant advance in microarray technology that utilizes contiguous probe sets to interrogate entire genomic sequences in exhaustive detail. The aim of this project is to develop novel tiling array based ribonomic profiling methods including high-throughput RBP foot-printing, which will in turn, facilitate the comprehensive mapping of RNA regulatory elements. [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
}
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The molecular characterization of neural cell surface antigens during development may give some clue as to their importance for cell recognition and interaction in the nervous system. Cell type specific antigens will be characterized as to their molecular nature. Their differential distribution on particular cell types will be determined using cell biological and immunohistological methods. Cell surface antigen expression will be analyzed at different developmental stages and their functional significance be probed for. Cell type specific antibodies will be used to separate cerebellar cells and cell hybrids using a fluorescence activated cell sorter and affinity chromatography with antibody columns.
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{
"pile_set_name": "NIH ExPorter"
}
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This proposal focuses on the development of an algorithm for determining the underlying factors responsible for predisposition to or protection from polygenic diseases. The algorithm relies on Markov chain Monte Carlo exploration of the space of possible genetic variants coupled to a Bayesian statistical test based on phenotypic ranks. The developed algorithm will improve our ability to reduce complex genetic interactions from the growing genotypic and single nucleotide polymorphism databases. The identification of interacting genetic variants placing individuals at risk for or providing protection from the development of polygenic diseases remains a problem both for our understanding of these diseases and for our ability to develop new treatments. Fundamentally, the problem involves the inability of standard statistical approaches to achieve power in the face of the enormous growth in our knowledge of genomics, brought about by the various genome projects and high throughput single nucleotide polymorphism (SNP) and genotype analyses. Similar "curse of dimensionality" problems have arisen in other fields, and Bayesian statistical approaches coupled to Markov chain Monte Carlo (MCMC) techniques have led to significant improvements in understanding, which has led to our focus on this technique here. Because polygenic diseases are much more widespread than single gene diseases, the potential impact on health is substantial, and many common diseases are believed to have a polygenic basis, including obesity, cardiac disease, and Type II diabetes. A method to dissect the complex genetic interactions underlying predisposition to or protection from polygenic diseases would have a substantial effect on improving health. We will disseminate the algorithm through publications and presentations at [unreadable] conferences. We will also through contact individuals in foundations focused on research in specific complex diseases, so that the algorithm can have the maximal impact on health. Upon successful completion of this project, we plan to develop the algorithm more fully. We would like to modularize the algorithm, allowing easier inclusion of different prior distributions and implement a more friendly interface with the ability to utilize the emerging bioinformatics standards for data exchange. [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
}
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In each IVF cycle, a decision must be made as to which embryo(s) will be selected for transfer. This decision has a far reaching impact on the outcome of an IVF cycle, namely whether the embryo will develop into a healthy child. It is estimated that at least 50% of human embryos are affected by chromosomal abnormalities such as aneuploidy, and implantation of such embryos can lead to undesired outcomes such as failed implantation, spontaneous abortion, or birth of a trisomic offspring. Reproductive specialists have been increasingly turning to pre-implantation genetic diagnosis (PGD) in efforts to identify embryos with the best chance of developing into healthy children. However, current techniques are expensive, unreliable and typically test only a small selection of chromosomes. GSN has developed an innovative technology termed Parental SupportTM (PS) whose output is an in silico reconstruction of the embryonic DNA at thousands of loci with confidence exceeding 99%. This technology will, for the first time, allow IVF physicians to screen embryos for chromosomal abnormalities including aneuploidy, translocations and deletions across all 23 pairs of chromosomes with an error rate below 0.1%. The Phase I objective of this application is to integrate our PS technology with a new, highly parallelized custom Infinium-based genotyping platform to dramatically reduce costs that will, in turn, enable GSN to offer PGD service with superior accuracy, scope and at a cost equivalent to current, less reliable FISH methods. The new customized platform will then be applied in Phase II where we propose to evaluate the concordance between a new trophectoderm biopsy technique on day 5, traditional blastomere biopsy on day 3, and the actual child. The results from these studies will allow us to assess the value of the new biopsy technique, evaluate the largely unstudied phenomenon of embryo self-correction between day 3 and day 5, and provide IVF physicians with powerful and far-reaching knowledge about the developmental potential of each embryo.
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{
"pile_set_name": "NIH ExPorter"
}
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The Administrative Core will be responsible for financial and scientific aspects of the grant, including administrative and secretarial services for the individual projects and Clinical Core, as follows: a) administration of the Program Project Grant with respect to Federal and University regulations; b) disbursement of funds; c) annual reports; d) facilitation of progress and completion of all projects, coordination of individual projects, and coordination between projects; e) organization of regular meetings between investigators In addition, the Administrative Core will interact with the Clinical Core personnel to ensure seamless provision of data to the Project P.l.s and to facilitate protection of the safety of research subjects. New to the PPG are establishment of both an Internal Advisory Committee and a Data Safety Monitoring Board. Both will meet on a regular basis to evaluate the progress and to assure safety and efficacy of the individual projects. The DSMB will report their conclusions to the Project P.I., the Administrative Core personnel, and to the Internal Advisory Committee. Of note, for a number of reasons (protection of patient privacy, quality control, prevention of duplication of effort) access to the database is limited. We have clearly defined (our IRB approved) policies for access to the database. The database has security measures so that there are different types of access no access, read only, data entry only, report writing (see attached manual). The Administrative and Clinical Core oversee access to the database and protection of safety of research subject. RELEVANCE (See instructions):
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{
"pile_set_name": "NIH ExPorter"
}
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Traditionally, the accurate description of the anatomic alterations and physiological sequellae of congenital cardiac malformations has depended upon cardiac catheteriztion and selective angiocardiography. It must be recognized that these techniques are associated with substantial risk in the neonatal period and lesser, although still significant risk, in small children. Thus, the development of accurate, non-invasive methods to assist in the diagnosis and management of heart disease is of particular importance in the pediatric age group. The specific aim of this proposal is to develop and refine three non-invasive modalities whose combined applications may be expected to significantly improve the clinical aproach to the infant or child with heart disease. The techniques to be employed include scintillation scanning of the heart and lungs, phased multiple crystal echocardiography, and pulsed Doppler echocardiography. Scintillation scanning will be employed primarily to assess the magnitude of intercirculatory shunting. The goal of the ultrasonic instrument development program will be to assemble an ultrasonic echographic system utilizing, in combination, the technologies of multi- scan echocardiography, pulse Doppler velocity indication, phased array beam steering, and moving target indicator techniques, in order to provide a high resolution image of internal soft tissue structures. Individually, each of these ultrasound approaches has been demonstrated to have advantage over traditional, single element, mechanically scanned pulse echo techniques for cardiac visualization. It is our conviction that these new approaches used in combination, and exploiting the unique advantages of each, will provide a significantly improved image of cardiac and vascular structures and thus, enhance the capability for atraumatic description of cardiovascular morphology and function in infants and children. It should be recognized that infants and children are especially well suited for ultrasound examinations since considerations of patient size, bone density, and acoustical attenuation all serve to enhance diagnostic accuracy.
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{
"pile_set_name": "NIH ExPorter"
}
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Sarcoidosis is a heterogeneous, granulomatous disease of world-wide prevalence, most commonly involving the lung, skin, lymph node and eyes. Although the etiology is unknown, molecular and immunologic investigations of sarcoidosis specimens suggest that host interactions with infectious antigens, particularly mycobacterial antigens, contribute to pathogenesis among some subjects. In addition, it has been recently demonstrated that broad-spectrum antimycobactehal therapy (CLEAR regimen) provides clinical improvement among pulmonary sarcoidosis subjects without fibrosis, but not among those with advanced fibrotic disease. It is unknown if distinctions in the microbial community exist among heterogeneous sarcoidosis populations. In a similar fashion, an association has been observed between disease severity among patients with Alpha-1-antitrypsin (AAT) deficiency and pathogenic bacteria. Alpha-1 antitrypsin (AAT) is a potent antiprotease with activity against neutrophil elastase (NE). AAT deficiency is associated with a greater neutrophil load, higher elastase activity, leukotriene-B(4) concentration, and serum protein leak than matched patients without deficiency; the resultant airways inflammation with neutrophil recruitment and elastase release is positively correlated with colonizing bacterial load. Microbiome analysis of both sarcoidosis and AAT specimens will enhance understanding of the contribution infectious agents provide to disease severity. Sarcoidosis is also characterized by cellular anergy; we observed improvement in sarcoidosis T cell biologic function among some sarcoidosis subject who completed this antimycobacterial regimen, suggesting the microbial virulence factors may contribute to sarcoidosis T cell anergy. We hypothesize the distinctions in microbiota correlate with disease severity among sarcoidosis and AAT populations, and that the microorganisms present possess the capacity to induce T cell anergy. We propose to define the microbiota within heterogeneous sarcoidosis and AAT populations, and to determine the effects of the microbiome on T cell biologic function. RELEVANCE: Sarcoidosis and AAT are important medical problems. The strength of this proposal is microbial characterization according to disease severity among heterogeneous sarcoidosis and AAT cohorts. This proposal will also enhance understanding of microbial induction of T cell anergy, for which there is a lack of appreciation in idiopathic lung diseases.
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{
"pile_set_name": "NIH ExPorter"
}
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Although protective armor and acute medical intervention allow soldiers to survive explosions, a growing number of veterans will have disability stemming from blast-related brain damage. Soldiers also return from combat with psychological disabilities caused by traumatic war events. The clinical presentations of individuals with blast-related brain damage and post-traumatic psychopathology markedly overlap and thus a clear description of the direct consequences of explosive blast is complicated by the emotional and cognitive sequelae of psychological trauma. The inability to clearly demonstrate the basis of presenting symptomatology impedes the development and implementation of effective treatments for traumatic brain injury (TBI). Objective tests that characterize the essential features of neural damage in blast-related brain injury are therefore required to advance clinical care. To date, very few studies have specificall investigated neural injury from blast. In a pilot investigation of Operation Iraqi Freedom and Operation Enduring Freedom (OIF/OEF) troops we found individuals with blast-related TBI (bTBI) exhibited diminished electroencephalography (EEG) phase synchronization between lateral frontal scalp sites of the two cerebral hemispheres (Sponheim et al., Neuroimage, 2011). Diminished synchrony was not accounted for by post-deployment psychopathology and was associated with the structural integrity of white matter tracts in the frontal lobes of bTBI subjecs. We are also near completion of a Department of Defense funded study investigating the use of EEG and Diffusion Tensor Imaging (DTI) measures to assess the functional and structural neural consequences of bTBI in a National Guard OIF/OEF sample. To date we have studied over 150 individuals divided into bTBI, bTBI+PTSD, PTSD, and control groups and found evidence of diffuse white matter abnormalities associated with bTBI. The question that the current proposal addresses is whether measures of neural structure (e.g., DTI) and function (EEG) have clinical utility in differentiating the effects of bTBI from post-traumatic psychopathology in veterans referred to a Regional VA Polytrauma Center. Hence, the goal of the proposed project is to validate measures of neural function and structure as means of separating the effects of bTBI from post-traumatic psychopathology in veterans being seen at a VA Polytrauma Center for evaluation. Specific Aim 1. We will determine whether EEG and DTI indices differentiate between bTBI and post- traumatic psychopathology in veterans who have screened positive for TBI and were referred to a Regional VA Polytrauma Center. Analyses will include testing for differential effects of bTBI and impact TBI (iTBI) on neural indices. We hypothesize that compared to individuals with only post-traumatic psychopathology, individuals with a bTBI will exhibit diminished EEG phase synchronization between brain regions and a greater percentage of white matter voxels with low fractional anisotropy (FA) as measured by DTI. Specific Aim 2. Given variability in the outcome of individuals affected by mild TBI (either blast or impact) we will determine how chronic EEG and white matter abnormalities predict long-term functioning in veterans who screened positive for TBI and were referred to a Regional VA Polytrauma Center. We hypothesize that greater abnormalities on EEG and DTI indices will be associated with functional impairment in subjects with mild TBI. Specific Aim 3. Because the adaptation of the brain to damage varies across individuals, we will carry out analyses to determine whether risk genes for neural injury mediate the association between the number and type (blast, impact) of TBI with the presence of chronic EEG and DTI abnormalities. We hypothesize that the ApoE 4 allele will be associated with greater abnormalities on neural indices in individuals who have sustained a mild TBI.
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{
"pile_set_name": "NIH ExPorter"
}
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All viruses tightly regulate cellular processes to create an environment supportive for replication. An understanding of the mechanisms viruses use to do this is vital for our knowledge of viral pathogenesis as well as the normal cellular controls of these pathways. Human cytomegalovirus (HCMV) is a large and slow-growing herpesvirus which must maintain the host cell in a metabolically and translationally active state and circumvent the inhibitory effects of cellular stress responses induced during infection. HCMV induces intracellular reactive oxygen species (ROS) generation soon after infection to facilitate its own gene expression. However, overproduction of ROS can lead to oxidative stress, and it is unclear how the virus controls the detrimental effects of ROS on cell growth pathways. The specific aims of this application are to (1) characterize mitochondrial respiratory chain activity and ROS generation in infected cells throughout a viral time course, (2) examine the effects of oxidative stress on cell growth pathways during infection, and (3) determine the mechanisms that confer resistance to oxidative stress in HCMV infected cells. The Research Design and Methods for this project involve identifying the basal respiration rate and ROS content of cells throughout an HCMV infection. Oxygen consumption is measured using a dark-type oxygen electrode and ROS levels are examined by microscopy and flow cytometry using several different indicators. Next, we will examine the ability of HCMV to resist the effects of oxidative stress. Infected cells are treated with exogenous ROS and cellular signals normally inhibited by ROS will be examined by Western. Finally, pathways important for the maintenance of redox homeostasis, including the antioxidant activities of p53 and Nrf2 and the proteasomal and autophagy pathways, will be examined in HCMV infected cells to identify the mechanisms responsible for protection from oxidative stress. The study of the generation and control of oxidative stress during HCMV infection may facilitate the development of effective therapeutic strategies against this and other viruses. Additionally, these studies may provide valuable insight into key cellular regulatory mechanisms essential for redox homeostasis and their perturbation in disease states.
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{
"pile_set_name": "NIH ExPorter"
}
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This study aims to determine whether usual blood pressure control is different from low blood pressure control in slowing the progression of hypertension-induced renal disease in African-Americans. The second objective is to determine whether a calcium channel blocker, an ACE inhibitor or a beta blocker are different in their effects to slow progression of hypertension-induced renal disease in African-Americans.
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{
"pile_set_name": "NIH ExPorter"
}
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In most eucaryotic genes, a common set of nucleotides has been found before the start site of RNA transcription. These nucleotides, referred to as either a TATA or Goldberg-Hogness box, are important determinants of transcription in vivo and in vitro. The control region for the late SV40 transcripts have been examined by site specific mutagenesis and by generation of deletion mutations. Mutations that enhance or suppress the transcriptional activity of a single start site have been identified and differ from the consensus Goldberg-Hogness box. An additional cluster of nucleotides that is located at base positions 75-96 also control the level of transcription. It maintains this enhancing effect when its position relative to the 5 foot start site is altered.
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{
"pile_set_name": "NIH ExPorter"
}
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We propose to facilitate the understanding of protein functions and metabolic processes in living species, by developing a computer system for completely automated prediction of biochemical or biological functions for large sets of protein sequences. Instead of the conventional approaches that typically rely on one round of database search and inherit the annotation from the best-scoring sequence match, we will construct a proprietary structured database of complete proteomes and develop an automated multistep strategy of database searches, feature prediction, and annotation. The cascade of analysis will include: selection of the reference databases, flexible filtering, domain dissection, iterative searches, motif analysis, and rule-based modification of the database annotations. The system to support this approach will be capable of analyzing one protein every five minutes, which is approximately a tenfold increase over the productivity of a trained analyst. In addition, custom heuristic algorithms will result in 20-30 percent greater accuracy in annotations compared to the existing automated approaches. The rapid and accurate prediction of protein functions at the industrial scale established in the proposed project will be applied for exhaustive annotation of newly sequenced proteomes and EST collections, in order to reconstruct in detail biochemical pathways targeted for modification in various organisms. PROPOSED COMMERCIAL APPLICATION: The development of the genome-scale functional annotation of protein sequences at high speed and accuracy will accelerate gene discovery, target validation and metabolism reconstruction. This will provide for rapid and reliable development of human therapeutic molecules, antimicrobials, agricultural products, and industrial enzymes. Many new functional predictions for proteins from different species will be made and the value-added sequence databases will become available.
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{
"pile_set_name": "NIH ExPorter"
}
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Introns are removed from eukaryotic mRNA precursors by a large, complex, and dynamic molecular machine known as the spliceosome. The fully assembled spliceosome consists of five small nuclear ribonucleoprotein particles (snRNPs) each containing a single small nuclear RNA (U1, U2, U4, U5, or U6 snRNA) together with a number of snRNP proteins. In collaboration with as many as 100 auxiliary proteins, the snRNPs assemble stepwise onto the mRNA precursor, and then participate in a changing network of interactions that catalyze the two chemical steps of the splicing reaction, release the spliced mRNA and excised intron, and prepare the five snRNPs for reuse. A major experimental problem in analyzing this complicated multistep process has been to identify reaction intermediates. Following the lead of Konarska, we have used oligonucleotides resembling splice sites, and antisense 2'-0-methyl oligonucleotides directed against certain snRNA sequences, to efficiently induce the assembly of spliceosome-like snRNP complexes in mammalian nuclear extracts. These artificially induced spliceosome-like complexes appear to freeze a variety of transient spliceosomal interactions (snRNP/snRNP, snRNP/substrate) that take place naturally over the course of an mRNA splicing reaction. Using this approach, we have identified two novel complexes: a U1/U4/U5 complex that appears to be an intermediate in displacement of U1 from the 5' splice site, and a U2/5' SS complex in which U2 bind the 5' exon in much the same way as the EBS2 element of a Group II intron binds the 5' exon sequence known as IBS2. We propose to characterize each of these complexes biochemically, to identify the sequence of events leading to formation of the complexes, and to develop assays to explore the role of these new RNA/RNA pairings in vivo. The experiments should shed new light on how the initial U4/U6 pairing comes undone early in spliceosome assembly; how U1 binding to the 5' splice site is replaced by U2, U5, and U6; how the active site is assembled from U2 and U6; and how the 5' exon is retained by U2 and U5 after the first chemical step of splicing, and aligned for ligation in the second step.
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{
"pile_set_name": "NIH ExPorter"
}
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PROJECT SUMMARY Rates of anxiety and depression in youth are substantial, causing a major unmet need for effective interventions. Although some progress has been made in preventing these internalizing problems in adolescents, further research is needed that specifically targets theoretically and empirically supported risk processes. An important and salient risk factor found to increase the likelihood of anxiety and depression is negative affectivity ? a partially heritable trait propensity to experience and express more frequent, intense, and enduring aversive emotional states. The proposed randomized controlled prevention trial builds on our finding from our longitudinal study that elevated levels of negative affectivity during adolescence prospectively predicted internalizing disorders in early adulthood (Zinbarg et al., 2016); moreover, this relation was mediated by changes in momentary negative affect (mNA) measured with ecological momentary assessment (EMA) (Adam et al., 2018). The first phase (R61) of the proposed selective prevention trial will test whether an app-based, coach-supported mindfulness intervention as compared to an assessment-only control reduces momentary negative affect, measured with ecological momentary assessment (EMA), in 120 adolescents (age 12-16) at- risk based on their having high levels of trait negative affectivity. EMA will be used to measure average daily mood, (the ?Target?) collected six times a day across three days at pre-, mid-, and post- intervention. ?Target? engagement will be defined as a medium effect size (>.40) in the comparison of youth randomized to MBI versus control on the target ? momentary negative affect ? at post-test, adjusting for pre-test levels. We also will assess the dose-response relation by testing the association between number of sessions and exercises completed with changes in momentary negative affect and weekly mood ratings. In the second phase (R33), we will conduct a replication trial with a new sample of 360 at-risk (i.e., high trait negative affectivity) youths (ages 12-16) randomized to one of three conditions ? MBI, a nonspecific control, or an assessment-only control. Youth will be evaluated with regard to the target (i.e., mNA), internalizing symptoms and disorders, and functioning (e.g., social, academic) at baseline and post-intervention (R61 and R33), and at a 6-month follow-up (R33). Finally, in the R33 we will test if significant reductions in momentary negative affect are associated with improvements (or less worsening) in internalizing symptoms and fewer onsets of internalizing disorders.
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{
"pile_set_name": "NIH ExPorter"
}
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Currently approved medications for the treatment of alcohol dependence are not overwhelmingly effective. Therefore, researchers continue to look for better medications to treat this serious illness. One approach to improve medication efficacy is to target specific medications for treatment resistant subpopulations such as Type B alcoholism, as described by Babor. Type B alcoholics are characterized by an early onset of alcohol problems, greater severity of dependence, and a poor treatment response. In contrast, Type A alcoholics are characterized by a later onset of problem drinking, less severe alcohol dependence, fewer alcohol-related problems, and a better response to treatment. We conducted a double-blind, placebo-controlled, pilot trial, involving 61 alcoholics (32 type A and 29 Type B) treated with quetiapine, 400 mg daily, or placebo. Significantly more quetiapine-treated patients remained abstinent during the trial, compared to placebotreated patients (31% vs. 6%). Quetiapine was significantly more effective in reducing drinking in Type B patients. In Type A patients, both the quetiapine and placebo treated patients significantly reduced their drinking so no effect of quetiapine was detected. Upon further analysis, we found that the characteristic most highly predictive of a good response to quetiapine was frequent heavy drinking as measured by a high average number of drinks per drinking day and a greater number of days of drinking to intoxication in the 30 days prior to the trial. It is likely that the Type A patients in our pilot trial represented a less severely addicted subgroup of alcoholics and this explains why they responded equally well to placebo or quetiapine. Quetiapine may be more effective in frequent heavy drinkers because of associated abnormalities of serotonergic and dopaminergic neurotransmission. Atypical antipsychotics target both the dopamine and the serotonin systems and have shown efficacy in reducing alcohol use in alcoholics with comorbid psychiatric illness. Quetiapine is an atypical antipsychotic that has a favorable side effect profile. The proposed 5-year project is intended to confirm and extend our initial findings regarding quetiapine for the treatment of frequent heavy drinkers in a double-blind, placebo-controlled, 13-week trial. The study will compare quetiapine (400 mg daily) to placebo in 180 DSM-IV alcohol dependent patients who are frequent heavy drinkers as determined by averaging more than 12 drinks per drinking day and having 15 or more days of drinking to intoxication in the thirty days prior to entering the trial. The primary hypotheses are that quetiapine-treated patients will have 1) more abstinent days from alcohol and 2) fewer heavy drinking days compared to placebo-treated patients as measured by the timeline follow back. Exploratory analyses will be conducted to further investigate other possible quetiapine / alcohol subtype interactions
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{
"pile_set_name": "NIH ExPorter"
}
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Retinoblastoma (RB) is the most common intraocular malignancy of childhood. Enucleation (eye removal) is effective treatment, but vision-preserving approaches, such as radiation and chemotherapy have become the standard of care. Despite these advances in treatment, many survivors of RB develop metastatic disease or secondary tumors. We propose that there are subpopulation(s) of RB cells that escape standard treatments to form new tumors. The cancer stem cell theory suggests that only a small percentage of cells within a tumor are capable of driving tumor growth and metastasis, whereas the majority of a tumor is actually made up of non-tumorigenic cells. Tumors recur because a small number of chemo-resistant tumor-initiating cells survive to repopulate the tumor. At present, there is a gap in knowledge regarding the existence and potential role of tumor-initiating cells in RB. Our central hypothesis is that RB tumors contain subpopulation(s) of tumor-initiating, chemo-resistant "cancer stem cells" that promote tumor progression. We base this hypothesis on our previous findings of heterogeneity in both RB tumors and cell lines, in which subpopulations of RB cells display a number of stem cell characteristics: the ability to exclude Hoechst 33342 dye as a "side population" (SP), expression of ABGC2 (a stem cell/chemo drug resistance marker), as well as immunoreactivity to human stem cell markers such as Oct3/4, Nanog and Musashi-1 (genes of self-renewal and pluripotency). Our overall goal is to isolate and characterize subpopulations of tumor-initiating RB cells via the following methods: With access to live human RB cells from over 200 tumors, we will identify potential tumor- initiating cells using fluorescence activated cell sorting based the following criteria: 1) the expression of cell surface markers that are expressed by tumor-initiating cells in other tissues;such as ABCG2, 2) the ability to efflux the fluorescent dye, Hoechst 33342 and 3) the expression of genes involved in stem cell renewal/pluripotency such as Oct3/4, Nanog and Musashi-1. We will assess specific features of these subpopulations that correspond with functional attributes of cancer stem cells, such as self-renewal and slow cell cycling. We will then compare the relative ability of cell populations with and without these properties to form tumors in immunodeficient NOD-SCID mice. We will identify potential new therapeutic targets in RB by comparing gene expression profiles of cell populations enriched for tumor-initiating cells to their non-tumor forming counterparts. Results from these studies will define the tumor-initiating phenotype(s) in RB, and assess their potential as novel therapeutic targets in retinoblastoma, as well as other types of malignancies. PUBLIC HEALTH RELEVANCE: Retinoblastoma, the most common intraocular tumor of childhood, appears to contain a small population of cells that behave as "cancer stem cells". The goal of our project is to identify and target cancer stem cells in retinoblastoma to show that they are primarily responsible for tumor progression and resistance to chemotherapy drugs. Results from our study will lead to new treatment strategies for retinoblastoma and other tumors that contain these small populations of cancer stem cells.
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{
"pile_set_name": "NIH ExPorter"
}
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The dorsal striatum is a brain region crucial for expression of habit and goal-directed behaviors that has recently been linked to addiction. I found that ex vivo or in vivo ethanol exposure and withdrawal causes a long-lasting increase in the activity of the NMDA receptors (NMDARs) that contain NR2B subunits (NR2B- NMDARs) in the dorsomedial striatum (DMS), and that inhibition of NR2B-NMDARs specifically within the DMS attenuates rat operant ethanol self-administration (Wang et al., J Neurosci 2007; Wang et al., J Neurosci 2010). NMDAR-mediated glutamatergic transmission is crucial for synaptic plasticity and learning and memory, as well as for expression of some goal-directed behaviors. The DMS receives a number of different glutamatergic inputs, including corticostriatal (CortSt) and amygdalostriatal (AmygSt), and also contains two groups of principal neurons: dopamine D1 or D2 receptor-expressing medium spiny neurons (D1 or D2 MSNs, respectively). Very little is known about the glutamatergic neurotransmission in ethanol-exposed animals at these different inputs and cell types. Thus, the goal of this application is to elucidate how different afferent glutamatergic inputs in distinct types of neurons within the DMS are altered by excessive ethanol intake, with the long-term goal of determining how such alterations contribute to ethanol-drinking behaviors. My hypothesis, based in part on my preliminary results, is that NR2B-NMDAR-dependent glutamatergic transmission and synaptic plasticity in different striatal circuits (CortSt vs. AmygSt afferents and D1 vs. D2 MSNs) are differentially regulated by excessive ethanol consumption, and that ethanol-related adaptations are predominantly expressed in AmygSt inputs and in D1 MSNs. This hypothesis will be tested by pursuing the following aims: 1) to determine whether excessive ethanol consumption causes a differential increase in the CortSt and AmygSt glutamatergic transmission in the DMS of rats, 2) to determine whether AmygSt transmission facilitates induction of NMDAR-dependent long-term potentiation (LTP) at CortSt synapses in the DMS of naive and ethanol-drinking rats, and 3) to determine whether excessive ethanol consumption alters glutamatergic transmission differentially onto D1 or D2 MSNs of transgenic mice where D1 or D2 MSNs express a fluorescent marker. This application is highly innovative because it applies state-of-the- art approaches including the combination of optogenetics and transgenic mice, allowing us for the first time to determine how ethanol regulates synaptic transmission in different afferent projections to the DMS and different cell types within the DMS, which are important questions that cannot be addressed using conventional methodologies. Knowledge generated from this proposal will greatly facilitate our understanding of the mechanisms by which ethanol-mediated neuroadaptations in glutamatergic transmission are central to promoting pathological ethanol-drinking behaviors.
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{
"pile_set_name": "NIH ExPorter"
}
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The long-term objective of this proposal, as in the previous project period, is to study placental angiogenesis (development of the placental blood supply), and its role in supporting utero-placental and fetal growth. The mammalian placenta is the organ through which nutrients, gases and wastes are exchanged between the maternal and fetal systems. Thus, size of the placental blood supply has a profound influence on the rate of transplacental exchange, and thereby on the rate of fetal growth. Factors affecting fetal development, in turn, have a significant influence on postnatal survival and growth. The specific goals of the proposed research are to isolate and characterize angiogenic factor(s) produced by maternal and fetal placental tissues. Based upon preliminary evidence indicating that endometrial tissues of ewes secrete heparin-binding angiogenic factor(s), immunoblotting and immunohistochemical techniques also will be used to evaluate expression of heparin-binding growth factors by placental tissues. To accomplish these goals, placental tissues will be obtained from ewes throughout early gestation (preattachment through definitive implantation stages; Exp. 1) and mid-late gestation (periods of maximal and then decreasing rates of placental growth; Exp. 2). An additional study (Exp. 3) is designed to examine in vivo effects of heparin-binding angiogenic factors on uterine angiogenesis. Ewes will be utilized in the proposed experiments because ontogeny of placental production of angiogenic factor(s) was well characterized during the previous grant period, and, in addition, placental vascular function has been well studied in this species. Additionally, ruminant placentas are comprised of highly vascular structures (placentomes) consisting of maternal caruncles and fetal cotyledons, as well as interplacentomal areas consisting of intercaruncular endometrium and intercotyledonary chorioallantois. This placental arrangement provides a unique opportunity to study differences not only between maternal and fetal portions but also between functionally specialized portions of the placenta. Knowledge of factors which influence placental vascular development should lead to a better understanding of how fetal and maternal placental tissues interact to support fetal growth. In addition, because placental vascular development is one of the few instances of angiogenesis in a non-pathological situation, isolation and characterization of angiogenic factors produced by placental tissues will be important not only for gaining a better understanding of placental function but also for understanding regulation of angiogenic processes in general.
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{
"pile_set_name": "NIH ExPorter"
}
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The Center for Analysis & Synthesis of Marcomolecules (CASM) at Stony Brook requests funding to acquire a robotic workstation for proteomics to process gel plugs for analysis by mass spectrometry. The institution has acquired an ABI Voyager research-grade MALDI mass spectrometer and a Micromass QToF tandem mass spectrometer to support proteomics research at Stony Brook. The present application seeks support for the acquisition of a Micromass MassPREP workstation to provide an automated, 96-well format digestion and spotting/sampling of gel plugs for detailed mass spectrometry. There are five major users who need the high-throughput capacity to enable their NIH-funded research. Dr. Danial Bogenhagen (Pharmacology) will employ the instrumentation in mitochondrial proteomics, an area of high interest, seeking to define the full complement of proteins that act in maintenance of mtDNA. Dr. Wen-Tien Chen (Medicine-Oncology) requires the instrumentation to enable his work on the proteomics of invasive cell protrusions involved in angiogenesis and metastasis. Dr. Malbon (Pharmacology) is using proteomics to deduce the composition of large macromolecular signaling complexes for G-protein-coupled receptors as well as in beta-catenin/Lef-Tcf pathways. Todd Miller (Physiology) requires this instrumentation in his proteomic analysis of tyrosine kinases, focusing upon the SH3-dependent partners of Hck and interacting protein partners for Jak2. William Van Nostrand (Medicine-Vascular Biology) is probing the processing of amyloid beta-protein precursor as well as the identification of new protein-protein interactions using proetomics. All of these investigators have published research based in protein chemistry and in mass spectrometry. To optimize the use of the instrumentation, the new workstation will be located in CASM, a protein core facility with more than a decade of experience. Three FTEs have been recruited to manage the mass spectrometry employed in proteomics. The institutional commitment includes the on-going support for these 3 FTEs as well as provision of funds to acquire a complementary automated gel scanner/spot picker jointly by the School of Medicine and the Vice-President for Research. The research proposals are high quality, well-funded by the NIH, and will benefit by the addition of this instrumentation. The institution has provided resources to enable the acquisition of two new mass spectrometers and this proposal builds on the investment.
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{
"pile_set_name": "NIH ExPorter"
}
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Mechanical forces play key roles throughout biology, from governing the adhesion of leukocytes in the immune response, to determining cell fate and tissue development. This emergent field of mechanobiology is providing vital insights into diseases such as bleeding disorders, cancer, and infectious diseases, where it is becoming clear that conventional biochemical and genomic characterizations are not sufficient to understand the rich behavior of living systems or how they fail. Rather, we must uncover how force can change the structure and function of molecules, and trigger mechanotransduction pathways to modify cell responses. Technological developments that enable precise manipulation of single molecules and cells (e.g. optical tweezers and AFM) have been a driving force in the development of the field. However, growth of the field is impeded by limited access to such technologies as they can be expensive, technically challenging, and low- throughput. These challenges have also limited the types of scientific questions that can be addressed. To overcome these challenges, we will develop high-throughput and accessible new approaches in mechanobiology that will (i) open up new areas of study through the introduction of new capabilities, and (ii) democratize single-molecule force measurements so that all biomedical researchers can make discoveries using these powerful tools. For example, we will accelerate single-molecule measurements by building upon an instrument that almost all biomedical researchers already have: the benchtop centrifuge. By developing a miniature microscope that fits into a standard centrifuge bucket, we will create an accessible and inexpensive benchtop instrument that will bring high-throughput single-molecule manipulation to non-specialists, offering a 1000 fold efficiency boost and 10-100 fold cost improvement over many other methods. We will also develop self-assembled DNA nanoscale devices that facilitate single-molecule studies of population heterogeneity, and that enable instrument-free force spectroscopy. Significantly, these projects will open the fields of mechano- biology and single-molecule manipulation to new researchers and systems, accelerating the pace of discovery. Additionally, we will apply our single-molecule approaches to answer key open questions in mechanobiology regarding (i) the mechanical regulation of hemostasis, (ii) adhesion molecules in the immune response, and (iii) mechanotransduction and the molecular basis for hearing and deafness. For example, we will perform massively-parallel force measurements using single-molecule centrifugation to study force-regulated enzymatic cleavage of von Willebrand factor, and investigate mutations related to von Willebrand Disease, the most common inheritable bleeding disorder. We will also study cellular adhesion of leukocytes, and investigate the molecular basis of hearing and deafness. Overall, these efforts should firmly establish force as a key parameter for understanding the basic processes of life, and provide a new handle for both understanding? and treating?disease.
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{
"pile_set_name": "NIH ExPorter"
}
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Low-dose Tamoxifen in Hodgkin Lymphoma Survivors for Breast Cancer Risk Reduction Mantle radiation has been a cornerstone of HL treatment;however, female survivors of HL treated with mantle irradiation before age 30 have a 20- to 55-fold increased risk of developing breast cancer (BC) - a risk that is comparable to that of BRCA mutation carrierrs Surgical prophylaxis is very effective in reducing the risk of BC, but such invasive strategies are not suitable for all women. Pharmacologic interventions exist, but only tamoxifen is approved for use in young women who have not yet reached menopause. Standard-dose tamoxifen (20 mg daily) is associated with undesirable side effects, but recent studies have laid convincing groundwork that tamoxifen at lower doses may be similarly efficacious in reducing BC risk with fewer side effects. We hypothesize that tamoxifen administered at a lower dose (5 mg daily) would be both an efficacious and safe non-surgical risk reduction intervention for female adult survivors of HL diagnosed during childhood or as a young adult. Thus, using a Phase IIb randomized, double-blind, placebo-controlled trial of low-dose tamoxifen (5 mg daily) in long-term female HL survivors treated with chest radiation, we aim to 1) Determine the impact of a two-year course of low-dose tamoxifen on well-established surrogate biomarkers of chemopreventive efficacy;2) Establish the safety and tolerability of low-dose tamoxifen in this population;and, as an exploratory aim, 3) Examine the modifying effect of several well-defined demographic and clinical characteristics associated with radiation-related BC risk on the risk:benefit ratio from this intervention. Eligible subjects who provide informed consent will be randomized to 5 mg per day of tamoxifen versus placebo for two years. Outcomes will include several surrogate biomarkers of efficacy, including mammographic breast density (MBD, primary endpoint), breast cytomorphologic and proliferation measures, and insulin growth factors. Subjects will be carefully followed for safety and tolerability using patient-reported outcomes as well as lipid profiles, clotting factors, and markers of bone turnover as objective endpoints. Risk modifiers that will be examined include age, menopausal status, prior hormone use, body mass index, personal history of benign breast disease, and family history of cancer, as well as chest radiation dose, age at exposure, and latency from chest radiation. A sample size of 127 per arm will be able to detect a 20% reduction in MBD with low-dose tamoxifen relative to placebo with 80% power. We have identified over 900 potentially eligible subjects within our consortium of five institutions that have well-developed infrastructure to follow childhood cancer survivors long-term, thus demonstrating that we will have a sufficiently sized pool to draw the eligible patient population from and complete the study. At completion of this study, we hope to identify a well-tolerated risk reduction option for HL survivors that are at high risk for developing BC. Low-dose Tamoxifen in Hodgkin Lymphoma Survivors for Breast Cancer Risk Reduction Public Health Relevance: Survival from Hodgkin lymphoma (HL) is excellent, but chest radiotherapy (RT) has been a cornerstone of treatment, and women with HL exposed to chest RT when they are young have a 20- to 55-fold increased risk of developing breast cancer (BC). Tamoxifen reduces the risk of BC by 50%, but at the cost of some undesirable side effects, while more recent studies suggest that lower dose tamoxifen may be similarly efficacious in reducing BC risk with fewer side effects. We believe that tamoxifen administered at 5 mg daily would be an ideal non-surgical risk reduction intervention for female HL survivors exposed to chest RT at high risk for BC;therefore, we plan to test this hypothesis in a Phase IIb clinical trial.
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{
"pile_set_name": "NIH ExPorter"
}
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Our group is interested in understanding the role of cytokines in the development of autoimmune diseases. This year we made progress in understanding old observations as well as discovering new, exciting mechanisms in B cell biology. Specifically, are studying the role of tumor necrosis factor (TNF) in both autoimmune models of lupus as well as in human autoimmune disease. We now understand why mice that are deficient in TNF develop anti-nuclear autoantibodies (ANA), but very rarely develop kidney disease. My students, Rachel Robbins and Carolyn Karafiath found that although loss of TNF drives autoantibody production that deposit in kidneys, no lymphocytic inflammation occurs due to the loss of TNF (which is a pro-inflammatory molecule). These data have important implications in the treatment of rheumatic diseases with drugs that inhibit TNF?s actions. Recently, we have turned our attention to a newly discovered cytokine, termed ?IL-21?. This protein is secreted only by activated CD4 T cells, and known to be important in T cell and NK cell activity. We have found that it also plays a critical role in B cell maturation and development. We have turned our attention to it?s possible role in autoimmune disease by studying how it may be involved in animal models of lupus as well as in human systemic lupus erythematosus and Rheumatoid arthritis. Lastly, Anna-Marie Fairhurst, a post-doctoral fellow, is investing the role of the Toll-like receptors (TLR) in the development in various murine models of SLE as well as in human B cell biology. She has found that TLR9 is differentially expressed on distinct human B cell populations which may suggest a role for this molecule in controlling B cell maturation and have important implications in human disease. Using mice deficient in TLR9 expression she is studying its possible role in controlling B cell maturation and development as well as its role in autoimmune disease. Our group is dedicated to understanding how cytokines are contributing to the development of systemic lupus erythematosus and rheumatic diseases. We feel that our research will lead to new therapeutics as well as to understanding the mechanisms of basic B cell biology.
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{
"pile_set_name": "NIH ExPorter"
}
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Project Abstract and Summary Falls and their consequences are among the major problems in the medical care of older individuals. With advancing age, the capacity to maintain balance after perturbations deteriorates due to a number of age- related sensorimotor deficits, and likely increases the risk for falls. The unexpected nature of falls triggers startle-like whole body postural responses. Startle responses are characterized by exaggerated whole body postural responses with increased muscle co-activity causing co-contraction during the first trial response (FTR) and normally diminish with repeated exposure due to behavioral habituation. Because all falls involve a downward motion of the body with gravity which may trigger a startle response, understanding whether these responses are also superimposed onto the landing response will be important to determine. Our central hypothesis is that age-related abnormalities of exaggerated startle responses and habituation will influence the initial landing response to a sudden loss of the ground support surface but can adapt due to the modulatory effects of motor prediction and participant awareness. Understanding whether the FTR during sudden drop perturbations influences the landing response will be important to determine whether or not it enhances balance recovery or is problematic and precipitates falls. Moreover, identifying whether the effects of age- associated abnormalities on landing movements during FTR can be modulated using motor prediction and participant awareness will help direct future interventions aimed at fall prevention. Whole body postural muscle activation patterns, movement kinematics, and landing impact forces will be assessed by electromyographic (EMG) recordings, motion capture, and force platform recordings. The specific aims are 1) Compare changes in landing responses to unexpected and expected drop perturbations in relation to age during a) FTRs and b) subsequent trials; and 2) Determine the modulatory effects of repeated expected drop perturbations on reducing FTR magnitude evoked by unexpected drop perturbations in young and older adults. Expected outcomes are that abnormal neurophysiological mechanisms causing excessive startle contributions to postural FTRs with older age will be linked with changes in landing control mechanisms. Moreover, demonstrating the capacity to adapt landing responses through behavioral habituation will direct future interventions aimed at improving landing strategies to attenuate the impact of falls.
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{
"pile_set_name": "NIH ExPorter"
}
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The long-term objectives of this work are to determine how sperm movement is regulated and how flagellar movement patterns affect the ability of sperm to reach the site of fertilization and penetrate the oocyte. We have focussed our efforts on a movement pattern termed hyperactivation, because it has been observed to occur in the oviduct near the site of fertilization. Our three specific aims at this point are as follows: (1) Investigate the functions of hyperactivation by comparing the ability of activated (the movement pattern of sperm in the ejaculate) and hyperactivated sperm to generate force for pulling away from the oviductal wall and for penetrating the zona pellucida. Pulling forces will be compared by determining the relative force required to hold activated and hyperactivated sperm onto suction micropipets. Zona penetration will be tested by comparing the rates at which acrosome- reacting activated and hyperactivated sperm penetrate zonae in vitro. (2) Characterize the mechanism that raises intracellular free Ca2+ ([Ca2+]in) to induce hyperactivation. Based on our observations, we hypothesize that a different mechanism regulates the calcium influx associated with hyperactivation than that associated with the acrosome reaction. Calcium channels can be characterized by their response to various organic agents and ions. We have the ability to estimate concentrations of [Ca2+]in in various regions of individual, moving sperm. These techniques can be used to determine whether an agent actually affects [Ca2+]in while it is affecting motility. (3) Determine whether levels of [Ca2+]in in sperm flagella may be regulated by the oviduct or zona pellucida. We have observed that hyperactivation is initiated in sperm that are attached to the oviductal wall and that only hyperactivated sperm flagella may be regulated by the oviduct or zona pellucida. We have observed that hyperactivation is initiated in sperm that are attached to the oviductal wall and that only hyperactivated sperm free themselves from the wall. It has also been reported that the pattern of flagellar beating alters when sperm bind to the zona pellucida. We hypothesize that elements in the oviductal wall and/or zona regulate sperm flagellar beating by affecting [Ca2+]in. [Ca2+]in will be measured in individual sperm as they attach to oviductal epithelium and to the zona pellucida. The information gained may be used to treat infertility, to monitor the effects of toxins and disease on fertility, and to develop methods of contraception that act prior to fertilization.
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{
"pile_set_name": "NIH ExPorter"
}
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Technical Abstract The incidence rate of pelvic fractures increases dramatically with age, from 5.4 and 3.8 per 10,000 person- years in women and men aged 65 to 69 years to 93.5 and 44.5 per 10,000 person-years in women and men aged 90 years and older, respectively.3 Pelvic fractures are accompanied by severe pain, chronic immobility and loss of function and independence in the elderly.14 Pelvic fractures consume substantial healthcare resources, and based on administrative claims data, they are one of the most costly osteoporosis related fractures.4 Un-healed fractures, occurring in one-third of pelvic fracture patients at 3 months,16 can cause continued pain and impact mobility. With aging of the population, and expected concomitant increase in the incidence of pelvic fractures, there is a pressing need to find effective treatments that will accelerate healing. Fracture of the pubic ramus is most relevant and practical for randomized double-blinded placebo controlled study as this fracture is accompanied by severe pain and immobility in elderly, is associated with delayed fracture healing, and is almost always treated non-operatively. The current standard of care for pelvic fractures includes pain management, patient mobilization, and the prevention of complications associated with comorbid conditions. We hypothesize that development of a successful adjunctive therapy to accelerate fracture healing would lead to improved care and reduce both direct and indirect costs from pelvic fractures. In the proposed trial we will recruit women and men >65 years of age with acute osteoporosis-related pelvic fractures and address 3 specific aims over 3 months of treatment in a placebo controlled double blind study to determine if standard care and teriparatide 20 mcg/day versus placebo for pelvic fractures: 1. Results in earlier evidence of cortical bridging on routine radiographs followed by confirmatory Focus CT, a novel method to reduce radiation exposure from CT scans (primary outcome). 2. Leads to a faster reduction in pain as assessed by both the Numeric Rating Scale and a reduction in the use of narcotics (secondary outcome). 3. Leads more rapidly to improved functional outcome using a short physical performance battery to assess lower extremity function (secondary outcome). We will extend this study with 9 months of open label TPTD to determine if any potential differences between the placebo and TPTD groups during the 3 months of treatment are evident and persist over time, even in patients who use TPTD after the three month placebo controlled intervention. If TPTD can improve fracture healing, this study will have an impact on the treatment of persons with pelvic fracture who are not surgical candidates and often face severe pain, chronic immobility, and loss of function in the elderly. A positive finding of accelerated healing of pelvic fractures would also encourage study of TPTD for treatment of other osteoporotic fractures.
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{
"pile_set_name": "NIH ExPorter"
}
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Alterations of the tumor suppressor p53 are the most commonly identified mutations in human cancers. In responses to various stresses, p53 protein level and its activity are greatly induced. However, the mechanism how p53 responses to various stresses are activated largely remains unclear. Recent studies have suggested that various phosphorylation events of p53 might regulate p53 stability and activity. However, the physiological roles of these phosphorylation events of p53 in regulating p53 responses to DNA damage and other stresses remain to be determined. To address this issue, I propose to employ homologous recombination and LoxP-Cre-mediated deletion to introduce missense mutations (Ser/Thr to Ala mutation) at several potentially important p53 phosphorylation sites, including Ser18 and Thr73/83, into the endogenous p53 in mice. Preliminary analysis of the p53(Ser18Ala) and p53(Thr73/83Ala) primary cells suggested that both phosphorylation events play important but distinct roles in regulating p53 stability and activity after DNA damage. The mechanism for the impaired p53 responses to DNA damage in these p53 knock-in mice and the effects of these mutations on the p53-dependent tumor suppression will be determined. In addition, employing the same approach, we will determine the potential functional redundancy between phosphorylation of p53 at Ser18 and ser23 in regulating p53 responses to DNA damage. Phosphorylation of human p53 at Ser46 has been suggested an important role in regulating p53 apoptotic function. However, Ser46 of human p53 is not conserved in mouse p53. Therefore, a human p53 knock-in mouse model, in which exons 4-9 of mouse p53 gene was replaced with exons 4-9 of human p53 gene, will be used to address the physiological roles of this phosphorylation event. Two observations indicated the feasibility of this strategy. First, the humanized p53 is functionally equivalent to the endogenous mouse p53. Secondly, the DNA damage-induced signaling pathways leading to the phosphorylation of human p53 at Ser46 is conserved in mouse cells. Identification of the phosphorylation events that regulate p53 stability and activity will indicate the signaling pathways involved and thus reveal the mechanism how p53 responses are activated during various stresses or cellular senescence.
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{
"pile_set_name": "NIH ExPorter"
}
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Ethanol produces a well recognized spectrum of neurologic sequelae following acute and chronic exposure. The development of tolerance/dependence to this drug, together with the consequent neurodegenerative effects of chronic ethanol abuse pose a major public health concern. Recent studies have documented that ethanol can modulate the expression of specific gene products. This might be the ultimate mechanism in cellular adaptation to chronic ethanol exposure. This proposal describes a comprehensive biochemical/molecular genetic approach to the study of ethanol effects on gene expression in the NG108-15 glioma x neuroblastoma cell line. This system has been well characterized as a model system for studying the biochemistry of ethanol effects on neurons. The goal of this study is the identification and isolation of ethanol responsive genes (ERGs). Initial experiments will extend current studies with Northern blot analysis of specific marker genes that have been shown to respond (induction and inhibition) to chronic ethanol. The effects of ethanol on NG108-15 cell mRNA will also be studied by 2-dimensional gel analysis of ## vitro translation products. These studies will then be extended to include a detailed analysis of protein synthesis in cells exposed to acute or chronic ethanol. Finally, molecular cloning will be used to isolate ERGs. Differential and/or subtractive hybridization screening will be used to select genes that are preferentially modulated by ethanol. The identification and isolation of ERGs may provide powerful molecular probes for the study of ethanol tolerance/dependency both experimental models and in humans. Finally, the cloning of specific ERGs may enable a molecular study of the genetics of alcoholism.
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{
"pile_set_name": "NIH ExPorter"
}
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The peroxisome proliferator activated receptor gamma (PPARgamma) is a transcription factor that plays a pivotal role in adipogenesis and adipocyte gene regulation. The overall goal of this proposal is to define the molecular mechanisms by which differences in PPARg activity, caused by different human PPARg mutations, impact on lipid, glucose, energy metabolism, cardiovascular function, and longevity. Human genetic studies have identified several natural PPARgamma variants (Pro12Ala, Pro115Gln, Pro495Leu and Val290Met) that affect both ligand-dependent (AF-2) and -independent (AF-1) activation functions of the receptor and that span a wide spectrum of PPARgamma activity. Unfortunately, human studies that address the relationship between differences in PPARgamma activity and the above complex traits have produced unequivocal answers, because known confounders, such as age, gender, genetic background and environmental factors such as diet, and exercise, obscure the in vivo effects of genetic variants that affect such complex traits. Therefore, we propose to study the impact of these mutations in mouse models where genetic and environmental factors can be well-controlled permitting careful analysis of gene-gene and gene-environment interactions. The specific aims of this proposal are: (1) to generate mouse models on two distinct genetic backgrounds for the human PPARgamma Pro12Ala, Pro115Gln and Pro495Leu mutations, thus spanning a wide range of activity for this transcription factor; (2) to assess the impact of these muations on: i/lipid, glucose, and energy metabolism, ii/ cardiovascular function; and iii/ longevity; and (3) to determine the molecular changes induced by these mutations with the goal to identify the regulatory pathways that underpin the pleiotropic physiological effects of PPARgamma. Particular attention will be given to the impact of various dietary and exercise regimens on PPARgamma activity. The comprehensive nature and scientific breadth covered by this research requires a multidisciplinary approach, a condition that is met by the collaboration of specialists in cardiovascular and metabolic facets of mouse physiology and molecular genetics. Understanding the physiological and molecular impact of these PPARg muations may ultimately not only give us a better knowledge of fundamental PPARg activity, but may also point the way to the discovery of improved PPARgamma therapeutics.
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{
"pile_set_name": "NIH ExPorter"
}
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The problem of proliferative diabetic retinopathy is addressed in experimental studies utilizing several types of neovascularization. Special attention will be given to the model of retinal neovascularization in the young beagle. Major emphasis is directed toward the development of effective inhibitors of this retinal neovascularization that develops following oxygen exposure to young, pure strain beagles. The beagle model will be used primarily because the retinal neovascularization at the disc and elsewhere in the retina of classical human diabetic retinopathy. The proliferative lesions can be precisely localized at the disc or elsewhere in the fundus and develop in 100% of the animals exposed. Preliminary studies suggest that adult rabbit vitreous contains an inhibitor of neovascularization. Further documentation of this inhibition will be done and extended to a comparison with other species. Other possible inhibitors of neovascularization, e.g., cartilage extract, will be tested in the above beagle model of retinal neovascularization after preliminary testing in the more economical and simpler assays, such as tissue culture and the corneal preparations to identify the most suitable materials to develop for future evaluation in selected patients with proliferative diabetic retinopathy.
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{
"pile_set_name": "NIH ExPorter"
}
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The etiologies of dental caries and periodontal diseases are closely associated with dental plaque, the biofilm that develops from microbial colonization of the tooth surface. Colonization is initiated by a limited number of gram positive species, primarily viridans group streptococci and actinomyces. These bacteria typically occur in vivo as members of a complex microbial community, the formation of which depends on an array of specific adhesive interactions. Interactions between different bacteria, detected by in vitro coaggregation, are thought to play an important role in biofilm development. Coaggregations between actinomyces and streptococci or between different streptococci generally depend on GalNAc- or Gal-sensitive adhesins present on the actinomycete or on certain streptococci and complementary receptor molecules present on other strains of at least four viridans group streptococcal species. The adhesins of actinomyces are associated with the type 2 fimbriae of these bacteria and the receptors of streptococci with a family of structurally related cell wall polysaccharides. Type 2 fimbriae interact with either of two host-like disaccharide motifs that occur within the oligosaccharide repeating units of different streptococcal receptor polysaccharides. These host-like motifs are relatively non-immunogenic and consequently, the antigenicity of receptor polysaccharides is more closely correlated with other features of the polysaccharide chain. Current studies are directed toward defining the genetic basis of receptor polysaccharide structure and function. Genes for polysaccharide biosynthesis have been identified at four distinct loci of the streptococcal chromosome. Information gained from these studies should contribute to the development of molecular strategies to further assess the recognition role of streptococcal receptor polysaccharides in oral biofilm formation and hopefully, improved approaches for the prevention of plaque-related diseases.
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{
"pile_set_name": "NIH ExPorter"
}
|
Chronic Obstructive Pulmonary Disease (COPD) is now the 3rd leading cause of death in the United States, and the only cause of death that is on the rise. Smoking is the most common risk factor for COPD, however thereareotherenvironmentaltriggerssuchasdust,smoke,sand,oraerosols.NearlyoneintenVApatients arediagnosedwithCOPDandthecostsassociatedwiththesepatientsaretentimeshigherthanforallother conditions. Despite the prevalence of COPD, our understanding of the immune mechanisms that drive developmentofthediseaseisincomplete.ThereisincreasingevidencethatcytotoxicCD8+Tcellsarecritical tothepathologyofCOPD.WorkdonebythePIandcollaboratorshasidentifiedmucosalassociatedinvariant T(MAIT)cellsasan?innate?-likelung-residentCD8+Tcellpopulationcapableofrecognizingairwayepithelial cellsinfectedwithCOPD-associatedlungpathogens.Althoughlowerairwaybacterialinfectionsareassociated with COPD and are correlated with airway function, the mechanisms by which infections develop and contribute to COPD pathogenesis is not yet clear. Because CD8+ T cells recognize and destroy target cells infected with pathogens, lung-resident CD8+ T cells like MAIT cells may be important for containment of bacterialinfections.Infact,wefindthatMAITcells,whicharehighlyenrichedinhumanairways,havereduced frequency in the peripheral blood of COPD patients. Furthermore, we find that MAIT cells, while capable of controllingthegrowthofbacteria,areimpairedintheirabilitytomakeIFN-g?inresponsetobacteriallyinfected airway epithelial cells from patients with COPD. The effect of COPD on MAIT cell function is not known and may be essential to understanding how lower respiratory infections contribute to the development of COPD andCOPDexacerbations. ThisproposalisfocusedondeterminingthemechanismsbywhichCOPDaltersthefrequencyandfunctionof MAITcells,particularlywithregardtorecognitionofbacteriallyinfectedairwayepithelialcells.Additionally,we willdeterminetheconsequencesofalteredMAITcellactivationonbacterialsurvivalandinflammation. ThisprojectcontainstwoAims: Aim 1. Define the frequency and functional capacity of MAIT cells among CD8+ T cells from COPD lungs. Aim 2. Define the mechanisms underlying MAIT cell-mediated killing of bacteria, and how these mechanismsaremodulatedinCOPD.
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{
"pile_set_name": "NIH ExPorter"
}
|
We are studying methods that may facilitate direct gene transfer into muscle. Intramuscular injections of plasmid DNA has been shown to express transgenes at relatively efficient levels. Experiments are currently underway to define the mechanism of DNA uptake by muscle cells. We would like to use the confocal microscope to examine the efficiency of nuclear uptake of DNA under various conditions after cytoplasmic microinjection. The confocal n-dcroscope is necessary to determine the exact location of injected DNA, i.e. inside or outside of the nucleus. We will also be testing other techniques for gene introduction such as digitonin permeabilization and in vitro transfection.
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{
"pile_set_name": "NIH ExPorter"
}
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We shall attempt to determine the biologic importance of a humoral immunosuppressive factor(s) found in cancer serum and cancer ascites in the pathogenesis of cancer. We shall isolate this suppressive factor from the protein present in the ascites of human cancer patients. We shall use the isolated macroglobulin factor to produce a radioimmunoassay for quantitative measurement of this factor, and for identification and determining relationship to other suppressive factors. This antisera will also be used for further purification using immunosorbent columns, and, potentially, for treatment of patients. We shall attempt to find antagonists for this factor, utilizing extracorporeal immunosorbent removal. We shall begin to use this substance to facilitate transplantation especially bone marrow transplants used for cancer treatments.
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{
"pile_set_name": "NIH ExPorter"
}
|
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