text
stringlengths 1
32.7k
| meta
dict |
|---|---|
Rationale: Glutamate Carboxypeptidase II (GCPII; EC 3.4.17.21) was identified as an enzyme that catalyzes in brain the hydrolysis of N-acetyl- aspartyl-glutamate (NAAG) to acetyl-aspartate (NAA) and glutamate with 200 nM Km for NAAG. Given the role of NAAG as an endogenous antagonist at NMDA receptors and an agonist at mGluR3 receptors in brain reduced GCPII activity would be predicted to attenuate NMDA receptor. Since the dissociative anaesthetics, phenylcyclidine and ketamine, produce the symptoms of schizophrenia by antagonizing the NMDA receptor, NAAG could serve as endogenous psychotogen. Indeed, post-mortem neurochemical studies indicated significant reductions in the activity of GCPII in temporal, cortex, hippocampus and frontal cortex in schizophrenics (Tsai et al., 1995). Consistent with this post-mortem finding, several studies have independently demonstrated by MR spectroscopy reductions in the levels of NAA, the product of GCPII activity, in temporal cortex and frontal cortex in schizophrenia, the very regions shown to have reduced GCPII activity. Folate reduction and its surrogate elevated homocysteine, have been shown to be common in schizophrenia. Studies of enzymatic activity and, more recently, molecular cloning have demonstrated that GCPII accounts for folypoly- gamma-glutamate carboxy peptidase activity in the small intestine in humans, which is responsible for effective absorption of folate. While we have demonstrated that the GCPII protein expressed in human cerebellum is identical to that of jejunal peptidase as cell as the cell surface marker for metastatic prostate cerebellum is identical to that of jejunal peptidase as well as the cell surface marker for metastatic prostate cancer, Prostate Specific Membrane antigen (PSM) a complete characterization of human brain GCPII, possible allelic variants and the nature of the loss of activity in schizophrenia needs to be performed in order to understand more fully its possible role in schizophrenia and how that may related to reported impairments in folate absorption in schizophrenia. Specific Aim 1. Identify common allelic variants of GCPII targeting both normal subjects and patients suffering from schizophrenia studied in Project VI. Specific Aim 2. Carry out Northern blots,, in situ hybridization, Western blots and immunocytochemical studies to define the expression for the GCPII in selected regions of control and schizophrenic brains as previously described in rat brain (Berger et al., 1999). Specific Aim 3. Measure the levels of NAA, NAAG, glutamate and aspartate in temporal cortex, hippocampus, frontal cortex and other regions in post-mortem brain studies from individuals suffering from schizophrenia and suitable controls. Through microdissection, we will distinguish between gray matter and white matter as NAA is present in axons. Levels of glycine, D-serine and folates will also be measured. Potential alterations in amino acid levels and folate levels will be correlated with the expression GCPII in these samples. Specific Aim 4. Develop a conditional knock-out of GCPII to determine its effects on behavior and hippocampal neurophysiology in Project III.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Description: There are three levels of planning and evaluation: Strategic planning, Internal Advisory Committee, and External Advisory Committee.
|
{
"pile_set_name": "NIH ExPorter"
}
|
In order to characterize the molecular and cellular causes of human disease, it will be essential to unravel the functional impact of genetic variation. However, we are currently unable to predict the impact of the majority genetic variants that lie in non-coding regions of the genome, where indeed most complex disease-associated variants are found. Additionally, recent evidence suggests that a significant fraction of the non-coding genome is likely to be functional, often playing a role in gene regulation. Therefore, our limited understanding of non- coding variation is a critical hurdle to characterizing the genetic basis of disease. The goal of this project is to develop methods for interpreting non-coding genetic variation: to provide a robust and extensible Bayesian method for predicting causal variants from full genomes, to identify and validate a large set of functional non- coding variants using CRISPR technology, and to predict disease-relevant traits likely to be affected by each variant. Our project will leverage a unique cohort from a founder population in Sardinia, with genome sequence and/or transcriptome data available from 3000 individuals, along with extensive phenotyping for hundreds of traits. We will combine advanced statistical modeling with experimental validation based on genome engineering to identify causal non-coding variants affecting biomedical traits in the cohort, along with predicting functional mechanisms through which these variants ultimately perturb the cell. In Aim 1, we develop computational methods for predicting causal non-coding variation from full genomes, incorporating informative genomic features including epigenetic data, sequence motifs, and conservation information into a Bayesian approach jointly modeling multiple transcriptomic signals. We will optimize and apply these methods on genome and transcriptome data available for the Sardinia cohort to identify a large set of variants predicted to causally affect gene expression. Based on these predictions, in Aim 2, we connect putative causal variants with the diverse set of disease-relevant traits measured in the cohort, using network inference to capture the cascade from genetic variation to gene expression to disease. We will develop methods to integrate across variants, using the models in Aim 1, to identify the common causal mechanisms related to each trait. In Aim 3, we validate the causal impact of non-coding variants predicted to affect high-level traits. We will us genome editing through CRISPR to introduce individual genetic variants into cell lines and use qPCR to validate the predicted effects on gene expression. Finally, a major goal throughout this proposal will be to provide the research community with convenient computational tools for the prediction of causal non-coding variants from individual genomes, updated on an ongoing basis to integrate the most recent genomic annotations and public data in order to provide the best possible accuracy in predicting causal variants and the traits they are likely to affect. Our projet will greatly advance our understanding of non-coding genetic variation, the specific mechanisms affected by causal variants, and the downstream consequences to the cell and individual health.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The NHLBI Trans-Omics for Precision Medicine (TOPMed) program will support the Institute?s larger precision medicine initiative by collecting and coupling whole-genome sequencing (WGS) and other -omics data (e.g., DNA methylation signature, RNA expression profiles, metabolite profiles) with molecular, behavioral, imaging, environmental, and clinical data from studies focused on heart, lung, blood and sleep (HLBS) disorders. The TOPMed program aims to: ?Collect and assemble -omics (RNASeq, methylation, metabolomics, epigenomics, and proteomics) data with WGS and clinical outcomes data across diverse populations including those traditionally underrepresented in research. ?Build a data commons repository that the scientific community can use for future research and to enable precision medicine. ?Stimulate systems medicine approaches that help organize data to ensure they are accessible and interpretable for health disease research. ?Promote discoveries about the fundamental mechanisms that underlie HLBS disorders. Establishing the WGS project was one of the first steps for the TOPMed program. The project?s goal is to collect WGS data for individuals who have well-defined clinical phenotypes and outcomes from earlier NHLBI-funded studies. Initially, the WGS project planned to sequence 20,000 genomes. The NHLBI has expanded the WGS project and it now will sequence about 62,000 individual genomes.
|
{
"pile_set_name": "NIH ExPorter"
}
|
There are three major objectives: First, to explore diaphragm pacing by electrical stimulation of the phrenic nerve as a method of artificial respiration in the treatment of certain patients with chronic ventilatory insufficiency including respiratory paralysis (quadriplegia), central alveolar hypoventilation (CAH) and chronic obstructive pulmonary disease (COPD). Investigations are designed to aid in the selection of patients for pacing and in the evaluation of the results. They include studies on respiratory control, airway resistance and hemodynamics. Second, to investigate newly developed electronic apparatus of diaphragm pacing including a totally implantable diaphragm pacemaker, a hermetically sealed capsule for electronic components, a demand diaphragm pacemaker, and a new electrode design. Third, to investigate ways to extend the safe period of continuous ventilatory support by diaphragm pacing. Tests designed to reveal the influence of altering physical and electronic parameters on the effectiveness of pacing will be employed.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Prior studies indicate that right brain function is altered in depressive disorders. Our long range goals have been to help clarify the nature of the right hemisphere abnormalities in different diagnostic subtypes of affective illness and to evaluate the effects of antidepressant medications on these abnormalities. We propose to extend and finish two ongoing studies that are using behavioral measures of perceptual asymmetry and event-related potential (ERP) measures of hemispheric asymmetry. Specifically, we aim: (1) to complete a comparison between unipolar patients vs. bipolar patients (including Bipolar I vs. Bipolar II) vs. normal controls in tandem with an atypical vs. typical distinction (see item 2); (2) to replicate in a new sample initial findings of differences in lateralized perceptual function between atypical depression (patients with features such as mood reactivity, hypersomnia or hyperphagia) and typical depression (patients without these features); (3) to test patients before and after treatment with antidepressants so as to determine whether alterations of lateral asymmetry on behavioral or ERP measures persist or disappear following treatment; (4) to evaluate whether pretreatment lateral asymmetries are predictive of therapeutic response to either a standard tricyclic antidepressant (TCA) or a monoamine oxidase inhibitor (MAOI); (5) to relate alterations of perceptual asymmetry in depressed patients to more direct ERP measures of hemispheric asymmetry; (6) to do a pilot study using measures of regional cerebral blood flow (rCBF). We propose to increase our sample sizes for Study 1 and Study 2 by testing an additional 30 unipolar depressed patients, 30 bipolar depressed patients (15 Bipolar I and 15 Bipolar II) and 30 normal controls. Study 1 uses a battery of dichotic listening and visual tachistoscopic measures of lateralized perceptual function. Study 2 simultaneously measures ERP measures of hemispheric asymmetry and behavioral measures of perceptual asymmetry in temporal and spatial discrimination tasks. A pilot study will measure the rCBF of 10 depressed patients and 20 normal controls in a resting condition and under activation conditions. Patients are tested after a minimum drug-free period of 10 days and are retested following 6 weeks of treatment with either a TCA, MAOI or placebo. The clinical significance of this research lies in the need for more precise diagnostic aids that facilitate the selection of antidepressants tailored to different types of depressive disorder.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Illuminating the arc of molecular events from chronic hepatic injury to fibrosis and then cirrhosis remains among the most fundamental of challenges in investigative hepatology. Today's epidemic of end-stage liver disease due to hepatic fibrosis has precipitated an urgent need for new therapies. The hepatic stellate cell (HSC), following activation during liver injury, plays a central role in the development of fibrosis. Our long-term goal is to understand how HSC activation unfolds as an integrated, homeostatic response to liver injury;this will lead to novel, targeted interventions for hepatic fibrosis. A seminal study published by others in NATURE describing loss of lipid droplets as a feature of autophagy sparked our idea that autophagy is a component of HSC activation. Indeed, the features of HSC activation are hallmarks of autophagy, which is a highly regulated cellular response that has evolved to maintain energy homeostasis during cellular stress or enhanced metabolic demand. The objective of this project, which is the next step towards our long-term goal, is to characterize the contribution of autophagy to HSC activation, thereby uncovering new regulatory pathways and predicting consequences of their inhibition. Our central hypothesis, therefore, is that autophagy is a critical and necessary component of HSC activation. We will test our central hypothesis through the following interrelated Specific Aims: 1. Define stimuli that provoke autophagy in HSCs using established culture models. This question will be addressed in models of quiescent and culture-activated HSCs in which autophagy will be documented by: Western blot to detect conversion of LC3-I protein to LC3-II, ultrastructure, and lipid &retinoid content assessed by HPLC. 2. Determine which features of HSC activation are autophagy-dependent. Following siRNA knockdown of the specific autophagic regulators, Atg7 or Atg5, we will assess the contribution of autophagy to HSC activation in culture-activated HSCs (rodent &human), and define autophagy-regulated pathways by quantitative PCR and Western for known activation markers, and exon arrays for novel targets. 3. Establish the contribution of autophagy to HSC activation in vivo and explore the therapeutic potential of blocking autophagy in HSCs. The response to TAA or bile duct ligation models of fibrosis will be assessed in: a) Mice with stellate cell-specific Atg7 deletion. Key endpoints will include histology, collagen content, fibrogenic mRNA expression, HSC activation markers (smooth m. alpha actin and 2-PDGF receptor), ultrastructure and retinoid content. Isolated HSCs from these mice will be characterized for autophagy and activation, complemented by analyzing cultured HSCs from Atg7LoxP/LoxP infected with a cre-expressing adenovirus;b) Mice treated with chloroquine, seeking specific evidence of attenuated autophagy in HSCs that cannot be explained by indirect effects of the drug on other cell types, or on the extent of injury. These studies should uncover fundamental new pathways of stellate cell activation, leading to innovative treatment approaches for patients with fibrosing liver diseases. PUBLIC HEALTH RELEVANCE: Chronic liver disease leads to scarring, or fibrosis, of the liver, and is a major public health threat, affecting hundreds of millions of individuals worldwide. We propose to study a specialized pathway called autophagy that drives activation of hepatic stellate cells in liver to make scar. New insights expected to emerge from these novel studies could significantly advance our understanding of how to block scar formation in chronic liver disease, accelerate liver repair, and prevent the end-stage of liver disease called cirrhosis, thereby improving the lives of patients throughout the developed and developing world.
|
{
"pile_set_name": "NIH ExPorter"
}
|
At the molecular level most of the new mutations occurring in a population are different from the alleles preexisting in the population and the classical concept of recurrent mutation is incorrect in the strict sense of the term. This new concept requires many new mathematical formulations of population dynamics of mutant genes, taking into account stochastic changes of gene or gamete frequencies. In the proposed study the following four problems will be studied from the standpoint of this idea. (1) Mathematical theories for the process of genetic differentiation of populations will be explored. The theories developed will be employed to study the factors which have caused the racial differences in frequencies of normal and deleterious genes in man. (2) With the aim of clarifying the relatedness and evolution of human races, gene frequency data for a large number of protein loci will be analyzed by using genetic distance measure. The pattern of gene substitution in evolution will also be studied by looking at each enzyme locus separately in various organisms. (3) The mechanism of maintenance of deleterious mutations in human populations will be studied in terms of stochastic theory of gene frequency change. This study is intended to contribute to a better understanding of the mutational burden on the human society. (4) The theoretical basis of the use of marker genes in genetic counseling will be studied. Special attention will be given to the relationship between the utility of marker genes and recombination value. BIBLIOGRAPHIC REFERENCES: Nei, M. (1976) Mathematical models of speciation and genetic distance. In: Population Genetics and Ecology, eds. S. Karlin and E. Nevo, Academic Press, New York, pp. 723-765. Nei, M. and R. Chakraborty (1976) Electrophoretically silent alleles in a finite population. J. Mol. Evol., in press.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The effects of cocaine and other psychomotor stimulants on a number of physiological parameters are being studied. Recent studies in squirrel monkeys have indicated that alpha-1 adrenergic mechanisms are importantly involved in the pressor effects of both cocaine and methamphetamine, while beta-1 adrenergic mechanisms are important for the tachycardiac effect of both drugs. Unlike with cocaine, dopaminergic mechanisms are involved in the cardiovascular effects of methamphetamine. The cardiovascular effects of cocaine in rats are completely antagonized by noncompetitive or mixed type autonomic ganglionic blockers, while only partially antagonized by the competitive ganglionic blockers. Thus, these results provide substantial evidence that the cardiovascular effects of cocaine in conscious rats are mainly centrally medicated. Acute lethality studies indicate that various adrenergic agents modify acute cocaine intoxication. These studies also indicated that convulsant activity, rather than either cardiovascular or respiratory failure, is primarily responsible for the acute lethal effects of cocaine in conscious animals. Additional studies clearly demonstrate that central stimulation of sympathoadrenal neural axis plays an important role in cocaine's cardiovascular effects. Further, the pressor effect of cocaine is mainly medicated by catecholamines of sympathetic neural origin, whereas the tachycardiac effect is mainly mediated by catecholamines of adrenal medullary origin. We have also recently begun investigations of the effects of the major cocaine metabolites in anesthetized rats. Like cocaine, the cocaine metabolites norcocaine and cocaethylene (a metabolite produced during co-administration of cocaine and ethanol) have potent local anesthetic effects. Benzoylecgonine, ecgonine methyl ester and ecgonine did not produce local anesthetic effects. Benzolyecgonine and ecgonine methyl ester did possess potent sympathomimetic effects however. These potent sympathomimetic effects in the absence of a counteracting local anesthetic effect may account for some of the toxicity observed hours following cocaine administration.
|
{
"pile_set_name": "NIH ExPorter"
}
|
In chemotaxis by Bacillus subtilis, the information that attractant has bound to chemoreceptors is transduced along a pathway, which includes loss of methyl groups from the methyl-accepting chemotaxis proteins (MCPs), and ultimately results in a transient increase of counterclockwise rotation by the flagella. It is very important to trace this pathway, to discover what proteins are involved and what their functions are. B. subtilis is perhaps the best organism for this work since we have, using purified methyltransferase and methylesterase, shown that attractants retard methylation and strongly enhance demethylation of MCPs in vitro. Only in B. subtilis have these enzymes been purified to homogeneity. We have also developed two complementation systems and have obtained evidence for over twenty new genes. Thus, the stage is now set to investigate proteins that modulate methylation reactions. Using this system, we will explore the mechanism of several novel events induced by attractant: transfer of methyl groups between MCPs, turnover of methyl groups on MCPs, and transfer to an intermediate acceptor before formation of methanol. The last step is blocked by repellents and is the first biochemical handle on their mechanism of action. Isolation and characterization of mutants, including those containing a transposon that may express Beta-galactosidase, and cloning the chemotaxis genes will greatly assist us. Once the genes are cloned, we will carry out a restriction analysis of chromosomal segments and correlate the genetic and physical maps. We will express the genes in E. coli minicells as radioactive proteins so that we can ascertain their size and hydrophobicity and search for protein/protein interactions, especially with the switch, which controls direction of flagellar rotation. By second-site reversion studies, we will also seek evidence for such interactions. Our understanding of the MCPs, which represent our major biochemical handle on chemotaxis, is hampered by lack of mutants and insufficient gene expression and, using the techniques of molecular biology, we exprect to remedy this. Our knowledge, not only of the MCPs, but of over twenty other chemotaxis proteins, will be enhanced by mapping and complementing the mutants and by varying levels of genes expression. Armed with a uniquely suitable in vitro system for evaluation the effects of chemotaxis proteins, we believe that a renewed emphasis on genetics and molecular biology will prove especially fruitful.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The human population is routinely exposed to a large number of environmental chemicals: some of them may initiate cancer while others, only slightly different in structure, are harmless. One prominent route by which carcinogens exert their effects is to react with DNA in a way that leads to a mutation in a vital cellular target. Insight into the mechanism by which a carcinogen-damaged DNA produces mutations is needed in order to identify potentially hazardous substances. In this project, intensive computer modeling is used to explore this process. Our efforts here are targeted particularly to frameshift mutations, whose contribution to carcinogenesis has perhaps been underemphasized. In particular, we will attempt to relate chemical structure to mutagenic effectiveness within the framework of the slippage/misalignment theory. This theory has successfully explained the sequence dependence of many frameshift mutations. We will work with four aromatic amines, members of a chemical class that has demonstrated an exceptional ability to induce frameshifts. Our selection includes acetylaminofluorene (AAF), chosen because of the extensive data based concerning its mutagenicity, 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (PhIP) and 2-amino-3-methyl-imidazo(4,5-f)quinoline (IQ), carcinogens that are formed during the cooking of protein-rich foods, and 1-aminopyrene (AP), the transformation product of a common pollutant present in diesel engine exhaust, urban air particulates, and a number of other sources. We will follow the behavior of modified DNA primer-template complexes as they proceed through the steps of extension, blockage, and/or misalignment within the active sites of selected polymerases for which suitable crystal structures are available. Our methods include the use of the programs DUPLEX (for molecular mechanics with modified DNA) and AMBER for molecular dynamics simulations with DNA in solution or in a polymerase. DUPLEX permits an extensive search of conformation space without the use of assumptions concerning the final structure. The molecular dynamics studies include explicit solvent and salt, and provide animation, but are more restricted in their search. Molecular dynamics trajectories yield ensembles of structures that will be used to compute free energy differences between conformers in solution, and binding free energies of polymerase-primer-template complexes.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The study of transcription in yeast has led to the identification and characterization of many conserved genes that have general roles as transcriptional regulators. The genetic selection known as the Spt selection has been particularly productive, identifying many transcriptional regulators including histones, chromatin modifiers, TATA binding protein (TBP), regulators of TBP, and transcription elongation factors. Because the Spt phenotype has been indicative of proteins with roles during transcription, we used a newly developed systematic yeast overexpression library to screen for genes that cause the Spt- phenotype when overexpressed. We recovered essentially all of the factors that were previously identified using random genomic libraries, but additionally recovered an uncharacterized gene, PSH1, which encodes a RING domain protein that was reported to co-purify with the FACT histone chaperone complex and histones in large-scale proteomic studies. We propose the following Specific Aims to characterize PSH1 and its potential role in transcription regulation. For specific aim 1 we will determine the domains of Psh1 that are required for its normal function and for its overexpression phenotype. Psh1 has two recognizable motifs that suggest functional roles for this protein;a RING finger domain, and a highly acidic stretch of amino acids. To test this, point mutations in crucial amino acids and deletion derivatives that include the identified domains of Psh1 will be created. These mutants will also serve to identify domains that are important for the biochemical activity of Psh1 as proposed in Specific Aims 2 and 3. For specific aim 2 we will determine if Psh1 interacts with the FACT complex and histones and whether it is associated with chromatin. The identification of PSH1 in the Spt overexpression screen suggests that it has a role in transcription. The goal of this Specific Aim is to gain a biochemical understanding of its specific functions. We will determine whether Psh1 co-IPs with FACT and histones, and chromatin-IPs will be used to evaluate the association of Psh1 with chromatin. Combined, these approaches will establish whether Psh1 is associated with FACT, histones and transcribed regions in vivo. For specific aim 3, we will determine whether Psh1 ubiquitinates the FACT complex or other SPT genes. Our very recent preliminary data shows recombinant GST-Psh1 to possess ubiquitin-protein ligase activity in vitro. Using missense and deletion mutants constructed in Specific Aim 1, we will determine the domains required for ubiquitin-protein ligase activity in vitro. Having detected E3 ubiquitin-protein ligase activity, the next goal is to identify the in vivo substrate(s) of Psh1. We will take a candidate approach by testing in vivo substrates such as the FACT subunits Spt16 and Pob3.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Tissue engineering has produced a clinically successful equivalent of human skin (Integra). Similar efforts, applied to the cornea, an avascular and relatively acellular connective tissue, should also yield beneficial results. Although two corneal constructs have been produced, there is little hope of their use in the clinic in the near term, even as temporary scaffolding. The problem lies in the reproduction of the simple but highly organized stroma. The ability of the cornea to admit light, provide strength and generate a smooth spherical surface depends critically on its nanoscale structure. In this Phase I proposal, specific aim 1 describes the design of a novel collagen reactor capable of producing thin sheets of aligned collagen fibrils. The system is based on the well-defined science of spin coating and is designed to create a microenvironment promoting collagen tibrillogenesis in extremely thin sheets. In specific aim 2, a slight modification allows the orthogonal stacking of these sheets of aligned fibrils, mimicking the primary corneal stroma. Specific aim 3 proposes to create a rudimentary three-layered corneal construct by culturing human epithelium and endothelium onto the engineered stroma. Such an effort, focused on the stroma, is critical to the eventual generation of a biomaterial suitable for use in corneal repair or replacement therapy. PROPOSED COMMERCIAL APPLICATION: Between 40,000 and 50,000 cornea replacement surgeries are performerd in the United States every year, with an average cost of harvesting donor corneas ranging between $1,000-$2,000, yielding a market of $100M. Donor shortages are expected to grow in the future, as European and third world markets depend more on US-based eye banks, demographic changes limit donor pools, and keratorefractive surgery becomes more prevalent. A market exists for an artificial cornea, particularly one that can potentially be plated with a patient's own cells, which will reduce the chances of organ rejection.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Perturbations in hormone levels, growth, body habitus, and behavior are generally believed to be related during human puberty. A central issue is whether increases in hormone levels directly influence behavior or whether the adolescent's perception of his/her secondary sex characteristics provides the major basis for behavioral change. The present proposal will attempt to determine the direct influence of sex hormones on behavior by performing sequential behavior testing in 55 sex-steroid-deficient adolescents before, during and after they receive therapeutic hormone replacement therapy. Boys and girls with primary or secondary hypogonadism are receiving three dose levels of intramuscular testosterone enanthate (boys) or oral conjugated estrogen (girls) alternating with placebo over 3-month periods. The study design is randomized, double-blind, placebo controlled cross-over, clinical trial. To assess longer term behavioral effects of sex steroids, subjects will be retested 12-18 months after completion of the initial 21 month protocol.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Mitochondria are essential organelles in nearly all eukaryotic cells. These membrane bound structures contain their own DNA, and maintenance of this DNA is vital for normal cellular function. Mutations in the human mitochondrial genome have been implicated in many diseases, including neuromuscular disorders, heart disease, and diabetes. Moreover, the accumulation of mutations in the mitochondrial genome has been shown to contribute to aging and aging-related disorders. Mutations in DNA occur as a result of errors in DNA synthesis and repair processes acting on damaged DNA. Ultraviolet (UV) light is a DNA damaging agent which efficiently induces mutations in mitochondrial DNA of both yeast and humans. The mechanisms involved in the repair and maintenance of mitochondrial DNA in response to UV exposure are poorly understood. The studies proposed here address fundamental processes involved in mitochondrial genome maintenance after exposure to UV light. The specific aims of this proposal are I. to characterize the REVersionless mutants in UV-induced mutagenesis of mitochondrial DNA using genetic reporters allowing for the determination of the frequency of spontaneous and induced mutations of different types, and II. to determine the role of the heterotrimeric Rad17p/Mec3p/Ddc1p checkpoint clamp of Saccharomyces cerevisiae. Both molecular and genetic approaches will be used to determine the function of these proteins in the prevention of mitochondrial dysfunction. Relevance: Mitochondria contain their own DNA. This mtDNA is prone to DNA damage by chemical as well as enviornmental agents. One such agent is ultraviolet light, which causes lesions in the DNA. As the cell attempts to repair or bypass these lesions, mutations are produced. It has been shown that accumulation of mutations in mtDNA contributes to many diseases and disorders. Studying recovery of mtDNA after UV exposure in the budding yeast may shed light on how these processes occur in human mtDNA.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The NIH Pediatric Allergy Clinic has provided residents in the NIAID Allergy and Immunology Clinical Fellowship Program with medical training and continuity clinic experience in pediatric allergic diseases including asthma, allergic rhinitis, food allergy, atopic dermatitis and urticaria. During the past fiscal year, patient care visits exceeded 350, with a total accrual of 260 new patients. The NIH Pediatric Allergy Clinic also provides clinical resources for the study of children with allergic inflammatory disorders. In addition to standard diagnostic testing for allergic diseases, which includes skin prick testing, radioallergosorbent (RAST) testing, patch testing and pulmonary function testing, the clinic has integrated other modalities in the evaluation of children with allergic diseases. To evaluate lung function we have emplotyed impulse oscillometry, a non-invasive, non-effort dependant technique that measures lung resistance and elasticity. This is being used in conjunction with standard pulmonary function testing in older children and by itself in infants as young as two years old. A manuscript reviewing the clinical application of impulse oscillometry in pediatric patients with asthma is being prepared for publication. Through consultation with the NHLBI pulmonology service, exercise challenge pulmonary function testing has been performed. In conjunction with other investigators in the LAD, expired breath condensate and exhaled nitric oxide measurements have been obtained for diagnostic and research purposes. Research blood has been collected for laboratory studies to measure markers of allergic diseases, develop approaches to culture mast cells from blood, and determine cytokine and signaling profiles. In particular, serum tryptase levels were measured in 44 non-atopic and 153 atopic clinic patients. Median and 95% prediction intervals of tryptase levels for non-atopics were 3.44, 0.64-6.77 and 3.56, 0.98-10.80 for atopics. This study, recently published in the Journal of Allergy and Clinical Immunology (9 August 2009 on-line) establishes normative serum reference tryptase values in atopic and non-atopic children. Among atopics, a statistically significant association with tryptase was found for gender and race but not for ethnicity, total IgE levels, weight/weight percentile or eczema status. Patient documentation for communication with referring physicians and for longitudinal analysis is entered into CRIMSON, the NIAID database system.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The nodes of Ranvier of myelinated axons are critical for action potential propagation by saltatory conduction and are a striking example of the exquisite domain organization characteristic of neurons. Nodes are comprised of a multimeric complex that includes voltage gated sodium and potassium channels, accessory beta subunits, cell adhesion molecules (CAMs), and a cytoskeletal complex that includes ankyrin G to which channels and CAMs bind. Our recent studies indicate that the node assembles from distinct sources. CAMs, notably NF186, accumulate at forming nodes by redistribution from existing pools on the axon surface via diffusion trapping from interactions with the Schwann cell. In contrast, channels and cytoskeletal proteins are delivered to the node primarily via axonal transport. Surprisingly, there also appears to be a pool of sodium channels that traffics and accumulates at the node independent of transport and of ankyrin G. Finally, our studies suggest there is an active program to clear nodal proteins from extranodal sites that further reinforces their selective enrichment at the node. To extend these findings and further elucidate the mechanisms of node assembly, we will investigate trafficking and assembly of components of the node, and how they are cleared from extranodal sites, by live imaging strategies in myelinating cocultures. In particular, we will: i) determine whether vesicles that transport components to the nodes segregate into those specific for CAMs and others for ion channels, and exhibit any domain specificity, ii) characterize transport-dependent and -independent trafficking of sodium channel components to the node, including the roles of ankyrin G and NF186, respectively, and determine whether the sodium channel complex assembles locally, and i) investigate how nodal components are cleared from extranodal sites, focusing on NF186 to examine further the role of its cytoplasmic segment and that of endocytosis and proteolysis in clearing the internodal, surface pool of NF186. Relevance: These studies will elucidate the mechanisms that regulate the assembly of the node of Ranvier, which is critical to the ability of nerve fibers to conduct electrical impulses appropriately. Findings in this study may therefore have important implications for our understanding of the pathogenesis of disorders of myelinated fibers, including neuropathies that result in aberrant nerve conduction, and may thereby lead to new therapeutic strategies for these neurological disorders.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Although it has traditionally been assumed that the CNS and the immune system are separate and independent, recent evidence increasingly suggests that these systems are interrelated. One line of evidence is based upon a few reports of behavioral conditioning of immunological responses, particularly immunosuppression. Since these reports have been largely demonstrational in nature, the question of generality remains unanswered. In the present proposal conditioned immunosuppression will be studied in a wide range of conditioning preparations, some of which involve complex associations. Immunological responses will be assessed with techniques which measure changes at the cellular rather than the serum level. Together, these procedures will enable us to correlate precisely the degree to which these systems chance together as a function of various independent variables. Mice will be exposed to Pavlovian conditioning procedures in which gustatory, olfactory, and exteroceptive conditioned stimuli (CS) are paired with an unconditioned stimulus (US), cyclophosphamide, which has immunosuppressive effects. Conditioned immunosuppression will be examined as a function of CS- and/or US-magnitude, CS-US delay, and in terms of resistance to experimental extinction. In other experiments, potentiation of conditioned immunosuppression will be examined in a compound stimulus preparation. In still others, external stimulus control of immunosuppression will be explored in a context-conditioning paradigm. In a final experiment, conditioned immunoenhancement will be attempted by employing an antigen as the US. These experiments have important theoretical implications, not only in terms of advancing our understanding of the immune system and its susceptibility to modulation by external factors, but also in terms of furthering our knowledge of the types of response systems which are subject to control by Pavlovian processes. They also have long range implications for treatment of a variety of neoplastic and immunologic diseases as well as internally- and externally-produced stress.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The centromere regulates the movements of the chromosomes in mitosis. Work from this lab, among others, has used antibody probes to show that the centromere is composed of several subdomains in addition to the one prominent subdomain recognized by electron microscopy: the kinetochore. The kinetochore has recently become the focus of wide interest since it may contain the mechanochemical motor responsible for anaphase movement of the sister chromatids to the spindle poles. The ultimate goal of our studies is to construct a structural and functional map of the outer domains of the human centromere. Using human autoantibodies, we identified a family of centromere proteins, CENP-A (17 kDa), CENP-B (80 kDa), and CENP-C (140 kDa) in previous work supported by this grant. We cloned, sequenced, and began an in depth characterization of the molecular structure and biological role of CENP-B. We also obtained preliminary clones of CENP-C. In the experiments proposed for the upcoming grant period we will continue our analysis of CENP-B function, we will carry out a detailed molecular and functional analysis of CENP-C, and we will continue to screen for new components of the centromere and kinetochore. CENP Antigens, General. We will continue to characterize the disruption of chromosome movements in mitosis caused by injection of purified autoantibodies into cultured cells. CENP-B. We will examine the association between CENP-B with alpha-satellite DNA by in vitro binding experiments using cloned proteins and crude chromosomal extracts and by immunocytological investigation of abnormal chromosomes with translocated alpha-satellite domains. We will attempt to determine the role of CENP-B in vivo by creating dominant disruptions of CENP-B function, using transfection and/or injection of cultured cells with plasmid constructs expressing different CENP-B subdomains. Such constructs will also be used to identify those portions of CENP-B required for correct targeting and assembly into the centromere. We will examine the interaction of CENP-B with other chromosomal proteins by chemical crosslinking and by affinity chromatography using various CENP-B subdomains expressed in bacteria. We will characterize the patterns of transcription and posttranslational modification of CENP-B across the cell cycle. CENP-C. We will isolate full-length cDNA clones for CENP-C, obtain the DNA sequence, and use an immunological approach to demonstrate that these clones encode bona fide CENP-C. We will map the distribution of CENP-C in the centromere by immunoelectron microscopy.We will perform analyses similar to those listed above in order to determine the biological role of CENP-C in centromere structure and function. New Centromere Components. We will use a newly designed shotgun cloning method to identify new components of the centromere and kinetochore. Either mitotic chromosome scaffolds or an isolated human minichromosome will be used as antigen. Newly identified antigens will be used to elucidate the network of protein-protein interactions within the centromere. Antigens of suitable interest will be characterized as above for CENPs B and C.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The overall objective is to investigate the physiology, including behavior, of the adult female mosquito, especially the endocrine control of digestion, egg maturation, sexual behavior, host attraction, and biting. The effect of aging on certain physiological events will be investigated, as we have found that some reproductive processes are slower as the female ages. This has important epidemiological implications, because multiple blood-feeding during each gonotrophic cycle may become obligatory as the female ages. The ovary must be regarded as an endocrine organ, since it produces a hormone that stimulates vitellogenin synthesis by the fat body of blood-fed females, and, when the ovary retains a large enough number of eggs, it secretes a yolk inhibitor that suppresses development of less advanced oocytes. The relationship of these ovarian hormones to those secreted by the neurosecretory system and corpora allata (CA) will be investigated by ovarian transplantation and surgical ablation of the endocrine tissues. We will try to develop a bioassay for CA hormones in circulation and to determine the duration of the dependence of developing oocytes on both the CA hormone and the neurosecretory hormone. We will continue to study the rate of oocyte growth and the factors that influence the rate. The sequence of deposition of the various major constituents of yolk will be determined by chemical analysis. The effects of various physiological states (including different known nutritional levels, reproductive conditions, and parasite infections) on host attraction and biting by the female mosquito will be tested in a "host response chamber".
|
{
"pile_set_name": "NIH ExPorter"
}
|
The purpose of these studies is to elucidate the mechanisms of human basophil and mast cell desensitization. The intention is to understand the basic mechanisms of humanbasophil and mast cell mediator release in hopes of providing a firm framework on which future therapies for allergic disease can be based. One aspect of the mediator release mechanism is these cells' capacity for autoregulation through the process termed desensitization, and it is on this aspect which we will focus our efforts. Four areas are targeted for study. 1) elucidating the mechanism by which diisopropylfluorophosphate inhibits desensitization 2) defining the role of large scale aggregation of antigen and IgE in promoting desensitization 3) comparing mast cell and basophil desensitization 4) determining the effects of desensitization on two known activation event biochemistries, calcium flux and phospholipid turnover DFP has been found to inhibit basophil desensitization and enhances histamine release supporting the concept that desensitization regulates release. Studies are proposed which will use radiolabeled DFP or DFP analogs to isolate the enzyme(s) involved in desensitization. Previous studies suggested that the size of cell surface IgE-antigen aggregates determine the characteristics cell mediator release and desensitization. Experiments to substantiate this hypothesis involve comparing several well defined antigens for their ability to induce desensitization. Binding assays and microscopic fluorescent photo-bleaching techniques will be employed to characterize surface IgE redistribution under the influence of these antigens. Mast cells are central to the pathophysiology of allergic diseases. They can now be purified to near homogeneity. Studies are proposed which will determine whether non-specific desensitization occurs in these cells and how they process antigen-IGE aggregates during stimulation and desensitization. Previous studies suggested that the size of cell surface IgE-antigen aggregates determine the characteristics cell mediator release and desensitization. Experiments to substantiate this hypothesis involve comparing several well defined antigens for their ability to induce desensitization. Binding assays and microscopic fluorescent photo-bleaching techniques will be employed to characterize surface IgE redistribution under the influence of these antigens. Mast cells are central to the pathopysiology of allergic diseases. They can now be purified to near homogeneity. Studies are proposed which will determine whether non-specific desensitization occurs in these cells and how they process antigen-IgE aggregates during stimulation and desenstitization. Finally, studies are proposed which will investigate the occurrence of calcium translocation and phospholipid turnover in both mast cells and basophils. The effect of desensitization will then be examined.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Toxoplasma gondii is an important opportunistic pathogen in patients with acquired and primary deficiencies in T cell functions. The overall aim of this proposal is to understand the factors that regulate the development and maintenance of T cell responses required for resistance to T. gondii. The early events that occur during infection have a profound influence on the development of NK and T cell production of IFN-gamma required for resistance to this pathogen. While there are many factors such as cytokines and the presentation of parasite antigen to specific T cells, that regulate the activation of these protective responses, the second signals provided by co-stimulation also play a role in innate and adaptive immunity to T. gondii. In order to fully understand how protective immunity to toxoplasmosis is initiated and maintained we need to understand which co-stimulatory pathways are important and the mechanisms that underlie their effects. Our previous studies have demonstrated a critical role for CD28 in the generation of CD4 v T cell responses required for protective memory responses and the proposed studies will utilize in vivo studies with transgenic mice to understand the mechanisms whereby CD28 mediated production of T cell growth factors or anti-apoptotic molecules underlies this effect. In addition, our studies have shown that there are CD28-independent mechanisms that regulate NK and T cell responses to T. gondii and we have identified the Co-stimulatory molecule ICOS as being important in CD28-independent resistance to T. gondii. The proposed studies will integrate molecular and cellular approaches to address the relationship between ICOS and CD28 and their role in innate and adaptive responses to T. gondii and determine whether agonists of ICOS and CD28 can be used therapeutically to prevent disease caused by this opportunistic pathogen.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The broad long-term objective of this proposal is to understand why memory loss occurs in Alzheimer's disease. The more immediate objective of our research is to investigate the hypothesis that the beta-amyloid molecule (AFJ) has dual roles in learning and memory, one beneficial and the other harmful. We have developed transgenic mice expressing various forms of the amyloid precursor protein that show either superior memory or an age-related deterioration in memory. We have shown that monomeric AB is present from infancy in neurons of the brain, but that abnormal, aggregated forms of AI_ appear only with aging. We propose to determine whether there is a normal form of AB, called AB*, which enhances memory. We have evidence that abnormal forms of AB are associated with memory loss, and propose to further delineate the corresponding AI3 species, called AB*.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The investigator will pursue her interest in the organization and management of Intensive Care in her dissertation, in which she examines the triage of ICU patients. Triage is a term used by clinicians and administrators to refer to decisions regarding the range and scope of services offered throughout the hospital and the selection of patients for hospital units deemed scarce. Triage of ICU unit patients is rapidly becoming a national policy issue as the costs and prevalence of ICU services grow. Using data from ten sites (one in-depth case study and nine other cases), the investigator will examine the relationship between the triage process, the organization of ICU'S, the organization of other hospital services and the utilization and efficiency of ICU services.
|
{
"pile_set_name": "NIH ExPorter"
}
|
This study is examining patterns of drug use and changes in these patterns as the sample of black youths passes through the adolescent and young adult stages of their life span. Drug use (non-medical) will be analyzed in relation to: 1) physical and emotional health status; 2) physical growth and development; 3) psychosocial attributes; 4) socio-cultural background--all measured at two points in time. Reinterviews (6-8 years later) have been completed with 89 percent of the original sample who were still alive and in the metropolitan New York Area. These 536 youths (259 Female, 277 Males--a ratio similar to the original sample) represent 80 percent of all the initial sample. In the coming year, analyses will be conducted of: 1) Age, sex, and other sub-cultural variations in drug use behavior in this high risk, urban black youth population. 2) Precursors or predictors--physical, developmental, psychosocial and social structural--of drug use (analysis of baseline data relative to subsequent drug behavior). 3) Health correlates of drug use behavior (current physical, emotional and psychosocial status relative to drug use behavior). 4) Longitudinal analysis of health consequences of drug use: changes in health attributable to drug use behavior (timing, extent, and type). BIBLIOGRAPHIC REFERENCES: Brunswick, Ann F. Indicators of Health Status in Adolescence. Int. J. Hlth Serv., Vol. 6, No. 3, pp 475-492, 1976. Brunswick, Ann F. Health and Drug Behavior: Preliminary findings from a study of urban black adolescents. Addict. Dis. 3 (2) in press, 1977.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Early work described the correlation between birthweights of mothers and their infants. The study of other fetal growth parameters, including length and head circumference demonstrated that infants of low birth weight mothers were both shorter and lighter than infants of larger mothers, but that the infants were normally proportioned. In ensuing studies, birth certificates of infants born in Tennessee between 1979 to 1984 were matched with those of their mothers, who were born in Tennessee between 1959 to 1966. Maternal and infant birth weights were again shown to be correlated. In addition, women who were themselves of low birth weight were up to four times as likely to have a small-for-gestational age infant as were women who weighed 4000-4499 grams at birth, but low birth weight women were less than twice as likely to have a preterm infant. A group of Swedish women, born from 1955 to 1965, was next studied. Women who themselves were small-for gestational age at birth were at increased risk of giving birth to both small-for-gestational age and preterm infants. Women who were preterm at birth were not at increased risk of either outcome. Among women born in the Danish Perinatal Study (1959-61), women born SGA were found to be at doubled risk of having an SGA child, but women born preterm were not at increased risk of giving birth to a preterm child. In addition, women who were small-for-gestational age at birth were at increased risk of developing hypertension during pregnancy. Independent of maternal size at birth, paternal size at birth was found to be associated with fetal growth, but not with duration of gestation. Girls who were born from 1959-66 as subjects in the Philadelphia and Providence cohorts of the Collaborative Perinatal Project were followed-up in adulthood, and their pregnancy outcomes ascertained. Data analysis from these women is currently underway. Initial analyses evaluated the relative contributions of the birth characteristics of the mother and those of older siblings on the birth characteristics of the youngest sibling. Both the mother's own SGA status at birth, and the SGA status of older siblings each predicted SGA status of the youngest sibling. Although having a preterm older sibling predicted preterm birth in the youngest sibling (odds ratio=5.5), having a preterm mother did not (odds ratio=1.0). There were no interactions between maternal and older sib birth characteristics.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Coronary heart disease (CHD), which causes heart attacks and angina, is the single leading cause of death in the United States. The strongest risk factors for CHD development include circulating levels of lipids and lipoproteins, both of which are strongly regulated by genetic factors. Variants with significant effects on lipid levels have been identified for a number of monogenic familial disorders, but these account for only a small proportion of CHD. In contrast, genetic determinants for common lipid abnormalities remain unknown. The overall plan for this project is to identify and characterize genetic variants that contribute to the regulation of fasting serum concentrations of total cholesterol. The focus of this study is a locus which has been initially mapped on the basis of linkage data to chromosome 19 in at least 15 studies, then subsequently narrowed to a 1-LOD support interval <15.7 cM in Pima Indians. The specific goals of this proposal are to first refine and prioritize localization of the QTL(s) for fasting serum TC concentration on 19p by genotyping a dense set of SNP markers in a study sample of 2,884 Pima Indians. All QTL-associated SNP alleles and haplotypes will then be genotyped in 701 African American individuals from the Genetics of NIDDM (GENNID) study, in whom we have previously observed linkage for TC concentration, and 740 Mexican American individuals from the San Antonio Family Gallbladder Disease Study (SAFGS), who are more likely to share greater genetic similarity with Pima Indians. Completion of this aim will provide validation of findings obtained in Aim 1. The next step will utilize an innovative bar-code approach to next-generation sequencing to fully characterize regions showing association with TC concentration, and identify specific variants that are likely to exert functional effects on cholesterol metabolism. Finally, we propose an exploratory characterization of functional effects associated with strong QTL-related alleles, which will provide a basis for the development of a full-scale investigation of the molecular mechanisms by which these specific variants affect regulation of cholesterol metabolism as the focus of an independent grant application. Combined, these aims will identify novel variants with critical effects on fasting total cholesterol concentration. Identification of the genetic mechanisms influencing cholesterol concentrations will advance our understanding of lipid metabolism, leading to an enhanced knowledge of the pathophysiology of the atherosclerotic process. PUBLIC HEALTH RELEVANCE: Total cholesterol levels are under the control of genetic factors. The goal of this study is to advance our knowledge of common lipid abnormalities, such as hypercholesterolemia, through the identification and characterization of genes that have been linked to chromosome 19 in a number of different populations. Identification and characterization of genes that regulate cholesterol metabolism will enhance our understanding of the inheritance of common lipid abnormalities, provide markers to target individuals at greatest risk for developing heart disease, and potentially lead to improved treatment strategies for hypercholesterolemia.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Dr. Silliman is a geriatrician and clinical epidemiologist whose research most broadly described, has focused on chronic disease management in older adults. Over the past fifteen years, her breast cancer studies have been the core of this work. They have identified aspects of patient-physician communication as important factors in treatment decision-making, even after tumor characteristics, comorbidity, and functional status are taken into account. Dr. Silliman's investigative work has also explored alternative methods of comorbidity measurement and has considered the implications of treatment variations for a range of health outcomes: (1) general and breast cancer-specific emotional health; (2) upper body function; and (3) breast cancer recurrence and mortality. Dr. Silliman has also added to the small but growing body of observational study evidence that suggests that variations in breast cancer treatment do influence rates of breast cancer recurrence and mortality. Her currently funded R01 is designed to: (1) compare the effectiveness of adjuvant tamoxifen therapy, versus no adjuvant tamoxifen therapy, in reducing breast cancer recurrence and breast cancer mortality rates among older women receiving recommended definitive primary tumor therapies and those not receiving such therapies; (2) quantify the prevalence of tamoxifen discontinuance and identify patient and physician characteristics associated with it during the recommended five years of therapy; and (3) identify risk factors associated with a decline in health- related quality of life outcomes, particularly upper body function, during five years after breast cancer diagnosis. Dr. Silfiman's specific career objectives for the proposed award are: (1) to provide scientific evidence about breast cancer care and its consequences in older women, especially those > 75 years of age; (2) to apply new analytic techniques to existing data to control for confounding and misclassification in observational studies of breast cancer treatment; (3) to develop the research database infrastructure to study age-related variations in cancer treatment and outcomes among older cancer patients, applying the methods and experience developed studying breast cancer in older women to other common cancers (e.g., lung, colerectal, and prostate cancer) and, (4) to develop young clinical and non-clinical investigators whose research will focus on cancer prevention and control in older persons.
|
{
"pile_set_name": "NIH ExPorter"
}
|
DESCRIPTION: Increased hepatic secretion of apolipoprotein B100 (apoB) is involved in the development of certain hyperlipidemias and atherosclerosis. Recent findings have demonstrated that hepatic apoB intracellular transport in the secretory pathway is a unique dynamic process that includes sorting of apoB molecules to secretion or degradation pathways. A candidate for the pathway that degrades apoB is the ubiquitin-proteasome system. The overall goal of this research is to understand the mechanism of hepatic apoB-lipoprotein secretion, including the precise pathway of intracellular transport of nascent apoB through the various compartments of the secretory pathway, and the pathway of degradation of apoB by the ubiquitin-proteasome system. The intracellular transport of endogenous and expressed apoB will be studied in HepG2 and McArdle Rh-7777 hepatoma cells. Studies will also be performed with primary pig hepatocytes to confirm observations made in cell lines and to study the physiological role of co- or post-translational apoB degradation. Aim I will investigate the location, orientation, and intracellular transport of nascent apoB in the endoplasmic reticulum (ER) and Golgi compartments of the secretory pathway using 1) puromycin-synchronized cells and immunocytochemistry of apoB and other proteins; 2) transport of apoB tagged with green fluorescent protein in live cells; and 3) nanogold immuno electron microscopy to more precisely localize apoB in or near organelles. Aim 2 will determine: 1) the mechanism of apoB transport to the proteasome; 2) the proteasome population (cytosol or ER-associated) involved in apoB degradation; 3) the role of the translocon in retrograde transport of apoB; and 4) the sites of ubiquitination in apoB. Aim 3 will take advantage of our newly developed permeabilized cell degradation assay for apoB and identify the protein(s) in rabbit reticulocyte lysate that stimulate the rapid proteasomal degradation of apoB. Significance: The entire secretory pathway of apoB will be elucidated in detail and proteins involved in the degradation of apoB will be characterized. A comprehensive understanding of the regulation of apoB secretion will shed light on hyperlipidemias involving overproduction of apoB-lipoproteins, possibly the result of inefficiency in the degradation of lipid-poor apoB by the proteasome pathway.
|
{
"pile_set_name": "NIH ExPorter"
}
|
A major goal in neurobiology is identifying functional neuronal pathways. Classical anatomical tracing techniques allow connectivity between brain regions to be determined. Lesion studies and anatomically restricted infusion of pharmacological agents have helped to identify the functional role of these brain regions at a gross level. The development of techniques for the genetic manipulation of the mouse during the past 15 years has greatly expanded our ability to probe the molecular mechanisms of biological function in a mammalian model system. However, one of the difficulties in the application of the genetic approach to the nervous system is the relative lack of ability to map molecular genetic changes onto the complex neuroanatomy of the brain. The goal of the current application is to develop a series of 48 driver lines for the generation of anatomically restricted and inducible gene knock-outs. For greatest genetic utility to the neuroscience community the strains will be developed on a pure C57BL/6 background. The most widely used tools for anatomically restricted and time dependent manipulation of gene function in the mouse are the CRE-recombinase and the tTA-transactivator, respectively, but very few driver lines are available to the neuroscience community. Furthermore, most CRE-driver lines do not allow strict temporal control, such as the ability to knock-out genes in adult tissues. We plan to use transgenic approaches to express CRE and tTA driven by the regulatory elements of 24 chosen genes. The default approach is targeting via homologous recombination in ES cells; vectors for pronuclear injections (including BACs) will also be utilized. The driver lines will allow us to achieve temporal and spatial control of recombinase activity in disparate regions of the nervous system. Driver lines will be validated by monitoring cell type and time dependent CRE activity. Behavioral studies will ensure that the transgenes do not affect neuronal function. The choice of the 24 driver loci represents the expertise of the four investigators involved in this application and are of relevance for the neuroscience community, covering sensory biology and pain, CMS and stem cell development, the limbic system, and learning and memory. From a clinical perspective the lines generated should be particularly relevant to the study of Neuropsychiatric disorders and addiction (eg. targeting the dopamine system) and neurodegenerative disorders (e.g. targeting the limbic system and adult stem cells). [unreadable] [unreadable] [unreadable]
|
{
"pile_set_name": "NIH ExPorter"
}
|
The Harvard Regional Clinical Center (RCC) ofthe current NHLBi Heart Failure (HF) Network, including a Clinical Research Skills Development Core, looks fonA/ard to expanding into a triangle where 3 uniquely aligned and supported HF programs will accelerate the mission ofthe NHBLI to improve the future for HF patients. Study patient recruitment within this RCC will be enhanced through utilization of real-time databases and diversity of site populations. Creation of new Network Assets is planned, using common data fields that will bridge individual Network trials to multiply the scientific yield ofthe Network. These Assets will inform current challenges in HF care, such as preventing hospitalizations, identifying new pathologic mutations in dilated cardiomyopathy, and characterizing patients with improved LVEF. The Harvard RCC will provide collaborative leadership for design and implementation of studies that address vital mechanistic and therapeutic questions that cannot be answered without NHLBI sponsorship. One example is a 3-arm comparative effectiveness study to determine Iron Repletion Effects on Exercise Capacity and Cardiac Function in Heart Failure (IRONOUT HF). This study will add to the understanding ofthe physiologic impact and explore treatments of iron deficiency in HF. The primary endpoint will be change in peak oxygen consumption after 24 weeks of intravenous iron, oral iron, or placebo. If benefit is shown, oral iron will be assessed for non-inferiority compared to the more expensive and inconvenient therapy with intravenous iron. This study will also assess skeletal muscle oxygen utilization during low level exercise, and relate changes in hemoglobin, iron stores, the iron-regulatory protein hepcidin, and novel iron-dependent metabolomic profiles to changes in exercise capacity in order to identify patients most likely to benefit from iron repletion. The Clinical Research Skills Development Core supports a rigorous course curriculum, hands-on-laboratory experience, and a Mentor Review Board to train and inspire new HF clinical investigators. Utilizing the Network Assets, an Apprentice Investigator Network is planned where fellows throughout the HF Network will collaborate to become the next generation of clinical investigators. RELEVANCE (See instructions): Heart failure is a growing burden on US health that can only be diminished by the collaboration of leading investigators through initiatives such as the NHLBI Heart Failure Network. The Harvard Regional Clinical Center proposes to join other leaders in the field in studies such as IRONOUT HF, testing whether treatment of iron deficiency in heart failure patients with either oral or intravenous iron will improve exercise capacity.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Chronic infection and intense inflammation are hallmarks of CF lung disease. Characteristics include colonization with bacteria and IL-8-driven, neutrophil predominant inflammation. However, a 'chicken and egg'conundrum regarding the chronology and causal relationship between inflammation and infection in the CF lung continues to plague the field. Existing mouse models of CF have helped advance our knowledge, but fail to develop the pulmonary phenotype characteristic of humans with CF. In this proposal a porcine model of CF (CFTR null) establishes the opportunity to study the onset of lung disease with approaches inconceivable in humans. By focusing on the prenatal and perinatal time periods, we may gain insights into underlying disease mechanisms without confounding variables. We will investigate key questions at the level of the intact animal and in specific cell-based models. We propose three aims. Aim 1. Does the fetal porcine CF lung exhibit inflammation in the absence of infection? The time of onset of inflammation in the CF lung is not known. We hypothesize that loss of CFTR function predisposes the lung to a pro-inflammatory state in the absence of infectious or inflammatory stimuli. We will thoroughly query the late fetal CF and non-CF lung in vivo for evidence of inflammation. Aim 2. Does the neonatal porcine CF lung develop inflammation spontaneously or in response to infection? We hypothesize that loss of CFTR function predisposes the newborn lung to spontaneous inflammation and causes enhanced or prolonged responses to infectious or inflammatory stimuli. In these studies the CF and non-CF piglets will be followed in clean housing, germfree isolators, or following specific microbial challenges for the development of spontaneous inflammation with or without infection as described in Aim 1. These approaches will allow a conclusive assessment of the spontaneous or inducible properties of the CF pig lung for developing inflammation. Aim 3. How does loss of CFTR function alter specific innate immune functions in the CF pig? We hypothesize that loss of CFTR alters specific innate immune functions. These studies will address the following questions. We will perform experiments using primary cells from CF and non-CF animals to address the following question: 1) Does loss of CFTR function drive inflammatory responses in CFTR-/- epithelia in vitro?, 2) Do submucosal gland secretions exert an anti-inflammatory effect on the surface epithelium that is altered in CF?, and 3) Does the loss of CFTR function in macrophages and neutrophils contribute to inflammation in the CF lung? Our focus on the early onset of inflammation in the late fetal, early postnatal period allows the potential to mechanistically understand primary events negatively affecting the CF lung.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Preliminary data indicate that fourteen of sixteen chronic schizophrenic patients are in good states of remission after sixteen weekly treatments utilizing hemodialysis. This study will divide twenty-four patients into experimental and control groups. The experimental group will undergo hemodialysis; the control group, sham dialysis, to determine the placebo effect, if any, of renal diaysis on patients who are chronically schizophrenic. The testing and evaluations will be done blindly.
|
{
"pile_set_name": "NIH ExPorter"
}
|
We have demonstrated the feasibility of prolonged (10 week) pancreas organ culture and growth in vitro in a defined medium. In addition we have determined that certain known carcinogens in this system produce alterations in morphology and growth. It is the purpose of this project: to determine whether the observed changes are neoplastic and if so, this should be an ideal system to study pancreas carcinogenesis. Following dimethylnitrosourea (DMNU) or dimethylnitrosamine (DMN) administration, ultrastructural alteration (EM), cell proliferation (DNA synthesis, autoradiographic) and enzymatic activity (i.e.: amylase, lipase, chymotropsin, etc.) will be studied. Morphologically transformed explants will be implanted in vivo to evaluate their biological transformation. In earlier work it was established that a relative methyl deficiency in organ culture allowed pancreatic growth but prevented differentiation. The separation of these two processes (proliferation and differentiation) will allow study of the actions of carcinogens under conditions of embryonic growth.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The objective of the proposed research is to secure definitive information at the molecular level about the mechanism by which ferritin sequesters and subsequently mobilizes metabolic iron. The experimental approach in this proposal is based on the unique structural features of apoferritin. It is proposed to study the selectivity of apoferritin with respect to what can pass into the interior of this hollow shell. The physical-chemical approach to be employed is to measure the rates of passive diffusion into apoferritin of a number of compounds of increasing Stokes radii and differing charge properties. Results from these measurements will yield information about the size and charge discriminatory properties of the apoferritin channels with respect to the passive diffusion of small molecules (such as potential iron chelates) in and out. Based on information from the aforementioned experiments, the kinetics of iron accumulation with various mononuclear chelates of Fe plus2 and Fe plus3 will be examined with respect to the "active" shuttling of iron into and out of the protein shell. Finally, chemical modification of selected amino acid side chains will be employed to determine the types of amino acid residues which participate in the passage of iron (or its chelates) through the channels. The results of these findings should not only better explain the role of ferritin in such widely divergent maladies of iron metabolism as anemia and hemochromatosis, but it may offer new options for their clinical treatment.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Several different cutaneous and deep somatic (intramuscular) laboratory pain-induction methods will be compared and evaluated for their applicability as objective, reliable and valid pain measurement instruments, and two or three of them selected as the most valid and practical (i.e. simple and fast) components of a pain "thermometer" or algesiometer. Chemical, thermal, electrical and mechanical noxious stimuli will be studied. Each method will yield four pain response parameters, namely pain threshold, drug request point, pain tolerance, and pain sensitivity range. Using paid, healthy normal subjects, the data will be correlated within and between techniques and response parameters to yield a correlation matrix. This matrix will be subjected to factor analysis in order to isolate a specific pain factor, termed pain endurance. It is hoped that this specific factor will greatly assist in the development of the pain measurement battery. Various of the above pain-induction methods will be studied under double-blind conditions with several analgesic drugs, such as codeine, propoxyphene and aspirin, and other treatments for pain relief, such as transcutaneous electrical stimulation, in order to validate the pain measurement battery. The advantage of complete cross-over designs to single dose designs in drug studies with healthy normal volunteers will be examined to determine future experimental approaches.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Because of reports that both barbiturates and ethanol interact with the GABA-benzodiazepine-chloride channel complex, we embarked on a series of studies to determine if agents which block various aspects of the complex might reduce toxicity from high doses of these sedatives. We reported last year tht IPPO, a chloride channel blocker, greatly reduced deaths from pentobarbital (PB) overdose and reduced loss of righting reflex from ethanol. We have continued these studies looking for agents which are both more effective, and less toxic when given by themselves. We found that TBPS, also a chloride channel blocker, was equally effective as IPPO but had less toxicity. We are currently investigating adenosine antagonists, in this same paradigm.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The purpose of this TL1 Clinical and Translational Science (CTS) Fellowship is to provide rigorous individualized training that encompasses core competencies in translational research methodologies and interdisciplinary professional skills. We will support the career goals of fellows through an intensive program of mentoring, scholarly activities, and career coaching. Over the past fifteen years, we have built an extensive infrastructure of career development and degree programs tailored to educate and train investigators at every stage of the career pipeline and across the spectrum of translational science. Our commitment to mentoring as an integral part of career training has resulted in well-established mentoring programs and an expansive pool of experienced senior investigators willing to serve as mentors to trainees and students. We also provide trainees access to extensive statistical, ethics, data management, design, analytic and editorial resources to further support their pilot research projects and grant proposals. For the CTS Fellowship, we will draw fellows and faculty mentors from the six Schools of the Health Sciences, as well other schools such as engineering, and social sciences. In addition, we are collaborating with 18 Minority Serving Institutions who send fellows to Pittsburgh for training. We also will utilize the more than 80 institutes and centers at the University that serve as hubs for multidisciplinary research. The immediate objectives of this TL1 CTS Fellowship are to: recruit outstanding fellows; train fellows in rigorous research methods; employ externships to provide real world research experience; build skills in team science, leadership, stakeholder engagement, and entrepreneurship; guide fellows in asking important translational research questions and implementing research projects; ensure close monitoring of fellows to guide their research career development; and help fellows learn about stakeholder engagement and develop relationships with stakeholders in their areas of research. Through rigorous methodological training and productive research, we aim to shorten the time to independence for fellows by providing them with the foundation for successful career development awards. Our long-term objectives are to ensure that fellows complete their translational research studies on schedule, disseminate their research widely, and are positioned to take full advantage of career options in translational research.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The aim of this proposal is to test the hypothesis that viral infections are associated with development of anti-virus antibodies cross-reacting with antigens present in cells of the CNS of normal animals. The possibility of such autoreactivity will be studied in vitro and in vivo using as a model mice spontaneously infected with Theiler's virus. The in vitro studies will be based on the isolation and characterization of antigens from uninfected human cells, cross-reacting with monoclonal Ab to purified measles virus or measles-infected cell lysates. The binding of monoclonal Ab to normal or infected cells will be established by immunofluorescence and immunoelectron microscopy, using the immunoperoxidase bridge technique or the immunocolloidal gold technique. These latter studies will be performed by Dr. Peter Lampert. Binding the monoclonal Ab will be performed by immunoprecipitation, Western blot analysis, involving separation of antigenic fractions in SDS gel, electrophoresis, reaction with monoclonal Ab, and identification of the reactive proteins with 123 I-labeled anti-mouse IgG; two dimensional gel separation; and high pressure liquid chromatography.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Project Summary/Abstract Minnesota HealthSolutions Corporation (MHS) proposes to develop an innovative all-new convertible child safety seat. Child safety seats are installed in vehicles by properly attaching and tensioning the main anchorage points. Proper use of the attachments is extremely important as these significantly reduce child head injuries in motor vehicle crashes. Attachments are inconveniently located on child safety seats and difficult to use. Several large studies have observed that only 10% to 20% of children are correctly harnessed into correctly installed seats. Improper use of child restraints substantially reduces their effectiveness and is a major public health concern. The proposed child safety seat will be extremely easy-to-use and will dramatically increase the visibility and usability of the attachments. We hypothesize that a child safety seat that provides a convenient, easy-to-use attachment will significantly improve the rate of proper child safety seat usage and reduce vehicle crash-related child injuries and deaths. An interdisciplinary team of researchers has been assembled to define, build, and evaluate a prototype system. The production development will be completed in a rigorous quality-controlled design and testing process to ensure a safe and effective product is produced.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Salsolinol (6,7-dihydroxy-1-methyl-1,2,3,4-tetrahydroisoquinoneline;SAL) is a natural derivative of dopamine. It has been reported that alcohol consumption increases the level of SAL. While R-SAL is formed from enzymatic condensation of dopamine with pyruvic acid, non-enzymatic condensation of dopamine with acetaldehyde, the oxidation product of ethanol, generates the racemic mixture of (R,S)-SAL in vivo. To test a possibility of SAL representing a marker for alcohol addiction, a reliable method for quantification of the (S)-SAL stereo isomer from biological matrices is being established. The previously reported method employed the GC/MS analysis after isolation by solid phase extraction and derivatization by silylation of hydroxyl groups and acylation of the secondary amine with a chiral reagent (R)-2-phenylbutyryl chloride. Upon examining the sensitivity and reproducibility of the method, we found that this approach suffers from drawbacks including poor recovery and stereoisomer conversion during the analysis process, preventing the reliable determination of the S isomer. To improve the sensitivity and stability of the SAL stereoisomer analysis, a new approach using pentafluorobenzyl (PFB) derivatization is being explored. By optimizing derivatization procedures, SAL was derivatized fully at both hydroxyl groups and the secondary amine and analyzed by LC/MS using atmospheric pressure chemical ionization or by GC/MS using electron capture negative ion chemical ionization technique. The resolution of the stereoisomers as well as the sensitivity of the technique with both approaches is being evaluated.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Molecular cloning has identified at least four E-Prostanoid (EP) receptor subtypes, designated EP1, EP2, EP3 and EP4 which mediate the effects of Prostaglandin E2 (PGE2). The EP3 receptor modulates water and ion transport in the kidney, gastric acid secretion, neurotransmitter release, and hepatic glucose metabolism. In the kidney PGE2 production modulates salt and water transport in the medullary thick ascending limb and collecting duct. Importantly, evidence suggests that some EP3 effects are mediated from the basolateral surface of the nephron, while others are mediated by apical receptors. The EP3 receptor is unique among the EP receptor family in that it exists as multiple isoforms generated by alternative splicing of a common precursor mRNA transcribed from a single gene. Although the EP3 receptor has classically been characterized as a Gi coupled receptor that signals by lowering intracellular cAMP levels, recent studies suggest that EP3 alternative splice variants couple through a variety of signal transduction pathways. The proposed studies will focus on the functional differences of the alternatively spliced PGE2 receptor subtypes thereby providing insight into the roles of the multiple PGE2 receptors in normal and pathophysiological states. The principal hypothesis of this proposal is that differential expression of EP3 receptor splice variants in individual cell types results in differential activation of signaling pathways that determine the physiologic response of a target cell to PGE2. In Specific Aim #1 the principal investigator will determine the repertoire and tissue distribution of EP3 receptor splice variant messenger RNAs. Immuno-localization will be performed, including subcellular receptor targeting of selected EP3 receptor proteins using receptor selective antibodies in relevant tissues as well as in cell culture models. In Specific Aim #2 selected cDNAs encoding rabbit EP3 receptor splice variants will be expressed in mammalian cell lines and their ligand binding and signal transduction properties characterized. The third Specific Aim will determine the effects of site-specific amino acid substitutions on G-protein activation receptor phosphorylation and receptor desensitization. Completion of these studies will not only elucidate the significance of these multiple EP3 splice variants, but will also increase our understanding of the structure and function of G-protein coupled receptors.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Abstract The Research Translation Core (RTC) will partner with the Community Engagement Core (CEC) to help implement the various data sharing and IIRT plans for the individual projects and cores. During the next funding cycle, the RTC will assist Dr. Gallagher (Project 1) in developing material and scheduling presentations on the results of his studies to community members and agencies responsible for site remediation. The RTC will assist Dr. Xia with her new biomedical project focused on the effects of cadmium exposure on adult neurogenesis (Project 2). We will build on Dr Xia's previous activities focused on support groups for patients with neurodegenerative diseases such as Parkinson's and Alzheimer's. Dr. Furlong (Project 3) is Co-director of the RTC and is our primary contact with the UW's Center for Commercialization (C4C). Dr.Furlong will continue to share his expertise regarding PON1 status assays with investigators involved in pesticide exposure research. He will also continue his work with ATSDR and CDC in developing new protocols for measuring OP exposures and will work with these organizations and EPA regarding the implications of the results of his studies regarding children's increased susceptibility to OP pesticides. We will assist the investigators of the two new site based projects, Dr. Racette (Project 4) and Dr. Neumann (Project 5), who have already developed ties with community organizations and regulatory agencies working on their study sites. For Project 4, we have added Dr. Nelson to our RTC staff to help build upon their successful work in the communities surrounding the Manganese (Mn) emitting smelter in Meyerton, South Africa. RTC staff will develop appropriate educational materials related to Mn exposure and effects and will share the results of the study as it unfolds. RTC staff will also work with Drs. Racette and Nelson to ensure the latest findings of the study are shared with the Department of Environmental Affairs. For Project 5, we will use our strong ties with the State Department of Ecology (the agency responsible for developing remediation strategies for the lake) and EPA Region 10 to provide lay-friendly material related to the objectives of the study and the potential implications of the results for site remediation and water quality standards. RTC staff will work with the Training Core to include trainees in activities, such as involvement in our Science and the Public journal club and developing materials for their community engagement rotations. The RTC will coordinate the activities of all investigators and cores in developing and executing our Continuing Education Courses in years 2 and 4. Advances in research in environmental exposures and neurodevelopmental and neurodegenerative disorders will be the focus of the initial course. Investigators will also be involved in the Workshops, Summits, and Webinars that the CEC will host. Finally, RTC staff will utilize the SRP Community Engagement/Research Translation Data Collection Form reporting system developed by the NIEHS to ensure all research advances and community engagement activities are reported.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Since 1988, an interactive and multi-disciplinary team of investigators at the Case Western Reserve University and the Tuberculosis Research Unit have effectively and creatively fostered the development of scientific, clinical, laboratory, and education programs in the area of infectious disease (tuberculosis, HIV-AIDS) with investigators at the Makerere University (MU) in Kampala, Uganda and the AIDS Control Program of the Ugandan Ministry of Health. The intent of this application is to continue to build on this program by expanding expertise within the center for AIDS research at CWRU and create an International Clinical Coordinating Center (ICCC). The core facility will provide an administrative structure with which to coordinate and considerably expand upon the international collaborative endeavors of CWRU and to foster realistic and meaningful clinical advances to clinicians, scientists, and other healthcare providers in Uganda with the emphasis on certain aspects of the HIV/AIDS pandemic. The creation of the ICCC core facility represents an unprecedented and exciting opportunity to join basic and clinical researchers from six-NIH designated centers at CWRU inclusive of: The Tuberculosis Research Unit, Dr. J.J. Ellner, Director; University Hospitals Ireland Cancer Center (UHICC), Dr. James Wilson, Director; Skin Disease Research Center (SDRC), Dr. Craig Elmets, Director; AIDS Clinical Trials Unit (ACTU), Dr. Michael Lederman, Director; Fogarty AIDS International Training and Research Program, Dr. Christopher Whalen, Director; and the Center for AIDS Research (CFAR), Dr. Stuart Le Grice, Director. The specific aims of this proposal arena to enhance interaction between Ugandan and CWRU clinicians and scientists in the HIV/AIDS are inclusive of tuberculosis, medical mycology, AIDS- related malignancies, and primary HIV vaccine development; to coordinate research activities in the field, clinic, and hospital settings in Kampala, Uganda and research programs/laboratories at CWRU; and to create an environment in Uganda for collaborative research suitable for realistic and meaningful technology transfer. Key research projects that would be funded by the International Core are DNA metabolizing enzyme targeted therapy (CA62502, J. Wilson), Viral etiology and cellular signals in AAKS (CA43703, J. Wilson), preparation for AIDS vaccine evaluation AI35173/CPV1, J.J. Ellner); Preventative therapy for TB in Uganda(CCU50671605, J.J. Ellner); Tuberculosis prevention and control unit (AI45244, J.J. Ellner); Impact of TB on HIV infection in Uganda (AI32414, C. Whalen).
|
{
"pile_set_name": "NIH ExPorter"
}
|
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. An essential element of the RFA entitled Institutional Clinical and Translational Science Award (CTSA) is the creation of an academic home that is a department, center, or institute. In response to the RFA, the University of Iowa will create The University of Iowa Institute of Clinical and Translational Science ("the Institute"). The director of the Institute[unreadable]and PI of this application[unreadable]will report directly to the provost of The University of Iowa. The Institute will serve as an overarching academic structure for all clinical and translational research and training at The University of Iowa. Its overall functions are entirely consistent with the goals of the CTSA and the expected institutional commitments outlined in the RFA. The Institute will be supported by a significant cost-sharing effort between the CTSA, The University of Iowa, and various entities within the State of Iowa community. It will have dedicated faculty who will serve as mentors for all trainees. The career development of these faculty and all trainees will be the primary responsibility of the Institute. The Institute proposes to unify existing training programs for clinical and translational research under one new program[unreadable]the Graduate Program in Clinical and Translational Science. This program will have a K12, T32, one-year, and summer components. In addition, there will be training for nurse " coordinators and collaborating researchers in the community. A major feature of the program is a new community education and research network. Initially this network will start in four large centers and ultimately will involve the entire State of Iowa. The Institute will provide and support key functions to facilitate clinical and translational research at Iowa. This includes a large investment in information technology that is entirely supported by Institutional resources. This system should provide an ideal environment for research and training at The University of Iowa and translation of these research studies into best practices in the state. The application has the support of all relevant health care entities within the State of Iowa.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Short bowel syndrome (SBS), a major clinical problem affecting patients of all ages, results from the functional or anatomic loss of extensive segments of small intestine. SBS has an overall 5-year survival of 70%, and in newborn infants with less than 10% of expected intestinal length, 5-year survival is only 20%. Current treatment options are inadequate and associated with severe complications and death. Patients with SBS require total parenteral nutrition (TPN) to survive. Currently, over 20,000 SBS patients per year are on home TPN in the US alone. Annual costs of home TPN exceed $300,000 per year per patient ($6 billion/year). In addition the use of TPN is associated with numerous central venous catheter-associated infectious and thrombotic complications, resulting in additional high costs. Thus, SBS represents an extremely costly, and deadly, burden to society. At present, the treatment for SBS is mainly supportive. Although small bowel transplantation is an option, the results are suboptimal, with 1-year and 4-year survival rates of 90% and 60%, respectively, and the required life-long immunosuppression causes substantial secondary complications. Novel approaches for the treatment of patients with SBS are critically needed. Our approach to this unsolved medical problem is the production of tissue-engineered intestine (TEI) using the patient's own intestinal cells combined with a synthetic nanofiber-based scaffold, resulting in a novel solution to this unmet clinical need. In this project, we will accomplish three specific aims necessary to move towards commercialization: Aim 1) Determine the optimal cell isolation and cell seeding methodology for structural formation of TEI. Aim 2) Optimize enteric nervous system (ENS) development in TEI. Aim 3) Develop an in vivo culture standard operating procedure (SOP) for development of TEI. This Phase I SBIR project will allow us to collect the pivotal data needed to scale up to a large animal model in Phase II, and to license the technology to a commercial partner in Phase III for commercialization.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Close relations, especially romantic partners, can exert profound influence on alcohol use both in everyday life and in treatment contexts. However, the specific mechanisms underlying these effects are not well understood. Elucidating the nature of these interpersonal processes and their role in the maintenance or reduction of high- risk drinking behavior is crucial to addressing problematic alcohol use in romantic relationships. Based on previous models of social control (i.e., influence, regulation) of health behavior, the nature of the social control strategy (i.e., overt, covert), as well as the recipient?s emotional and behavioral responses to that strategy, are relevant constructs that will be examined. The proposed Mentored Research Scientist Award (K01) entails structured training and research activities that will enable the applicant (Dr. Fillo) to examine the nature, use, and relative effectiveness of the full range of social control strategies used among romantic partners aimed at reducing high-risk drinking behavior. A long-term career goal of the candidate is to establish an independent research program investigating the psychosocial mechanisms linking close relationships and high-risk alcohol use, and this award is a critical initial step toward that goal. The candidate will be mentored in professional development (i.e., responsible conduct of research, grantsmanship, conference presentations) and receive extensive training in three key areas: 1) alcohol-related influence processes in close relationships, 2) measure development and validation techniques, and 3) intensive longitudinal methods and data analysis. The candidate proposes a three-phase project using samples of heavy drinking adults in committed, romantic relationships. A comprehensive measure of social control strategies will be adapted from existing measures and prior qualitative research, supplemented with items derived from focus groups (Study 1), and validated using online survey methodology (Study 2). In Study 3, the use and relative effectiveness of overt and covert social control strategies among romantic partners will be prospectively examined as they naturally unfold using daily diary methods. It is hypothesized that covert social control strategies can effectively influence partners? high-risk drinking behavior by avoiding the negative emotional and behavioral recipient responses often associated with overt strategies. By examining the role of interpersonal processes in the pursuit of identifying and measuring mechanisms of behavior change, the proposed research aligns well with NIH?s cross-cutting program on the Science of Behavior Change. The training environment of the Clinical and Research Institute on Addictions, combined with a knowledgeable and experienced mentorship team (Drs. Leonard, Testa, McCrady, Tucker, Lucke, Derrick), will provide the candidate with unparalleled resources and support during the award period. Furthermore, the proposed training and research experiences will facilitate the development of critical skills and enable the candidate to successfully launch an innovative program of independent research investigating psychosocial mechanisms linking close relationships and problematic alcohol use.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Co-Principal Investigator: Rev. Micah Park, Young Sang Church PROJECT SUMMARY Colorectal cancer (CRC) is the second most commonly diagnosed cancer and the third highest cause of mortality in Asian Americans. Despite a decline in the incidence of CRC in the general population, its incidence in the Korean community is rising. Korean men ranked the second highest in incidence and mortality from CRC compared with those of non-Hispanic White men. Among Korean women, CRC is the second most commonly diagnosed cancer. Although the efficacy of routine and timely screening has been demonstrated as a viable preventive measure against the development of the disease, various studies have shown that CRC screening rates among Koreans are extremely low for every type of CRC screening, and when screening is obtained, it is more likely to reveal a late stage of the disease and poor prognosis for 5-year survival. Our studies ofthe Korean community in PA and NJ indicated consistently low rates of CRC screening (13% to 17%, in Koreans vs 52% in general populations) and multiple barriers to health care. About 80% Koreans we serve are medically underserved and low income, lack of knowledge about CRC risks, screening benefits and health system, and have limited or no access to culturally and linguistically appropriate health care. There is a lack of CRC screening intervention program tailored to the needs of Koreans. The proposed project will address this gap by evaluating the effectiveness of a multifaceted and culturally appropriate Korean church-based intervention using community-based participatory research (CBPR) approach to significantly improve CRC screening among medically underserved members of Korean churches (30 church sites) in PA and NJ. The project will be built on an established long-standing infrastructure of community-academic-clinical partnership that has collaborated successfully on several CBPR guided projects in Korean and other Asian communities, as well as a pilot CBPR CRC intervention among Koreans. The overall goal of the study is to maintain and ensure sustainable partnerships using CBPR approach to increase CRC screening by reducing healthcare access barriers for underserved Koreans. Specifically, the study aims to examine whether a culturally appropriate CBPR CRC intervention is more effective in increasing screening, knowledge of, perceived risks of and susceptibility to CRC and benefits of screening, as well as reducing health system barriers to screening. CBPR principles and logic model will be used to guide all phases of the study in developing and implementing project plans, procedures, intervention, evaluation and dissemination. The study design is a 2-arm group randomized trial with baseline and post-intervention assessment and a 12-month follow-up. A total of 30 Korean churches will be randomized to either an immediate intervention or control group (general cancer education plus delayed intervention). The proposed intervention, guided by an integrative framework of Health Belief Model and Social Cognitive Theory, addresses both individual and healthcare system barriers through multifaceted innovative approaches. Key components include: (1) group education by trained bilingual community health educators and church health workers;(2) patient navigation by Community Health Educators (CHEs) and Community Health Workers (CHWs);and (3) engaging community bilingual physicians in CRC screening and referral. The proposed study is expected to yield important and new data on the inten/ention effects. If this CBPR CRC intervention proves effective, it can be used as a model program that has potential transportability to and sustainability in Korean and other Asian communities nationally, hence make a substantial contribution toward reducing health disparities.
|
{
"pile_set_name": "NIH ExPorter"
}
|
It is proposed to evaluate permeability of electrotonic junctions between embryonic cells. Use of ion sensitive microelectrodes should allow accurate correlation of junctional resistance with permeability to appropriate ions injected iontophoretically. Permeability to other small molecules will be tested by iontophoretic or pressure injection. Fluorescence, histochemical staining or isolation of coupled uninjected cells will be used to assay transjunctional movement. Preparations suitable for determining permeability in adult tissues will be developed, and comparisons made with embryonic cells. The role of cell movement in junction formation will be assayed by use of specific inhibitors. Existence of precursor molecules for junction formation will be studied by determining the cell's ability to form junctions in the presence of inhibitors of protein synthesis. The mobility and segregation of potential sites of coupling in the membrane will be investigated by marking specific regions of the cell surface. The goal is to determine the structure of the hypothesized channels connecting the cell cytoplasms and to analyze the mechanisms involved in their formation. This information should be uniquely accessible in the experimental systems to be employed and it should provide insight into the role of these junctions in development as well as in functioning of the adult tissues in which they are so prevalent.
|
{
"pile_set_name": "NIH ExPorter"
}
|
PROJECT SUMIVIARY (See instructions): Cell culture technology plays an essential role in modern cancer research. The Cell Culture Facility (CCF) serves as a customized pay-for-service Facility that provides expertise, technical support, reagents, equipment, liquid nitrogen storage, and training to increase the effectiveness and efficiency of using mammalian cell culture in Fox Chase Cancer Center (FCCC) laboratories. This Facility provides technical assistance and consultation services in tissue culture techniques, centralized liquid nitrogen banking, mycoplasma screening services, the preparation of custom and standard culture media and supplements, performance testing of fetal bovine serum, cell propagation, and technical support and supplies for mouse embryonic stem (ES) cell transfection and culture. Major efforts by the CCF include carrying out technically challenging techniques, such as the establishment of primary cell cultures from patient or animal tissue samples, hybridoma fusion and cloning services to generate monoclonal antibodies, and establishing gene knockout or knock-in Embryonic Stem (ES) cells to generate genetically manipulated mice. Since the last review the Center established a governing body for the CCSG supported Shared Resources called the Facility Parent Oversight Committee (FPOC). The FPOC recommended in 2008 that the CCF incorporate the Hybridoma Facility, which was previously approved as a distinct CCSG Facility. This merger has provided operational efficiencies by centrally locating the merged staff. Economic efficiencies have been achieved by reducing total staff by 2.05 FTE, and by presenting a reduced overall request from the CCSG. The merged CCSG budget request is $19,724 less than the combined levels approved for previously two distinct cores at the last review. In 2009, the Facility served 47 investigators with peer-reviewed funding. Service was provided to all five of the Center's CCSG Research Programs. Peer-reviewed, funded research accounted for 95.3% of CCF usage in calendar year 2009. Based on an annual quality survey administered by the FPOC, the Facility is highly regarded among faculty users, and received an Outstanding to Excellent rating at the last CCSG review. In the past five years, the demand for CCF services has remained strong. Importantly, the Facility supplies the FCCC community with valuable on-site expertise for a host of established cell culture protocols, consultation in the design and evaluation of experiments, training in cell culture methodologies, high quality culture media and supplements, and adaptable assistance delivered in a cost-effective manner.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Of the many immunotherapy approaches under development, the ability to use bispecific antibodies or chimeric antigen receptors (CARs) to direct T-cells to selectively kill tumor cells has demonstrated significant early success. Typically, bispecific antibodies or bispecific T-cell engagers (i.e., BiTes) cross-link T-cells by binding to CD3 and to a target cancer cell surface antigen, usually through a monovalent interaction. An alternative approach is to genetically engineer a cancer patient?s T-cells to express a single chain antibody (scFv)-CD3? fusion protein that can target the cancer cell surface antigen. After re-introduction into the patient, CAR-expressing T-cells have been able to selectively eliminate the target cancer cells. While successful, the genetic engineering of cell surfaces is time consuming and irreversible and the use of BiTes requires continuous infusion. Furthermore, the resistance to both approaches due to antigen loss has been observed. Our group has shown that two dihydrofolate reductase molecules (DHFR2) fused to an ?CD3 single chain antibody (scFv) can be engineered to spontaneously self-assemble upon the addition of the chemical dimerizer, bis-methotrexate (BisMTX), into either highly stable octavalent chemically self-assembled nanorings (CSANs). CSANs have been prepared with BisMTX containing a third arm, thus enabling it to be conjugated to oligonucleotides, fluorophores, radiolabels and drugs. Recently, we have prepared ?EpCAM/?CD3 CSANs and ?CD133/?CD3-CSANs. The bispecific CSANs rapidly (min) and stably (days) bind to CD3 on T-cell membranes, thus forming chemically self assembled prosthetic antigen receptor (PAR) T-cells. Upon incubation of the PAR T-cells with EpCAM+ and/or CD133+ cancer cells, rapid and selective killing of primary and tumor initiating cancer stem cells (CSC) was observed. We have also demonstrated with an orthotopic murine cancer model that ?EpCAM and ?CD133 PAR T-cells are non-toxic and able in combination to eradicate tumors in vivo. A unique safety feature of our approach is the ability to remove the CSANs from the T-cells by dosing with the FDA-approved non-toxic antibiotic trimethoprim at clinically relevant concentrations, thus allowing us to deactivate the cells pharmacologically and reduce cytokine release. Consequently, as an alternative to current approaches, we determine the generality T-cell induced killing and eradication of TNBC tumors with ?EpCAM/?-CD3-CSANS and ?CD133/?CD3-CSANS. In addition, we will develop a tripspecific ?EpCAM/?CD133/?CD3 CSANs that will allow the simultaneous elimination of TNBC primary tumor cells and CSC. The successful completion of the project milestones should result in the elucidation of the key features governing a chemical biologically based non-genetic and reversible method for T-cell targeting, as well as the importance of CD133 on TNBC proliferation. These rules will be applicable to the clinical development of anti-cancer immunotherapy with greater selectivity, lower toxicity and a reduced ability for the development of resistance.
|
{
"pile_set_name": "NIH ExPorter"
}
|
We developed a suite of four transcript prediction algorithms collectively called "FEAST" (Fast Empirical Algorithms Suggesting Transcripts), which are conceptually independent of the two established classes of gene discovery algorithms, namely "ab initio" and database search methods. The main goals of this proposal are (1) to develop further this independent third class of gene prediction algorithms, (2) to apply them to the dentification of novel genes in the genome, and (3) to test the hypothesis that non-coding transcripts are prevalent in the genome, and are the medium for the expression of small RNA genes and other functional genomic elements. We will extend the statistical model and develop the software towards a fully integrated gene prediction tool capable of discovering genes in genomic sequences of one species, or in multiple species simultaneously for higher precision. We will use the new tool to produce a comprehensive catalog of predicted genes. This is the genetic "parts list", that is required for the construction of metabolic and regulatory models of cell function. We will correlate the transcript predictions to expression data from hybridization array technology, and validate novel genes experimentally by RT-PCR and sequencing. We identified an unusual class of genes (which we call "stencil" genes) in which the exons play no other role than the production of introns as precursor material for deriving one or more functional RNA molecules, like miRNAs and snoRNAs. We will put special emphasis in obtaining a comprehensive catalog of such "stencil" genes and will study computationally their prevalence, their modes of regulation and how they evolve. We expect many of the novel transcripts to be central to the genetic regulation of development, and therefore of direct importance to cancer research. [unreadable] [unreadable] [unreadable]
|
{
"pile_set_name": "NIH ExPorter"
}
|
Abstract Colorectal cancer is one of the most common cancers in both men and women in the United States and accounts for approximately 140,000 new cases annually. Hepatic metastasis, the dominant feature of life- limiting disease, occurs in approximately 20?50% of patients with colorectal cancer. Although regional treatment options, including hyperthermic isolated hepatic perfusion (HIHP) and percutaneous IHP, offer the benefits of both aggressive local treatment and limited systemic toxicity, the management of unresectable colorectal liver metastases remains a major unsolved issue and more effective novel regimens are needed. During the grant period, we will develop a novel strategy for targeted HIHP therapy. We hypothesize that a combinatorial treatment of targeted hyperthermia, the biologic agent Fc-TRAIL (immunoglobulin Fc domain fused tumor necrosis factor-related apoptosis-inducing ligand), and the chemotherapeutic agent oxaliplatin will be effective in treating unresectable colorectal liver metastases. The specific aims of this project are to (1) elucidate the mechanism of synergistic anti-tumor efficacy caused by Fc-TRAIL-based targeted multimodal treatment, and (2) investigate the preclinical efficacy of this combinatorial treatment in an IHP rat model. The proposed studies for the first aim will employ biochemical and molecular techniques to investigate multimodal treatment. For the second aim, we will employ a syngeneic hepatic metastasis rat model of colorectal carcinoma to study the efficacy of the multimodal treatment. We believe that our proposed regimen will provide information on the potential therapeutic advantage of an Fc-TRAIL-based multimodal treatment on patients.
|
{
"pile_set_name": "NIH ExPorter"
}
|
PROJECT SUMMARY Candidate: Dr. Kelli Ryckman is a Postdoctoral Research Fellow for the Carver College of Medicine at the University of Iowa. Her primary research focus is on the genetics and epidemiology of complex reproductive disorders including preterm birth and related complications. Her short-term career goals are to enhance her knowledge of the genetics of complex reproductive and perinatal conditions in newborns and to gain additional skills in analytical methods and techniques for elucidating risks of complex diseases. Her long-term goals are to become a funded independent scientist in the fields of genetics, statistics and epidemiology as tenure-track faculty at a University. Her research plan is innovative in combining genomic and metabolomic profiles to understand complex disease risks. This research will utilize the Iowa Newborn Metabolic Screening Program (INMSP) where a deep resource of phenotypic, environmental and genetic components is available. This research has the potential to identify new biological pathways involved in neonatal disease and to have direct clinical impact on universal newborn screening programs. Mentors and Consultants: Dr. Jeff Murray will serve as primary mentor on this award. He is a well-funded full professor at the University of Iowa with 25 years of experience in training successful graduate and postdoctoral students. Dr. Mary Marazita, an expert in the field of statistical genetics, will serve as co-mentor of this award. Drs. Stanton Berberich and Kristi Borowski will serve as consultants on this project by providing clinical and statistical genetic expertise. Drs. Sara Copeland, John Dagle and Jonathan Haines will serve as external advisory members and will consult with, advise and evaluate the progress of Dr. Ryckman throughout the award period and have complementary expertise in the metabolomics of the neonatal biochemical milieu, the complications of preterm birth and genomic analysis. Environment: The University of Iowa has an established reputation for the successful training of graduate and postdoctoral students, particularly in research and clinical areas. The University offers a range of learning and development seminars as well as other programs focused on career development and advancement and particularly benefits from training opportunities deriving from its ICTS (Institute for Clinical and Translational Sciences) support by a CTSA award. Career Development Plan: Dr. Ryckman plans to meet regularly with her mentors, consultants and external advisory members to gain the knowledge necessary to obtain her career and research goals as well as attending tutorials, courses, seminars and relevant conferences. Research Study: The focus of this study is to use the Iowa newborn screening data as a model for studying the metabolomic and genomic profiles of newborns and relate the profiles to human disease, particularly prematurity and its complications.
|
{
"pile_set_name": "NIH ExPorter"
}
|
According to the 1992 National Adult Literacy Survey, 25 percent of US adults are functionally illiterate. Because written health information is frequently provided at above the tenth grade level, health messages are not reaching low literacy patients jeopardizing their health status. Low literacy has been found by researchers to have a detrimental effect on general health and the use of medical services. Evidence from research in medical settings consistently highlights the importance of comprehension and literacy for patient compliance and increased positive health outcomes. Although much is known about medical health literacy, there are no studies that have systematically studied oral health literacy. The purpose of this current application is to examine how a low dental literacy population interprets dental health prevention information, navigates the dental health system and whether participation in a large comprehensive public health program (WIC) can be effective in improving this process for pregnant women and their children and reduce health disparities. Specifically, we will: 1) Determine the dental health literacy rates among a low income, high risk population, 2) Examine the role of the Women, Infant, and Children's Program in understanding preventive dental health information such as consent forms, oral hygiene instructions and compliance to fluoride instructions among a low income/low dental health literacy population 3) Examine the role of the Women, Infant, and Children's Program in navigating the .dental health care system among a low income/low dental literacy population including compliance for dental appointments and appropriate use and 4) Evaluate the effects of the Women, Infant, and Children's Program on cumulative treatment and costs of dental services provided by Medicaid as well as oral health related quality of life in a population with low dental health literacy. [unreadable] [unreadable] [unreadable]
|
{
"pile_set_name": "NIH ExPorter"
}
|
The systematic combination of anatomy and nomenclature is an essential component of all computer processes that seek to visualize, relate, manipulate, and compare three-dimensional human anatomic structures. Structure names must be elemental, categorized and associated with standard comparison images in order to: create computer visualizations at multiple levels of detail; associate structures spatially and symbolically; assign anatomic units their physical characteristics; and categorize new patient- derived anatomic data. This project will demonstrate that the Systematic Combination of Anatomy and Nomenclature (SCAN) can enable semi-automatic identification and segmentation of structures in radiological data. The first long-term goal of this project is to relate three-dimensional anatomic knowledge in the Visible Human Data (VHD, a full digital description of the human being) to the symbolic anatomic knowledge in the Systemized Nomenclature of Human and Veterinary Medicine (SNOMED, a data table relating terms to codes), which is a component of the Unified Medical Language System (UMLS, a relational database). This project will identify elemental gross anatomic structures within the nomenclature of SNOMED and relate them to registered, segmented slice image sets and high quality Virtual Reality Modeling Language (VRML) surface models derived from VHD. The second long-term goal of this project is to link the standard anatomic knowledge to new three-dimensional radiological data for any patient. This project will generate a semi-automated process for combination of patient- specific anatomic data and nomenclature using mutual information for automatic multimodality image fusion (MIAMI Fuse, a process which integrates three- dimensional images of different modalities into a single image set) to link new data to standardized data and subsequently to nomenclature. By integrating standardized three-dimensional and symbolic anatomy, coupled with semi-automated fusion of new radiological data to these standards, SCAN will provide the foundation for widespread advances in biomedical visualization, simulation, medical education, diagnostics, and treatment planning.
|
{
"pile_set_name": "NIH ExPorter"
}
|
This project concerns identification of unusual patterns of cancer within specific occupational groups that may be indicative of exposure to hazards in the work environment. Study groups are selected for a variety of reasons including 1) known or potential exposure to established or suspect carcinogenic substances, 2) prior case reports and case-control studies suggesting an unusual cancer experience for the group, 3) availability of existing sources of data for defining a population-at-risk. Data sources include employment and death records maintained by companies, membership and death listings maintained by labor unions and professional organizations, and state licensing records. Studies underway include cohort mortality studies of persons employed in dry cleaning establishments, machinists, foundry workers and pest control operators. Proportionate mortality studies are being conducted for workers in the petrochemical industry and pharmaceutical and biological manufacturing, chromate painters, leather workers and metal workers.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Feline leukemia viral infected cats are a unique model to study immunosuppression, acquired immunodeficiency syndrome and immune mechanism(s) involved in the initiation and progression of neoplastic growth. In this proposal we plan to take advantage of the dramatic unequivocal reductions of tumor size, changes in marrow and peripheral blood, neoplastic cell populations and clearance of viremia following treatment of virus infected and/or malignant cells with SpA therapy (ex vivo immunoadsorption or injection). We will treat additional animals to determine whether the responses are biostatistically significant. We will examine both humoral (Complement (C) and C activation products) and cell mediated immunity. We will first identify T cell subsets which are poorly defined in the cat. Monoclonal antibodies (Mabs) to kitten thymocytes have been initiated and we have established for the feline system a reverse plaque hemolytic assay for evaluating B cells and roles of T helper or suppressor on induction of Ig synthesis. We now have a Mab which identifies T-S cells. A major goal is to mimic in vitro immunosuppressive FeLV effects observed in vivo i.e. to study the effects on non infectious FeLV and its purified components on T cell functions. We have already shown that U.V. FeLV has a direct suppressive effect feline IFN production and IgG secretion in vitro. Furthermore p15E peptide homologous to retroviral envelope proteins has a dramatic suppressive effect on polyclonal B cell activation by Staphylococcus Protein A. If we can show immunomodulatory influences of retrovirus and its peptides on animal or human cells, not only will we obtain a greater understanding of the in vivo potent immunosuppressive effects of retroviruses, particularly in acquired immunodeficiency syndrome, but also the immunoregulatory role of viruses may become sources of powerful pharmacologic agents.
|
{
"pile_set_name": "NIH ExPorter"
}
|
DESCRIPTION (provided by investigator): With increase in life expectancy, there has also been a parallel increase in chronic co- morbidities that impair functional status and quality of life in older age. Converging lines of evidence link sleep quality and quantity with cardiovascular health. A considerable body of literature demonstrates that short sleep duration (<7 hours) is associated with increased levels of cardiovascular disease (CVD) risk factors including obesity, hypertension, and diabetes in population-based studies. Further, evidence suggests that poor health status also increases the prevalence of sleep disorders. This relationship may be particularly prominent in older adults, who often have the burden of both sleep disturbances and medical co-morbidities. The proposed study is uniquely able to examine these matters cross-sectionally (in older people), and prospectively -- in particular the relation of early adult CVD risk factor levels to sleep quality and sleep disorders in older age by examining a pre-assembled cohort of the Chicago Heart Association Study with CVD risk status characterized approximately 40 years ago. The specific aims are to: a) determine the relationship between CVD risk status, especially low risk (LR), at ages 25-44 with subsequent sleep quality, levels of daytime sleepiness, and sleep apnea in older age (65-79);b) assess whether change in risk factor status from ages 25-44 to older age relates to sleep quality, daytime sleepiness, sleep apnea, neuropsychological performance, and cardiovascular health status in older age, and c) explore relationships among current CVD risk factors, CVD, sleep quality, sleepiness, sleep apnea, daytime neuropsychological performance, and other variables in older age. Both objective and subjective measures of sleep are to be used in conjunction with measures of CVD risk (BMI, blood pressure, lipids, glucose, smoking, C-reactive protein), clinical and subclinical CVD (coronary artery calcium, ankle brachial index), obtained by the recently funded parent grant (R01 HL081141;Principal Investigator, Martha L. Daviglus). 1,380 participants, identified at baseline as LR for CVD or not LR for CVD, are to undergo subjective and objective sleep measures (questionnaires and actigraphy). A subset of 150 participants (50 LR who do not become high risk by older age, 50 LR who became high risk, and 50 high risk at baseline who remain so in older age) are to attend the laboratory for more detailed physiological studies of sleep. By leveraging resources of the parent grant, this ancillary study is uniquely positioned to study these multiple relationships among CVD risk at ages 25-44, sleep variables, and other traits in older age. Results of this study can have important implications theoretical and practical, e.g., for prevention and treatment of sleep disorders and CVD in later life. PUBLIC HEALTH RELEVANCE: Recent studies indicate a strong link between sleep and sleep disorders, such as sleep apnea with cardiovascular health. A considerable body of evidence clearly demonstrates that short sleep duration disrupts metabolic and cardiovascular function and has been associated with increased levels of cardiovascular risk factors, such as C-reactive protein, BMI, blood pressure, lipids, coronary artery calcium, ankle brachial index, glucose, and smoking. Therefore, an improved understanding of the relationship between sleep and cardiovascular health will lead to preventive approaches and treatment strategies to improve function, health, and overall quality of life in the growing population of older adults.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Severe combined immunodeficiency disease (SCID) is a fatal congenital disease of children. Approximately 1/3 of cases are associated with the deficiency of adenosine deaminase (ADA) and some of the remaining cases with the X-linked variant have mutations of the IL-2 gamma receptor gene. The severe nature of this illness, the deficiency of single peptide proteins, and the ability to correct the disease by allogeneic bone marrow transplantation have led many investigators to explore the use of gene transfer technology as a therapeutic option for the treatment of SCID. The long term goal of the work proposed here is to utilize gene transfer technology to effect efficient transfer into reconstituting hematopoietic stem cells and stable expression in progeny of these cells of genetic sequence defective in SCID in order to provide life-long cure of this disease. The basis of the approaches to gene transfer proposed in this grant include the realization that published data, including from our own laboratory, demonstrate that current protocols for infection of reconstituting stem cells of large animals species are not optimal. Furthermore, a wealth of experimental data suggests that the hematopoietic microenvironment is a source of both positive and negative regulators of hematopoiesis and these signals may be important for successful and efficient transduction of primitive hematopoietic stem cells. The hypothesis to be tested in this proposal is that transduction of long term reconstituting stem cells can be accomplished in large animal transplant models by retroviral infection in the presence of critical components of the hematopoietic microenvironment. We will utilize our previously characterized simplified retroviral vector which expresses the ADA cDNA and both in vitro and in vivo assays for human stem cells to analyze transduction efficiency and expression of introduced sequences. In addition, we will continue to use primate transplants to evaluate gene transfer technology in a large animal autologous transplant model.
|
{
"pile_set_name": "NIH ExPorter"
}
|
This study addresses the development of young children's sex identity and knowledge of culturally prescribed sex roles, and the process by which maternal childrearing behaviors contribute to this development. Previous research has demonstrated that by the time children are 5-6 years of age, consistent sex differences appear in children's choices of activities, their motives when engaged in an activity, and their performance styles. There are few detailed data on factors contributing to these differences. The goal of this study is to fill this gap. Maternal behaviors with girls and boys are being observed over a range of activities. This study is a by-product of the laboratory's Rearing Study which provides extensive interactional data on 100 mothers and their children, and thereby permits a focus on sex of child as a determinant of rearing environment. The data sources are videotaped observations of mothers and their 2-3 1/2-year-old children in a variety of naturalistic settings. Similar observations are obtained two years later. In the follow-up observations, a series of gender related probes have been incorporated which will provide information about the mother's conscious preferences for her child as well as the child's own preferences for her/himself and knowledge of the cultural sex stereotypes.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Detailed transcriptional maps and translational maps of selected portions of the vaccinia virus genome were constructed. More than 12 early transcripts were characterized and two early genes were sequenced. These studies revealed that early mRNAs are colinear with the genome, unspliced, and have multiple closely spaced 5' ends. The nucleotide sequences for about 60 base pairs upstream of transcriptional initiation sites are extremely A T rich and contain long runs of As and Ts with some homology to prokaryotic and eukaryotic consensus sequences. The eukaryotic A2TA3 poly(A) signal sequence was not found near the ends of the genes. Marker rescue techniques were developed to map functional vaccinia virus genes. In this manner, thymidine kinase was mapped to the HindIII J fragment located near the center of the DNA molecule.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The overall goal of this proposed research is to develop a multi-well insert apparatus that will facilitate rapid automated detection of reporter genes in living fish from embryonic to juvenile stages of development. This system will be relevant for any zebrafish-based assay which utilizes fluorescent or luminescent reporter genes as a means of evaluating a biological question of interest. Zebrafish are becoming an important model for understanding human disease and for discovery of new therapeutic compounds. In particular, zebrafish are well-suited to large-scale screening procedures, providing an in vivo validation system that approaches high-throughput capacities. Full realization of this potential will require the development of automated assay platforms, such as reporter based systems utilizing fluorescent or luminescent read outs, and the inclusion of fish at more mature stages of development. During Phase I, components of a system will be developed that will enable embryos to be maintained in individual chambers for up to 1 month of development;and enable a high-throughput primary screen. During Phase II a second component of the system based on the PSI SideView imaging technology will be developed that will enable a more rigorous, image-based secondary screen. PUBLIC HEALTH RELEVANCE: This research will develop technologies that will enable the use of zebrafish models of human disease in high-throughput screens for active pharmaceutical compounds. One key area that this technology will benefit is regenerative medicine with a particular focus in this application on regeneration of insulin-producing pancreatic cells for diabetes treatment.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Our goal is to eventually develop site-specific inhibitors of the HIV-1 protease. To that end we continued our investigations on the effect of heavy metals on protease activity. Also, other related areas on the biochemistry of the HIV-1 protease were investigated. We found that peptide copper chelates as well as nonpeptidic copper complexes were able to strongly inhibit the protease activity similar to that of free copper. The chelates appeared to be working by delivering copper to the site of action in the protease. Data on the inhibition of the protease by copper in the presence of dithiothreitol indicated that it, too, may function as a copper chelate inhibitor. Therefore, we may be able to use delivery systems like these to specifically target the protease. An antibody was produced which recognized the HIV-1 protease on Western blots. This antibody will be used to further probe the mechanism of copper inhibition. It was also found that albumin could significantly stabilize protease activity and was useful in studies of the protease run under native gel filtration. Activity of the protease was recoverable under conditions where albumin was present in the running buffer or in the collection tubes. The HIV-1 protease is essential for the production of infectious virions from infected cells. A clear understanding of the biochemistry of the HIV-1 protease should allow for the development of novel inhibitors of this important target for the prevention of AIDS.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Amyloid deposition in cerebral vessels (CAA) is a constant feature in A[unreadable] and non-A[unreadable] cerebral amyloidosis. Notably, vascular and not parenchymal deposits appear today to be more sensitive predictors of dementia. Unrelated amyloid subunits produce similar lesions in medium/small cerebral vessels and, as often neglected, in capillaries, suggesting common pathogenic mechanisms. To date, it is neither clear why specific mutations predominantly associate with cerebrovascular pathology nor understood why the main phenotype for some vasculotropic variants is hemorrhage/ischemic stroke while for others is cognitive impairment. Co-localizing with the amyloid deposits there is also a group of so-called amyloid-associated components, among them markers of cellular stress and inflammation whose role as active key players in the processes of amyloidogenesis and cell toxicity is far from being defined. Complicating the picture, oligomerization/aggnegation but not fibrillization of amyloidogenic peptides appear to be a pre-requisite to exert both, their cytotoxicity and their ability to trigger an inflammatory response. Although these effects are partially studied in neurons and glial cells, little is known about these mechanisms in the cells forming the vessel wall. Intriguing as well is whether the presence of CAA genetic variants exerts enhanced detrimental effects on cell toxicity and differentially alters endothelium properties, e.g. monolayer permeability. We hypothesize that conformational transitions of amyloidogenic peptides are key elements responsible for the pathogenic vascular features characteristic of the CAA including, among others, protein oligomerization and amyloid formation, cell toxicity, complement activation and the induction of inflammation-related mechanisms. Accordingly, oligomeric and fibrillar assemblies of synthetic A[unreadable] variants in the presence and absence of selected amyloid associated components will be tested for their ability to induce a cytotoxic effect on cerebral endothelial and smooth-muscle cells and elicit concomitant permeability alterations in endothelial monolayers (aiml) as well as to trigger complement activation and the release of vascular and glial inflammatory components which, in turn, may worsen the toxic phenotype (aim 2). When feasible, results will be validated with similar assemblies extracted from well typified CAA/AD cases. Whether or not specific membrane receptors / binding molecules are mediators of the cellular responses will be explored in aim 3.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Genetic polymorphisms in the CCR5 locus impact susceptibility to HIV-1 infection and rates of disease progression. This project investigates the biologic basis for these impacts. Promoter polymorphisms have been shown to impact the rate of CCR5 receptor recycling, and may have the effect of reducing or increasing the number of available target cells for HIV-1 entry, a process that depends uupon CD4 and CCR5 expression. GCRC Normal Blood Donors are essential to this project.
|
{
"pile_set_name": "NIH ExPorter"
}
|
PSL representatives have tested and evaluated software packages as part of the Personal Workstation Project. Eleven word processing packages have been evaluated to determine the best ones for support by DCRT. Many criteria were used to decide that DisplayWrite2 is the most convenient for NIH needs at this time.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The objective of this four year project is to develop improved techniques for diagnosing, determining the risk for, and monitoring the course of treatment of osteoporosis. To achieve this goal we will undertake four studies. These include ultrasound and DEXA measurements, on human subjects, the construction of a new radiological scanning device for measuring bone density, the construction of a new ultrasound scanner for determining Broadband Ultrasonic Attenuation (BUA) and the Speed of Sound (SOS) in cortical bone, hypothesis of a composite model of bone and of fracture risk. Our human subject studies will test the hypothesis that high levels of correlation can be obtained between cortical and trabecular bone measurements using ultrasound. Human studies will also allow us to follow the course of treatment of patients taking osteoporosis medications. Our new radiological scanner will use the highly sensitive coherent to Compton photon scattering ratio (CSR) technique to measure the "true" volumetric. We will test the hypothesis that right-left differences in the calcaneus and distal radius are good indices of osteoporosis. We will test our new CCSR and ultrasound scanners on human subjects. A model of bone as a composite material made of collagen and mineral will be used to calculate fracture and osteoporosis. We will test our new CCSR and ultrasound scanners on human subjects. A model of bone as composite material made of collagen and mineral will be used to calculate fracture and osteoporosis risks and to plan treatment for those diagnosed with or at significant risk for osteoporosis.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Research on the influence of gender on sexual behavior has tended to focus on women, leading to a relative lack of knowledge of the male side of gender, and the development and content of men's perspectives on their sexual behavior, prompting NICHD to issue PA-05-033, Men's Heterosexual Behavior and HIV Infection. In addition to the paucity of research on men's perspectives, HIV-prevention research has employed limited theoretical models that focus primarily on cognitive factors as predictors of decision-making and behavior. In the proposed project, we will develop and test an expanded model of heterosexually active young men's sexual risk taking behavior, explicating mechanisms as well as outcomes. Using a sexual scripts perspective, we will incorporate intrapsychic, interpersonal and cultural levels into this new model, which will expand on the social cognitive models that have dominated sexual risk behavior research in the first two decades of HIV research. We will examine gender role norms and ideologies, concurrent partnerships, sexual violence perpetration and victimization, and emotional arousal. The project will contribute both to empirical knowledge of young adult men's perspectives on sexual behavior and sexual risk- taking, and to development of theory that incorporates cultural, emotional, and interpersonal factors as predictors of sexual behavior. The research will proceed in three phases: Initial interviews will elicit young men's sexual scripts and examine their relationship to sexual risk and protective behaviors. These interview data will then be used to describe the sexual scripts embedded in these stories, and to create synopses of the scripts to identify script elements that are of theoretical or empirical interest in relationship to sexual risk. In the second phase, we will use these script elements to develop scales to refine a survey that will be administered to a larger sample of men in the third phase. The goal of the third phase is to survey a sample of 500 men to assess the degree to which these scripts act as mediators between hypothesized predictor variables and HIV risk/protective factors. Sufficient numbers of African-American, Asian-American, and white men will be included for analyses and comparison of these ethnic groups. [unreadable] [unreadable] PUBLIC HEALTH RELEVANCE: The lack of knowledge of the male side of gender, and the development and content of men's perspectives on their sexual behavior limits our ability to design effective HIV- and STI-prevention interventions. This project will extend our understanding of heterosexual men's behavior and provide a base on which to develop new theories and interventions. [unreadable] [unreadable] [unreadable]
|
{
"pile_set_name": "NIH ExPorter"
}
|
Progression of renal disease leads to common consequences including interstitial fibrosis, glomerulosclerosis, vascular narrowing, and renal failure, ultimately requiring dialysis or renal transplantation. The kidney myofibroblast is the cell type most responsible for matrix accumulation during renal fibrosis. Although there is a large amount of data on mechanisms of interstitial myofibroblast matrix synthesis late in the course of renal fibrosis, information on the earliest cellular events involving myofibroblast encroachment into the perivascular and interstitial spaces prior to matrix accumulation is lacking. Experiments are proposed to examine mechanisms of myofibroblast encroachment (migration and proliferation) in a model of accelerated renal fibrosis in which myofibroblasts first originate from perivascular and periglomerular regions. Our central hypothesis is: Activation of PDGFR-2 and TGF-beta receptor induces fibroblast migration and proliferation into the peritubular interstitium via Rho/ROCK modulated ROS generation and transduction of PI3-kinase/Akt, MAPK (ERK1/2) signaling pathways early during the progression of renal fibrosis. The following aims are set: Aim 1. NAD(P)H oxidase-derived ROS are involved in myofibroblast interstitial encroachment early during the course of kidney fibrosis; Aim 2. ROS-induced interstitial myofibroblast migration, proliferation and matrix synthesis is a consequence of PDGFR-2 and TGF- beta receptor activation; Aim 3. ROS generation through PDGFR-2 and TGF-2 receptor regulate SMAD2/3, Rho/ROCK, ERK1/2 and Akt signaling cascades, initiating interstitial myofibroblast migration, proliferation, and matrix synthesis. PDGF BB and TGF2-1 initiate ROS generation in vascular disease. Experiments are designed to critically examine a role for NAP(P)H oxidase and associated phox subunits in ROS generation, PDGFR-2 and TGF-2 receptor activation and signaling pathways on myofibroblast migration, proliferation and matrix synthesis in vitro and in vivo. Specifically, a role for the NAD(P)H oxidase pathway (Nox2 and Nox4 and phox subunits) will be examined in PDGFR-2 and TGF-2 receptor transduction of SMADs, Rho/ROCK, PI3K, and ERK1/2 regulation of activation of kidney myofibroblast. These studies will provide much needed insight on mechanisms of myofibroblast activation during the early stages of renal fibrosis.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Specific somatic mutations in hematopoietic stem and progenitor cells (HSPCs) provide a proliferative advantage, leading to clonal hematopoiesis and the development of myeloproliferative disorders (MPDs) and acute myeloid leukemia (AML). Studies of patients with TET2 or DNMT3A mutations show that hematopoiesis can be clonal without pathology, suggesting that clonal expansion may be a prerequisite for disease development. It is unclear which combination of these mutations are necessary or sufficient to promote clonal dominance or whether the order of the mutational events are critical for disease progression. The zebrafish has emerged as an excellent model for hematopoietic malignancies, notably acute lymphoblastic leukemias. Models of MPDs and AML show low penetrance with no overt AML, although it is not clear in these models whether a non-pathological clonal dominance is established. It would be interesting to evaluate clonality of normal HSCs as genetic perturbations occur in the premalignant state. Viral clonal marking would be helpful, but the relatively small number of HSCs present in the zebrafish kidney marrow require a less complex clonal labeling system. To investigate HSPCs at the clonal level, we have optimized the Brainbow system for use in zebrafish hematopoiesis. This system extends clonal analysis to the erythrocyte lineage, which is nucleated in zebrafish but not in mammals. Zebrafish that ubiquitously express the Brainbow construct and contain a blood specific CreERT2 were treated with tamoxifen at various embryonic stages. Treated fish were grown to adulthood, and peripheral blood and marrow samples were subjected to confocal and FACS analysis to observe color barcodes. Using automated cluster analysis on normalized intensities of the Brainbow fluorescent proteins, we detected multiple and unique barcoded cells (average of 4-8) in all lineages of the blood as far as 8 months post treatment. The efficiency of recombination in this system allows for labeling of 20-80% of the mature blood pool. Transplants of Brainbow-labeled whole kidney marrow near limiting dilution into irradiated recipients showed strong monoclonal engraftment, suggesting that HSCs are being labeled. Using this system, we will investigate mutations that are necessary or sufficient for generation of clonal dominance. Our studies will provide insight on early clonal events that regulate normal and premalignant hematopoiesis.
|
{
"pile_set_name": "NIH ExPorter"
}
|
This is a resubmission of a competing renewal proposal of grant RO1 EB000246-17 for the continuation of our studies on the fundamental understanding of transmucosal delivery of drugs, peptides and proteins using a series of novel biopolymeric complexation carriers with unique properties and in advanced controlled release systems. It has generally been believed that peptides and proteins such as insulin cannot be administered via the oral route because of their degradation by the proteolytic enzymes in the gastrointestinal tract and their extremely slow rate of transport across the mucosal membrane. A successful oral delivery system for protein drugs could lead to the development of new therapeutics with increased patient compliance and efficacy. Our group has developed a new class of polymeric complexation hydrogels comprised of poly (ethylene glycol) (PEG) chains grafted on poly (methacrylic acid) (PMAA) backbone chain. The new systems, henceforth designated as P (MAA-g-EG), have shown extreme promise as oral delivery vehicles for insulin, calcitonin, and growth hormone. Prior studies have established that these systems are promising candidates for oral delivery of insulin. Yet, our more recent work and attention is now focused on continued strategies for enhancing the bioavailability of oral delivered proteins, in addition to insulin. The methodologies we intend to pursue in this competing renewal proposal include: (i) developing further strategies for protection of the protein in the GI lumen;(ii) enhancing transport of the proteins across the GI lumen;and (iii) increasing the residence times in the upper small intestine through enhanced bioadhesion. While some strategies have been previously investigated with moderate success using insulin, in this work, we will pursue novel strategies that may provide significant advances over our prior studies with insulin. Additionally, we intend to further develop these complexation hydrogels for oral delivery of other therapeutic proteins such as calcitonin and growth hormone. PUBLIC HEALTH RELEVANCE: There is a general scientific belief that oral delivery of protein drugs such as insulin, calcitonin, and growth hormone is impossible due to the protective nature of the GI tract and extensive proteolytic activity of enzymes in the GI tract. We have developed a novel, complexation hydrogel system that has been engineered to protect the protein drugs in the GI tract and enhance transport of the drugs into the bloodstream. A successful oral delivery system for protein drugs could lead to the development of new therapeutics with increased patient compliance and efficacy.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The objective of this work is to study in mice the molecular basis of teratomas that arise from the germ cells. One aspect of the investigation requires that knowledge of the mechanisms of X chromosome inactivation be acquired since inactivation of the X chromosome seems to be a fundamental step in differentiation and possibly the development of teratomas. Our approach to this problem is to assess whether differences in the amount of an enzyme, phosphoglycerate kinase, which is coded for by a gene on the X chromosome, can be detected in primordial germ cells from female, male and X/O embryos. Another aspect of the problem is that since phosphoglycerate kinase, coded for by an X-linked gene, seems to be regulated coordinately with phosphoglycerate mutase and triosephosphate isomerase, two enzymes that are probably coded for by genes on autosomal chromosomes, information on the interrelations between genes on the X chromosome and those on autosome can be obtained by investigation of the basis of this coordinated regulation. The initial approach to this problem will be to produce specific antibody against the purified enzymes and determine their relative rates of synthesis and degradation.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The skeleton is one of the main targets of estrogen (E) action, as E regulates bone growth and remodeling. Although decreased E levels are known to be one of the main causes of osteoporosis, the specific molecular pathways by which E regulates bone metabolism are not fully characterized or understood. This Project uses a combination of novel mouse and cell models to define, at the cellular and molecular level, the signaling pathways by which E regulates bone turnover and bone mass, focusing on two modalities by which the estrogen receptor-alpha (ERa) functions: the classical mode of action, where ERa directly interacts with estrogen response elements (EREs) on DMA, and the non-classical mode, where ERa indirectly functions through protein-protein interactions with other transcription factors. Preliminary data demonstrates that mice containing an ERa mutation that eliminates classical ERa signaling (NERKI) exhibit osteopenia and impaired bone formation in both cortical and trabecular bone in males, but only in cortical bone in females. We hypothesize that due to increased E levels in female mice, ER(3 functions to mitigate the negative effects of the NERKI receptor in trabecular bone. In Aim 1, we directly test this by assessing whether loss of ERB leads to greater skeletal deficits in female, but not male NERKI mice. We will examine bones from wild type, NERKI//ERB+/+, and NERKI//ERB-/- mice using bone densitometry and histomorphometry. Aim 2 tests this hypothesis at the cellular level by assessing the ability of bone marrow stromal cells from these mice to commit and differentiate along the osteoblast lineage. Using quantitative polymerase chain reaction assays, we will test the hypotheses that expression of the NERKI receptor leads to impaired responses to Wnts and BMPs, and that ERP modulates these effects. Aim 3 examines loss of classical ERa signaling on the recruitment of the NERKI receptor to non-classical DNA binding sites using chromatin immunoprecipitation assays. Finally, Aim 4 uses a novel transgenic approach to examine the skeletal consequences of selective replacement of the endogenous ERa with the NERKI receptor only in osteoblasts. Collectively, these studies will provide a more detailed understanding of ER signaling pathways in bone.
|
{
"pile_set_name": "NIH ExPorter"
}
|
A year primarily of writing and re-analysis of New Guinea records has resulted in completion or near completion of a paper on pacemaker involvement in the free run flashing of Pteroptyx cribellata and manuscripts on artificial driving of the flashing of P. cribellata and of two other, rather different, species. The work has illuminated particularly the cycle-by-cycle independence of both spontaneous and driven flashing and the several devices, including reversion to free-run flashing and abrogation of flashing, by which the pacemaker copes with entrainment to rhythms markedly shorter than or longer than free run. In addition Mrs. Buck and Dr. Lesley Ballantyne have a paper in progress on a remarkable non-synchronizing species which lives in the surf zone.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Fungal pathogens are responsible for a variety of opportunistic diseases related to AIDS. This proposal describes a high throughput screen for identifying new antifungal agents specifically targeted against protein translation by screening for growth in two yeast mutants, one overexpressing yeast elongation factor EF-3, the other underexpressing the same factor.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Follicular helper T (TFH) cells are critical for development of germinal center (GC) B cells responses following immunization with TD antigens and in systemic autoimmune responses; however, the development and function of these cells have been less clearly defined in both normal individuals and in patients with systemic lupus erythematosus than other CD4 T cell effector subsets. These issues have been a focus of study for the past several years, with characterization of CD4 T-B helper cells in murine models of lupus and in conventional immune responses, with the idea that information gleaned from the latter studies could be applied to the understanding of autoimmunity. The proposed work builds on these observations, and the more recent finding that TFH cells require the expression of the transcriptional repressor Bcl6 (B-cell CLL/lymphoma 6) for their development and for their function in B cell maturation in GCs - when upregulated in CD4 T cells, Bcl6 initiates a program of gene expression that drives a TFH cell phenotype. Preliminary studies have demonstrated that Bcl6 is also required for downregulation of P-selectin glycoprotein ligand-1 (PSGL1), a surface protein on T cells that engages the chemokines CCL19 and CCL21 expressed in the T cell zone of secondary lymphoid organs. Indeed, TFH cells in normal and systemic autoimmune responses are uniformly PSGL1lo. These observations suggest that downregulation of PSGL1 would disengage binding to these chemokines, allowing T zone egress with migration towards the B cell follicle where TFH cells are functionally required. This would support a scenario in which Bcl6, in its role as an initiator the TFH cell developmental program, is required for both T zone egress and for development of functional TFH cells. During these studies on TFH cell development and function, the transcription factor, Ascl2, a conserved member of the Drosophila Achaete scute-like 2 complex, was also found to be selectively upregulated in this T cell subset compared to other CD4 T cell effector subsets and compared to B cells. Expression of Ascl2 also is sufficient for development of a TFH cell phenotype. Based upon these findings, two hypotheses will be addressed in this proposal. First, PSGL1 acts as a retention signal for T cells in the T zone, and that in addition to driving TFH cell development, Bcl6 expression initiates downregulation of PSGL1 as a critical first step in T cell migration to the B cell follicle and ultimately in TFH cell function. Second, the transcription factor Ascl2, like Bcl6, plays a necessary role for TFH cell development and/or function. These hypotheses will be explored in two specific aims: 1) determining the roles of Bcl6 and PSGL1 in the initial events that lead to TFH cell migration and development; 2) asking if the transcription factor Ascl2 is required for TFH cell differentiation and function, and if so, a dissection of the mechanism of its T cell-specific effects in immune responses.
|
{
"pile_set_name": "NIH ExPorter"
}
|
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 is focused on understanding the interaction between human and schistosoma proteins, with the aim of identifying drug targets and/or vaccine candidates for protection from schistosomiasis. Previous work on this project has been to analyze active secretory proteins from Schistosoma mansoni during initial host exposure to invasive larvae as a method for identifying functional proteins involved in invasion and host immune evasion. This work has expanded to analyze the the natural substrates for the proteases of invasive schistosome larvae in skin, where Schistosome cercariae are incubated with sections of human skin and products of degradation extracted. In a parallel track, the active proteases across multiple species including Schistosoma mansoni, Schistosoma japonicum and Schistosomatium douthitti were characterized for proteolytic classification and activity.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Poor adhesion is the single most severe problem related to the long term viability of implantable hydroxyapatite coatings. An innovative room temperature electrophoretic process is proposed to fabricate nanostructured hydroxyapatite ("n"-HAP) coatings, eliminating problems related to amorphous phase formation and delamination. To date, only micron-sized hydroxyapatite materials have been deposited using electrophoresis. The n-HAP coating will exhibit increased toughness, bond strength, and a high degree of crystallinity. The nature of the bond is metallurgical, resulting in dramatically improved bond strength when compared to mechanical bonds obtained in thermal spray coatings. Electrophoresis also enables multicomponent codeposition. This program consists of (1) synthesizing n-HAP particles, (2) depositing the n-HAP coating, and (3) coating evaluation. while the n-HAP is expected to yield a several4old increase in bond strength, a calcium phosphate cement (CPC) additive will be cc-deposited with the n-HAP to further increase bond strength. A dense graded TiO2 bond coat will be introduced (a few microns thick) between the n-HAP and the titanium substrate in the deposition process, so that body fluids will have no opportunity to attack and degrade the titanium. Inframat is collaborating with Dr. Antoni Tomsia of the Lawrence Berkeley National Lab to evaluate the nanostructured coatings. PROPOSED COMMERCIAL APPLICATIONS: Potential commercial applications include advanced coated implants for hips, knees, and other prostheses in humans and animals. In the US alone,the annual number of hip and knee operations using HAP coated implants is estimated to already produce an annual revenue of $3 billion. With the increasing age of the larger population, the estimated demand for implants will rise significantly in the near future. A comparable European market doubles the potential revenue.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Techniques are currently available for propagating foreign DNA segments in mammalian cells using SV40 and other papovaviruses as cloning vectors. The foreign DNA may be inserted into the late region of SV40; 1. between two appropriate restriction endonuclease sites; 2. by the poly(dA:dT) joining procedure; or 3. by joining at one endonuclease site and allowing for circularization in vivo. If the size of the inserted DNA segment is less than or equal to the fragment excised from the late gene region, one may co-infect permissive cells with a tsA helper viral DNA and hybrid and helper genomes will be packaged into virions at the non-permissive temperature. An analogous approach to this involves insertion of the foreign DNA into the early gene region of SV40 DNA and the subsequent cloning with a late gene ts mutant helper (tsB or C). Alternatively, one may construct hybrids larger than wild-type SV40 and propagate these molecules as free-replicating episomes in appropriate permissive or semi-permissive host cells. A limitation with this approach at present is the difficulty in the selection of cloned cell lines which retain viability, express the SV40 T-antigen and continue to replicate non-rearranged recombinant viral genomes. We have commenced work aimed at development of improved eukaryotic cloning vectors. Our approach is to insert the thymidine kinase gene of Herpes Simplex virus type I into the late gene region of polyoma virus or SV40 and then use tk-host cells for selection of these vectors. We have attempted to propagate an SV40-argF hybrid which contains approximately 1100 base pairs more than wild-type SV40 DNA by insertion of the 1650 base pair argF segment from E. coli between the EcoRI and Bam HI sites on the SV40 genomes. A significant fraction of transfected cells (5%) became T-antigen positive and were presumably replicating the argF-SV40 hybrid. However, this episome was lost from these cells after a few weeks.
|
{
"pile_set_name": "NIH ExPorter"
}
|
This is a parallel application from researchers at the Population Research Centers of the University of Wisconsin and University of Maryland. The overall goal of this application is to examine patterns, transitions, and determinants of health in a comparative framework within countries of the Latin American and Caribbean region. The study seeks to contrast patterns within the region and also with those found in the U.S., in particular with populations of Mexican origin, taking advantage of recently gathered, high- quality, comparable, and as of yet under-analyzed survey data on elderly people. The goal is to exploit the available cross-country data to identify conditions that promote (inhibit) the emergence of relations between health status and disability and well-specified determinants, and to assess how recurrent and generalizable are relations found in any one country. The specific aims of the project are: to estimate profiles and determinants of health status and disability of elderly populations in Mexico, and in seven capital cities of Latin America (Argentina, Barbados, Brazil, Cuba, Chile, Mexico, and Uruguay); to identify determinants of health and disability status using cross-sectional data to contrast health and disability profiles of populations in Mexico with that of Mexicans and Mexican-origin populations living in the U.S.; to estimate correlates of health status changes, disability transitions and mortality in Mexico and the U.S. with particular emphasis on the role of early childhood conditions, experiences with illness, and on the influence of income and wealth. Because of their increasing importance in the region, the project focuses more narrowly on diabetes and obesity, and proposes to estimate models for incidence and duration of diabetes as well as health-related costs associated with these conditions. In pursuing these goals the study adds to and fine-tunes existing methods and procedures to help address two issues: health selection among immigrants to the U.S., and estimation of couple-models to assess the effects of select determinants on health and mortality net of shared attributes.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Cytoplasmic inclusions of the TAR DNA-binding protein of 43 kDa (TDP-43) is frequently found in degenerating neurons of patients with Amyotrophic lateral sclerosis, Frontotemporal Dementia, and Alzheimer?s Disease. TDP-43 is a ubiquitously-expressed, tightly-regulated, and predominantly nuclear DNA/RNA-binding protein and the cause and mechanism underlying the formation of TDP-43 protein inclusions remains unclear. Furthermore, modeling these inclusions has proven challenging and it is unclear if these are neurotoxic or neuroprotective. To address these issues, we developed an optogenetic approach to induce TDP-43 inclusions in live cells. We show that these inclusions mimic the properties of TDP-43 inclusions found in patient neurons that are neurotoxic. We further show that RNA binding status dictates the propensity for TDP-43 for form inclusions in live cells. In this grant, we propose to identify the characteristics of neurotoxic TDP-43 assemblies employing our light-induced system. Specifically, we will describe the post- translational modifications, localization, and material state of neurotoxic TDP-43 assemblies. Since our preliminary data indicates that RNA binding state determines whether TDP-43 proteins can oligomerize into a pathogenic conformation, we propose to study whether bait oligonucleotides designed to bind RNA deficient TDP-43 proteins confer neuroprotection in cortical neuron cultures, and in vivo using a TDP-43 Drosophila model system developed in our labs. These studies have the potential to address pervasive questions regarding the neurotoxic state of TDP-43 assemblies and the development of a new use for oligonucleotide therapeutics.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The proposed research will be a physical/chemical investigation of protein-protein interactions involved in regulation of contractile activity in skeletal, cardiac, smooth, and non-muscle cells. A survey of the shape of regulatory and essential myosin light chain subunits and the effects of phosphorylation on the interaction between the regulatory light chain and the heavy chain of myosin and on the conformation of both the isolated light chain and the whole myosin and its regulated proteolytic subfragments using sedimentation velocity and equilibrium ultracentrifugation, analytical gel permeation chromatography, fluorescence anisotropy decay measurements, circular dichroism, nuclear magnetic resonance and electron spin resonance spectroscopy. This work will help answer the question of how the regulatory subunits interact with each other after being activated by calcium ion to allow actin-myosin interaction, ATP hydrolysis and, ultimately, contraction to occur.
|
{
"pile_set_name": "NIH ExPorter"
}
|
[unreadable] Under funding for the original U19, we addressed several hypotheses that have provided new insights into the fundamental, mechanistic understanding of regulatory T cells and how these pathways are altered in animal models of autoimmune disease and in the human diseases type I diabetes and multiple sclerosis. In addition, we have developed novel immunotherapeutic approaches to induce regulatory T cells using oral anti-CD3. The grant will continue to focus on these fundamental aims, reflecting the discoveries we have made over the past five years. In this renewal, the overall goals of this Autoimmunity Prevention Center Project are: [unreadable] 1) To determine, in human autoimmune disease, which of the CD4+CD25+ subsets of DR+ and DR" regulatory T cells are defective; [unreadable] 2) The identification of a "core set" of genes and proteins that are expressed on the "innate", CD4+CD25+ regulatory T cells; [unreadable] 3) To determine which of the "core sets" of genes and proteins are altered in the regulatory T cells found in blood and lymph nodes in patients with diabetes and MS; [unreadable] 4) To understand the mechanism of oral anti-CD3 and GRAIL transfection in T cells that allows the translation into human clinical trials by year five of the grant. The key goal is the development of specific drug targets and methods to induce the function of defective regulatory T cells in patients with autoimmune disease. [unreadable] [unreadable] Lay Summary: Diseases such as type I diabetes and multiple sclerosis are complex genetic diseases thought to be initiated by an autoimmune response directed against self-proteins in the inflamed tissue. Why autoreactive T cells attack the insulin producing islet cells in diabetes or the myelin in multiple sclerosis remains a major question. We recently demonstrated there is a loss of an important "regulatory" immune cell in the circulation of patients with autoimmune disease. This grant assembles a group of investigators to understand why these regulatory cells are dysfunctional, and what can be done to restore their function. [unreadable] [unreadable] PROJECT 1: Mechanism of action of CD4+CD25+ T cells (FATHMAN, GARRISON C.) [unreadable] [unreadable] DESCRIPTION (provided by applicant): [unreadable] This is a competing renewal as a U19 for a previously funded U01 entitled, "CD25+ Regulator CD4+ T Cells." Under funding for the original U01, several specific aims were addressed: initially (1) that expression of GRAIL (a recently identified anergy gene), following peptide administration iv, blocks IL-2 transcription and induces anergy, a form of tolerance, and (2) that GRAIL expression provides a novel and effective screen for the anergic phenotype in mice (and in man), and (3) that CD4+CD25+ suppressor T cells were involved in this form of tolerance induction, and a fourth (4) GVHD will be blocked by adoptive transfer of Tregs, was added in the second year of support. Ten articles were published on these four specific aims and this proposal will extend these studies as follows: Using microarray and RNAi technology for mRNA expression and gene silencing, the "core genes" that define Treg core transcriptome will be identified. In this proposal, these "core Treg" genes, identified by cDNA microarray studies, will be validated by functional genomics (RNAi) and tested in vitro in T cell proliferation assays and in vivo in a model of GVHD, and the "peripheral Treg subset," provisionally defined as CD4+ antigen specific T cells that contact antigen under anergy inducing conditions in the periphery, will be further characterized and studied as proposed in the following four specific aims: [unreadable] * Specific Aim 1: Identification of a "core set" of CD4+CD25+ Treg genes, the Treg core transcriptome. [unreadable] * Specific Aim 2: Characterization of peripherally induced Tregs and core transcriptome identification. [unreadable] * Specific Aim 3: GRAIL transductants as Tregs for immunotherapy. [unreadable] * Specific Aim 4: To evaluate the role of specific genes on Treg function in an in vivo model of graft vs. host disease. [unreadable] [unreadable] The use of adoptive cellular therapy, in particular use of Tregs is rapidly gaining credibility as a useful potential therapy for immunoregulation, in particular in the setting of GVHD, a form of adoptively transferred autoimmune disease. Studies proposed in this project will attempt to identify the "core" set of genes that define Tregs and then using RNA silencing techniques, attempt to identify functionally relevant genes through knock down and loss of function assays in vitro and using a model of GVHD in vivo. All of the techniques required for these studies are currently practiced in our labs. [unreadable] [unreadable] [unreadable] [unreadable]
|
{
"pile_set_name": "NIH ExPorter"
}
|
Project Summary This RFA is aimed at bringing together interdisciplinary teams to focus on novel, transformative and integrative efforts that will revolutionize our understanding of the biological and bioinformatics content of the data collected from non-invasive human functional brain imaging techniques. Our proposal does exactly this. We are a multidisciplinary team of scientists with combined expertise in optogenetics, two photon Ca2+ imaging, biomedical engineering, molecular biology, animal and human fMRI, network theory, data analysis and modeling. In this work, we will use a novel imaging device that combines mesoscopic imaging of genetically encoded Ca2+ indicators with very high (50?m) spatial and high temporal (25ms) resolution across the entire cortex and simultaneous fMRI in transgenic mouse models. These animal experiments are designed to complement similar experiments in healthy human subjects. The results from the animal experiments will answer several long-standing questions about the source of the fMRI signal. Specifically, using imaging, we will quantify the contributions of different cell populations (excitatory neurons, inhibitory neurons, and glial cells) to the fMRI signal observed. We will be able to test and validate, for the first time, the application of graph theory approaches to the analysis of human fMRI data, and we will develop and test a new approach based on control theory for extracting more information from the fMRI signal. A powerful set of carefully controlled imaging experiments in mice will inform several aspects of analysis of human data. The human data will contain a test/retest component to ensure replication of the results and to allow predictive models to be built in one data set and tested in another. This work truly bridges scale and modalities and the simultaneous nature of the animal experiments will allow unprecedented clarity on the underlying source of the signal changes observed in fMRI. These animal studies are essential for providing new insights into the basis of human fMRI signals and data of this nature has not previously been available. The work in this proposal is novel in that it will directly inform measures of both evoked and spontaneous activity in terms of the underlying cell signal sources revealing the relative contributions of excitatory, inhibitory and glial cells to the fMRI signal. The implications of the work are multifaceted. This work will provide a platform for evaluating neurological models of disease. For example, mouse models of disease can be used to link to human data in diseases such as PTSD, depression, and autism, to name a few. It will also provide a firmer biological basis for understanding the node and network measures used in assessing the functional organization of the brain and will have important implications for the design of therapeutic interventions across a range of diseases.
|
{
"pile_set_name": "NIH ExPorter"
}
|
We have been studying F-incompatibility, a function of the E. coli F factor related to the control of its replication. Incompatibility exists between an integrated F factor and an autonomous F factor and between two autonomous F factors. In order to determine the mechanism(s) of incompatibility, we will try to isolate more inc- mutants and to analyze and map these and the mutants we have already isolated by genetic and physical methods including DNA heteroduplexing and electron microscopy. In studying incompatibility between 2 F' factors we have found that selection for the presence of both F' factors often results in fusion of the 2 F' factors to form a single genetic and physical unit. To elucidate the process of fused plasmid formation we shall use heteroduplex analysis. We also plan to isolate phi 80 transducing phages carrying inc genes. These will be used to do physical mapping of the inc region by heteroduplex analysis and to study transcription and tranlation of F DNA in an in vitro system. In addition, we will study plasmids which carry genes for the production of enterotoxin in enteric bacteria (ent plasmids) and Pseudomonas plasmids. Their relationships to other known plasmids will be investigated by heteroduplexing techniques and by testing for incompatibility between these and other plasmids belonging to known incompatibility groups.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The proposed research will continue a systematic investigation of probable in vivo regulatory mechanisms, which control the establishment, maintenance and reversal of enzyme specific latency during cellular aging in the slime mold, Dictyostelium discoideum using trehalase as a model system. The specific aims to be achieved will be: (1) to determine the basis for the increase in trehalase specific activity during germination; (2) to localize intracellular trehalase in the dormant spore by application of the techniques of immunoferritin and/or indirect immunofluorescent staining using purified specific antibody elicited against this enzyme; (3) to determine the basis for the decrease in trehalase specific activity during the aggregation process and (4) to resolve, purify and characterize the multiple forms of trehalase from aging, mature sorocarps and to determine the basis for changes in their specific activity during sorocarp formation.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Neurophysiological studies have shown how disparity-tuned neurons in primary visual cortex make the initial disparity measurements. However, the response of these neurons bears little relation to human depth perception. While the disparity-tuned neurons code absolute disparity, and are little affected the context of stimuli falling within their receptive fields, depth perception depends heavily on relative disparity, and is greatly influenced by the surrounding disparity field. How does this transformation from the disparity signals to the signals that generate perceived depth occur? One aspect of human stereopsis that is probably limited by these primary neurons is stereoacuity. Yet, stereoacuity may also be influenced by stimulus conditions that profoundly affect perceived depth. Stereoacuity can thus serve as a psychophysical probe for how the signals generated by disparity-tuned neurons are combined in subsequent processing stages. In the proposed research, we will measure how the surrounding context (long-range interactions and depth contrast) affects stereoacuity, since context has important effects on perceived depth. In addition, we will examine use stereo transparency measurements to estimate the pooling area for disparity signals, as well as how these signals are combined. The experiments in this proposal will examine stereo processing in normal human observers. About 5 percent of the population suffers from oculomotor (strabismus) or refractive (anisometropia) disorders that threaten the development of normal stereopsis. While most individuals can cope with the loss of stereopsis, abnormal binocular development is also frequently associated with a deficit in the acuity of one eye (amblyopia) -- a more serious problem. These studies of normal observers will provide basic knowledge about normal stereopsis, a prerequisite for understanding abnormalities of the binocular system.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Low circulating levels of vitamin D [25-(OH)D] have been associated with various adult onset cancers. However, studies of childhood cancer have not been conducted despite compelling experimental and epidemiologic evidence suggesting that reduced vitamin D during gestational development may be a risk factor, particularly for childhood brain tumors (CBT). For CBT, the in utero period of development has been shown to be a critical time window of exposure. Neonatal dried blood spots (DBS) collected and archived as part of newborn screening programs represent a unique resource for examining pre-diagnostic 25-(OH)D in relation to childhood cancers and allows for focus on the impact of vitamin D during the in utero time period. The long-term goal is to understand the pathogenesis of CBT so that interventions can be developed to reduce the burden of this disease among children. The overall objective for this application, which is a step toward attainment of the long term goal, is to examine the association between neonatal 25-(OH)D, as measured in DBS, with risk of CBT. The central hypothesis of the proposed study is that vitamin D deficiency during pregnancy is associated with increased risk of CBT. The hypothesis will be tested by pursuing two specific aims 1) Evaluate among 250 childhood brain tumor cases and 250 controls the association between 25-(OH)D, as measured in DBS as a marker of gestational circulating vitamin D levels, and the risk of CBT; 2) In exploratory analyses, determine if the association between 25-(OH)D and brain tumor risk differs by sex, race/ethnicity, prenatal smoking, maternal SES, brain tumor histology, birth weight, gestational age and month and season of birth. This study is innovative because it will, for the first time, examine circulating vitamin D levels as predictors of CBT risk by using measurements in DBS as markers of in utero vitamin D levels. The proposed research is significant because it is expected to provide valuable data that will ultimately lead to efficacious targets for interventions to mitigate the occurrence of CBT.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Calcium signaling plays a central yet surprisingly flexible role in the function of neurons, contributing to neurotransmission, synaptic plasticity, ad neuronal death. Calcium-calmodulin (CaM)-dependent protein kinase II (CaMKII) is a multifunctional Ser/Thr protein kinase that is essential for normal hippocampal synaptic plasticity and spatial learning in mice. CaMKII is believed to decode the frequency of calcium transients (i.e. neuronal activity) in to graded levels of activity. Unlike the role of CaMKII in physiologica calcium signaling, its role in pathological calcium signaling is largely unknown. Aberrant calcium signaling accompanies multiple disease states associated with glutamate, the major excitatory neurotransmitter in the brain. Excessive glutamate release accompanies acute disease states (e.g. ischemia and brain trauma) as well as chronic neurodegenerative disorders (delayed neuronal death with ischemia and epilepsy). Exactly what CaMKII is doing in excitotoxicity is unknown; however, there are clues worth noting. First, CaMKII is highly expressed (1-2% of total protein) in regions of the brain highly susceptible to excitotoxicity. Second, ischemic duration correlates to CaMKII inactivation and neuronal death. Third, preventing CaMKII from activating during excitotoxicity prevents neuronal death; a process that also prevents CaMKII from undergoing activity-dependent inactivation and aggregation. Fourth, we have recently published that inhibiting CaMKII directly induces neuronal apoptosis via calcium dysregulation and hyperexcitability to aberrant glutamate signaling. In this application, we propose to understand novel mechanisms underlying CaMKII substrate phosphorylation, inactivation and aggregation in both highly controlled biochemical experiments and in living cells. In addition, we propose to determine if CaMKII inactivation in astrocytes disrupts normal glial-neuronal communication; a process our preliminary data indicates leads to astrocyte degeneration. These experiments will not only advance our understanding of CaMKII signaling during pathological calcium signaling, but they will also shed new light on basic mechanisms related to CaMKII structure, substrate phosphorylation, and protein aggregation. Conclusions from these studies may also identify novel therapeutic targets and mechanisms to disrupt neuronal and glia death induced by glutamate excitotoxicity. PUBLIC HEALTH RELEVANCE: CaMKII is serine/threonine protein kinase that undergoes aggregation and inactivation following excitotoxic calcium signaling associated with ischemia and epilepsy. The long-term goal of these studies is to identify molecular mechanisms and therapeutic strategies to disrupt CaMKII's vulnerability to excitotoxic glutamate-signaling; a process we also believe will reveal novel insights into synaptic plasticity and the function of CaMKII in physiological calcium signaling.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Generalized resistance to thyroid hormone (GRTH) is an inherited defect of reduced responsiveness to thyroid hormone often associated with growth retardation and learning disabilities. Our aim is to assess responses to thyroid hormone in non-resistant and resistant subjects in order to standardize a clinical procedure for the diagnosis of GRTH.
|
{
"pile_set_name": "NIH ExPorter"
}
|
An isometric virus-like particle (VLP) approximately 20nm in diameter has been isolated from three strains of Neurospora crassa: a wild type (OR23-1A), "poky" (1578), and P-147. Based on co- sedimentation with known standards, the sedimentation constant for the particle is about 86S. The particles from the three strains appear morphologically identical, but no immunological tests of relatedness have yet been conducted. Purification of the particles involves precipitation with polyethylene glycol followed by differential centrifugation and separation on sucrose gradients. Absorption spectra are consistent with the particles being a nucleo-protein containing 5-15% nucleic acid. The particles when purified appear to be unstable since following dialysis and sedimentation through a second sucrose gradient, their sedimentation constant is lowered (about 73S). Concomitant with this change the UV absorption spectrum approaches that of a protein. Purification in the presence of EDTA reduces the instability of the particles. The quantity of VLP (based on UV absorption) extractable from wild type is markedly less than that from "poky" and P-147. It is not yet clear whether the higher titers of VLP's in "poky" and P-147 are causally related to any physiological parameter such as the slow growth characteristic of those strains. During the coming year, our efforts will be directed at determining the nucleic acid contained in the 20nm particle and attempting to correlate quantities of VLP's with slow growth. Methods for the purification of a second VLP in the P147 strain will be developed. In addition, a VLP carried by the unknown (un) mutant of Neurospora is being investigated.
|
{
"pile_set_name": "NIH ExPorter"
}
|
A cooperative study to determine if treatment of Sickle Cell Anemia is improved with Erythropoietin.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Abstract. The research program of the Latham Laboratory centers on understanding the relationship between structure, dynamics, and function. We are particularly interested in examining this relationship in large macromolecular assemblies, which have for the most part been recalcitrant to detailed structural and dynamics studies. We couple high-resolution methyl-based nuclear magnetic resonance (NMR) spectroscopy, which is capable of probing macromolecules up to ~1 MDa in size, with other biophysical (e.g., lanthanide resonance energy transfer ? LRET) and biochemical techniques to better understand function. Currently, the focus of our program is on two protein complexes that bind to nucleic acids: the universally conserved and essential DNA double strand break repair complex Mre11-Rad50-Nbs1 (MRN) and the mRNA cleavage and poly-adenylation assembly cleavage stimulation factor complex (CstF). In both cases, existing structural and biochemical studies have suggested a role for protein motions in the function of the complex. However, many questions still remain about the interplay of structure and dynamics and how these relate to and control MRN and CstF activity. For the MRN complex, X-ray crystallography, cryo-electron microscopy, and small angle X-ray scattering studies have shown large structural changes within the complex upon ATP binding and have also hinted at local domain motions upon dsDNA binding; however, the timescale of these movements, other key structural states, their importance in function, and local dynamics have not been described. Similarly for the CstF complex, X-ray crystallography and NMR spectroscopy on isolated domains have given views of local structure and dynamics and provided a clue for nascent mRNA recognition; yet, the architecture of the full complex and complete structure of the RNA-bound state is missing. Additionally, disease associated mutations have been noted for both the MRN and CstF complexes. Thus, a main goal of the parent grant is to describe the structures and dynamics of these two nucleic acid binding complexes to better understand the how these characteristics affect function.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Calcium channel inhibitors (CCI's) and calcium channel activators (CCA's) are thought to affect calcium flux through membrane bound channels as their major site of action. There are reports of clinical trials involving over 150 patients suggesting that CCI's also have beneficial effects in neuropsychiatric disorders. There are four major subclasses of CCI's, and each appears to have different biochemical and behavioral properties. Additional studies have suggested a complex interaction between dopamine receptor function and calcium channels. We are currently investigating verapamil and nifedipine as treatments for schizophrenia and tardive dyskinesia. Patients on these protocols are tested with regional cerebral blood flow and computer analyzed brain electrical activity mapping. The platelets of these patients are also studied for 3H-nitrenidipine and 3H-yohimbine binding, as well as for differential patterns of protein phosphorylation.
|
{
"pile_set_name": "NIH ExPorter"
}
|
In APCs, antigen processing compartments often have the phenotypic characteristics of multivesicular bodies (MVB). In studies examining MHC-II trafficking to antigen processing compartments we found that activated (but not resting) B cells secrete significant amounts of their total pool of pMHC-II on MVB-derived vesicles termed exosomes. Surprisingly, it was predominantly Ii-free surface pMHC-II that internalized and trafficked to these MVB, highlighting a previously unrecognized transport pathway followed by pMHC-II. We also found that interaction of antigen-loaded B cells with antigen-specific T cells stimulates exosome release from B cells, and these exosomes in turn can stimulate primed (but not nave) T cells to proliferate. Our results support a model that T cell stimulated exosome release from activated B cells serves to augment T cell responses.The movement of proteins and lipids from one intracellular compartment to another is carried out by a well-orchestrated process of transport vesicle formation, vesicle docking with a target compartment, and finally vesicle-target membrane fusion. The proteins that catalyze membrane fusion are termed SNAREs. We have been investigating the role of distinct SNARE isoforms in regulating secretory granule exocytosis from immune cells using a variety of SNARE knock-out mice and have found that VAMP-8, but not VAMP-2 or VAMP-3, regulates mast cell secretory granule exocytosis. Surprisingly, VAMP-8-deletion only affects serotonin exocytosis but not histamine or TNF-alpha exocytosis, showing for the first time that secretory granule heterogeneity exists in mast cells. We are currently using these mice to examine the role of specific SNARE proteins in regulated exocytosis from other immune cells.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The goals of this project are (1) to understand better the dynamic interactions between the voltage-gated Na channel and specifically bound ligands and (2) to characterize the unique conformational transitions of the ligand-modulate voltage-gated Na channel. We plan to examine the pharmacological properties of local anesthetics, Alpha-, and Beta-scorpion toxins on the voltage-gated Na channel at the level of macroscopic currents in axons and, subsequently, at the level of single channel events in cultured cells. The kinetics of these current records reflect the conformational transitions among resting, open, inactivated, and several intermediate states of the Na channel. The existence of many conformational states requires complicated structural rearrangements of the Na channel polypeptide following membrane depolarization. Perturbation of these structural rearrangements upon binding with a ligand may stabilize certain conformations and thus provide enlarged opportunity to study the properties of these conformations. Reciprocally, the ligand binding site may change its configuration during the structural rearrangement of the channel and thus alter its binding affinity with the ligand. This general description of channel-ligand interactions is now known as Hille's modulated receptor hypothesis. The usefulness of this hypothesis will be tested by using three different classes of ligands as molecular probes: local anesthetics which block the Na currents, Alpha-scorpion toxins which specifically modify Na channel inactivation, and Beta-scorpion toxins which primarily modify Na channel activation. Both the actions of the native ligands and of their various derivatives, including the photoactivatable derivatives which can be covalently bound to the channel by phase-locked photoactivation procedures, will be studied during voltage-clamp steps of varying magnitude and duration. The resulting dynamics of ligand-receptor interactions and their underlying molecular mechanisms should provide novel information about the microscopic processes occurring during channel gating. Furthermore, affinity coupling of these ligands or their derivatives on the primary sequence of the channel polypeptide may in the future permit the direct identification of a known region of the channel with a special function.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The Preclinical Magnetic Resonance (MR) Imaging Resource is a new core facility at Roswell Park Cancer Institute (RPCI) that offers individualized MR protocols designed to answer specific questions related to cancer detection, drug/gene delivery, animal phenotyping and therapeutic efficacy. The goal is to provide readily available access to state-of-the-art noninvasive in vivo imaging technologies yielding high-contrast, serially-acquired images of visceral tumors at an affordable cost. In RPCI's previous Cancer Center Support Grant renewal application, the Resource was included as a developing core facility. It is now fully operational and available 24 hrs/day, 7 days/wk offering services and training to all Program members. Three general types of services are offered: (1) instrumentation and expertise to acquire high resolution (more than 50 mu m[2] in-plane spatial and more than 100 ms temporal) heteronuclear imaging and spectroscopy data sets; (2) instrumentation, expertise and software (commercial and custom designed source code) for analysis/image processing of digitally acquired data sets; and (3) visualization and surface/volume rendering of 2D and 3D data sets for quantitation of morphologic/physiologic relationships. Instrumentation available for use includes a 4.7 T wide bore (33 cm) magnet incorporating Bruker's Avance platform with Paravision (R) 2.1 OS, shielded Accustar gradients and digital system electronics. Several customized heteronuclear RF transceiver coils, removable gradient coils and animal positioning devices are available for use with gas anesthesia equipment, a computer-controlled MR-compatible injector and surgical microscope. Instrumentation is available for physiologic monitoring of animal respiration/cardiac output, tissue temperature, oxygenation, and blood flow using fiberoptic-based laser doppler technology. Ancillary equipment includes image/data processing workstations (PC and Unix based), image analysis/visualization software (AnalyzePC, Version 4.0) and high resolution color printers. The MR system represents the highest magnetic field strength scanner available within 200 miles of RPCI.
|
{
"pile_set_name": "NIH ExPorter"
}
|
Erectile dysfunction (ED) affects 52% of men between the ages of 40 and 70. 30-87% of prostate cancer patients treated by prostatectomy experience ED and PDE5 inhibitors are ineffective in 29-86% of prostatectomy patients who experience ED, depending on their nerve injury status. The reduced efficacy of treatments in this population makes novel therapeutic approaches to treat ED essential. Significantly increased apoptosis of penile smooth muscle is common in both animal models and human patients with ED. We propose that abundant apoptosis observed in penile smooth muscle when the CN is cut is a major contributing factor to ED development. If apoptosis could be prevented following prostatectomy while the CN regenerates, then resumption of normal erectile function would occur more quickly, and irreversible morphology changes in the penis that cause ED would be prevented. Understanding the mechanisms that regulate smooth muscle apoptosis in the penis is critical for development of new therapeutic approaches for ED treatment and prevention. Sonic hedgehog (SHH) is an essential regulator of penile smooth muscle. When SHH is inhibited in the penis, there is a 12-fold increase in smooth muscle apoptosis that results in ED. SHH protein treatment is able to suppress CN injury induced apoptosis, indicating that SHH has significant potential to be developed as a treatment to prevent ED by suppressing smooth muscle apoptosis. The Affi-Gel bead technology used in these studies is not applicable to humans, so we propose to develop nanoparticle delivery of SHH protein to the penis and hypothesize that SHH delivery via nanoparticles will be effective in suppressing apoptosis induction caused by CN injury. This novel technology has substantial potential to be developed into a therapy to prevent apoptosis in patients at the time of prostatectomy, so has significant clinical relevance. The mechanism of how SHH itself is regulated in the penis and how decreased SHH protein induces apoptosis is poorly understood. It is likely that neural input/integrity regulates SHH in the penis since SHH protein is significantly decreased in two models of neuropathy, the CN injured rat and in the BB/WOR diabetic rat. Since SHH protein is decreased in diabetic human penes in parallel with observations in the rat, this lends clinical significance to how decreased SHH protein can induce apoptosis in the penis. Our results suggest that HIP out competes PTCH1 for SHH binding after CN injury. Thus we hypothesize that loss of neural input decreases SHH protein in the penis and induces apoptosis in penile smooth muscle through a PTCH1 and HIP dependent mechanism.
|
{
"pile_set_name": "NIH ExPorter"
}
|
The project is a coordinated clinicopathologic investigation of ocular and orbital tumors. The specific objectives of this project are: (1) To improve preoperative methods for the diagnosis of intraocular neoplasms. (2) To expand and maintain an ongoing ocular tumor clinic. (3) Thorough clinicopathologic case documentation on patients with ocular neoplasms. (4) To correlate the behavior in individual tumors in patients with various morphologic and nonmorphologic features of the neoplasm. (5) To test the possibility that retinoblastomas produce nerve growth promoting factor and if this is correct, devise procedures for correlating its production with the presence of this tumor or its metastases. Furthermore, to explore the possibility of treating retinoblastomas with NGF or anti-NGF. (6) To preserve strains of tumor cell clones so that the comparative effect of drugs and other therapeutic agents can be evaluated. The research methods include sophisticated ophthalmologic diagnostic procedures (e.g. fluorescein angiography, light and electron microscopy, as well as tissue and organ cultures).
|
{
"pile_set_name": "NIH ExPorter"
}
|
Subsets and Splits
No community queries yet
The top public SQL queries from the community will appear here once available.