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The Shaker gene of the fruit fly encodes voltage-dependent K+ channel proteins. The long-term goals of the research are to derive a biochemical model for the structure of the channel, and to correlate its functional properties with its structure. The specific aims are: 1) to study the function of the conserved S4 sequence, which has been proposed to be the voltage-sensor of the channel. Single amino acid mutations will be made in the S4 basic amino acids, acidic residues that might form ion pairs with the S4, and control residues. The mutant channels will be expressed in Xenopus oocytes and analyzed electrophysiologically. 2) to study the disposition of the Shaker proteins in the membrane. The modification of potential sites for posttranslational glycosylation will be studied in in vitro translation reactions and in vivo. If these sites are modified in vivo, their topological location can be inferred. 3) to study the subunit structure of the channel. The Shaker gene will be expressed in a tissue culture system to begin biochemical experiments with the eventual goals of purifying the channel and determining the number of Shaker subunits per channel. K+ channels in the nervous system are implicated in the basic mechanisms of epileptogenesis. K+ channels in smooth muscle are promising pharmacological targets for the control of hypertension and stroke. K+ channels in lymphocytes are altered in a mouse model for lupus erythematosus. Therefore, studying the structure and function of K+ channels will contribute to our understanding of the etiology and treatment of a variety of diseases.	{ "pile_set_name": "NIH ExPorter" }
Studies are to be conducted to explore the mechanisms by which cell proliferation is controlled by a specific mitogenic agent - epidermal growth factor (EGF). Attempts will be made to isolate and define the biochemical properties of membrane molecules mediating the cellular response to EGF. These molecules include the EGF receptor, the EGF sensitive protein kinase, and the endogenous substrates for the EGF enhanced phosphorylation of proteins.	{ "pile_set_name": "NIH ExPorter" }
The Administrative Core provides the organization and structure for planning and implementation of ADRC core and research activities. The core supervises all ADRC functions and the Clinical, Data Management & Statistics, Neuropathology & Genetics, Education & Information, and Neuroimaging Cores. The Administrative Core monitors ADRC budgets to ensure fiscal responsibility; organizes the review and funding process for pilot studies, and monitors ADRC research projects, pilot projects and studies utilizing ADRC subjects or resources. This core determines new research directions, including the recruitment of new investigators to work in Alzheimer's Disease (AD). Establishment and expansion of outreach efforts to the medically underserved and minority communities are directed and stimulated by the Core. The Core also oversees participation in multicenter studies, data transfer to the NACC and other NIA data repositories, tissue and DNA samples to NIA repositories, cross-ADC collaborations, and other national and international initiatives. Through the establishment of a comprehensive communications system including Website and e-mail ListServes, the Core Directors, project investigators, and staff will maintain an integrated, coherent and multidisciplinary focus. The goals of the Administrative Core are achieved by the Executive Committee, which is comprised of the Director, Co-Director, Associate Director, Center Administrator, and Core and Project Leaders. It is aided in these tasks by advice of the Internal and External Advisory Committees, and input is solicited from our investigators. The Internal Advisory Committee meets once per year, is chaired by the Senior Vice Chancellor/Dean of the Medical School, and includes the former Senior Vice Chancellor for the Health Sciences, the Chair of Psychiatry, the former Chair of Epidemiology at the University of Pittsburgh and the Co-Director of the Center for the Neural Basis of Cognition. They provide guidance on policy, administration, and scientific direction. The External Advisory Board is comprised of nationally respected researchers and clinicians with expertise in disciplines relevant to AD. They provide scientific critiques of research and operations of the ADRC as well as recommendations for new initiatives.	{ "pile_set_name": "NIH ExPorter" }
The object of the proposed research is to identify and characterize the normal low density lipoprotein (LDL) receptors on the plasma membranes of fibroblasts from normal subjects and the defective LDL receptors from plasma membranes of patients with familial hypercholesterolemia (FH). To accomplish this we will use new, very sensitive techniques which we have recently developed to screen membrane proteins for small alterations in primary structure. Our preliminary results suggest that these techniques are capable of detecting differences as small as the modification of a single tryptic peptide residue in two otherwise identical proteins. Only microgram quantities of each protein, isolated as a discrete polyacrylamide gel band, are required. Critical to this approach is our new method for double label thin layer autoradiography. Following identification of the normal and defective LDL binding proteins, each will be characterized structurally and, to some extent, functionally. Structural analysis will include peptide mapping, amino acid composition, amino acid sequencing in the regions of dissimilarity between normal and defective proteins, and analysis for possible post-translational modifications such as phosphorylation or glycosylation. Functional analysis will focus primarily on development of antibodies specific for normal and defective LDL binding proteins. The antibodies will then be used to localize the binding protein within the membrane, to follow its fate after exposure to LDL, and to study its biosynthesis and insertion into the membrane in normal and FH cells. The immediate purpose of this work is to help elucidate the nature of the control mechanisms for regulation of cholesterol biosynthesis with particular emphasis on events occurring at the plasma membrane. Detailed structural comparison of normal and FH binding proteins should answer the question of whether altered LDL binding function in FH is the result of a direct modification of the gene coding for this protein or an indirect effect due to post-translational modification. An additional immediate goal will be to provide antibodies for clinical investigation and early detection of familial hypercholesterolemia. In a more general sense, however, the purpose of this research is to utilize pathology as a probe of membrane protein structure-relationships and to demonstrate the general applicability of our new techniques to similar problems involv (Text Truncated - Exceeds Capacity)	{ "pile_set_name": "NIH ExPorter" }
Children hospitalized for psychiatric conditions are at risk for chronic and severe behavioral dysfunction after discharge. They also use mental health and social services in greater quantities and intensities than nonhospitalized child psychiatric patients. The principal purposes of acute psychiatric inpatient care of children are stabilization, evaluation, and assessment of which treatments and services after discharge can help them function in the least restrictive setting. Knowledge of postdischarge clinical course can help hospital care more efficiently achieve these goals. First, knowing whether stabilization attained in hospital persists after discharge is an important consideration in determining the level of improvement hospital care should aim for. Second, evaluation includes consideration of factors that may influence prognosis, for which longitudinal data are clearly necessary. Third, accurate appraisal of postdischarge needs requires valid prediction of functioning in several settings. Because no longitudinal study has evaluated these patients' functioning across settings at several in the year after discharge, decisions about postdischarge services lack an empirical basis. This project's long-term objective is to enable clinicians to estimate postdischarge clinical course based on knowledge of selected patient, family, hospital course, and aftercare characteristics, leading to more efficient use of hospital care and improved service delivery after discharge. The study will examine two questions relating to the postdischarge functioning of child psychiatric inpatients: 1. What is the postdischarge clinical course of child psychiatric inpatients? 2. What characteristics of the child, family, hospital course, and aftercare services predict these patterns of postdischarge clinical course? More specifically, this is a longitudinal, prospective follow-up of approximately 120 child psychiatric patients with three predischarge assessments: at admission, during initial hospitalization, and right before discharge. In the year after discharge, there will be three follow- up assessments, at three, six, and twelve months after discharge. Assessments of preadmission and postdischarge condition will evaluate functioning at home and at school. The same of similar instruments will be completed by hospital staff, teachers, and caregivers during hospitalization. Data analyses will evaluate changes over time for the sample as a whole, and evaluate predictors of follow-up status suggested in previous research. These analyses will be supplemented with methods to develop individual growth curves of clinical course over time, and to discern how these curves are correlated with patient, family, course-in- hospital, and aftercare characteristics.	{ "pile_set_name": "NIH ExPorter" }
The development of organs and their function is governed by the differentiation and patterning of cells through the coordinated expression of specific genes. This competitive renewal for an O'Brien Urology Research Center is focused on the genetic, molecular and cellular events that regulate genitourinary (GU) tract development. Four projects by established senior scientists and two pilots by junior investigators spanning two clinical and one basic science department with a ten-year record of productive collaboration are proposed. Project 1 (Gomez) examines the role of a novel kidney-specific gene in proximal tubular cell maturation and function. Whether sequence elements in the promoter region of this gene determine organ and cell specificity will be investigated. Project 2 (Chevalier) will study the mediators of both salutary and injurious effects in a newly developed model of neonatal partial ureteral obstruction. A translational research component will search for urinary markers for obstructive nephropathy. Early detection may enhance recovery from obstructive renal injury. Pilot 1 (Roth) will dovetail on Project 2 by examining the embryonic origin and lineage of ureteral smooth muscle. It will be determined whether clonal expansion versus synchronous migration of differing cell types contributes to ureteral expansion and will offer insight into hydronephrotic disorders. Project 3 (Turner) investigates the developmental and regulatory (androgenic), compensatory (vasectomy), and genomic events leading to the synthesis of natural antibacterial peptides, including defensins, in the GU tract. Project 4 (Flickinger) examines development and novel consequences of epididymal segmentation involving sonic hedgehog pathway genes. Pilot 2 (Lysiak) investigates the role of caspase 2 and downstream pro-apoptotic members of the Bcl-2 family in establishing the number of germ cells in the testis. Investigators in this P50 share common themes and study similar mechanisms yet direct attention on different organs within the GU tract. The proposed investigations will advance the development of novel diagnostic tests and therapies for disorders ranging from renal insufficiency to male infertility building on strengths in reproductive and cell biology, urology and pediatric nephrology at the University of Virginia.	{ "pile_set_name": "NIH ExPorter" }
A better understanding of the molecular basis of enzymatic breakdown and removal of lipids in lipoprotein complexes is necessary in order to elucidate the pathogenesis of disordered lipid metabolism. A lipase from human post-heparin plasma which is different from lipoprotein lipase from that source has been purified to homogeneity and a hormone-sensitive lipase from adipose tissue has been highly purified. Both of these enzymes appear to have phospholipase activity and this proposal concerns a more detailed analysis of the phospholipase activity of these two enzymes and a pure phospholipase A2 from cobra venom (Naja naja naja). These enzymes will be studied in a Triton X-100--phosphatidylcholine mixed micelle system usina a "surface-as-cofactor" model previously developed to explain the action of the cobra phospholipase A2 (Deems, R.A. Eaton, B.R., and Dennis, E.A. (1975) J. Biol. Chem. 250, 9013-9020). The main studies will be conducted on the cobra phospholipase because of its structural simplicity and small size (molecular weight 11,000) (Deems, R.A., and Dennis, E.A. (1975) J. Biol. Chem. 250, 9008-9012), compared to the human enzymes. These studies are aimed at developing a better understanding of the kinetic analysis of the action of phospholipases and lipolytic enzymes toward substrates in aggregated forms such as lipoprotein complexes, intracellular fat droplets, mixed micelles, and membranes. Kinetic studies on the action of the cobra enzyme toward red blood cell membranes and sonicated vesicles above and below the thermotropic phase transition as well as sequence and structural studies are also proposed.	{ "pile_set_name": "NIH ExPorter" }
Project Summary/Abstract. This is a competing renewal application of the highly successful UC Davis T35 STAR (Students Training in Advanced Research). The program has trained 225 DVM students over the last 20 years. Students are encouraged and mentored to submit hypothesis-driven research proposals during their first, second, and occasionally third year of the professional curriculum. The objective of the STAR program is to provide DVM students, whose application is approved by the review committee, stipend support to join experienced training faculty members and their productive research teams in 10 weeks of structured research mentoring and hands-on modern biomedical research. The program emphasizes 5 fundamental research objectives: 1) how to gain knowledge and understanding o f one?s field of science; 2 ) h o w t o formulate a scientifically sound and testable hypothesis; 3) identify specific objectives, conduct controlled methodical experiments, and develop technical expertise; 4) analyze results, derive conclusions, propose additional experiments, and anticipate new directions; and 5) convey research findings succinctly and convincingly to others. Responsible conduct of research and scientific rigor are key components of the training plan. STAR program has consistently received 55 to 63 applicants annually, and in 2015 expanded to 15 positions supported by the NIH T35 mechanism to meet growing demand from students with interest in participating in NIH-relevant research. Students with broad interests ranging from molecular and cellular medicine, to biomedical engineering, to vector borne diseases, to epidemiology will be trained. The program maintains 57 faculty trainers with proven successes in undergraduate, predoctoral and postdoctoral training. Thus, students have access to research projects conducted not only on a variety of lab animal species (C. elegans, zebrafish, rodents, non-human primate), but also clinical research on companion and food animals. Increasingly students express interest in transdisciplinary ?One Health? research experiences with hypotheses targeting complex questions about disease emergence and transmission at the interface of animals, humans and their environment. Accordingly, we have dynamically expanded our training faculty to meet student interests. The greatest strengths of our short-term training program include the outstanding quality and motivation of our DVM students, the strong highly collaborative multidisciplinary nature of our research programs, and student access to translational research projects that use innovative approaches. Trainees will have access to advanced technologies, such as epi/genomics and metabolomics, proteomics, state-of-the-art imaging, BSL3 labs, genetically modified organisms, and gnotobiotic and inhalation facilities. This five-year competitive renewal application requests to maintain support for a total of 15 DVM students per year for each of five years (a total of 75 students), in order to maintain T35 program missions; spark the interest of the next generation of DVM clinician-scientists that will take leadership positions in academia, government, and the private sector.	{ "pile_set_name": "NIH ExPorter" }
Eosinophils are innate immune leukocytes associated with asthma, allergies and other diseases. Despite the long recognition of eosinophilic involvement in asthma, specific, non-redundant roles for eosinophils in disease pathogenesis had been elusive. However, recent studies from several laboratories have revealed new functions of eosinophils in immunomodulation and airway remodeling, changing the classic paradigm of disease pathogenesis and reinvigorating the field with a promise of more specific therapeutic targets. Our overall goal is to identify and mechanistically define effector functions of eosinophils vital to the initiation and exacerbation of asthma. In line with this goal, this proposal builds upon our novel discovery that mature human eosinophils express fully functional Notch ligands and receptors, indicating eosinophils utilize Notch signaling pathways, both as signal-receiving "target" cells and as signal- sending "signaling" cells. Our preliminary data establishes our overlying hypothesis that Notch signaling underlies eosinophil functions critical to asthma. Our studies will specifically investigate two hypotheses: 1) Notch receptor activation is required in parallel with cytokine signals to achieve full and sustained activation of eosinophils in asthmatic airways;and 2) Eosinophils promote a Th2 milieu in asthma by Notch ligand-mediated juxtacrine interactions with T cells. While experimental approaches utilize predominantly human eosinophils, proposed studies also take full advantage of the manipulative benefits of mouse models. Our proposal may provide vital insights into the mechanistic basis for eosinophil functions pertinent to asthma, and by extension other inflammatory diseases of the lung involving eosinophilia, relevant to development of novel, targeted therapeutic approaches. PUBLIC HEALTH RELEVANCE: Asthma is a highly prevalent and costly disease. Eosinophils, innate immune leukocytes associated with asthma, allergies and other diseases, are the predominant cellular infiltrate in asthmatic airways. This proposal is designed to elucidate the mechanistic basis of eosinophil functions leading to asthma exacerbation. These studies may lead to the development of novel, targeted therapeutic approaches.	{ "pile_set_name": "NIH ExPorter" }
The overarching aim of this application is to greatly expand knowledge about preclinical AD and specifically to characterize the chronology of biomarker changes, determine their path biological signatures (if any), and identify cognitively normal persons who are at very high risk for developing symptomatic AD. Successful treatment strategies will require biomarkers that can identify individuals at high risk for AD and at the earliest clinical stages in order to target them for clinical trials, disease- modifying therapies nd to monitor therapy success. The overall Specific Aims in this renewal application are to: 1. Follow the current participants in ACS and add new enrollees to maintain the sample size at ~300. 2. Obtain longitudinal data from the ACS participants at 2 year intervals with the following measures: a. Clinical and cognitive assessments (Clinical Core) b. Amyloid imaging with PET PIB (Project 1) c. Assays of amyloid-beta (A[unreadable]), tau, phosphorylated tau181 (p- tau181), and novel analytes in CSF and blood (Project 2, supported by the Biomarker Core) d. Attentional control battery and task-related functional MRI (fMRI) (Project 3) e. Structural MRI, resting state functional connectivity MRI (fcMRI), diffusion tensor imaging (DTI), and cerebral blood flow using arterial spin labeling (ASL) (Project 4) 3. Analyze associations among rates of change of all disease markers from all Cores and Projects (Data Management and Biostatistics Core). PUBLIC HEALTH RELEVANCE: The results of clinical assessment, studies of biomarkers (PET imaging, MRI of the brain, cerebrospinal fluid analysis, and tasks which place a high load on attentional systems) will be correlated to identify the earliest brain changes of AD, determine the evolution of these changes over time, and assess their predictive power for the eventual development of symptomatic AD by studying participants in two groups-one with a family history of AD and the other with no family history of AD. REVIEW OF INDIVIDUAL COMPONENTS OF THE PROGRAM PROJECT CORE A: ADMINISTRATION;Dr. John C. Morris, Core Leader (CL) DESCRIPTION ( provided by applicant): The Administration Core acts to ensure that the research and programmatic goals of the Adult Children Study program project grant (ACS PPG) are met. The administrative leadership consists of the Director (Morris) and the Executive Director (Buckles). They are assisted by the Executive Committee that includes these individuals, leaders of Cores and Projects, and other senior faculty. The Administration Core supports, monitors, and coordinates the activities of all components of the ACS PPG. It will annually convene an External Advisory Committee to review activities and progress. The specific aims are to: 1. Coordinate and integrate the Cores and Projects and provide administrative and budgetary support and oversight, ensuring appropriate utilization of funds 2. Monitor the effectiveness of the PPG toward achieving the stated goals 3. Arrange for periodic external review and advice concerning PPG goals and progress 4. Coordinate and oversee data integration with Washington University Center for Biomedical Informatics to develop, maintain, and monitor an integrated database PUBLIC HEALTH RELEVANCE: The Administration Core ensures that goals of the Adult Children Study (ACS) are met, including identifying the earliest brain changes of AD, determining the evolution of these changes overtime, and assessing their predictive power for the eventual development of symptomatic AD. With the assistance of an External Advisory Committee and an Executive Committee including the leaders of the Projects and Cores, the Administration Core supports, monitors, and coordinates the activities of all components of the ACS.	{ "pile_set_name": "NIH ExPorter" }
A study of the kinetics and mechanisms of metal ion transfer reactions involving peptide complexes is proposed. Copper transport in the blood involves its transfer to and from peptide coordination in serum albumin. Other metals such as nickel and cobalt may also be transported by serum albumin. Amino acid complexes are believed to play an important role in mediating the transfer of copper to low molecular weight complexes which can be readily transported across biological membranes. In the treatment of Wilson's Disease, chelating agents are used to remove copper from the body. The factors which control the pathways and rates of copper, nickel and cobalt transfer reactions will be examined. The effects of various functional groups, in combination with peptide bonding, on the properties of metal complexes will be studied, in particular the effect of sulfur-containing peptides where sulfhydryl, disulfide, or thioether groups are available for metal coordination. Factors which kinetically and thermodynamically stabilize Cu(III), Cu(II), and Cu(I) oxidation states will be examined. The major areas of investigation of metal peptide complexes include: (1) proton-transfer mechanisms, (2) metal ion transfer mechanisms to other multidentate ligands, (3) characteristics of new peptide complexes with unusual oxidation stages or metal ions not previously studied, (4) reactions of sulfur-containing ligands, (5) nature of copper transport in blood and reactions of copper in proteins where there are possible conformational changes of the proteins. A variety of stopped-flow techniques are critical to these studies and we need to extend our capabilities to include anaerobic, non-ambient systems and flow epr cells with rapid spectral scanning.	{ "pile_set_name": "NIH ExPorter" }
Experiments have shown that hemoglobin associated acetaldehyde (HBAA) and plasma associated acetaldehyde (PAA) as measured by a fluorigenic high performance liquid chromatographic assay normalized to protein concentration can serve as markers in humans for ethanol intake with different biological kinetics. Additional experiments in animal models (C57 mice and miniature swine) have confirmed these observations and indicated that use of the same assay system to measure platelet associated acetaldehyde (PLTAA) and whole blood associated acetaldehyde (WBAA) provides assessments of ethanol intake as well. The present proposal is designed to define the clinical utility of the above four assays as markers for ethanol intake. Ongoing studies with Schick Health Services assure availability of samples such that assays can be evaluated in well characterized groups of teetotallers, social drinkers, alcoholics, and pregnant women. We have also applied for and received over 10,000 plasma samples from the NICHD-Diabetes and Early Pregnancy Study to help in the definition of the utility of these assays in this latter population. Thus it will be possible to determine norms for the assays involved by age and gender, the statistical significance of discrimination for each assay between drinkers, nondrinkers, and alcoholics as well as correlation with another measure of drinking behavior (the SAAST), and also evaluate special subgroups. Specific studies are designed to determine the chemical nature of the adducts measured by the assay, the biological on and off rates of each assay, as well as precision, recovery, optimum conditions for sample storage and shipping. The development of two animal models will be continued in our own laboratories (the C57 mouse) and in collaboration with investigators at the University of Missouri (miniature swine). The availability of these animal models will allow definition of questions concerning the assays which are difficult to perform in humans including gender differences in response to quantified intakes of ethanol, tolerance, on and off rates, and responses of recidivist drinkers.	{ "pile_set_name": "NIH ExPorter" }
The humble fruit fly, Drosophila melanogaster, is an important model system, boasting facile genetics, complex organ systems, complex behaviors and a sequenced genome. Our long-term interest has been in determining how the sex of the germ line is established. In other words how a germ line stem cell gives rise to either sperm or eggs. Stem cell development occurs in a defined niche. Our work suggests that when a male stem cell develops in a female niche a tumor results, while when a female stem cell develops in a male environment, stem cell proliferation or survival is poor. Microarray analysis shows that stem cell tumors forming due to inappropriate juctaposition of male germ cells in a female soma are similar to tumors resulting from mutations in any of several genes required for female fertility. These result confirm our longstanding proposition that the proteins encoded by these genes play important roles in germline sex determination. We are also using microarray technology to identify diagnostic markers for germ line stem cells of the two sexes and are exploring the peculiar nature of the X chromosome. We have also expended considerable effort on the detailed analysis of the ovo gene, which is required for the viability of female germ line stem cells regardless of niche environment. The regulatory circuit controlling ovo expression is devilishly complex. The locus encodes both positively and negatively acting transcription factors from alternative promoters. These alternative promoters are cross-regulated by the antagonistic ovo transcription factors. Additionally, the mechanism of ovo biochemical function is unusual, in that ovo proteins bind and function directly at transcription start sites of target genes (locations normally occupied only by basal transcription factors). This analysis has been extended using comparative genomics (recently sequenced Drosophila species). One of the advantages of working on a model organism at the NIH is exposure to scientists interested in a range of topics, from basic research, to insect disease vectors, or translational and clinical studies. We have also participated in the NIH multiple endocrine neoplasia type-1 (MEN1) consortium led by Francis Collins (NHGRI), Steve Marx, and Allen Spiegel (both NIDDK). We have determined that Drosophila MEN1 can act to negatively regulate the jun/fos complex, AP1. It is our hope that fruit fly studies will provide a skeletal view of gene regulation pathways to be fleshed out by research on mammalian systems.	{ "pile_set_name": "NIH ExPorter" }
?-secretase inhibition remains a viable anti-amyloidogenic strategy. In addition to previous reports that novel ?-secretase inhibitors (GSI) significantly reduced A[unreadable] production both in vitro and in vivo, Comery and colleagues recently reported that similar GSIs may even improve cognitive functioning in a transgenic mouse model of AD (Tg2576). These finding have functioned to further the vigorous search for potential candidate GSIs. Among the many promising potential candidates are the glycogen synthase kinase 3 (GSK-3) inhibitors. These compounds target this tonically active serine/threonine kinase, which has been implicated in several disorders of the CNS. With regard to Alzheimer's disease (AD), both isoforms of GSK-3 (a and [unreadable]) have been found to directly phosphorylate tau on residues specific to hyperphosphorylated paired helical filaments (PHF), GSK-3[unreadable] has been shown to phosphorylate APP and to contribute to A[unreadable] mediated neurotoxicity, and GSK-3[unreadable] has been found to phosphorylate PS1, which may act as a docking site for subsequent tau phosphorylation. Therefore, GSK-3 inhibitors are especially attractive as they may not only oppose A[unreadable] generation but also neurofibrillary tangle (NFT) formation. Moreover, Phiel and colleagues reported that inhibition of the GSK-3a isoform may regulate ?-secretase cleavage of APP in a substrate-specific manner. Accordingly, this selective inhibition of GSK-3 might provide the maximal therapeutic benefit while reducing the potential for toxic side-effects. The intense search for small-molecular compounds that may modulate AD pathology has advanced the analysis of specific dietary derived substances from fruits and vegetables, which epidemiological study suggest are beneficial against the neurodegeneration and aging processes. In this light, recent focus has been given to a group of polyphenols categorized as flavonoids, which have been found to be potentially anti-amyloidogenic. In our preliminary studies we demonstrate that treatment of both murine N2a cells transfected with the human "Swedish" mutant form of APP (SweAPP N2a cells) and primary neuronal cells derived from Alzheimer's "Swedish" mutant APP overexpressing mice (Tg2576 line) with the flavonoid luteolin results in a significant reduction in A[unreadable] generation. Furthermore, data show that luteolin treatment apparently achieves this reduction through a selective inactivation of the GSK-3a isoform. As in vivo validation, we find that administration of luteolin and a glycoside of a structurally similar flavonoid, diosmin, to Tg2576 mice similarly reduces A[unreadable] generation potentially through GSK-3 inhibition. Importantly, this reduction in GSK-3 activation increases phosphorylation of presenilin 1 (PS1), which forms the catalytic core of the ?-secretase complex, and may suggest a mechanism whereby these small-molecular compounds (GSK-3 inhibitors) modulate AD pathology. In this proposal, we will evaluate this small-molecular compound's therapeutic effect on improving cognitive impairment and correlation of this effect with reducing cerebral A[unreadable] pathology in a transgenic mouse model of AD via an oral administration. Flavonoid-diosmin (DiosMemTM) will be orally administered to Tg2576 mice before (prophylactic treatment group) or after (therapeutic treatment group) development of AD-like pathology. Groups of untreated non-transgenic littermates will be compared to the transgenic treatment groups. Oral administration of DiosMemTM to Tg2576 mice will be performed for 6 months. Aim 1, following the administration, cognitive testing will be done at several ages. Aim 2, we will sacrifice these mice at several ages to examine histological and biochemical endpoints and correlate pathological changes with improvement of cognitive impairment. In this study, we plan to evaluate two time points comparing DiosMemTM to control. Groups will be compared by their effects on opposing cognitive impairment and reducing AD-like pathology. These studies could lay the foundation for AD clinical trials with DiosMemTM diet supplementation in the near future. PUBLIC HEALTH RELEVANCE: Relevance to public health in this proposal, we will fully characterize the effect of flavonoid-diosmin (DiosMemTM) on being beneficial to cognitive impairment and correlate its therapeutic effect with reducing cerebral amyloidosis in a transgenic mouse model of Alzheimer's disease (AD). These studies will lay the foundation for AD clinical trials with DiosMemTM in the near future.	{ "pile_set_name": "NIH ExPorter" }
Strategies to change risk perceptions are a models of health decision making. Both are based on an assumption that may not always be true - that individuals know their risk for a given illness. Preliminary work from our and other laboratories has shown that a non-trivial portion of the population is uncertain about their risk for common illnesses. Moreover, we have shown that uncertainty about one's illness risk is particularly likely among U.S. populations who suffer the burden of health disparities. However, the mechanisms underlying risk uncertainty remain unknown, which limits our ability to develop effective intervention strategies to change behavior, especially in those who remain uncertain about their risk despite the wide-spread dissemination of risk-based health messaging. The objective for this R01 program of research is to explicate the mechanisms leading individuals to answer do not know in response to risk perception questions. In a program of research encompassing both observational and experimental work, we will examine how factors relating to knowledge and health literacy as well as factors related to defensive information avoidance influence do not know responding. This objective will be achieved by addressing two specific aims: 1) To explore two distinct, theoretically-informed explanations for do not know responding, and 2) To test whether experimentally manipulating the mechanisms hypothesized to cause do not know responding, reduces do not know responding. Aim 1 will be achieved with a national survey examining if low health literacy and defensive risk information avoidance are related to do not know responding to colorectal cancer and type 2 diabetes risk perception items. Achieving Aim 2 will be accomplished by experimentally testing 1) whether giving do not know responders with low health literacy health messages tailored to a low health literacy audience increases risk knowledge and reduces do not know responding and 2) whether giving do not know responders who tend to avoid health information the opportunity to self-affirm prior to receiving health messages increases attending to, and comprehension of these messages and reduces do not know responding. This research is significant in that it examines a key, but understudied phenomenon related to a central decision making and behavior change construct, uncertainty about perceived risk for illness. Successful completion of the proposed research will generate tested strategies for developing more effective behavior change messages for individuals who are uncertain about their risk for common illnesses. Given the degree to which uncertainty about illness risk is more common in groups experiencing health disparities populations, our work has the potential to address behavior change factors that contribute to these ongoing health disparities.	{ "pile_set_name": "NIH ExPorter" }
The mission of the NIEHS Microarray Core (NMC) is to utilize advances in genomic and bioinformatic technologies to enable Intramural NIEHS scientists to pursue studies that address how the environment influences human health. The NMC continuously strives to develop and implement new analytical and computational approaches to investigate genome-wide changes in gene expression. Specific technical accomplishments include testing and optimization of RNA isolation and amplification protocols to perform microarray experiments with RNA from limited tissue sources e.g. laser capture microdissected and embryonic samples. The NMC utilizes Affymetrix, Agilent, and Illumina platforms to explore genome-wide analyses such as DNA methylation, gene expression, and microRNA (miRNA) applications. Additionally, the group utilizes Nanostring technology to further enhance its capabilities. The installation and implementation of software packages for emerging and existing genomic technologies has greatly improved the bioinformatics capacity of the NMC. In addition, continuing modifications to the NIEHS Microarray Core custom in-house MicroArray Project System (MAPS) have ensured that the NMC can sufficiently track and manage sample and experimental information related to microarray and Nanostring analyses. Finally, the NMC has continued to refine a rigorous QA/QC protocol incorporated into a standard deliverable that is distributed upon completion of a project.	{ "pile_set_name": "NIH ExPorter" }
The objective of this study is to determine the effects of varying degrees of renal dysfunction on the pharmacokinetics of suramin in cancer patients; to elucidate the toxicity of suramin in cancer patients with varying degrees of renal dysfunction as assessed by spontaneous reports of adverse events and by clinical laboratory measurement; & to document antitumor activity of suramin in the patients treated.	{ "pile_set_name": "NIH ExPorter" }
Maryam Valapour, MD, is an Assistant Professor of Medicine in the Pulmonary and Critical Care Division;and Center for Bioethics, University of Minnesota (UM). Her research focuses on policies that govern the national distribution of lungs for transplantation therapy of end stage lung diseases. The institutional environment at UM provides very strong support through the UM Lung Transplant Program (one of the world s most active, and one of very few that currently perform living donor (LD) lung transplants) and the Center for Bioethics, which has an active interest in ethical issues related to LD organ transplantation. This proposal is designed to: 1) promote Dr. Valapour s development as a scholar in the field of lung transplantation ethics and policymaking by extending her prior training in pulmonary medicine and bioethics, and by adding formal training in health policy and clinical research methodologies. This will give her additional skill sets to become an independent researcher in the area of LD organ transplantation outcome assessment and policy development;and 2) inform the policy debate regarding LD lung transplantation. LD lung transplantation offers the only therapy for the many individuals unlikely to survive until a deceased organ becomes available. The research program of this grant will identify barriers that have prevented LD lung transplantation from becoming widely available by comparing it to the more widely practiced enterprise of LD kidney transplantation. The specific aims of this proposal are to: 1) Compare perceptions about LD transplantation of lungs versus kidneys in key stakeholder groups, including recipients, donors (related or non-related), nondonating family/friends, and transplant professionals;2) Compare policies and practices related to LD transplantation of lungs versus kidneys at a group of institutions that currently perform, stopped performing, and never performed LD lung transplantation (Aim 2A). Using the information from Aims 1 and 2A, she will develop policy recommendations for improvement of LD lung transplantation (Aim 2B). These aims will be accomplished by utilizing quantitative and qualitative data collection and analyses, along with ethical and policy analyses, thereby putting into use the new knowledge acquired during didactic components of this training program.	{ "pile_set_name": "NIH ExPorter" }
As highlighted at a recent NIMH Research Roundtable, the field of psychiatry is in dire need of research addressing the special needs of children with severe mood and behavioral dysregulation who meet diagnostic criteria for bipolar disorder. Although studies have addressed clinical characteristics, family history, and treatment, no study to date has addressed the longitudinal course of children and adolescents with bipolar disorder. To help fill this gap and contribute important information about the validity of the diagnosis of pediatric-onset bipolar disorder, we propose a five-year follow-up study to assess the outcome of children and adolescents with DSM-IV bipolar disorder. At the completion of the current Mentored Clinical Scientist Development Award period, we will have ascertained 107 children and adolescents (73 children, 34 adolescents; 24 girls, 83 boys) with bipolar disorder along with 296 first-degree relatives. This sample has been well characterized along multiple domains of functioning (psychiatric, cognitive and psychosocial) as well as phenotypically (with photographs of face and hands) and genotypically (with blood samples of probands and first-degree relatives). Each proband and his or her parent(s) has been assessed in direct clinical interview by the Principal Investigator. At the follow-up assessment, we will examine the outcome of the probands and siblings in three domains of functioning: psychiatric, neuropsychological and psychosocial. Our intention is not only to determine which children with bipolar disorder continue to meet criteria or not, but to assess functioning in multiple domains over time. Our main aims regarding pediatric-onset bipolar disorder are to assess: 1) patterns of persistence and remission; 2) course; 3) patterns of comorbidity, neuropsychological functioning and psychosocial disability; 4) the course and outcome of high risk siblings; 5) the association with attention deficit hyperactivity disorder; 6) the association with conduct/antisocial disorder; 7) the association with substance use disorders; 8) the utility of the Child Behavior Checklist as a screening instrument. This follow-up study of children and adolescents with bipolar disorder seeks to contribute valuable information regarding the longitudinal course and outcome of pediatric-onset bipolar disorder in multiple domains.	{ "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. Participants will be 14 adults between the ages of 20-22 years and 16 teens between the ages of 13-14 years. The scanning session will begin with a sagittal scout for purposes of prescribing subsequent images. Sagittal T1 MPRAGE images will be aquired for coregistration of functional images. Whole brain EPI BOLD images will be acquired in three runs while subjects perform the behavioral task below. An EPI fieldmap will be acquired at the same resolution and slice locations to allow for offline correction of field inhomogeneities. The behavioral combines a go/nogo task with emotionally arousing background images selected from the International Affective Picture System. In this task, letters are presented sequentially in a small box at the center of the computer screen. Subjects are asked to respond as quickly as possible with a button press to every one of a series of letters except for a specific target: an 'X'. The Xs appear on 25% of trials such that participants acquire a prepotent tendency to press and need to actively inhibit their response during nogo trials. The behavioral task consists of three runs of twelve blocks. Responses (press or no press) and reaction times will be recorded using an MRI compatible button box. We anticipate the entire session taking one hour per subject. This estimate includes 15 minutes for equipment setup and positioning of the participant, 11 minutes of structural scanning, 30 minutes of functional scanning and 10 minutes of clean-up and data transfer.	{ "pile_set_name": "NIH ExPorter" }
At least one-fourth of elderly persons in the United States live in rural areas and may, as a result of specific occupational and environmental exposures, socioeconomic characteristics, racial and ethic makeup, availability of infrastructure and resources, and cultural milieu, represent a unique group. The proposed Exploratory Center Grant on the Health and Effective Functioning of Older Rural Populations will establish a scholarly environment which will foster the development of rural gerontology and geriatrics. The proposed Center will offer administrative, biostatistical, data management, geographic, and demographic/sociologic consultation services through a centralized core. The Center's executive staff will regularly receive input from an External Advisory Board, which will be charged with the elaboration of a research agenda. A Steering Committee will assist with the implementation of the research agenda through administration of a pilot grant program. The Center will also sponsor at least one workshop, publish a regular newsletter, develop a local seminar/journal club, and engage in other outreach activities. At least three tracks of research will be maintained, focussing on the older agricultural worker, special rural populations, and social demography of rural areas. Specific proposals for each of these tracks are offered here.	{ "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. Nicotinamide adenine dinucleotide (NAD) is a molecule that is ubiquitous throughout all of life and is essential not only as a cofactor in redox reactions, but can also act as an adenosine diphosphate ribose (ADPR) donor. 'NAD consuming'enzymes, including ADP-ribosyl transferase, poly-ADP-ribosyl polymerase, cADP-ribose synthetase, and Sir2 protein deacetylase, can rapidly deplete cellular NAD stocks and in the process produce nicotinamide. Not only is it necessary for the NAD pool to be replenished, but the nicotinamide produced can act as a feedback inhibitor of these NAD consuming enzymes. While mammals can use nicotinamide directly to recycle NAD, most lower organisms must first convert it into nicotinic acid using a nicotinamidase enzyme. Not only does nicotinamidase play an essential role in NAD recycling, it can also act as a regulator of the NAD consuming enzymes by controlling cellular levels of nicotinamide. In addition, this enzyme has been shown to catalyze the conversion of the tuberculosis prodrug, pyrazinamide, into its active form, pyrazinoic acid. Structural studies completed to date have provided little information about substrate binding in the active site of this enzyme. A thorough study of ligand binding will provide crucial information about this enzyme's active site allowing for a deeper understanding of the mechanistic enzymology and providing essential details for structure-guided drug design efforts.	{ "pile_set_name": "NIH ExPorter" }
This study will use nuclear imaging techniques, analysis of lung lymph, and in vivo electron dense tracers to examine trans- epithelial movement of solutes among the air, interstitial, and vascular spaces of the lungs of anesthetized sheep. The relationship between the site of solute movement through the epithelium and alveolar edema will also be studied. Our studies to date show that nuclear imaging of aerosolized 99Tc-labeled albumin clearance may be a valuable technique for detecting changes in epithelial permeability because it is non- invasive, appears to be sensitive to lung injury, and its clearance rate is unaffected by changes in lung volume. To further evaluate this technique, pulmonary and systemic lymph will be collected to monitor the appearance of the albumin in the interstitial spaces. We will compare labeled albumin to the currently popular 99Tc- DTPA (diethylenetriamine pentaacetate) for detecting physiologically relevant increases in pulmonary epithelial permeability by measuring their clearances from the lungs of sheep with lung injury induced by intravenous infusions of air or oleic acid. We will also measure the effects of increases in lymph flow and alterations in the distribution of blood flow to the tissue on the clearance of labeled albumin. To determine if the site of solute clearance is through the alveolar or extra-alveolar epithelium, colloidal gold or horseradish peroxidase will be instilled in the airspaces of anesthetized sheep that receive intravenous oleic acid and lung tissue samples will be examined under an electron microscope. Additional sheep will receive one of the tracers intravenously during the development of oleic acid-induced alveolar edema to determine if the pathways for solute clearance are the same as those through which circulating solutes enter the airways during alveolar flooding. These studies on the measurement of increased epithelial permeability and the routes of solute movement during alveolar flooding will aid in the development of improved methods for the detection, prevention and treatment of alveolar edema.	{ "pile_set_name": "NIH ExPorter" }
Our long-term goal is to develop an inexpensive, simple, portable means of screening for oral cancer in the field by basic level healthcare workers in low-resource environments. Screening will identify whether a person needs to travel to a center with higher levels of expertise in oral cancer for further tests and potential cancer therapy. The objective of this application is to re- engineer existing technology to provide an inexpensive very small, robust portable diagnostic system for oral cancer based on Optical Coherence Tomography (OCT) and a simple diagnostic algorithm that indicates further diagnostic and treatment needs for each individual. Briefly, under the first Specific Aim a robust portable diagnostic system for oral cancer based on Optical Coherence Tomography (OCT) will be constructed and undergo clinical testing at the university of California, Irvine. Under Specific Aim 2, the mini-OCT device and a simple diagnostic algorithm will be tested in India in a specialty Head and Neck Cancer Clinic, and by low-level healthcare workers in the field. The proposed collaborative work between researchers at the University of California Irvine and the Mazumdar-Shaw Cancer Center in Bangalore is directly responsive to PAR 11-044 as the proposed device will provide point-of-care cancer screening by basic level healthcare workers. The low-cost, portable, robust imaging system will have the potential for integration into a networked mobile diagnostic network. Relevance. Annually 275,000 oral cancers (OC) and 125,000 deaths due to OC occur worldwide, and OC accounts for up to 50% of cancer cases in India. Because over 2/3 of OC lesions are detected late, treatment outcomes and prognoses are extremely poor. Currently there exists no portable, low-cost, robust, inexpensive simple means of screening for oral cancer in the field by basic-level healthcare workers. We propose to develop and validate a diagnostic modality that will overcome this critical barrier to widespread screening of high-risk populations in low-resource environments. The low-cost, portable end product will provide the basis for future networked and mobile technology for remote cancer diagnostics via minimally educated healthcare workers in the field.	{ "pile_set_name": "NIH ExPorter" }
Project Summary Traumatic brain injury (TBI) is a devastating neurological injury afflicting over 1 million people annually, including a large number of young adults and military personnel. Cerebral edema is associated with increased intracranial pressure (ICP) and a poor clinical outcome following TBI, although the cellular mechanisms underlying this process remain unknown. This gap in the understanding of cerebral edema formation contributes to the lack of clinically- effective therapeutics for TBI patients. Recent work by our laboratory demonstrates that acute neuronal necrosis stimulates the passive release of high mobility group box protein 1 (HMGB1), which in turn induces glial swelling and cerebral edema. Specific Aim 1 will establish whether activation of individual NMDA receptor subunits increase neuronal injury and cerebral edema following experimental TBI. The incorporation of NR2A and NR2B knockout mice will determine whether individual NR2 subunits contribute to HMGB1 release, brain swelling, and neurological outcome using following head trauma. Specific Aim 2 will determine whether toll-like receptor-4 (TLR4) mediates the pro- inflammatory and cerebral edema promoting effects of HMGB1. The ability of HMGB1 to stimulate the astrocytic water channel, AQP4, will also be addressed in TLR4 mutant mice. Specific Aim 3 will determine whether HMGB1 may represent a novel biomarker to predict the development of cerebral edema folowing head trauma in humans. Measurement of HMGB1 levels within the cerebrospinal fluid (CSF) and serum of neurotrauma patients will be correlated with acute neuronal injury and neurological outcome. Together, the proposed studies will investigate the novel possibility that HMGB1-TLR4 signaling contributes to the development of cerebral edema and increased ICP following TBI. The results of these studies may support the future development of novel therapeutics directed against this pathway to limit neurological injury following head trauma.	{ "pile_set_name": "NIH ExPorter" }
In this preclinical trial, we propose to target rho-associated protein kinase (ROCK) using fasudil in models of acute focal cerebral ischemia followed by reperfusion. ROCK is a major regulator of actin cytoskeleton controlling numerous functions in vascular smooth muscle, endothelium, neurons, glia, leukocytes and blood cells. Many of these are relevant for the pathophysiology of stroke, making ROCK a unique pleiotropic target with multiple converging and synergistic mechanisms, including cerebrovasodilation and reduced blood viscosity improving collateral flow in hyperacute stroke, and anti-inflammatory and anti-edema effects in acute to subacute stages. We and others have targeted ROCK in models of ischemic stroke in exploratory studies, vast majority of which showed improved tissue and functional outcomes, in multiple species and stroke models, and using various inhibitors and therapeutic paradigms. In a systematic review and stratified metaanalysis of 25 experimental studies, ROCK inhibition reduced infarct volume by 37% and improved neurological scores by 41%. Fasudil is the most commonly used ROCK inhibitor in experimental stroke and is off-patent to be procured in any formulation for both preclinical and clinical trials without delay. Fasudil has been the clinical standard of care for two decades in acute SAH in China and Japan, with a clean safety record. There have been numerous clinical trials of systemic fasudil in both US and abroad in healthy volunteers, and in patients with chronic cerebral infarcts, coronary artery disease, heart failure, and pulmonary hypertension among others. Indeed, in a small trial in acute stroke fasudil was efficacious. Nevertheless, none of these trials revealed any serious adverse events either with acute single doses or chronic daily dosing. We propose to confirm this promising profile in a preclinical trial using transient middle cerebral artery occlusion. Aim 1 will establish the optimal dose (Exp 1), confirm efficacy on long-term outcome (Exp 2), test whether fasudil extends the therapeutic window for recanalization (Exp 3), examine the efficacy of intraarterial delivery (Exp 4), and confirm efficacy using clinically-relevant MRI indices (Exp 5). Aim 2 will then test efficacy and safety in a clot model with or without tPA (Exp 6), and safety in comorbid hypertension and diabetes as well as combination treatments (Exp 7) and in permanent ischemia (Exp 8). The proposal comes from an experienced group of investigators with proven expertise, capable of adjusting to the needs of the network. Collectively, we have published ~100 manuscripts using various stroke models and neurocognitive and tissue readouts suitable for both acute (<72h) and chronic (up to 2 months) assessments in mice and rats. Besides the proposed experimental models, our expertise covers hemorrhagic transformation, combination treatment with tPA and ischemic brain edema. Altogether, we passionately support the concept of an unbiased multicenter preclinical testing platform such as SPAN for experimental stroke therapeutics. We are fully committed to support this network.	{ "pile_set_name": "NIH ExPorter" }
Coagulation proteases, in addition to their role in the regulation of blood coagulation, can modulate intracellular signaling events by activating a subfamily of G-protein coupled receptors named protease-activated receptors (PARs) expressed on the cell surface of various organs. In vitro studies have indicated that while the anticoagulant protease, activated protein C (APC), in complex with endothelial protein C receptor (EPCR) elicits a barrier protective response via activation of PAR-1 in vascular endothelial cells, thrombin elicits a barrier disruptive response via the activation of the same receptor. APC has been approved by the FDA for treatment of severe sepsis. It has been hypothesized that the activation of PAR-1 by APC on endothelial cells is responsible, at least partially, for its protective properties in preventing organ failure (i.e., lung and heart) which is caused by septic shock in severe sepsis. Thus, it is highly important to understand how the activation of PAR-1 by either APC or thrombin elicits paradoxical protective and disruptive cellular responses, respectively. We recently discovered that the activation of PAR-1 by thrombin can also elicit barrier protective cellular responses if EPCR is occupied by the Gla-domain of protein C. We demonstrated that the unoccupied EPCR together with PAR-1 are both associated with caveolin-1 within lipid-rafts of endothelial cells. Under these conditions, PAR-1 appears to signal via Gq and/or G12/13 subfamily of G-proteins, and thus thrombin activation of PAR-1 leads to disruptive cellular responses. However, we discovered that the occupancy of EPCR by protein C results in dissociation of EPCR from caveolin-1, thereby recruiting PAR-1 to protective pathways by likely coupling it to Gi-protein. Based on our recent preliminary data, we hypothesize that the occupancy of EPCR by protein C can also change the signaling specificity of PAR-2. Thus, it appears that the cell surface occupancy of EPCR determines the signaling specificity of activated PAR-1 and PAR-2 not the proteases that are cleaving them. To understand the PAR-dependent signaling specificity of coagulation proteases, we have prepared several mutant proteases and receptors and set up appropriate in vitro and in vivo models to investigate the following four Specific Aims. In Aim 1, we will investigate the PAR-1- and PAR- 2-dependent signaling specificity of coagulation proteases under conditions in which endothelial cells have been pre-incubated with vitamin K-dependent coagulation zymogens so that to occupy EPCR or other potential receptors by their natural ligands. In Aim 2, we will investigate the contribution of the Gla-domain of vitamin Kdependent coagulation proteases to the recognition and cleavage specificity of PARs localized into the lipidrafts. In Aim 3, we will develop novel APC variants which have improved PAR-1-dependent protective signaling properties, but have reduced anticoagulant activities. In Aim 4, we will characterize the cardioproperties of the improved APC variants or variants possessing only signaling activity or only anticoagulant activity in a mouse ischemia/reperfusion injury model. PUBLIC HEALTH RELEVANCE: The studies of this application will focus on the mechanism by which coagulation proteases activate a subfamily of G-protein coupled receptors called protease-activated receptors to alter the properties of vascular endothelial cells. The activation of these receptors by coagulation proteases is known to be important for the regulation of a range of (patho) physiological processes including coagulation, inflammation and angiogenesis. Understanding this role of coagulation proteases can lead to development of a new generation of therapeutic drugs for controlling thrombotic and inflammatory disorders, including heart attack and severe sepsis.	{ "pile_set_name": "NIH ExPorter" }
The long term objective of this proposal is to develop a diagnostic kit for monitoring nonenzymatically galactated proteins in galactose. Classical galactose is a disorder of galactose metabolism caused by a deficiency of galactose-1-phosphate uridyltransferase. The resulting severe impairment of galactose metabolism has been recognized for half a century and the frequency of this disorder is estimated to be 1 in 40,000 births. Generally, the biochemical phenotype includes elevated concentrations of galactose, galactose-l- phosphate and galactitol in tissue and body fluids. While the disease is generally fatal, treatment consisting of severe restriction of dietary galactose has proved life saving; nevertheless, most patients develop neurological abnormalities despite this diet. Earlier studies demonstrated the occurrence of elevated levels of glycated hemoglobin/albumin in galactosemia prompting the speculation that, akin to diabetes with glycated proteins, the monitoring of galactated proteins may prove useful for managing galactosemic patients. During Phase I, we will attempt to develop a monoclonal antibody probe directed to nonenzymatically galactated albumin that by standard immunological techniques will prove specific to galactated albumin. This probe will be then used in ELISA to establish its sensitivity and range of detection. Phase II efforts will focus on evaluating this ELISA in a clinical setting and optimizing its performance for the management of galactosemic patients. Phase III efforts will focus on the commercialization of this assay. Vandalia Research, Inc. is committed to manufacture, market and sell the diagnostic kit that will result from this research effort. Pediatricians and internists have long been waiting for assays to monitor galactosemic patients; a method to quantify galactated proteins may provide a viable approach for preventing or delaying many of the complications of the disease including the onset of neurological damage during childhood and post adolescent years. [unreadable] [unreadable] [unreadable]	{ "pile_set_name": "NIH ExPorter" }
The initiation of unhealthy risk behaviors such as premature sexual activity, alcohol use, and potentially violent acts like bullying and physical fights is a hallmark of adolescence. It is often the case that these behaviors occur in combination with another, increasing the potential for adverse social and health outcomes. Research has shown that exposure to media content that features depictions of a specific risk behavior is a key influence on adolescents' performance of that behavior. However, studies have failed to account for the effect of multiple risk character portrayals in popular media, and how it may affect risk behavior patterns. This project uses an innovative conceptualization of exposure to risk behavior to study the longitudinal association between multiple risk portrayals in popular movies and television shows and multiple risk behavior in youth. We will content analyze top-grossing mainstream and African American oriented films and television shows for depictions of single and multiple risk behaviors, and will link the results of the content analysisto media exposure and behavior data collected over 3 waves from a nationally representative sample of 14-17 year old teens. Because some adolescents are more susceptible to media effects than others, we will consider background factors (race) and personality characteristics (sensation seeking and impulsivity) as moderators of the exposure-behavior relationship. With the inclusion of African American media titles in our content analyses, we depart from more conventional approaches that rely solely on mainstream media despite the popularity of African American oriented media among African American youth. The findings will advance our understanding of how media contribute to youth engagement in dangerous combinations of risk behaviors and have implications for risk and disease prevention efforts.	{ "pile_set_name": "NIH ExPorter" }
This is a continuation of our studies of the electrical behavior of sympathetic preganglionic neurons (SPN's) in vitro and of transmitters activating or modulating the activity of these neurons. Experiments will be performed on SPN's contained in thin (300-400 Mum) sections of thoracic spinal cord removed from neonatal (10-15 days old) rats and from age-matched spontaneously hypertensive rats (SHR); intracellular current or voltage recording techniques will be used. Fluorescence labeling of SPN's in thin slices will be accomplished by placing for 1-2 hours a drop of 3% lucifer yellow CH solution on ventral rootlets. Spinal cord slices will be superfused with a physiological solution and viewed under a fluorescent microscope. Glass microelectrodes will be inserted under visual guidance into fluorescent SPN's. The proposed research has five major objectives. First, single electrode voltage clamp will be employed to voltage clamp SPN's with the aim to characterize the membrane currents that can be recorded from soma of SPN's. Second, synaptic potentials will be evoked by bipolar stimulating electrode placed 100-200 Mum away from the impaled SPN, and their characteristics will be analyzed under current or voltage clamp conditions. The data obtained will serve as a basis for identification of the transmitters responsible for synaptic potentials. Third, pharmacological antagonists to amino acids, biogenic amines and peptides will be employed to identify which transmitters may be involved in the generation of evoked synaptic potentials; furthermore, attempts will be made to mimic the synaptic potentials by application of suspected transmitters. Fourth, effects on SPN's and on evoked SPN transmission of centrally acting antihypertensive drugs such as clonidine and methyldopa, will be evaluated for their possible spinal site of action. Fifth, electrical membrane properties and chemosensitivity of the SPN's of spinal cord slices obtained from SHR will be investigated and compared with those of SPN's obtained from normal rats. The significance of this research is threefold: (1) this study will lead, for the first time, to characterization of the transmitters and modulators acting on SPN's; this will give insight into circuitry of supra-spinal control of the sympathetic nervous system; (2) comparison of electrical membrane properties of SPN's of normal rats and SHR and of their chemosensitivity will help our understanding of hypertension; and (3) identification of the site and mechanism of action of clonidine and methyldopa will establish the spinal cord as a potential site of action of antihypertensive drugs.	{ "pile_set_name": "NIH ExPorter" }
Four subjects will be studied for 3 drug dosages and a placebo in a double-blind three way cross-over design. Effects of injected IV drug on lower esophageal manometry tube and pH electrode for 6 hrs after IV injection. After overnight fast an esophageal manometric catheter with an attached pH probe will be passed transnasally. After 30 minutes of baseline recording zacopride will be injected intravenously. LES pressure monitored for 6 hours after injection.	{ "pile_set_name": "NIH ExPorter" }
This project funding is in support of informatics activities conducted by the National Cancer Institute's (NCI) CBIIT. CBIIT offers critical open-source infrastructure components that others can use to develop valuable databases and software tools to meet specific research needs. NCI's expanding suite of tools is built from these foundational components. Our projects bring tools and partners together to tackle key challenges.	{ "pile_set_name": "NIH ExPorter" }
Project Summary World-wide sequencing efforts have identified recurrent novel genetic lesions associated with disease progression, inferred patterns of common co-occurrence, and characterized clonal evolution in response to treatments in chronic lymphocytic leukemia. Despite the wealth of available genetic information, these analyses all rely on statistical inference, thus limiting the possibility to link the genotype to the phenotype. Currently, understanding functions of high frequency genetic lesions and how they cooperate with their co-occurring mutations to cause CLL initiation and progression are not available. In this proposal, we aim to understand how two of the most recurrent genetic lesions in CLL (SF3B1 mutations and ATM deletions) mechanistically impact CLL initiation and progression with the overarching goal to understand CLL biology, generate novel murine model, and find new strategies for treating CLL with these lesions. This proposal is based on our finding that co-expression of mutated Sf3b1 with Atm deletion resulted in the development of clonal pathognomonic CD19+CD5+ B cells in blood, marrow and spleen at low penetrance in aged (18 months) mice, that can be propagated by in vivo passaging. Interestingly, whole-genome sequencing of DNA from murine CLL revealed recurrent chromosome amplifications, suggesting chromosome instability as a mechanism contributing to CLL in the mice with Sf3b1 mutation and Atm deletion. We now propose to investigate the hypothesis that SF3B1 mutations promote genomic instability through RNA splicing related R- loop formation while ATM deletion further augment genomic instability through decreasing R-loop associated DNA damage repair (Aim 1). We further hypothesize that modulation of the R-loop formation is contributing to the acceleration of CLL (Aim 2). Thus, targeting both RNA splicing and DNA damage response checkpoints are likely to provide synthetic cytotoxicity for CLLs with these lesions (Aim 3). Completion of the proposed work will create a fundamental foundation to explain how splicing factor mutations impact genomic instability and contribute to oncogenesis, and provide rationale for designing novel clinical trials in CLL patients with both SF3B1 mutations and ATM deletions.	{ "pile_set_name": "NIH ExPorter" }
Bradykinin (BK) is a tissue hormone with vasodilatory properties, whose role in regulating systemic blood pressure or local perfusion and metabolism of certain organs is still poorly understood. Its participation in cardiovascular physiology has been explored in the past via use of selective antagonists of the B2-type BK receptors, which are believed to mediate its clinically significant actions. The development of genetically engineered mice whose B2-receptor gene has been disrupted (B2-knockouts) should permit more accurate evaluation of the role of BK. The hypothesis to be tested in this proposal is that BK, via its B2 receptor-mediated actions, tends to counteract hypertensive mechanisms and to protect the perfusion and metabolism of sensitive organs from hypertensive or ischemic tissue damage. The following Specific Aims are proposed: 1. Development of experimental hypertension of the renin-dependent (renovascular) or salt- dependent (DOC-salt, end-stage renal disease) types in B2-receptor gene knockout mice to be compared with wild type mice. 2. Development of ischemic cardiomyopathy post myocardial necrosis via: a) angiotensin II infusion and b) coronary artery ligation in mice: comparison of cardioprotective role of ACE inhibition or Ang II blockade in wild type vs B2-receptor gene knockout mice. 3. Study of insulin-mediated glucose metabolism via euglycemic insulin infusion clamp to compare glucose utilization in wild type vs B2 receptor gene knockout mice at baseline and during ACE inhibition. 4. Study of the relationship of selected local vasodilators with BK in maintaining blood pressure in B2 receptor gene knockout mice (i.e., NO, substance P, ANF) via use of their specific inhibitors. Our laboratory has had extensive experience in the past with experimental studies using rats. We have successfully adapted our methodology and equipment to the study of mice, and have acquired expertise in inducing and quantitatively monitoring cardiovascular diseases in diminutive animals.	{ "pile_set_name": "NIH ExPorter" }
Cystic airspace enlargement and spontaneous pneumothorax are major pathological manifestations Birt-Hogg- Dube (BHD) syndrome. Although the etiology of this disease is not known, lung cysts in BHD are linked to autosomal dominant mutational inactivation of tumor suppressor gene folliculin (FLCN). FLCN, a 64-kDa ubiquitously expressed protein with high homology throughout species that lacks apparent functional domain, through adaptor proteins FNIP1/2, binds the 5'-AMP-activated protein kinase (AMPK). Little, however, is known about a role of folliculin (FLCN) in lung cell survival. Our in vitro and in vivo studies demonstrate that FLCN is required for lung cell survival by acting within 5'-AMP-activated protein kinase (AMPK) - tuberous sclerosis complex 2 (TSC2) - mammalian target of rapamycin complex 2 (mTORC2) signaling pathway. To fill a gap in the current understanding of airspace enlargement, we have restricted the scope of this proposal to one testable, central hypothesis that FLCN is required for lung cell survival. To test our hypothesis, in Aim 1, we will determine whether FLCN is required for lung epithelial cell survival in vivo using new transgenic FLCNf/f:SP-C- Cre mice generated in PI's lab. Targeted conditional inducible deletion of FLCN in SP-C-expressing lung epithelial cells will allow us to identify the specific lung epithelial cell type affected by FLCN loss that promotes alveolar space enlargement. In Aim 2 we will examine whether FLCN is required for maintenance of epithelial cell morphology and metabolism. In Aim 3, we will determine FLCN role in regulating activities of Akt and AMPK kinases, and whether pharmacological activation of AMPK will rescue lung epithelial cell survival with targeted inducible deletion of FLCN in FLCNf/f:SP-C-Cre mice. These studies will identify the role of FLCN in lung cell survival, provide insights into the cellular an molecular mechanism of lung epithelial cell-cell contacts, actin cytoskeleton, and metabolism regulated by FLCN. These studies will also establish a mechanistic link between loss of FLCN and cystic airspace enlargement by using our unique novel FLCNf/f:SP- C-Cre transgenic mice and identify potential molecular target(s) for novel therapeutic approaches to treat BHD.	{ "pile_set_name": "NIH ExPorter" }
The objective of the proposed research program is to develop a low-cost nanopore sequencing technology taking advantage of the latest advances in solid-state nanopore technology and utilizing the recently proposed concept of hybridization-assisted nanopore sequencing (HANS). It has been proposed by the PI and coworkers that by hybridizing short matching strands of oligonucleotides, known as DNA probes, and using nanopore ionic conductance to detect the positions of the probes, one can construct DNA sequences using the ionic current profiles, thus avoiding the classical "repeat problem" in the standard sequencing-by-hybridization (SBH) method, and bypassing the harsh requirement of single-base spatial resolution (0.4nm) in the original nanopore sequencing proposal of Kasianowicz et al. The proposed HANS sequencing technology has the potential of being fast, low-cost and portable. The proposed research is to test the feasibility of the HANS concept. The specific aims of the project are: (1) Determine the spatial resolution of the ionic conductance method using a combined optical tweezers and nanopore setup. (2) Parallel manipulation of DNA translocations in multiple nanopores. (3) Detect hybridization of single DNA probes on a single-stranded ssDNA using optical tweezers and ionic conductance of the nanopore. (4) Re-sequencing a sample DNA sequence to determine accuracy. (5) Develop non-specific DNA-bead binding technology for de novo sequencing. The proposed research will greatly advance the field of bio-nanotechnology and could directly result in a low-cost DNA sequencing technology, which in turn will have far-reaching benefits to the public health of the US in disease detection and in the studies of the molecular processes underlying diseases. The research program provides excellent training opportunities for students and postdoctoral researchers in preparing them to join the workforce of the emerging bio-nanotechnology economy.	{ "pile_set_name": "NIH ExPorter" }
The objective in this study is to elucidate the basic mechanisms involved in the regulation of erythropoiesis and to apply this knowledge to human erythropoietic diseases. The techniques of marrow cell culture will be used to study the effect of erythropoietin (Ep) and endotoxin on marrow erythroid precursor cells. The effect of endotoxin on the number, state of cell cycle, velocity sedimentation at unit gravity and Ep sensitivity of murine day 3 and day 7 BFU-E will be determined. The relation of endotoxin, as a contaminant in most Ep preparations, to the development of day 7 BFU-E in vitro will be determined. These experiments should clarify the developmental characteristics and capacities of erythroid precursor cells by their response to endotoxin and Ep, and should aid in the interpretation of prior experiments up to this time that have been performed with contaminating endotoxin present in Ep preparations. The polycythemia produced by the Friend virus will also be studied. Friend virus markedly increases erythropoiesis in mice by a mechanism that does not require the presence of added Ep. We have shown that Friend virus produces erythroid bursts when added to mouse marrow cells in vitro. We will determine if the erythroid bursts produced by Friend virus in vitro are clones arising from a single cell by mixing male and female cells and determining if the bursts have a homogeneous karyotype. Studies will be performed to identify the in vitro target cells for Friend virus-induced erythroid bursts and to determine if Ep has a role in the production of erythroid bursts by the Friend virus in vitro. The relative roles of the spleen focus-forming virus and the murine leukemia virus complex will be investigated. The study of the effect of Friend virus on the early erythroid precursor cells will greatly increase our knowledge of the different developmental stages and capacities of these cells and should provide a model for neoplastic transformation in vitro. Finally, we will attempt to construct an animal model for polycythemia vera.	{ "pile_set_name": "NIH ExPorter" }
This study will determine whether certain factors in the blood are associated with the severity of diabetic retinopathy	{ "pile_set_name": "NIH ExPorter" }
Studies of Project 1 showed that protein kinase A isozyme switching, via eliciting differential cAMP signaling in the cell, might provide a tumor target-based gene therapy for cancer treatment [1]. PKA isozyme switching can be achieved in many ways, including use of site-selective PKA activators such as 8-Cl-cAMP, viral and non-viral vector-mediated gene overexpression, antisense DNAs, siRNAs, and targeted gene repair/replacement therapy methodologies. Our study clearly warrants translational research. GEM 231 (RIa antisense) is under Phase I-II clinical studies. 8-Cl-cAMP for Translational Research This study is performed in conjunction with NIH FY2005 Bench-to-Bedside Awards to Yoon S. Cho-Chung, M.D., Ph.D., and Constantine A Stratakis, M.D., NICHD, on the project entitled, "Site-Selective cAMP Analogs for Treatment of Carney Complex." Carney complex (CNC), the complex of spotty skin pigmentation that can accompany multiple endocrine neoplasia, is attributed to the mutational loss of PRKARIA, the gene that codes for the RIa subunit of cAMP-dependent protein kinase type I (PKA-I) [2]. Patients often have tumors of two or more endocrine glands, including primary pigmented nodular adrenocortical disease (PPNAD), pituitary adenoma, thyroid adenoma or carcinoma, testicular neoplasms, and ovarian tumors. The skin is affected by multiple lesions that are either pigmented (lentigines, caf-au-lait spots, common and blue nevi) or not pigmented (mainly myxomas and other skin tags). The genes responsible for CNC have been mapped to chromosome 2p16 and 17122-24 [2]. In affected subjects, tumor-specific loss of heterozygosity (LOH) has been found within the 17q region [2]. The presence of tumor-specific LOH suggested that the CNC gene is a tumor suppressor gene. PKA exists in two isoforms, PKA-I and PKA-II. The PKA isozymes are expressed in a balance of cell growth and differentiation. It has been shown that the PKA-I to PKA-II ratio is reversed in primary clinical tumors and transformed tumor cell lines compared to their normal counterparts, and that the experimental approaches that induce PKA-isozyme switching in cancer cells result in tumor cell growth arrest and induction of tumor reversion [1] (Project 1). The objective of this study is to restore the PKA-I to PKA-II ratio in the adrenal glands and other related tumors of Carney complex to that of normal tissue by the following experimental approaches: (1) Use of a site-selective cAMP analog, 8-Cl-cAMP [3], to restore normal PKA activity in adrenal glands and tumors of CNC patients in the setting of an 8-Cl-cAMP phase II-III clinical study; (2) Use of unhydrolyzable Rp-8-Cl-cAMPS and Sp-8-Br-cAMPS with 8-Cl-cAMP to enhance PKA-II activation [4]; (3) Use of the cAMP (CRE response element)-transcription factor decoy to inhibit tumor growth without harming normal cell growth [5]. A long-term goal is the RIa gene transfer/gene therapy of our patients; in the context of this grant we will test this in CNC tumor cell lines. In summary, Carney complex is an incurable disease and currently there is no treatment method for it other than the individual surgeries for the various types of tumors associated with it. We will attempt to restore the abnormally inactivated PKA activity back to the normal physiological state. Because of the loss of PRKARIA gene, the gene coding for RIa regulatory subunit of PKA-I is considered causative of Carney complex, we will attempt to restore the unbalanced PKA-I to PKA-II ratio back to its normal state by using the site-selective cAMP analog, 8-Cl-cAMP, which selectively activated PKA-II in a phase I clinical setting. Goals of the Study Clinical Goals I. Natural history of CNC and related tumors and skin lesions.	{ "pile_set_name": "NIH ExPorter" }
Drug metabolism, programmed cell death, DMA biosynthesis and repair, respiration, and photosynthesis all occur via electron-transport (ET) mechanisms. Theoretical and experimental advances indicate that both protein structure and dynamics control ET reaction kinetics, and that changes in the kinetics of these reactions may lead to disease states or may be used for therapeutic purposes. In the case of ET within proteins, structure and dynamics both appear to control the reaction mechanism. For inter-protein ET, minority population conformations appear to dominate the kinetics. Seminal recent experiments provide key structural and kinetic data for intra-protein and inter-protein ET that should allow us to develop a detailed understanding of the reaction mechanisms. Additional chemical modification and mutation studies are revealing details of the reaction mechanisms. Surprising rate dependencies on protein structure, docking mode, and solvent isotopes have been reported, and these observations are not anticipated by the simple static theoretical models. This proposal aims to develop molecular-level descriptions of ET within and between proteins with realistic fluctuating structures. The studies will target the influence of protein and solvent dynamics on intra-protein and inter-protein ET kinetics, with the aim of establishing rules to define how protein structure and interracial water control this kinetics. Reaching our long-term goal of developing a molecular-level understanding of how proteins control ET reaction rates should enable new strategies to disrupt or enhance redox processes; these strategies could lead to new therapeutic schemes derived from controlling the flow of electrons along essential biological electron transfer chains. [unreadable] [unreadable] [unreadable]	{ "pile_set_name": "NIH ExPorter" }
Colorectal cancer (CRC) is one of the most common cancers in both men and women in the United States, and comprises about 10% of new cancer cases. Colorectal cancer is an age-related epithelial cancer that often takes over twenty years to progress from initial benign adenoma (polyp) to invasive adenocarcinoma. The majority of colon cancers are characterized by genetic mutations in adenomatous polyposis coli (APC) or B-catenin, two key components of the Wnt signaling pathway. In the absence of Wnts, B-catenin is normally targeted for degradation by the GSK-3/Axin/APC complex. In the presence of Wnts, this degradation pathway is inhibited, B-catenin accumulates in the cytoplasm and nucleus and binds and transactivates Tcf/Lef proteins. Polyps arise when there is inappropriate activation of Wnt signaling, and B-catenin protein accumulates in the nucleus. We have recently developed a new mouse model of colon cancer where mice develop colon tumors that are consistent with those found in human FAP patients. As part of this proposal, we will further characterize this model using molecular tumor mariners. Additionally using mouse genetics, we will rigorously test via gain and loss of function studies, the role of Tcf4 on colon tumor formation in this mouse colon tumor model. Tcf4 is a member of the Tcf/Lef family of transcription factors that is essential for small intestinal cell proliferation and is expressed in human colon cancer cell lines and primary colon tumors. Taken together, results from these studies will provide important new insights into the role of Tcf4 during colon tumor formation, and may reveal novel downstream targets of Tcf4 function.	{ "pile_set_name": "NIH ExPorter" }
The candidate (Kenneth J. Ottenbacher) holds a faculty position at the University of Texas Medical Branch (UTMB) in Galveston, Texas, that involves several administrative roles including: Vice Dean in the School of Allied Health Sciences, Director of the Division of Rehabilitation Sciences, and Associate Director of the Sealy Center on Aging. The K02-Award will allow Dr. Ottenbacher to reduce his administrative responsibilities and focus additional time on research. K02 funding will permit him to expand his examination of the disablement process in older adults. His current research is funded by grants from the National Institute on Aging and, more recently, the American Heart Association. Specifically, Dr. Ottenbacher will systematically explore the relationship between functional status and two components of the disablement process associated with quality of life - patient satisfaction and participation in community and social/personal activities (as defined in the World Health Organization's, International Classification of Functioning, Disability and Health). The immediate goals for the K02 include: 1) reduce administrative responsibilities to a less than 25% time commitment, 2) increase publication rate by 20% per year for the next four years, and 3) increase amount of externally funded grant dollars by 100% by end of K02-award. These goals will be accomplished by permanent resignation of his administrative role as Vice Dean in the School of Allied Health Sciences and reassignment of other responsibilities, including transferring management of a Health Services Resources Administration training grant, for which he is currently PI, to another faculty member. These changes will allow Dr. Ottenbacher to devote a minimum of 75% time to research and achieve his long term goals of establishing a program of externally funded research supported by multiple R01 type grants that contributes to the understanding of older adults with disabilities.	{ "pile_set_name": "NIH ExPorter" }
Hospitalized patients faced with sudden inability to speak (e.g., patients with tracheal intubation) are unable to verbalize even the most basic comfort and care needs, or their urgent safety needs. These patients and nursing staff must consequently resort to alternative nonverbal communication strategies such as gestures, mouthing words, or the use of alphabet boards, which are slow, energy-draining, and ineffective. These strategies are also inadequate for expressing emergent, life-threatening needs such as inadvertent disconnection of ventilators/oxygen or a blocked airway. Currently, no standards of practice, commonly used best practices, or software/devices exist to facilitate the care of suddenly speechless (SS) patients. Clearly, current practice lacks effective approaches to prevent and rapidly treat dangerous situations in this highly vulnerable population and to lessen patients' frustration, anxiety, fatigue, and dissatisfaction with provided care. The purpose of this innovative project is to (1) advance development initiated during our Phase1 SBIR grant # 1R42NR010842-01 of communication software uniquely designed for this population, conduct a research study to determine whether GatorVoice improves SS patients ability to communicate with nursing staff, and produce a commercially viable communication device. Based on our Phase I project, GatorVoice is the first solution of its kind tailored specifically to communicate the needs of SS patients. Our pilot study demonstrated the feasibility of GatorVoice in the hospital setting. In addition, it will be the first system to provide data capturing from patient-staff interactions that can be organized as summary reports and printed out by nurses for use during shift reports. The project's emphasis is consistent with NINR's research focus including: 1) seeking ways to reduce the burden of illness and disability by understanding and easing the effects of acute and chronic illness and 2) technologies to be used in the hospital... that improve symptom evaluation in persons with chronic conditions (Omnibus Solicitation for SBIR/STTR Grant Applications, 2010). A study is proposed to examine the effectiveness of GatorVoice in facilitating the communication process for SS patients. A quasi-experimental study will be conducted with SS participants using GatorVoice and a control group of SS participants using current communication methods, e.g., paper/pen, gestures (and an Urgent Button developed for this study) to communicate. Outcome measures will be ease of communication, frustration with communication, and satisfaction with communication method. In a qualitative study, we will examine nurses' experiences with communicating with SS patients, and the impact that communication methods have on that process. An iterative process of product development and clinical testing will be used throughout Phase to advance maturation of GatorVoice software and integration into one or more hardware platforms, thereby bringing the system to the commercial stage.	{ "pile_set_name": "NIH ExPorter" }
ATG16L1 was recently identified in genome-wide association studies as a Crohn's disease susceptibility gene, thereby implicating autophagy in the pathogenesis of Crohn's disease. While the strongest genetic association results in a threonine to alanine transition at codon 300 (T300A) that may alter function of the resultant protein and initial studies have not yet identified Crohn's disease-associated changes in ATG16L1 expression, studies using hypomorphic mice expressing low ATG16L1 levels indicate that the reduced expression of ATG16L1 may significantly alter its function in intestinal epithelial cells. Overall, there is a lack of knowledge of how the expression of ATG16L1 and other autophagy genes are regulated. MicroRNAs are 21-24 nucleotide noncoding RNA molecules that negatively regulate gene expression by binding to complementary sequences in the 3'untranslated region (3'UTR) of mRNA transcripts and directing either mRNA degradation or protein translational inhibition. We have previously identified microRNAs that are differentially-expressed in Crohn's disease and ulcerative colitis tissues. An in silico analysis found that the ATG16L1 3'UTR contains over 25 putative microRNA binding sites and 3 naturally occurring SNPs that map to 3 distinct putative microRNA binding sites. Our preliminary results indicate that the ATG16L1 3'UTR plays a significant role in regulating ATG16L1 expression. Our long-term goal is to determine the role of microRNAs in the pathogenesis of inflammatory bowel diseases, such as Crohn's disease. The objective here is to determine the influence of 3'UTR microRNA binding sites in the regulation of ATG16L1 expression and function. Our central hypothesis is that microRNA play a significant role in regulating ATG16L1 expression and that microRNA-mediated alterations in ATG16L1 expression significantly impact autophagy in intestinal epithelial cells. We will address this central hypothesis with the following two specific aims: (1) To identify microRNA binding sites regulating ATG16L1 expression and subsequent autophagy and xenophagy;(2) To determine microRNA binding site SNP effects on ATG16L1 expression and subsequent autophagy and xenophagy. These aims will be pursued using established microRNA and autophagy functional assays and generating intestinal epithelial cells harboring the microRNA binding site SNPs using somatic cell gene targeting. Such results are expected to have an important positive impact because the determination of microRNA regulation of ATG16L1 expression and autophagy provides a physiologic mechanism by which autophagy can be regulated at a cellular level and influence disease processes. These results will confirm the utility of somatic cell mutations to study SNP influences on 3'UTR microRNA binding site function and may provide future microRNA-associated avenues by which autophagy-associated diseases may be targeted. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because the identification of microRNA-mediated mechanisms of ATG16L1 expression and function will increase our understanding of the role of microRNAs and autophagy in the pathogenesis of related disease, like inflammatory bowel disease, and provide potential mechanisms for future targeted therapeutics. The proposed research is relevant to the part of NIH's mission that pertains to applying innovative research strategies and fostering fundamental creative discoveries.	{ "pile_set_name": "NIH ExPorter" }
Loss of E-cadherin function is a critical event that is associated with epithelial-to-mesenchymal transition, invasiveness and the metastatic phenotype in human colorectal cancer and other carcinomas. Inappropriate activation of the canonical Wnt pathway is another critical event in the transition from normal epithelium to the neoplastic phenotype in colorectal cancer. Although it is well accepted that perturbation of these pathways represent important hallmarks of neoplastic tranformation, their mechanisms of transduction and regulation under normal and pathological conditions are poorly understood. We have developed highly reliable and sensitive assays for re-expression of E-cadherin in the SW620 colorectal cancer cell line that normally expresses very low levels of E-cadherin. Similarly, we have also developed a robust biochemical assay that recapitulates activation of the canonical Wnt pathway by the Wnt coreceptor, LRP6. Both of these assays have been adapted for a 384-well format. We now propose to interrogate regulation of E-cadherin expression and canonical Wnt signaling using a chemical genetics-based approach in a high-throughput screen to identify compounds that induce expression of E-cadherin in SW620 cells and that perturb degradation of beta-catenin and Axin, two key regulators of signaling through the canonical Wnt pathway. Our initial screen of 6,400 small molecules has already identified several lead compounds in both of these assays and we propose screen a total of 160,000 compounds. We propose to validate lead compounds from this initial screen in a variety of in vitro and in vivo assays. Our preliminary studies indicate that the small molecule trichostatin A, a histone deacetylase inhibitor, acts to induce E-cadherin expression in SW620 cells. We will explore its role in E-cadherin expression as well as its potential for regulating Wnt signaling. Finally, we will identify the mechanisms of action and potential targets of validated compounds by testing their effects on candidate cellular pathways as well as in biochemically reconstituted reactions.	{ "pile_set_name": "NIH ExPorter" }
Invasive Aspergillosis caused by azole resistant A. fumigatus has an alarming 12% survivability making this a clinical problem of acute significance. Early work on azole resistant isolates of A. fumigatus suggested that azole resistance was a relatively rare occurrence and that the genetic basis of resistance was most often due to changes in a gene (cyp51A) encoding the azole target protein, lanosterol ?-14 demethylase. Extensive studies from a group in the Netherlands provided compelling evidence that in that country, most azole resistant A. fumigatus isolates contained a single compound mutation consisting of a duplicated region of the cyp51A promoter region (TR34) and a change in the coding sequence altering the amino acid sequence of the protein (L98H). However, more recent studies from a group in the United Kingdom indicated that alterations in the cyp51A gene were unlikely to explain all azole resistance in A. fumigatus. More than 50% of azole resistant isolates from UK patients have wild-type versions of cyp51A, inconsistent with changes at this gene explaining drug tolerance. In 2013, the UK group identified an ATP-binding cassette (ABC) transporter-encoding gene called cdr1B as being associated with azole resistance in several patient isolates. Expression of cdr1B was elevated in highly azole tolerant clinical isolates while the cyp51A remained wild-type in sequence. Overexpression of ABC transporter-encoding genes resulting in azole tolerance has been the predominant route of azole resistance in the Candida species and we believe this will emerge as a key feature of drug resistance in A. fumigatus. This application will focus on the contributions of the cdr1B gene as expression of this ABC transporter is elevated in azole resistance isolates and loss of cdr1B produces an azole sensitive phenotype. We will directly compare the azole resistance phenotypes of wild-type and cdr1B? cells containing various cyp51A mutations that are associated with clinically relevant drug resistance. We will also evaluate expression of cyp51A using immunological probes to determine if these mutant alleles influence protein levels; information that is currently unavailable in A. fumigatus. We will carry out a forward genetic screen using impala transposon mutagenesis to identify genes that modulate cdr1B expression. Together, these approaches will provide important new information into the molecular basis of azole resistance in A. fumigatus and provide the direct assessment of drug resistance contributions from ABC transporters and the cyp51A gene.	{ "pile_set_name": "NIH ExPorter" }
The proposed research addresses two high priority areas identified by the Prostate Cancer Progress Review Group: the role of novel therapeutic agents directed at new targets and the use of relevant biomarkers to monitor clinical response. The studies proposed will be performed in the context of phase I a clinical trial of 17-N-allylamino-17-demethoxy geldanamycin (17-AAG), the first ansamycin antibiotic to enter clinical testing. Ansamycin antibiotics are a new class of compounds that induce the selective degradation of proteins which bind to the Hsp 90 family of chaperones. Hsp90 is required for protein refolding after stress, and for the conformational maturation of nuclear steroid receptors, certain protein kinases and other regulators of cell cycle control. Preclinical work by us and others has shown that malignant cells that express the androgen receptor (AR), the estrogen receptor (ER), HER2 and which overexpress cyclin D1 are sensitive to these agents, making prostate, breast and colon cancers of particular therapeutic interest. Approval to conduct a phase I clinical trial using of 17-AAG using an accelerated titration design has been obtained (LOI-98-355, Approval Date 4/15/99) and approved by the MSKCC Institutional Review Board (Protocol No. 99-37, Approved 5/25/99). In addition to the safety assessments and pharmacokinetic studies that are a routine part of the phase I study, we propose to conduct clinical and laboratory investigations to understand which tumors may be most sensitive, and which target proteins are affected in vivo. We will focus on the expression of the AR (prostate), ER (breast), HER2 and cyclin D1 in tumor biopsies obtained prior to and following treatment, proteins we have shown to be decreased by and which mark for sensitivity to the compound in vitro. These same proteins will be evaluated in circulating tumor cells isolated from the peripheral blood. To assess response, we will explore serial changes in FDG uptake using positron emission tomography. In vitro studies of established cell lines will also be conducted to explore the timing of the effects on specific proteins, and for other markers of drug effect. Preliminary data suggest that the accumulation of lipid droplets (prostate cancers) or fat (breast cancer) may mark for a differentiation effect of the drug.	{ "pile_set_name": "NIH ExPorter" }
Thrombogenicity of foreign substances is one of the leading issues limiting long term use of circulatory support devices and other devices having direct contact with blood. Numerous studies indicate that thrombogenic properties of a foreign surface can be significantly reduced by surface modifications. One very promising new technology is a biomimetic surfactant polymer coating developed by Marchant et. al. This coating has demonstrated excellent non-thrombogenic properties on graphite substrates, but under low shear stress environments. In Phase I, we will evaluate the feasibility of this novel coating for use in an extracorporeal blood pump, which is currently under design at BIOMEC. We will apply the surfactant polymer coating to critical surfaces of the pump, which have been coated with diamondlike carbon film. To evaluate the stability of our coating under the high shear stress conditions of the blood pump, we will subject test samples to high shear stress in a rotating disk apparatus. If successful, this will demonstrate the feasibility of surfactant polymer coatings for blood pumps and other circulatory devices, such as heart valves, cannulas for membrane oxygenators, and ventricular assist devices, and will improve the outcome of transplantations and other cardiac procedures. PROPOSED COMMERCIAL APPLICATIONS: The biomimetic coatings to be developed in this proposal are applicable to devices with blood contacting surfaces, including blood pumps, ventricular assist devices, artificial heart valves, and cannulas for extracorporeal membrane oxygenators.	{ "pile_set_name": "NIH ExPorter" }
A pilot/feasibility study of the efficacy and safety of secretin, a polypeptide neurotransmitter, to treat children with Autistic Disorder. Given the growing interest in secretin, a desire to serve area families with children with autism and a concern that this treatment receive scientific scrutiny the investigators have gathered an interdisciplinary group of researchers to address this issue.	{ "pile_set_name": "NIH ExPorter" }
Linkage analysis was performed on the synthesized glucans. The samples were methylated by the method of Ciucanu and Kerek. Powdered NaOH was added to the sample, and after stirring, methyl iodide was added. The reaction was quenched by addition of water after 20 min. The solution was extracted with chloroform. The permethylated sample was then hydrolyzed using 2 M trifluoroacetic acid. Glycosyl residues were reduced and then acetylated. GC/MS analysis was performed using a Hewlett Packard 5890 GC using a Supelco 2330 fused silica capillary column coupled to a 5970 MSD. The results showed the total ion chromatogram for the sample and the mass spectrum of the individual carbohydrate peaks.	{ "pile_set_name": "NIH ExPorter" }
The overall goal of this project is to elucidate the role of coagulation proteins in platelet function. Current work is aimed at studies of interaction between platelets and thrombin. Work in progress suggests that platelets contain a storage pool of microtubular protein in dense granules. Thrombin can produce a mobilization of the stored microtubular protein to form new microtubules upon platelet activation. This unique aspect of platelet-thrombin interaction may play a functional role in irreversible platelet activation and the release of platelet lysosomal enzymes. Further investigation will be aimed at developing a procedure to isolate and purify plasma membrane from thrombin-treated platelets followed by chemical characterization of these membranes. The proposed work is also intended to study the functional role of platelet fibrinogen in thrombin-induced aggregation.	{ "pile_set_name": "NIH ExPorter" }
Recent studies by the applicants have strongly indicated that immunologic phenomena are intimately involved in the pathogenesis of Mooren's ulcer and cicatricial pemphigoid. A Clinical Center is proposed to study and treat patients with cicatricial pemphigoid, Mooren's ulcer and other corneal diseases suspected of having autoimmune components. Because the diseases we propose to study are relatively uncommon and because laboratories with capabilities for extensive varied immunological testing are also uncommon, it is suggested that the proposed Clinical Center is the best way to gather significant data on the course, pathogenesis, and treatment of these painful, blinding diseases. Specifically, the Center will enable us to characterize the diseases as to their genetic aspects, possible etiologic factors, course of the diseases, and relate these findings to in-depth immunological studies. Finally, we will evaluate treatment modalities based on the clinical signs and synptoms of the diseases and on the immunopathological parameters in the tissues and sera of the patients.	{ "pile_set_name": "NIH ExPorter" }
The rat is a valuable animal model for the study of transplantation immunology, especially organ (kidney) allograft transplantation. The RT1 histocompatibility system in the rat is considered to be analogous to the major histocompatibility complex (MHC) of man. Studies using recombinant rats have demonstrated two gene regions of the MHC. One region codes for the major serologically defined alloantigen (RT1.A) and the other region codes for genes which control stimulation in mixed-lymphocyte-reaction (MLR), immune-associated (Ia) antigens, and immune response (Ir) genes. However, we and others have reported that certain rat strains (RT-11) identical at the MHC can exhibit immune responses usually associated with major histocompatibility differences such as MLR, rejection of skin allografts and graft vs host reactions. These observations suggested that these immune responses in MHC compatible rat strains may be due to non-RT-1 or minor histocompatibility antigens. An MLR model in which rats histocompatible at the MHC are alloimmunized with each other's lymphoid tissue will be employed to study the biological function of these non-RT-1 or minor histocompatibility antigens. Alloimmunization between RT-1 MHC compatible rats were performed for the following strain pairs: Maxx vs BN, RT-1n; Da vs ACI, RT-1a; Lew vs F344, RT-11. Alloimmunization between the two latter rat strain pairs produced altered reactivity in MLC suggesting differences in non-RT-1 or minor histocompatibility antigens. Experiments with congenic rat strains (Lew . B3 and F344 . B3) will be used to differentiate these two possibilities. Furthermore, the modulatory effects of alloimmunization, e.g., suppression of MLC reactivity may have important biological implication in the modification of the immune response in tissue/organ transplantation.	{ "pile_set_name": "NIH ExPorter" }
Structural priming occurs when speakers reproduce recently experienced sentence structures. Structural priming experiments with adults have demonstrated that the adult production system is sensitive to abstract representations of sentences. Moreover, the existence of cross-modal priming suggests that the representations are abstract enough to be shared between comprehension and production. This proposal seeks funding to investigate structural priming with first language learners aged 2;6-4, both within modality (from production to production) and across modalities (from comprehension to production). Results are expected to shed light on the organization of the child's developing language comprehension and language production systems and the relationship between them. Three hypotheses are tested: 1) the hypothesis that structural priming stimulates learning; 2) the hypothesis of continuity between child and adult language, specifically, that the processing systems of children and adults are organized in qualitatively similar ways; and 3) the hypothesis of shared representations, that a subset of the representations used in language production are shared with language comprehension.	{ "pile_set_name": "NIH ExPorter" }
Management of chronic wounds represent a major healthcare challenge responsible for over $15 billion in expenses annually. Wound surface sepsis, in particular biofilm formation, is a significant factor in nonhealing wounds. In chronic wounds, >60% have been observed to have clear evidence of a biofilm. Further, NIH has estimated that 65-80% of microbial infections in humans are biofilm-mediated. Biofilms are characterized by resistance to host defenses and to therapeutics that would otherwise have efficacy against the organisms planktonic state. Thus, biofilms present two distinct challenges: 1) the biofilm must be removed or dispersed to reduce bacterial defense mechanisms; and 2) effective antimicrobial agents applied to suppress the resident bacteria. In this application, we seek to address this problem by investigating a novel intervention with potential to prevent chronic colonization of wounds and disperse pre-existing biofilms. The innovation underlying our approach revolves around engineering the surfaces of wound beds to both promote the dissolution of biofilm bacteria back to a planktonic state, where they are more susceptible to antimicrobial agents, and immobilize antimicrobial agents at the wound surface where biofilm formation occurs. Our approach involves incorporation of absorbable nanobeads loaded with antibiofilm agents in a nanoscopic thin film, manufactured of polyelectrolyte multilayers containing silver nanoparticles, that is used to re-engineer the wound surface to increase its resistance to microbial colonization and biofilm formation. This structured engineered approach allows effective antimicrobial action with very low non-toxic concentrations of active agents. The innovative combination of these approaches in a single nanostructured film has the potential to markedly increase anti-biofilm and antimicrobial efficacy in vivo. In particular, use of nano-beads permits precise control over concentrations and release rates of the antibiofilm agent. They also penetrate and create microdomains within the biofilm, increasing surface contact by >80%, to increase adsorption of antibiofilm and antimicrobial agents concurrently and generating progressive dispersion-kill zones emanating from the beads at a nanoscale level. The central hypothesis of this study, supported by exciting preliminary data, is that incorporating select D- and L- amino acids into the wound bed will reduce bacterial biofilm formation and increase biofilm dissolution. A secondary hypothesis is that this approach will enhance the antimicrobial activity of silver nanoparticles immobilized at the wound surface. To address these goals we propose 3 Aims. In Aim 1, we will evaluate the ability of select D- and L- amino acids immobilized in polyelectrolyte thin films and loaded into absorbable PLGA beads to prevent a biofilm from forming and to stimulate dissolution of existing biofilms of Pseudomonas aeruginosa and Staphylococcus aureus in vitro . In Aim 2, we will optimize the integration of select amino acids and nano-beads onto model wound surfaces (full and partial thickness skin wounds), using polyelectrolyte thin film immobilization methods, and evaluate their efficacy in preventing biofilm formation and biofilm dissolution in vivo; and in Aim 3, we will test the hypothesis that combined application of silver nanoparticles and select amino acids further reduces biofilm formation and minimizes microbial bioburden in wounds in vivo. At the conclusion of this study, we expect to provide proof of concept that a two-armed approach to wound biofilms contained in an integrated nanoscale wound bed engineering platform will have increased efficacy against biofilms with reduced cytotoxicity in the wound bed. This approach is labile and generalizable to immobilization of other antibiofilm and antimicrobial agents. Thus, upon successful completion of this R21 application, we will seek support for research (via the R01 mechanism) that will broaden the scope of these investigations in optimizing these strategies and in evaluating the efficacy of an array of antimicrobial and antibiofilm agents with the goal of maximizing the ability to suppress wound bed sepsis and improve healing of chronic open wounds. PUBLIC HEALTH RELEVANCE: This application seeks support for an exploratory investigation of a conceptually novel approach to suppression of wound microbial bioburden and biofilm formation. Simple amino acids have been shown to be effective in preventing biofilms from forming and in promoting biofilm dissolution. These compounds, when integrated into a nanostructured wound surface engineering platform, offer a two-armed approach to dissolution of biofilms and killing of resident bacteria by their combination with a non-antibiotic antimicrobil such as silver in nanoparticulate form. The engineered immobilization platform allows use of low non-toxic concentrations of active agents. Such an approach provides a highly innovative attack of a major health problem without the side effects of cytotoxicity or microbial antibiotic resistance inherent to current conventional wound treatments. We seek to test this hypothesis using a wound engineering approach involving polyelectrolyte multilayers with absorbable nanobeads to deliver both antibiofilm and antimicrobial agents together at the wound surface. Approaches such as this that facilitate prevention of wound sepsis that do not impair wound healing would represent a major healthcare advance.	{ "pile_set_name": "NIH ExPorter" }
The overall objective of the proposed research is to enhance understanding of the relationship between risk and protective factors for binge drinking and alcohol use disorders in Mexican American young adults. This project will be an extension of our previous cross-sectional study to a longitudinal study in this population. Hispanics are the second largest ethnic minority group in the U.S. and the largest in the city of San Diego, yet biopsychosocial vulnerability factors for alcohol problems in this population remain relatively unexplored. We now have preliminary data that suggests a distinct cluster of risk factors may be associated with binge drinking and alcohol use disorders in Mexican American young adults. These associated factors include: age of onset of drinking, sleep quality, electrophysiological and behavioral measures of pre-pulse inhibition of the startle response, co-morbidity with other psychiatric disorders, measures of stress and acculturation, and distributions of alcohol metabolizing enzymes unique to this population. We propose to increase the participant population in order to have enough power to fully evaluate our findings. Additionally, we will be using our data to translate the findings to the development of valid animal models. In addition we will be testing, in humans, the overall theoretical construct of the grant related to stress, the development of binge drinking, and alcohol dependence in this population of Mexican Americans. This study has the potential to provide critical information for understanding how select genetic and environmental factors might interact in the development of alcohol use disorders among Mexican American men and women living in San Diego County.	{ "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. Cardiovascular Disease (CVD) and related disorders remain the leading cause of death in the nation1. Hypovitaminosis D has been linked not only to several cardiovascular (CV) risk factors including hypertension, diabetes, obesity2,3, but also to increased rates of CVD4,5,6. Thus, hypovitaminosis D presents a common pathway for a select subgroup with a clustering of CV risk factors in a profile that is predominant among ethnic minorities. Indeed, hypovitaminosis D is highly prevalent with an estimated 55% of the US adult population having levels at or below 30 ng/ml, and over 80% of African Americans having suboptimal values.7 A large body of evidence supports the premise that therapeutic agents that have salutary effects on vascular function, often translate into therapeutic benefits on 'harder'clinical endpoints such as adverse CV events and mortality. Exploring the potential vascular effects of vitamin D repletion presents a unique opportunity to examine both clinical outcomes and specific mechanistic pathways of a potentially modifiable intervention that could result in significant impact at a public health level. Thus, we propose a twelve week randomized doubleblind, placebo controlled pilot trial to assess the effect on vascular function and CV risk factors of 100,000 IU Vitamin D3 given every 4 weeks to overweight, hypertensive African-Americans with hypovitaminosis D. To our knowledge, the proposed project is the first to assess the effect of 'high-dose'Vitamin D3 administration on vascular function. We believe this study is also the first to examine the impact at a molecular level of Vitamin D3 repletion on the key mediators of cardio-metabolic pathways in humans. If our study results support our working hypothesis, we will be positioned to propose a larger scale study to detect a therapeutic effect on more definitive, clinical cardiovascular endpoints across a more diverse population. Hypothesis: Hypovitaminosis D activates select vasconstrictive and hypertrophic mediators that lead to endothelial dysfunction, elevated blood pressure and CVD;and Vitamin D3 repletion (using evidenced-based repletion strategies), in comparison to placebo, will ameliorate these abnormalities.	{ "pile_set_name": "NIH ExPorter" }
The long-term goal of this project is to determine the basic mechanisms by which signals generated through integrin receptors regulate chondrocyte function. The overall hypothesis driving this work is that changes in the cartilage extracellular matrix (ECM), including production of ECM protein fragments, are recognized by chondrocyte integrins and initiate a cascade of events intended to remodel the ECM but which in arthritis result in further matrix destruction (chondrocytic chondrolysis). The focus of this proposal is to determine the basic cellular mechanisms that control signals generated through the ?51 integrin which regulate production of catabolic mediators including cytokines and matrix metalloproteinases (MMPs). During the previous funding period, we have defined the signaling pathways that mediate MMP-13 production in response to fibronectin fragment (FN-f) stimulation of the ?51integrin and discovered reactive oxygen species (ROS) are necessary second messengers. Using an innovative proteomics approach we found that the MAP kinase family member JNK2 is oxidized in FN-f stimulated cells forming a Cys-SOH (sulfenic acid) intermediate. Sulfenic acid formation serves as a major mechanism by which ROS regulate cell signaling but its in chondrocyte signaling has not been investigated. In his competitive renewal, we propose to determine the mechanism by which sulfenic acid formation regulates JNK2 activity in chondrocytes. We will determine the role of JNK2 activation in OA in vivo by studying the development of surgically-induced OA in JNK2-/- mice. Finally, we will determine if HB-EGF, upregulated and released when chondrocytes are stimulated by FN-f, promotes Rac activity to activate a co-signaling pathway that augments MMP-13 expression and cartilage matrix destruction. These studies will have significant impact on the field by defining key hubs in a signaling network that mediates cartilage matrix destruction. By discovering novel mechanisms by which ROS regulate this signaling network, the information can be used to develop a unique approach to altering redox-regulated catabolic signaling networks in arthritis that targets specific protein modifications. This represents a significant advance over the general inhibition of ROS production which not proven successful in treating conditions promoted by excessive ROS, including arthritis.	{ "pile_set_name": "NIH ExPorter" }
To determine the safety and feasibility of deferoxamine and gallium nitrate in patients with hormone-refractory metastatic prostate cancer.	{ "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. SPECIFIC AIMS We plan to describe changes in lymphocyte populations which occur as a result of cardiothoracic surgery in childhood. Specifically we plan to measure: [unreadable] Whether thymus tissue is present or absent in individuals who have had cardiothoracic surgery in the past (for individuals who are having surgery or have had MRI or CT scans of the chest). [unreadable] Whether individuals have a change in frequency or type of infections after cardiothoracic surgery in childhood. [unreadable] Presence of symptoms of autoimmune disease and measure production of specific T cells which are related to autoimmune problems. [unreadable] Rate of T cell production, T cell function (in vitro), lymphocyte and T cell subset number, and antibodies to common viral illnesses and vaccines from blood samples. [unreadable] Response (T cell function in vitro and antibodies) to Hepatitis A vaccine before and after immunization. [unreadable] The above parameters in young children both before and then after cardiothoracic surgery so that we may follow the timing of the changes	{ "pile_set_name": "NIH ExPorter" }
Summary of work: Cockayne syndrome (CS) belongs to the category of premature aging disaeses where the individuals appear much older than their chronological age. Cells from CS patients are sensitive to UV light, exhibit a delay in recovery of DNA and RNA synthesis following irradiation, and are defective in preferential repair and strand-specific re-pair of active genes. Complementation studies demonstrate at least two genes involved in CS, designated CSA and CSB. CSB protein, by sequence comparison, belongs to the SNF2 family of proteins, which have roles in transcriptional regulation, chromosome stability and DNA repair. The cellular and molecular phenotype of CS include a significantly increased sensitivity to a number of DNA-damaging agents including UV irradiation. Studies in CS cells were initially confined to DNA repair in the general, overall genome, where no defect was found. However, CS cells are defective in the preferential repair of active genes and in the preferential repair of the transcribed strand of such genes. This defect in transcription coupled repair (TCR) in CS is not only found after UV exposure but also after exposure to certain forms of oxidative stress. Transfection of the CSB gene into hamster cells with the CS-B phenotype completely restores TCR and UV resistance to normal levels, demonstrating that the defect in TCR in CS-B is due to mutation in that gene. The complex clinical phenotype of CS, however, suggests that DNA repair may not be the primary defect. We have reported a defect in basal transcription in CS both in vivo and in vitro. This transcription defect is seen in CS-B lymphoblastoid cells and fibroblasts without any exposure to stress such as UV light. A previous study found that expression of a metalloprotease was reduced by 50% in CS cells, and recently it was reported that the purified CSB protein stimulates transcription, presumably as an elongation factor. We have used an in vitro assay to measure the incision event of the DNA repair process. During the first step of BER, there is an incision in DNA 5' to the lesion. The incision can be quantitated in cell extracts by using oligonucleotide duplexes that contain a single 8-oxoG lesion at a defined site. In primary CS-B cell lines we observe a deficiency in incision. This deficiency can be complemented by transfection of the CS1AN (CS-B) cell line with a plasmid containing the intact CSB gene, suggesting a role for CSB in the recognition of 8-oxoG. The CSB protein apparently functions at the crossroads of DNA repair and transcription. It has been reported to interact with the structure specific incision endonuclease XPG, CSA protein, and RNA polymerase II. It has considerable homology to the SWI/SNF complex, which in yeast is associated with RNA polymerase II. The SWI/SNF complex is involved in the initiation phase of the transcription process. We have also observed that CS-B cells appear to have a more open chromatin structure than normal cells, and this would be compatible with a function that involves a role in chromatin structural assembly. It would appear that the CSB protein has more than one function and is most likely involved in a large number of protein-protein interactions in transcription and repair pathways. One or more of these is likely to be very important for the assembly of the DNA repair and transcription factory at the nuclear matrix. A functional analysis of the CSB gene has been undertaken in our laboratory to better understand the nature of the molecular deficiencies observed in CS. Mutants, generated by site- directed mutagenesis have been tested for genetic complementation of CSB null cell lines by cell viability and RNA synthesis recovery upon exposure to UV light and other genotoxic agents. Point mutations in ATPase motifs I and II of CSB dramatically reduce CSB function in vivo suggesting that ATP hydrolysis by CSB protein is required for transcription-coupled repair of DNA damage. This mutant also shows dramatically increased apoptosis, suggesting a role for the CS protein in the apoptotic pathway. In contrast to the ATPase point mutations, deletions in the conserved acidic domain do not appear to interfere with the repair capacity of CSB protein. This suggests that this domain may be conserved in the SWI-SNF family for some other function. We have now made a series of point mutations in the helicase domain of the CSB gene in stably transfected human cell lines. We find that some of these are defective in the incision of plasmid with 8-oxoG, suggesting a defcet in base excision repair. We can conclude that the CSB protein is involved in the general genome base excision repair process, and that different domains of the helicase region play different roles in this process.	{ "pile_set_name": "NIH ExPorter" }
This project is concerned with the determination of protein conformation using the technique of X-ray crystallography. The major problem currently under investigation concerns the catalytic mechanism of the enzyme lactose synthase; a unique enzyme system formed by the interaction of a water-soluble milk protein, alpha-lactalbumin, with a membrane bound galactosyl transferase. The approach to be used involves determination of the tertiary structures of both alpha-lactalbumin and galactosyl transferase by X-ray crystallography. Significant homology in the amino acid sequences of alpha-lactalbumin and hen egg white lysozyme suggests that these two proteins have a common genetic origin. Determination of the X-ray crystal structure of alpha-lactalbumin would permit a comparison of the tertiary structures of these two proteins. This would provide an opportunity to reveal how, in one case, structural changes during evolution have given rise to a totally new biochemical function. Galactosyl transferase has been identified as a marker protein present in the serum of patients with certain types of malignant tumor. Structural studies of metabolic control of this enzyme at the molecular level could have implications for the advancement of cancer research. A second project is concerned with the structures of chlorophyll-containing proteins. The amino acid sequence of a bacteriochlorophyll-protein complex, with known tertiary structure is being determined in order to investigate the specific chemical interactions between the chlorophyll and the protein. Further studies of chlorophyll-protein interactions will involve determination of the tertiary structure of a water-soluble peridinin-chlorophyll-protein from marine dinoflagellates by X-ray crystallography. It is hoped that the results of this work will be useful in explaining the structural basis for the efficient transfer of energy between peridinin and chlorophyll in this complex. Preliminary crystallization studies on the enzymes, myeloperoxidase, AMP-deaminase and fumarse are also proposed.	{ "pile_set_name": "NIH ExPorter" }
The purpose of the proposed project is to examine the potential physiological role(s) for gamma-aminobutyric acid and substance P in regulation of excitability of physiologically and morphologically identified primary afferent fibers and interneurons on the superficial parts of the mammalian spinal dorsal horn (laminae I-III of Rexed). Conventional approach will be to use cat spinal cord in situ and apply GABA and substance P iontophoretically or by micropressure injections into the laminae I-III of the dorsal horn, while testing the electrical excitability of single sural A- and C-afferent fibers or performing intracellular recording from the large diameter primary afferents or functionally identified interneurons or dorsal root ganglion cells. Ionic mechanisms of presynaptic and postsynaptic actions of GABA and substance P will be investigated in vitro by using the rat spinal cord slice preparation. Morphological identification of physiologically and pharmacologically characterized dorsal horn interneurons and primary afferent neurons will be done by intracellular labeling with horseradish peroxidase.	{ "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 entails a detailed study of the interaction of the methyltransferases with the substrate DNA. Though the details of catalysis by DNA cytosine-5 methyltransferases are currently known in great detail, the conformational changes that happen before the chemistry takes place have not yet been described. The mechanism by which DNA cytosine-5 methyltransferases target a particular cytosine for extrusion and later methylation is unknown. Hence our goal is to obtain structures of early intermediates of M.HaeIII bound to DNA before the catalysis occurs.	{ "pile_set_name": "NIH ExPorter" }
New prophylactic and therapeutic materials are studied for the prevention and control of infectious diseases in volunteers and patients. Current studies include the evaluation of cellular and humoral immunity in individuals immunized with live rubeola and rubella vaccines.	{ "pile_set_name": "NIH ExPorter" }
The purpose of this study is to compare two different methods for securing endotracheal (ET) tubes: adhesive tape (Wet-Pruf [Trademark]) versus a tube fixation device. Within 4 hours of intubation adult patients requiring oral intubation will be randomly assigned to one of the two methods of securing ET tubes. The dependent variables, tube stability, skin integrity, patient comfort, nurse satisfaction, cost, and family fear associated with the appearance of their loved ones face will be evaluated at fixed intervals for up to 2 weeks. Data will be summarized with descriptive statistics, and multivariate data analysis techniques will be used to compare group differences. Due to slow accrual of subjects, the protocol was amended to allow data to be collected at the national Naval Medical Center. Summary of Findings: Data collection continues slowly and to date 23 subjects have been accrued. National Naval Medical Center has been added as a second data collec-tion site. Data collection continues.	{ "pile_set_name": "NIH ExPorter" }
Many vital organs, including the kidney, form as a result of a mesenchymal-epithelial interaction. A mediator of these interactions is hepatocyte growth factor, a. k. a. scatter factor, (HGF\SF). HGFISF is mesenchymally-derived growth factor and ligand for the epithelial c- rnet receptor. Though HGFISF is not the only inducer of renal development and tubulogenesis, it induces tubulogenesis both in vivo and in vitro ii Madin-Darby canine kidney (MDCK) cells, which are derived from canine renal tubular epithelium. Generation and maintenance of cell polarity is essential to epithelial cell function. To establish and maintain their polarity, epithelial, including MDCK, cells must send plasma membrane (PM) proteins to th( correct apical or basolateral PM. Preliminary results show that when HGF/SF induced tubulogenesis occurs MDCK cells lose their polarity as they branch out and then regain their polarity as the new tubules form. A major discovery in recent years is that almost all intracellular membrane traffic uses a common machinery for membrane fusion. Syntaxins are a gene family and component of the membrane fusion machinery that appear to specify correct delivery to the different PM surfaces. Syntaxins 2, 3, and 4 are abundant in epithelial organs, e.g. kidney. Syntaxins 2, 3, and 4 were found to be differentially expressed in MDCK cells, with syntaxins 3 and 4 having a completely non- overlapping distribution. Syntaxins are the first molecules that are part of the membrane fusion machinery, whose isoforms are differentially localized and whose overexpression has differential effects on polarized membrane traffic. Hence, they are the leading candidates for containing at least part of the information needed for the maintenance of cell polarity. The hypothesis is that transient loss of cell polarity i crucial for renal tubulogenesi: and syntaxins are involved in controlling cell poladty and therefore tubulogenesis. The expression and localization of syntaxins will be examined during HUF/SF induced tubulogenesis in MDCK cells, normal development of rodent kidneys, and abnormal development in disease states such a polycystic kidney disease and renal cell carcinoma. To directly test the hypothesis, the expression an function of different syntaxins will be perturbed by overexpression, inhibition of expression, and dominar negative mutants. If the expression of syntaxins is important for tubulogenesis, consideration can be given t finding ways to alter syntaxin expression in disease states, perhaps by gene therapy using viral vectors.	{ "pile_set_name": "NIH ExPorter" }
PROJECT SUMMARY Mr. Lance Murphy is an applicant for a diversity supplement under PA-18-586 for Dr. Clark Hung's NIH grant AR068133-A1 for an individual that falls under the designation of Baccalaureate and Master's Degree Holders. The Project Period: 05/16/2016 ? 04/30/2021. Mr. Murphy, who graduated on May 16, 2018 with his B.S. in Biomedical Engineering from Columbia University, is African American, a US citizen, and will be taking a 2- year gap year before attending medical school. As Mr. Murphy is currently planning a 2 year gap, we will be requesting support for 2 years and provide a validation metric (e.g., demonstration that the applicant has been making progress toward applying for medical school) with the progress report for year 1. The aforementioned has been discussed with Ms. Justine Buschman, NIAMS program official. As permitted in the PA guidelines, Mr. Murphy will be paid a salary equivalent to our graduate research assistants (GRAs), $39,133 plus fringe (30.3%) and inflated by 3% for year 2. Additionally, we will be requesting $3000 annually to support supplies and travel for Mr. Murphy, as allotted in the PA.	{ "pile_set_name": "NIH ExPorter" }
This program project application will enable Ordway investigators to further studies of a new paradigm for robustly determining doses and schedules of drugs to treat patients infected with the Bioterror pathogens Bacillus anthracis, Yersinia pestis and the Pox viruses Vaccinia Virus and Cowpox Virus. An interdisciplinary program project will facilitate concurrent investigation of this paradigm in several pathogens and enable investigators to learn more about the pharmacodynamics of drugs intended for therapy. Because of the seriousness of these illnesses, drug doses and schedules have to be right the first time. In addition to being able to set drug exposure targets employing a validated (in bacteria as well as in multiple HIV studies) in vitro hollow fiber infection model, the investigators have developed a robust approach employing Monte Carlo simulation for translating the insights from the in vitro system and animal models to man. Ordway has the expertise locally, with our New York State Health Department relationships, and in our US Army Medical Research Institute of Infectious Diseases (USAMRIID) collaborators as well as the infrastructure support to help solve this serious national problem. In order to correctly determine the right dose for man, there are four pieces of knowledge required. The first is a drug exposure target (ADC/MIC ratio, Peak/MIC ratio, Time > MIC, etc). The second is to have a measure of the variability of the pharmacokinetics of the agent in the population. The third is to have a measure of the distribution of MIC's for different strains of the pathogen of interest for the drug in question. The final piece of knowledge is to have a measure of the impact of protein binding on the microbiological activity of the drug. The program project application will develop this required knowledge through four integrated projects: (1) Determining therapeutic regimens for Bacillus anthracis in the hollow fiber system, (2) Determining therapeutic regimens for Yersinia pestis in the hollow fiber system, (3) Determining therapeutic regimens for the Variola major surrogates Vaccinia Virus and Cow Pox in the hollow fiber system and in an animal model, and (4) Examining the predictions of the hollow fiber model system in inhalational animal models of Anthrax and Plague. These projects will be supported by analytical, computational, and administrative cores to facilitate thematic integration. The Ordway investigators have already had significant success in addressing these issues. One example is the identification of an exposure to a fluoroquinolone antimicrobial that would achieve the desired goal of therapy for Bacillus anthracis. Indeed, the predictions from the hollow fiber system were evaluated in a Balb/c mouse model at USAMRIID, as well as in a rhesus monkey challenge and shown to be correct.	{ "pile_set_name": "NIH ExPorter" }
The aims of this core are to develop PCR-based assays to measure selected transcript concentration change in animals or tissues treated with botanicals, extract and store DNA from botanicals, identify variation in nucleotide sequences of botanicals, quantify DNA in botanical samples, and use "TaqMan" technology to devise method to identify impurities in the DNA extracts. Most of these aims will be accomplished using the new ABI TaqMan technology and an ABI Prism 7700 Sequence Detector and traditional sequence analysis.	{ "pile_set_name": "NIH ExPorter" }
A major focus of the Koch Institute is the development of new materials systems and biomedical devices for biomarker detection, imaging modalities, and targeted therapies and vaccines. Center Members are employing a wide range of materials for these goals, including synthetic polymers and copolymers, synthetic and naturally derived peptides and proteins, polysaccharides, inorganic and organic nanoparticles, and colloidal systems. It is essential that Center Members can conduct high resolution and surface imaging, and chemical and physical analyses of these materials. In the current period, a Nanotechnology Materials Core was established with the support of CCSG Developmental Funds and non-renewable support from MIT. The Core is located in custom designed space within the new Koch Institute building, and has a wide range of state-of-the-art instrumentation for imaging, physical, chemical and mechanical analyses, and an expert staff. In the upcoming period, the Nanotechnology Materials Core will continue to offer a wide range of state-of-the-art characterization capabilities that are essential to the Koch Institute mission. Thus, CCSG funds are requested to support the continued operation of the Nanotechnology Materials Core as an established Shared Resource.	{ "pile_set_name": "NIH ExPorter" }
Project Summary In response to RFA-MH-16-410, this R34 addresses key NIMH priorities, 1) focusing on health disparities, and 2) development and preliminary testing of an innovative services intervention. We propose to refine and test the efficacy of a brief meta-intervention for young adults when they begin treatment at adult mental health clinics (hence, treatment and meta-intervention run concurrently). The meta-intervention is designed to reduce treatment dropout, a documented problem of significant public health concern, by helping young adults put treatment into perspective, better understand the rationale and goals of care, and address barriers to continued treatment as they build attendance routines and relationships with providers. To date, research has focused on `transition interventions,' namely programs within children's systems that serve transitioning youth into young adulthood, some of which have extended services into the early twenties. These programs create temporary additional support for youth while they are still in the children's systems. Our research suggests a complementary strategy that outreaches to youth who have formed tentative and initial engagement in services in the adult system, but who are at high risk of disengaging due to the failure of services to take into account the unique developmental needs and orientations of young adults. Our team developed a mid-level theory to understand the underlying mechanisms of disengagement. The theory integrates theories of service use with theories of decision-making, and suggests a set of factors (individual and contextual) impact disengagement among young adults with serious mental illness. Our intervention uses novel communication strategies to maintain the attention of young adults and impact these factors and thus, overall engagement. We use narrative communication (e.g., strategic storytelling as a means of conveying health information) to keep young adults connected to care and, in turn, improve functioning. We use a recovery role model who is a person who has had similar difficulties that young adults have as they first engage the adult mental health system. S/he leads activities and is a key resource. Although the recovery role model is related to approaches of peer support, it is distinct from and appeals to different mechanisms. One hundred ninety five young adults (3 groups, 65 per group) will be randomly assigned to either the intervention group, an active control group (curriculum on safe relationships), or the treatment as usual control group when they first enroll in services. Assessments are taken at baseline, 2-week posttest, 4-week follow-up and at a three month follow-up. The study aims are (1) to develop and preliminarily test the efficacy of a novel, 2-session engagement meta-intervention aimed at engagement and functioning outcomes for marginalized young adults with serious mental illness, and (2) to preliminarily identify the underlying mechanisms of change for disengagement in mental health care, i.e., mediators and moderators of intervention effectiveness. We will advance theory as well as strengthen the proposed intervention.	{ "pile_set_name": "NIH ExPorter" }
New evidence indicates that distinct mutations cause familial Parkinson's disease (PD) by mechanisms that may also operate in sporadic PD. These new data point to the importance of cell dysfunction preceding cell death and to the involvement of non-dopaminergic neurons in the disease. Accordingly, identifying mechanisms of cellular dysfunction that are common to multiple causes of PD may offer new therapeutic targets to halt or reverse the course of the disease. This renewal application for the UCLA UDALL Parkinson Disease Center of Excellence focuses on studies of progression of dysfunction, in complementary models expressing PD-causing mutations, and in a well characterized patient population. The center consists of 5 projects supported by an administrative core and a mouse genetics core. In the first three projects we propose to continue coordinated multidisciplinary work supported by the current award to characterize the progression of motor and non-motor behavioral anomalies and neuropathology (project 1), anomalies of neurotransmitter release (project 2) and of synaptic function (project 3) in genetic mouse models of PD, including novel models based on BAG technology. These projects will be complemented by the addition of cellular models (project 4) to analyze the mechanisms of cellular dysfunction leading to the phenotypes observed in the mouse. Studying progression of dysfunction will also be the focus of the new patient oriented component of the Center. In this project (project 5), we will conduct clinical longitudinal studies of disease phenotype after diagnosis, including psychiatric and cognitive co-morbidities. This will be coupled to the development and validation of an improved health-related quality of life assessment tool. These patient oriented studies will provide crucial clinical data for future analyses of genetic material from the same patients and for the translation of our basic research efforts into improved patient care. To identify the cellular alterations leading to neurodegeneration in PD, the UCLA UDALL Parkinson Disease Center of Excellence will focus on early manifestations of the disease occurring before the onset of motor symptoms and their progression. Integrating experimental models and clinical studies, the goal of our center is to understand the mechanisms of these cellular dysfunctions in order to spur the development of therapeutic strategies able to stop the disease process. Project 1 Title: Progression of Behavioral and Pathologic Defects Preceding DA Cell Death in Mouse Models of PD PI: Marie-Francoise Chesselet, MD, PhD DESCRIPTION (provided by applicant): New evidence indicates that distinct mutations cause familial Parkinson's disease (PD) by mechanisms that may also operate in sporadic PD. During the current funding period we have identified cellular dysfunction without cell death in mice expressing various mutations known to cause PD in humans. In this renewal application we will test the hypothesis that common mechanisms of dysfunction may be induced by distinct mutations and offer therapeutic targets for treatment at early stages of the disease, before further cell death causes irreversible damage. Project 1. 2. and 3 form the continuation of the current award and will continue to use a multidisciplinary approach to uncover the mechanisms of neuronal dysfunction in existing and novel mouse models. In specific aim 1 of project 1 we will determine the progression of behavioral deficits and neuropathology in already available mice overexpressing alpha-synuclein under different promoters and parkin KO mice. We will use a battery of sensitive motor tests we have developed to assess the motor phenotype of the mice, and will extend this analysis to non-motor behaviors because related symptoms can have a major impact on patient quality of life, as examined in project 5 of the Center. We will use immunohistochemistry to determine the progression of alpha-synuclein pathology and glial activation throughout multiple brain regions known to be affected in PD, and to detect anomalies in the expression of proteins involved in neurotransmitter release and examined in project 4 of the Center. We will use ligand binding and molecular approaches to identify dysregulation of dopaminergic transmission that will be further examined with neurochemical approaches in project 2 and with electrophysiology in project 3. Finally, eventual cell loss in brain regions that are affected in PD (locus coeruleus, ventral medulla, nigrostriatal dopaminergic neurons) will be assessed in older animals with unbiased stereology. In specific aim 2, we will extend this analysis to novel mouse models generated in the Mouse Genetics Core with state of the art BAG technology. These include a mouse model expressing a parkin mutation shown to cause the loss of dopaminergic neurons in Drosophila. These mice will be characterized with the same methods described above, before being further analyzed in projects 2 and 3. Studies in project 1 will parallel longitudinal clinical studies of project 5 that examine disease progression in PD patients, including psychiatric and cognitive comorbidity. They will provide critical information on the time course of the deficits and new models for projects 2 and 3 and test in an in vivo mammalian system the hypotheses generated in cellular models in project 4 of the Center.	{ "pile_set_name": "NIH ExPorter" }
The single-gene reassortant (SGR) virus possessing a PB2 gene from an avian influenza A virus and the remaining genes from a human influenza A virus failed to produce plaques on mammalian cell monolayer culture and was restricted in replication in monkeys. Revertant viruses, which replicated and produced plaques efficiently on mammalian cells, possessed an amino acid substitution at residue 627 (glutamine in the avian PB2 gene -> lysine in the revertant) and replicated to moderate levels in monkeys. the amino acid at position 627 in every avian virus PB2 gene sequenced to date is glutamic acid and in very human virus gene it is lysine. Thus the amino acid at position 627 on the PB2 protein is an important determinant of host range of influenza virus as well as virulence of these viruses for primates. Gull-human influenza A virus reassortants derived from three different gull influenza A viruses were attenuated for squirrel monkeys or chimpanzees and, therefore, are candidate live attenuated viruses for use as vaccines in humans. Ten distinct amino acid substitutions in the human influenza A virus PB2 protein that each separately specify the temperature-sensitive phenotype were identified. Previous studies had demonstrated that influenza A viruses with ts mutations were attenuated in humans. Techniques are being developed to introduce one or more of these ts mutations into a synthetic copy of the PB2 gene and to rescue this synthetic gene into an infectious virus.	{ "pile_set_name": "NIH ExPorter" }
Obesity is a major health problem that predisposes people to diabetes, cancer, and heart disease. Despite considerable effort, the precise molecular mechanisms that regulate body weight are not well characterized. Neuropeptides are thought to play a critical role in regulating feeding, but these molecules are often missed by proteomics techniques due to their small size and low abundance. We have recently developed a method to isolate neuropeptide-processing intermediates from fat/fat mouse brain, and have used differential isotopic labeling to provide relative quantitation of peptides from two different animals (or pools of animals). Using this quantitative neuropeptidomics method, we have detected a large number of peptides in mouse hypothalamus, of which 20-30% are substantially up- or down-regulated by 2 days of food deprivation. In Aim 1, we will identify these peptides using tandem mass spectrometry. At the same time, in Aim 2 we will develop additional isotopic labels that allow for the quantitation of a larger number of peptides, and which provide more information from each experiment. Aim 3 will use either the new labels developed in Aim 2 (if successful) or the existing labels currently in use to examine additional paradigms of food deprivation and re-feeding. It is anticipated that these studies will lead to the identification of novel neuropeptides that are altered by food deprivation and/or re-feeding after fasting. In further studies, it would be important to test if these peptides are similarly regulated in wild type mice, and whether microinjection of the peptides influence feeding and/or body weight; these additional studies are beyond the scope of this R21 application, but would be continued in a subsequent R01 application. The studies in this initial R21 will provide an unbiased survey of neuropeptides that are altered by food deprivation, and which may therefore be involved in regulating food intake and/or body weight.	{ "pile_set_name": "NIH ExPorter" }
Excitatory synaptic transmission underlies all cognition, and changes in this transmission are thought to play a role in many neurological disorders. The fast component of excitatory transmission is mainly carried by AMPA-type glutamate receptors. These receptors are tetramers assembled in neuron- specific stoichiometries from four types of subunits (GluR1-4). It has recently been discovered that AMPA receptors in neurons are also associated with auxiliary subunits called TARPs (transmembrane AMPA-R regulatory proteins), a family of four proteins of which stargazin (STG) has been most widely characterized. STG has two distinct effects on AMPA receptors. One is to promote trafficking to the plasma membrane and to anchor AMPA receptors to the synapse via PSD-95. The other one is to modulate the kinetics of AMPA receptors by lowering the EC50 for glutamate and by slowing receptor deactivation and desensitization, effects which have immediate impact on the waveform of synaptic transmission. However, most studies have been carried out by coexpressing STG with GluR1 and 2, and much less is known about STG's effects on other receptor subunits and about the actions of the other three TARPs which are far more prevalent in cortex and hippocampus. We have recently discovered that the receptor subunit GluR3, unlike GluR1 and GluR2, exhibits almost no kinetic modulation by STG. The TARPs gamma-4 and gamma-8, on the other hand, caused a slowing of deactivation and desensitization in GluR3 that was larger than in GluR1 and 2. Interestingly, surface expression of GluR3 was strongly promoted by STG, which indicates (i) that STG did associate with GluR3 and (ii) that modulation of receptor kinetics and trafficking are independently regulated. In this project we will determine how and why the four TARPs differ in their effect on receptor subunits. In Specific Aim 1, we will complete our analysis of their impact on GluR3 and extend our analysis to include GluR2 and GluR4. Moreover, we will examine the effects of the four TARPs on heteromeric subunit combinations that are prevalent in brain. These data will have immediate relevance for understanding the waveform of synaptic transmission in different brain areas. In Specific Aim 2 we will then determine which amino acids in GluR3 are responsible for the differential effects of the TARPs. For this we will systemically exchange segments between GluR2 and GluR3, starting with the S1-S2 domain thought to be central, and then use point mutations to identify the critical amino acids. These experiments should allow us to identify the contact points between AMPA receptor subunits and TARPs that are important for kinetic modulation. Recent evidence suggests that both GluR and TARP expression is altered in disorders such as schizophrenia. Thus, the results of this study will be relevant also to understand aberrations in excitatory transmission that are thought to underlie many neurological and psychiatric disorders. Much evidence suggests that disturbances in excitatory synaptic transmission contribute to the pathogenesis of various neurological and psychiatric disorders. Such disturbances in synaptic transmission may arise from changes in AMPA-type glutamate receptors, but recent studies have found that proteins associated with AMPA receptors may similarly contribute to pathology. For instance, proteins called TARPs which modulate AMPA receptor function have been shown to be altered in the brains of schizophrenic patients. In this project we will study the impact of TARPs on different types of AMPA receptors. Given that AMPA receptors vary across neurons, the results from these studies are expected to offer new explanations as to why synaptic transmission varies in different parts of the brain, and they may also provide new insights about aberrations in excitatory transmission in various brain disorders. [unreadable] [unreadable]	{ "pile_set_name": "NIH ExPorter" }
A central characteristic of alcoholism and other addictive behaviors is the chronically relapsing nature. Despite continuing care's potential for reducing relapse, its implementation is not wide spread due in part to cost, geographic distance, lack of time and peer support. This proposal builds upon the premise that these barriers can be reduced with computer based support systems. We are the national program office for the Network for Improvement of Addiction Treatment (NIATx) and developers of the Comprehensive Health Enhancement Support System (CHESS), a non-commercial eHealth system shown to improve quality of life and produce behavior change. We propose to adapt CHESS for alcohol-dependency relapse prevention and test it in a randomized clinical trial. Addiction CHESS (ACHESS) will be delivered through a new modality (smart-phone) to broaden access, focus on new patients (alcohol dependent patients leaving residential care), offer optional audio delivery to provide access to the functionally illiterate and be enhanced with services tailored to relapse prevention: 1) communication with peer support groups and addiction experts; 2) monitoring of relapse risks, 3) alerts and reminders to encourage adherence to therapeutic goals; 4) addiction-related educational material and tools tailored to individual patients and 5) one-touch communication with a care manager. Research hypotheses include: Primary Outcome. ACHESS + standard care (vs standard care control group) will reduce days of risky drinking over a 12-month period. Secondary Outcomes. ACHESS will reduce proximal outcomes of cravings, negative affect, withdrawal symptoms and temptation events, and produce distal outcomes of increased total abstinence, days of abstinence and reduced negative consequences of substance use. Mechanism of effect. We hypothesize a mediational model where ACHESS improves competence, relatedness and autonomy, which will improve outcomes. We will recruit a final sample of 280 alcohol dependent patients (30% minorities) from two NIATx addiction treatment agencies. Data sources will be interviews at pretest, 3, 6, 9 and 12 months, chart reviews, lab tests and automatic collection of computer use data. This is a resubmission of 1R01 AA01792-01.	{ "pile_set_name": "NIH ExPorter" }
Malignant melanoma has one of the most rapidly increasing incidence rates in the U.S. Early detection and surgical excision can be curative, but once the tumor spreads beyond the skin it is one of the most deadly forms of cancer. There are currently no completely effective therapies for advanced (metastatic) disease and 10 year survival rates for these patients are very low. The objectives of this proposal are to conduct phase I clinical trials of a new gene therapy approach for the treatment of metastatic melanoma, to monitor for tumor responses attributable to treatment, and develop and characterize in vitro assays which may enable us to identify which patients are most likely to respond to this form of combined immuno-gene therapy. The therapeutic genes selected for use in the clinical trial are the genes for human granulocyte/macrophage colony stimulating factor (GM-CSF) and the gene for the superantigen staphylococcus enterotoxin B (SEB). In on-going preclinical trials conducted by one of the coinvestigators of this proposal, in a private veterinary oncology clinic, this combination of therapeutic genes has been shown to be more effective at inducing clinically significant tumor immunity than either gene used alone and more effective than other combinations of genes tested. The method used for gene transfection, polycationic lipid mediated gene transfection, has been tested extensively in the preclinical trials and found to be safe and effective. The phase I clinical trial is a dose escalation study designed to determine the safety and potential toxicities associated with the direct combination DNA injections of cutaneous melanoma metastases. Although the primary goal of the trial is to assess safety and toxicity, careful clinical analyses of treated and untreated tumors will allow the determination of whether the proposed treatments have an effect on local and distant metastases. In addition, in vitro immunologic assays which have proven to be indicative of development of protective tumor immunity in preclinical studies will be conducted concurrently with the clinical evaluations to determine if they have predictive value in identifying those patients which will benefit from this treatment approach. An improved understanding of how this treatment leads to the elimination of tumor tolerance and the development of immunity may enable us design more effective strategies for the use of these treatments in the clinical setting.	{ "pile_set_name": "NIH ExPorter" }
The project proposes a program of studies in speech perception and auditory psychophysics to examine the hypothesis that many of the predominant difficulties in speech understanding for elderly listeners are related to underlying problems in auditory temporal processing. Of the many forms of degraded speech observed to be difficult for elderly listeners, those that involve an alteration of speech rate appear to produce the largest age-related recognition deficits. Alterations of speech rate may be viewed as a form of degradation in temporal aspects of speech segmental cues and speech prosody, and these types of distortions are the focus of investigation in the next project period. Moreover, psychoacoustic results demonstrate that large age-related difficulties in temporal processing exist for the perception of auditory tempo and rhythmic characteristics of sequential stimulus patterns. Listener processing difficulty could be attributed to peripheral and/or central processing mechanisms, as well as various cognitive factors, including the degree of familiarity with prosodic features of different native languages. The project examines the relative contribution of peripheral hearing impairment, type of stimulus temporal complexity and cognitive demand on the processing of temporal segmental and prosodic cues in speech and non-speech stimulus patterns. The project comprises four stages of investigation, each consisting of a series of speech and non-speech discrimination and recognition tasks. Stages 1 and 2 investigate the sources of age-related differences in temporal sensitivity for discriminationand identification of speech and non-speech segments presented as isolated target stimuli or as embedded in sequential stimulus patterns. Stage 3 measures the influences of listener age on discrimination of temporal cues for prosody that characterize different languages. Stage 4 will examine psychoacoustic models of temporal processing and identify stimulus parameters contributing to age-related differences in understanding accented English. Participants in the project will include groups of subjects that differ by age and hearing sensitivity.	{ "pile_set_name": "NIH ExPorter" }
This K08 Career Development Award (CDA) application details the plan for development of an independent clinician-investigator skilled in the application of developmental genetics tools in the area of sarcoma biology. The candidate has recently completed rigorous clinical training in orthopaedic surgery and musculoskeletal oncology, during which aspirations toward science have been on hold. The candidate has nonetheless demonstrated sustained desire for involvement in science beginning during secondary education and culminating in the generation of a unique conditional knock-out mouse in his spare time in the midst of surgical training. Now beginning an academic appointment as a surgeon-scientist in a busy clinical center for pediatric sarcoma treatment and under the research mentorship of a recent Nobel Laureate with extensive experience in the study of pediatric sarcoma mouse models, the candidate is poised to receive intensive preparation for an investigative academic career. After gaining expertise in the reverse translation of hypotheses generated from patient-oriented research to developmental genetic models in mice, it is anticipated that the candidate's career work will not only provide insight into the pathogenesis of pediatric sarcomas-specifically osteosarcoma-and improvements in their treatment, but also expand the currently tiny pool of exemplary models for future would- be orthopaedic surgeon-scientists. Osteosarcoma is a leading cause of cancer death among adolescents and young adults. Further, it has one of the youngest ages of onset among genetically complex non-balanced-translocation-type cancers, making it likely to have patterned pathogenesis and likely to provide a useful window into the understanding of genetically complex cancers in general. A major challenge in the study of the pathogenesis of osteosarcoma is that no benign, low-grade, or even intermediate-grade precursor lesions are recognized. Observations in human tumors and mouse models have elevated p53 and the retinoblastoma susceptibility protein (pRb) as critical gene disruptions for osteosarcomagenesis. Conditional disruption of these genes at varied osteoblast precursor differentiation states and varied mouse host developmental stages will characterize the window of opportunity for osteosarcomagenesis. Transposon-mediated mutagenesis in association with pRb and p53 conditional disruption will identify the cooperative gene disruptions and amplifications needed to complete osteosarcomagenesis and attain the metastatic phenotype. PUBLIC HEALTH RELEVANCE: The development of model orthopaedic surgeon-scientists is crucial to the future attraction of those with scientific aspirations to the field of orthopaedics, which includes care for some of the most frequent, most disabling, and least understood problems in medicine. Improved understanding of osteosarcoma is needed as this remains an unacceptably deadly and disabling cancer in a young population. Further, osteosarcoma is a model for other genetically-complex cancers, many of which are major public health burdens.	{ "pile_set_name": "NIH ExPorter" }
The functions of potassium channels are central to the properties of all excitable mammalian cells. Electrophysiological and pharmacokinetic studies have defined many different classes of potassium channels. However, the molecular basis of this diversity, and the relationships between channel structure and function remain unknown. Through the use of molecular cloning and electrophysiology, we have isolated, characterized, and expressed in frog oocytes a mammalian potassium channel from rat hippocampus. Surprisingly, the properties of this channel do not fit any single kind of channel as defined by classical methods. The major goal of the research proposed here is to further our understanding of the molecular basis of variations among the different classes of mammalian potassium channels. First, we will clone, characterize, and express in frog oocytes mammalian potassium channels, both voltage and ligand gated, with distinct structural and functional properties. The different kinds of channels will be compared to those known by classical methods. Second, structure-function studies, using site directed mutagenesis, will be performed using a range of potassium channels with distinct properties. Site directed mutant studies of whole, reconstructed channels will be conducted in oocytes. Third, ancillary factors which may influence channel functions will be investigated. Fourth, subunit-specific probes will be generated, based on established nucleotide sequences, and employed in in situ hybridization studies to map the expression patterns of potassium channels in the brain. By studying the structure and function of mammalian potassium channels in this way, we hope to answer questions regarding the molecular basis of potassium channel diversity and the significance of this diversity to the basic properties of excitable mammalian cells.	{ "pile_set_name": "NIH ExPorter" }
Our long-term goal for the VNED is to create a cadre of mental health researchers studying suicide prevention who are members of, or interested in working with, diverse populations. To do so requires attracting early career investigators with these backgrounds and interests, and providing them with the mentorship, knowledge, and skills needed for successful academic careers. Our first objective is to engage members of diverse populations in the scientific enterprise by supporting their study of a major public health problem of relevance to their communities, such as suicide. Our second is to explore innovative ways of drawing people into virtual and face-to-face activities that will encourage and support mentoring, research collaborations and training experiences. The most cost-effective and efficient way to provide nurturing, consultation and support is through the use of virtual technologies to supplement more traditional, face-to- face (F2F) encounters;that is, to create a virtual network. The specific aims of this proposal are (1) to recruit a critical mass of suicide prevention research teams consisting of mentors and proteges;(2) to create an Academic Development Plan (ADP) for each protege that reflects expected participation and contributions by all the team members;(3) to provide opportunities for experiential learning through activities designed to disseminate content on the current state of suicide research in diverse populations;and (4) to evaluate the effectiveness of the network to recruit, retain and educate its members and thereby develop a robust supportive virtual community of suicide researchers that represent the heterogeneity of the US population. Measurable outcomes are to produce two webinar series, for network members to submit a manuscript outlining lessons learned with regards to suicide research with diverse populations, for each protege to construct a well thought out research proposal outline and an individualized ADP, and for the proteges and mentors to participate in a F2F focus group session. This resulting mentoring network will provide the infrastructure to facilitate learning and engagement with geographically-dispersed suicide research scientists who represent the diversity in the US population. PUBLIC HEALTH RELEVANCE: Through specific virtual and face-to-face interactive programs, career development opportunities, and tailored mentoring VNED proteges from diverse backgrounds will develop skills and knowledge necessary for establishing a career in the study of a major public health concern: suicide in diverse populations.	{ "pile_set_name": "NIH ExPorter" }
Improved quantitative planar and single photon emission tomographic (SPECT) imaging techniques permitting more accurate assessment of regional myocardial perfusion and viability in patients with coronary artery disease are needed. The objectives of this research proposal are, (1) to acquire further knowledge of myocardial transport kinetics of thallium-210 (Tl-210) and technetium-99m methoxy isobutyl isonitrile (MIBI) under normal conditions, during transient or sustained ischemia, after total coronary occlusion and after coronary reperfusion in both animals and humans, and (2) to develop and test more optimum imaging methods for quantitation of in vivo risk area and distinguishing viable from irreversibly injured myocardium. In the experimental protocols, the following hypotheses are to be tested: (1) quantitation of Tl-201 uptake and redistribution at rest will be more accurate for identifying viable but underperfused myocardium distal to a coronary stenosis, than MIBI uptake; (2) MIBI uptake patterns will underestimate degree of myocardial salvage during experimental infarction when administered after reperfusion in the presence of a severe residual stenosis; (3) MIBI given too early after reperfusion will reflect reperfusion flow more than viability resulting in overestimation of degree of myocardial salvage; and, (4) risk area in the distribution of an occluded coronary artery in intact animals can accurately be quantitated by SPECT MIBI imaging, and dual simultaneous imaging of MIBI risk area and Tl- 201 redistribution should enhance detection of viability. In animal experiments, regional flow is measured by radioactive microspheres, in vivo risk areas by MIBI autoradiography and anatomic risk area infarct size by histochemical techniques. Parallel clinical research protocols are designed: (1) to differentiate ischemia from scar during exercise scintigraphy using split dose injections of MIBI 2 hrs apart; (2) to develop dual simultaneous imaging techniques with Tl-201 and MIBI to better quantitate in vivo risk area and provide more reliable detection of viable but jeopardized myocardium; (3) to develop quantitative SPECT methods for MIBI imaging to critically compare with similar planar techniques for detecting and determining extent of physiologically important CAD; (4) to improve detection of myocardial viability with Tl-201 by quantitative serial assessment of early exchange kinetics and to compare this approach with 24-hr delayed redistribution imaging; and, (5) to determine the value of simultaneous quantitation of regional wall motion and systolic thickening on ECG-gated MIBI perfusion studies using Principal Components Analysis. It is expected that this research will lead to improved applications of Tl- 201 and MIBI imaging for noninvasive detection of ischemia and assessment of myocardial viability.	{ "pile_set_name": "NIH ExPorter" }
The Administrative Core (Core A) will provide the organizational management to assure that the program project meets its objectives effectively. The specific aims are: Aim 1. To establish an organizational structure that assures effective scientific progress. A major goal of the Administrative Core is to establish an organizational structure that allows effective scientific progress. A system will be established to promote synergy amongst investigators and that resources are appropriately allocated. In addition, a mechanism will be established to allow ample communication amongst the scientific team members and with External Scientific Advisory Panel and NIH program staff. Aim 2. To establish and maintain an infrastructure to support the fiscal monitoring and reporting requirements of the project. A critical role of the Administrative Core will be to provide strong financial and administrative management of the program. This goal will be accomplished through the efforts of an outstanding administrative team comprised of a Program Manager, a Financial Administrator, and an Administrative Assistant. Aim 3. To establish and maintain a system to promote coordination of communication, scientific interchange and collaboration. A major goal of the Administrative Core is to enhance scientific interchange and collaboration among investigators with the goal of rapidly developing new vaccine delivery platforms that meet scientific objectives of novelty and the practical objectives of efficacy, and evaluating B cell immune responses in detail against vaccine candidates. To ensure sharing of resources and research output, the Core will be responsible for creating and maintaining a consortium website as well as scheduling regular meetings of the participants.	{ "pile_set_name": "NIH ExPorter" }
There is a concern by some who use chlorine in chemical production facilities that there is an occupational hazard of inhaled chlorine. The purpose of this project is to determine the distribution of inhaled CL2 in healthy men and women during oral and nasal breathingat respiratory flows characteristic of rest and exercise. The specific aims are: Develop a bolus inhalation method to safely and indirectly measure Cl2 distribution in the respiratory tract. This method will be tested on 3-5 subjects who are breathing quietly through their nose. Determine the effect of different inhalation concentrations from 0.3 to 3.0 parts per million on the Cl2 distribution in a group of five subjects who are breathing quietly through their nose. Determine the change in Cl2 distribution during oral breathing when respiratory flow increases from a resting level to a level characteristic of light exercise in a group of ten subjects who are breathing through their nose. (The subjects will not be exercising) Determine whether a change from nasal to oral breathing increases the fraction of inhaled Cl2 that reaches the lower airways in a group of ten subjects who are breathing quietly.	{ "pile_set_name": "NIH ExPorter" }
The purpose of this proposal is to identify critical idiotopes in anticentromere (ACA) and antitopoisomerase I (antitopo I) from SSc absent in RD that may predict which patients will develop SSc. In addition, in long range it would be possible to target these idiotopes and control the expression of SSc. The hypothesis of this grant is that ACA and antitopo I Ids from SSc differ from the RD Ids. This proposal plans to 1) determine whether the SSc antitopo I and ACA utilize preferential Ig gene families with or without somatic mutations 2) determine whether RD antitopo and ACA are the product of randomly used genes, 3) locate a critical idiotope that defines the binding of antitopo Id to its anti-Id and to antigen, and 4) identify the role played by this idiotope in the idiotypic network dysregulation in mice. In order to do this, ACA and antitopo I secreting B cell clones from SSc and RD will be established and used to determine their immunoglobulin heavy chain variable (VH) region genes. The specific SSc-VH sequence will be expressed in baculovirus to determine the % inhibition of binding of polyclonal SSc autoantibodies to antigen and anti-Id compared to RD autoantibodies. The sequences of the framework (FR) and complementary determining regions (CDRs) from RD will be exchanged for the corresponding SSc-VH sequences. This RD-modified VH will be used as control to identify an idiotope critical to the antigen and anti-Id binding specific for the SSc autoantibodies. Mice will be immunized with a) the recombinant SSc-VH or b) the recombinant RD-modified VH, and emergence of autoantibodies or SSc manifestations will be studied. The proposed work will be done at the University of Connecticut Health Center, Division of Rheumatic Diseases where all the facilities and equipment necessary for this work are available, as well as advice from senior immunologists and rheumatologists who are part of the Division.	{ "pile_set_name": "NIH ExPorter" }
Any closed continuous plane curve can be expressed as an isomorphic transform which relates all points to a unique central description called the symmetric axis. Algorithms have been developed and implemented which permit a digital computer to reversibly transform data derived from cephalometric radiographs into such a description. When applied to the average shape of mandibles extracted from the Broadbent-Bolton series, axis points produce a characteristic shape which appears to be independent of age. Analysis of higher order relationships exhibited by the symmetric axis functions provides quantitative bases for characterizing the curvature of boundaries and segmentation of shape into mathematically tractable elements.	{ "pile_set_name": "NIH ExPorter" }
Immunologic and physiologic parameters of isolated pancreatic islet transplantation will be investigated in several directions. First, the mechanism of islet allograft rejection will be elucidated by determining the relative roles of cell-mediated vs. humorally-mediated factors in immunologic destruction of islet grafts. In conjunction, several methods directed at either altering islet immunogenicity or host immune mechanisms will be studied in an attempt to prolong islet allograft survival. Whole organ pancreatic allografts will also be compared to isolated islets with regard to the ability of these various regimens to prolong survival. Second, islets in intrasplenic and intraportal (intrahepatic) implantation sites will be compared with regard to: 1) differences in islet allograft survival in each site using standardized immunosuppressive protocols; 2) physiologic differences in the function of islets and their ability to maintain normal glucose homeostasis in each site; and 3) their effects on the inappropriate hyperglucagonemia of experimental diabetes. All of the foregoing will be performed using the inbred rat models. Third, short term preservation requirements for pancreatic fragment grafts will be studied using a large animal model (dog). Using various media and culture techniques, an attempt will be made to extend survival of these grafts ex vivo to 48-72 hours followed by successful function after transplantation. Metabolic studies will be performed on recipients of these preserved grafts.	{ "pile_set_name": "NIH ExPorter" }
The overall goal of this proposal is to further define the relationhips between dietary carbohydrate and lipid, glucose and glucoregulatory hormone concentrations in normal subjects and patients with insulin-dependent diabetes. Four distinct, but interrelated research areas are included in the nine specific questions to be addressed: 1. Plasma glucose response to dietary carbohydrate, 2. Plasma lipid response to dietary carbohydrate, 3. Relationship between glucose control and 24 hour plasma triglyceride patterns in insulin-dependent and 4. Relationship between glucose variation or stability and growth hormone integrated concentrations in insulin-dependent diabetics. The questions will be answered in a series of seven diet studies which are based on similar studies conducted in our laboratory over the last three years. The diets in each study will vary only in the source or amount of dietary carbohydrate to prevent confounding results from multiple simultaneous changes. The diets will be alternated in a Latin Square sequence to allow estimation of residual influence, period effect, and the effect of subject differences on observed results. Dietary influences on glucose, lipid and ingestion of the test diet by the continuous withdrawal of blood. Lipid responses will be further evaluated by lipid fractionation by ultracentrifugation of samples collected following a fast and at two and four hours post-prandially. Our overall hypothesis, based on eariler results in control subjects, is that appropriate dietary changes in insulin-dependent diabetics will result in improved glucose control, lower 24 hour triglyceride concentrations, and a reduction in the mean 24 hour integrated concentrations of growth hormone.	{ "pile_set_name": "NIH ExPorter" }
Amajorissueinschizophrenia(SZ)istobaccosmoking,whichleadstolungcancer,poorqualityoflife, andnegativehealthoutcomes.Theadenosinesystemrepresentsanovelpharmacologicaltreatmenttarget, becauseadenosinemaycontributetobothpsychosisandsubstanceabusecomorbidityinSZ.AdenosineA2A receptorsformaheteromericcomplexwithdopamineD2receptorsinthebrainthatismutuallyinhibitory. AdenosineA2AagonistsreduceD2receptorsensitivity,whereasD2agonistsreduceA2Areceptorsensitivity. Thisisimportant,becauseincreasedD2receptorsensitivityunderliespsychosisinSZ.Thisapplicationcenters aroundarodentmodelofSZofD2sensitivity,withratsneonatallytreatedwiththedopamineD2/D3agonist quinpirole(neonatalquinpiroletreatment,NQ).NQtreatmentresultsinincreasedD2receptorsensitivity throughouttheanimal?slifetimewithoutalteringD2density,consistentwithSZ.Insum,preliminarydata revealedthatNQenhancedtherewardingassociativeaspectsofnicotineaswellasthebrain-derived neurotrophicfactor(BDNF)andglialcell-linederivedneurotrophicfactor(GDNF)responsetonicotine.These responseswerereducedbyadenosineA2AagonistCGS21680aswellastheantipsychoticclozapine.In addition,NQtreatmenthasbeenshowntoresultinauditorysensorimotorgatingdeficits,abehavioralhallmark ofSZ.TheoverallhypothesisisthatNQ-inducedenhancementofthebehavioralresponsetonicotineand auditorysensorimotorgatingdeficitscanbemitigatedbyadenosineA2Aactivationincombinationwith clozapineandchangesinBDNFandGDNFsignalingcascades.Aim1willanalyzepotentialsynergistic effectsoftheA2AreceptoragonistCGS21680andclozapineonnicotineCPPandauditory sensorimotorgatinginadolescentNQ-treatedmaleandfemalerats.Aim1awillthentestthehypothesis thatCGS21680combinedwithclozapinewillmosteffectivelyreduceNQ-inducedenhancementofnicotine CPP.Aim1bwilltestthehypothesisthatCGS21680alonewillreduce,andincombinationwithclozapinewill alleviate,sensorimotorgatingdeficitsproducedbyNQ,showingthatadenosineA2Areceptoractivationis effectivetotreatbothnicotineabuseandpsychosis-likesymptoms.Aim2willinvestigatewhetherCGS 21680willreduceenhanceddopamineandglutamatereleaseintheNAccinadolescentNQ-treated maleandfemalerats.WehypothesizethatA2Areceptoractivationviareversemicrodialysisinfusionintothe brainwillreduceenhanceddopamineandglutamatereleaseinresponsetonicotineinNQ-treatedratstested inadolescence.Aim3willanalyzecommondownstreamneurobiologicaltargetsofBDNFandGDNFin NQ-treatedanimalsadministeredCGS21680inadolescence.WehypothesizethatA2Areceptoractivation combinedwithclozapinewillmitigatethedownstreamresponseofproteinscommontoboththeBDNFand GDNFsignalingcascadesinthenucleusaccumbensandhippocampus,brainareasthatareimplicatedindrug addictionandpsychosisrespectively.	{ "pile_set_name": "NIH ExPorter" }
Abstract. This Conference Grant application is to request funding to support three annual conferences aimed at sharing state of the art scientific research and best professional practices to address the mental health disparities of racial/ethnic minorities and immigrants. The successive themes for the three years are (a) addressing the intersections between race and culture in scientific research and mental health service delivery for children, adolescents, and families; and (b) examination of the intersection of culture, race, and trauma and mental health across the life span; and (c) racial identity and cultural factors in treatment and service delivery (Each conference builds upon the recommendations of Dr. David Satcher's supplemental report, Mental Health: Culture, Race, and Ethnicity; the President's New Freedom Commission on Mental Health Report: Achieving the Promise: Transforming Mental Health Care in America; the mission of NIMH; as well as the longstanding objectives of the Institute for the Study and Promotion of Race and Culture (ISPRC) at Boston College. In an effort to reduce mental health disparities among People of Color relative to non-minorities, each conference involves sharing innovative, pragmatic, and empirically based strategies for delineating and addressing the mental health burdens of ethnic and linguistic minorities. The primary common goals and objectives of each of the conferences are to: (1) Share state of the art mental health theory, research, and practice with respect to the roles of race and culture in the etiology, maintenance, and remediation of mental health disparities; (2) Use strategies to engage scientist- practitioners and mental health service providers in interdisciplinary communication and collaboration to address the role of culture and race in their research, practice, and service delivery. (3) Develop and disseminate information regarding race and culture and mental health disparities. Speakers for the conference will be nationally and internationally acknowledged researchers and clinicians presenting in their areas of expertise via interactive formats. Each of the thematic 2-day conferences will take place at the Diversity Challenge conference at Boston College in successive years, October of 2007, 2008, and 2009. Each thematic conference will be characterized by varied formats consisting of plenary sessions, individual presentations, concurrent workgroups, roundtable discussions, and informational poster and exhibit sessions intended to achieve the stated aims. Proceedings from the conference will be disseminated through published monographs, and on the ISPRC web site. Thematic books based on each conference will be co-edited by the co-PIs. The conference grant addresses mental health disparities of People of Color in the United States. This focus is consistent with the Surgeon General's Supplemental Report on Race, Culture, and Mental Health, the President's New Freedom Commission report, and the mission of NIMH, which identified these populations as being at significant public health risk and called for interventions to eliminate the health disparities. [unreadable] [unreadable] [unreadable]	{ "pile_set_name": "NIH ExPorter" }
Proteases of the endosomal/lysosomal system play important roles in a number of physiological and pathological processes, including the immune response, arthritis, bone remodeling and tumor growth. Although the enzymes are primarily lysosomal, they have been detected in endosomes, on the plasmamembrane and in extracellular fluids. The origin and activity of the non- lysosomal forms of the enzymes are not clear. The long-term objectives of this project are to develop procedures that enable the active enzymes to be identified so that they can be selectively inhibited to control their activity in pathological and physiological processes. A new method has been developed using an active-site directed, covalently binding inhibitor to target intracellular compartments. This technique has been used in one cell culture system to show that active proteases are found in both endosomes and lysosomes, but are not secreted or membrane bound. A new series of reagents will be developed to enable evaluation of the unversality of this observation. The specific aims are: 1. To determine whether newly synthesized lysosomal cysteine proteases are preferentially delivered to, and activated in endosomes. Biosynthetic pulse-chase experiments will be used in concert with a transferrin-conjugated covalently reacting peptidyl inhibitor to identify newly synthesized proteases. 2. To determine whether delivery of endocytosed proteins to protease-positive compartments occurs similarly in different cell types. A range of cell types will be examined using a protein conjugated inhibitor to identify active proteases in cells. 3. To determine whether molecules taken up by different endocytic processes are delivered to similar protease- positive compartments. A conjugation system will be developed to permit inhibitors and targeting proteins to be interchanged using biotin-avidin chemistry. These studies will show which molecular forms of the lysosomal proteases are capable of degrading endocytosed proteins, enabling the generation of more specific reagents to regulate endosomal proteolysis.	{ "pile_set_name": "NIH ExPorter" }
This application represents the second competitive renewal of the CALGB institutional grant submission from the University of North Carolina at Chapel Hill (UNC-CH) and its affiliates. UNC-CH has been a member of CALGB since 1986. Our accrual has been maintained at approximately 40 treatment cases (including 6-10 minority accruals/year) plus companion studies. We also maintain an affiliate network responsible for another 60-90 accruals/year while maintaining excellent data quality control. Moreover, our institution now has broad leadership participation in the Group in Breast, Cancer Prevention and Control, GI, Lymphoma, Molecular Pathology, Radiation Oncology, and is the research base for a UNC based Minority Initiative Program and a VCU based Minority CCOP. Moreover, our leadership efforts in the Group have also continued to expand. We now enjoy the distinction of having one committee chair, five committee vice-chairs, seven committee or working group core or liaison members, eight modality committee members, one Executive Committee member, two members of the Board of Directors, five administrative committee members and seven protocol principal investigators. Members from UNC and its affiliates have held 56 positions on various scientific, administrative, and leadership committees of CALGB during the past 5 years. Additional faculty at UNC-CH will expand our commitment and Group leadership in the Breast, Surgery, GU, GI, Lymphoma and PET committees (F. Detterbeck, M. Graham, C. Sartor, L. Carey, J. Mohler, R. Pruthi, B. O'Neil, R. Orlowski, P. Watkins, and C. Dees). We therefore believe that this application reflects the growing and crucial contribution that UNC-CH investigators provide to the Group in leadership, science and service as well as our continued efforts to increase accrual through affiliate members while maintaining rigorous control of quality data submission. We respectfully submit that this application demonstrates both the current value of this institution to the Group and the contributions that: the membership of UNC-CH and its affiliates will continue to provide CALGB over the next five years.	{ "pile_set_name": "NIH ExPorter" }
Our research focuses on natural compounds that possess chemotherapeutic and/or chemopreventive properties with respect to cancer. In the proposed study, we will characterize the mechanism of action of the compound Withaferin-A (WA), a bioactive compound of Withania somnifera, which is extensively used in Asian and African traditional medicine. WA displays impressive and selective activity against androgen- dependent and androgen-independent prostate cancer (ADPC and AIPC, respectively), of which the latter is refractory to all current forms of treatment. Our preliminary in vitro and in vivo data indicate WA targets ADPC and AIPC cells by inhibiting the activity of Akt, a protein kinase that activates cell survival pathways. Our preliminary data demonstrate that, concomitant to Akt inhibition in ADPC and AIPC cells, WA activates FOXO3a, which in turn activates prostate apoptosis response-4 (Par-4), a protein that selectively induces apoptosis in cancer cells Based on our results, we hypothesize WA overrides the negative effects of Akt on CREB/Par-4 signaling to achieve chemopreventive and/or chemotherapeutic effects on AIPC. To address this hypothesis, we propose the following aims: investigate the interaction between Akt/CREB and Par-4 signaling, (Aim 1); and how WA modulates the pro-apoptotic signaling in AIPC (Aim 2); evaluate the effects of WA on androgen- -independent prostate tumor growth in the transgenic adenocarcinoma of mouse prostate (TRAMP) model (Aim 3); and examine the mechanisms of WA-mediated tumor inhibition in TRAMP and/or TRAMP cell lines (Aim 4). For the in vitro arm of our studies we will use a variety of approaches (immunoprecipitation, siRNA strategies, immunofluorescence, Western blot, Northern blots, Chip assays, siRNA strategies, pull down assays, pharmacological blocking) in order to determine the molecular junction at which Akt/CREB 3aand Par-4 converge. For our in vivo studies, we will conduct tumor regression studies, yet we will also examine histopathology, cell death, expression patterns of pro-survival proteins and pro-apoptotic proteins in the tumor tissues, and WA levels in the serum and prostate tissue. Our long term goal is to promote natural compounds to a clinical environment, where these agents can be thoroughly assessed for their chemopreventive and chemotherapeutic properties. Our preliminary data indicate WA is a potential candidate for such clinical evaluation, and our proposed studies will elucidate at a mechanistic level its potency against prostate cancer, both from a chemopreventive and chemotherapeutic standpoint. PUBLIC HEALTH RELEVANCE: Prostate cancer (PCa) remains a major cause of cancer-related death, and at advanced stages current therapy is ineffective. Intervention with dietary compounds for the prevention and/or treatment of prostate cancer is a relatively new area of research focused on signal transduction pathways, which has resulted in the development of important new therapeutics for specific malignancies. The proposed study of dietary supplement Withaferin-A (WA), the major bioactive compound from Withania somnifera), is designed to provide essential insights at the molecular level into the mechanism of action of Withaferin-A against prostate cancer and may ultimately lead to clinical evaluation of its potential for chemoprevention and/ or chemotherapy in prostate cancer.	{ "pile_set_name": "NIH ExPorter" }
DESCRIPTION: (Applicant's Abstract) Parkinson's disease is a progressive debilitating neurodegenerative disorder for which there is no cure. The objective of this research project is to determine if neural grafting can improve the fixed neurological deficits of parkinsonism. The MPTP Parkinson-like syndrome is an excellent model to test surgical techniques for neural transplantation. Two major difficulties with application of neural transplantation are the poor graft survival (5-10 percent in most series) and the relatively poor degree of host reinnervation due to axonal migration of only 1-2 mm. from the graft. We have been able to demonstrate considerably higher survival rates and neuronal transformation when adrenal tissue was co-grafted with Schwann cells which presumably supplied the necessary growth factor(s). We propose to study the use of growth factors to determine whether fetal dopaminergic neurons can be promoted to more successfully survive and more robustly reinnervate the host striatum to produce functional recovery. These experiments should indirectly address the controversy that following transplantation, improvement in parkinsonian symptoms in the MPTP model is due to the reinnervation of the host. Motor activity will be evaluated by clinical examination, computer image analysis of spontaneous caged activity, drug-induced rotation, and performance on learned visual motor fore-limb tasks. The use of standard anatomical assessment, as well as Positron Emission Tomography assessment, should allow an investigation of (a) whether the extent of improvement is dependent upon the degree of dopaminergic reinnervation of the host, (b) whether the dopaminergic reinnervation changes with time paralleling motor improvement, and (c) whether dopaminergic reinnervation can be enhanced through the use of growth factors. Anatomical evaluations will allow not only determination of the survival of dopaminergc neurons, but also the fate of GABAergic neurons which also are concentrated in the ventral mesencephalon. Using a technique we are developing, all grafted cells can be identified. One of the most important questions are how well the noninvasive tests parallel the anatomical studies and correlate correlate with the degree of behavioral improvement. If the severe neurological symptoms of parkinsonism can be improved in this nonhuman primate model, the implications ar extremely important not only for ongoing transplantation clinical trials for parkinsonism, but also suggest possible hope for repair and restoration of neurological deficits following disease or injury.	{ "pile_set_name": "NIH ExPorter" }
Serotonin1A (5-HT1A) receptors mediate diverse signaling cascades via different G-proteins that are involved in various physiological functions and play a role in the etiology and/or treatment of mood disorders. Mood disorders are associated with increased activation of the HPA axis that is normalized by antidepressants. In neuroendocrine neurons in the adult hypothalamus, fluoxetine (Prozac(R)) and other serotonin-selective reuptake inhibitors (SSRIs) desensitize 5-HT1A activation of plasma hormones such as ACTH and oxytocin. The long-term objective of this proposal is to develop better treatment strategies by understanding the mechanisms invoked by various classes of agonists in mediating 5-HT receptor signaling of multiple intracellular pathways upon acute versus long-term repetitive drug administration. Because the clinical effectiveness of various drugs including the SSRIs is associated with adaptive changes in 5-HT1A receptor signaling, it is critical to understand the mechanisms by which 5-HT1A receptors mediate signaling in neuroendocrine neurons in response to the acute and chronic exposure to different classes of 5-HT agonists. Based on our initial findings that 5-HT1A receptors can couple to different populations of G proteins to activate both MAP kinase and hormone responses via pathways that act independently upon acute receptor activation but exhibit "cross-talk" resulting in desensitization of 5-HT1A mediated neuroendocrine response upon repetitive drug administration, we HYPOTHESIZE: MAP kinase activation plays an integral role in the neuroadaptive changes in 5-HT1A receptor-mediated hormone signaling in oxytocin and CRF containing neuroendocrine neurons in response to SSRIs and SNRI drugs. This will be tested by the four aims of this project;Aim 1 will determine effectiveness of different classes of agonists to traffick 5-HT1A receptors to Gai/o proteins and activate hypothalamic MAP kinase and/or Gaz-proteins to stimulate hormone responses;Aim 2;will determine the specific Ga-protein subtypes that mediate 5-HT1A receptor agonist activation of MAP kinase in the hypothalamic PVN and the localization of 5-HT1A activated MAP kinase (pERK) to oxytocin- and CRF-positive neurons;Aim 3 will determine the mechanisms by which serotonin/norepinephrine reuptake inhibitors (SNRIs) desensitize 5-HT1A receptor signaling of hormone responses in oxytocin and CRF-containing neuroendocrine neurons and the requirement of MAP kinase in mediating SNRI-induced desensitization, and Aim 4 will determine the efficacy of repetitive administration of different classes of serotonergic agonists to desensitize hypothalamic 5-HT1A receptor mediated MAP kinase signaling. These studies will provide important new information regarding the novel mechanisms of 5-HT1A receptor signaling pathways in hypothalamic neurons and the mechanisms by which clinically used antidepressant drugs may produce adaptive changes in signaling pathways mediated by different G proteins. These studies will elucidate role of MAP kinase in regulating the responsiveness of neuroendocrine neurons to various classes of drugs used clinically. This information is critical to identifying the mechanisms that may contribute to delays in the onset of clinical efficacy or the "side effects" of SSRIs, SNRIs or other drugs developed to treat various psychopathologies and mood disorders involving 5-HT1A receptors.	{ "pile_set_name": "NIH ExPorter" }
Validation of transformative MALDI-MS-based inhibitor screening technologies for cancer targets. Mass spectrometry (MS) has a long history as a transformative technology. Two specific examples include quantitative MS to measure the fate of drug compounds in biological systems and the development of proteomics techniques for protein identification and characterization. Recently MS-based applications have been demonstrated to be highly effective as the readout for high throughput screening (HTS) assays. The major advantage reported over the commonly used fluorescence and chemi-luminescence readout is the paucity of false positive or false negative readout. Other added benefits include a greatly reduced (>80%) reagent cost by using label-free substrates and the ability to multiplex assays such that multiple therapeutic targets can be screened for inhibitor hits with one pass through the compound repository thus saving millions of dollars in reagent and personnel costs. We have developed MALDI-MS based readout methods for measuring enzyme activity and inhibition for a variety of enzyme classes. However, the limited use of these methods with small test libraries has been insufficient to validate the overall utility of this readout for large screening campaigns. Thus the primary goal of this proposal is to systematically validate the reliability of the MALDI-MS readout compared to a traditional method of HTS using a library of 50,000 compounds. The measurements will include hit rates from the primary screen, the validation rate of the hits in secondary screening and other standard quality assessments common for HTS including signal to background, coefficient of variance (CVs) and Z'values. The target enzymes for these comparative assays will include two related kinases, PKC-zeta and PKC-iota, which appear to regulate two directly opposite effects on cancer initiation and development. Thus as a secondary aim of this proposal, the MALDI-MS readout will be assessed for its effectiveness to distinguish inhibitors from the compound repository that have selectivity for PKC-iota over PKC-zeta. We have shown that MALDI-MS readout is amenable to enzyme assays and inhibitor screening on a small scale with major advantages over existing readout methods. If, the MALDI-MS readout can be scaled to true HTS levels while maintaining all the advantages seen in the proof-of-concept studies, then this MS-based technology would likely transform the way we approach HTS, in much the same way as MS-based technologies have changed bioanalytical and proteomics applications over the past 15-20 years. Furthermore, by targeting kinases (a key class of regulatory enzymes whose dysregulation is often associated with cancer development) to validate the MALDI-MS readout approach, we can be assured that with the success of these studies, it will be clear that this technology can be readily reapplied to other cancer relevant kinases as well as expanded into other enzyme classes and disease categories. PUBLIC HEALTH RELEVANCE: Dysregulation of cellular function is a hallmark of cancer development and progression and can often be traced to the actions of one or a few cellular enzymes, thus these enzymes may act as primary targets to halt cancer development and progression. As such, rapid and accurate methods to evaluated inhibitors of these target enzymes represent an initial phase in the new drug discovery and development process. In this proposal, we investigate a new readout technology, based on mass spectrometry, to rapidly screen for inhibitors of a class of enzymes known to be involved in cancer development and progression.	{ "pile_set_name": "NIH ExPorter" }
In vivo wall shear rate in the thoracic aorta of dogs is measured with a flat edged hot film sensor mounted flush with the inner vessel wall. Probe position with respect to the vessel wall is monitored with a pulse echo ultrasonic system. The pulse echo probe is alligned directly opposite the hot film probe to measure relative hot film probe and vessel wall positions. A secondary hot film probe is placed downstream of the primary probe in close proximity to measure blood temperature in the thermal boundary layer established by the primary probe. Reverse flow components are then monitored by the secondary probe. A complete description of blood velocity near the vessel wall, including possible reverse flow is critical to an accurate quantization of wall shear stress. Wall shear stress is computed from shear rate data utilizing a Newtonian model for blood. The relation of in vivo peak and mean wall shear stress values to reported levels enhancing transendothelial lipo-protein flux will aid in the determination of the role of hemodynamic factors in atherogenisis.	{ "pile_set_name": "NIH ExPorter" }
The research theme of the Indiana Alcohol Research Center (IARC) is Genetic Determinants of Alcohol Ingestion. We hypothesize that between- individual variability in response to the reinforcing/aversive actions of ethanol, in development of tolerance and dependence, and in ethanol and acetaldehyde metabolism underlies much of the individual differences in ethanol ingestive behavior and susceptibility to alcoholism. Furthermore, much of this variability is genetic in origin. The IARC has unique resources to pursue this kind of research. A team of investigators with broad-based expertise ranging from molecular genetics and cellular neurobiology to physiological and behavioral studies in experimental animals and humans will address the central theme of the Center. Research for the 5 core components, 4 research components and a pilot projects component will include: the association of genetic polymorphisms of human alcohol and aldehyde dehydrogenases to alcoholism and its complications; the heritability, sensitivity and repeatability of a variety of responses to ethanol (e.g., electroencephalogram, event related potential, neuropeptide secretion, stance stability, voice pattern, eye movement and various other physiological and cognitive measures) in mono- and dizygotic twins; the value of such responses as predictors of subsequent abusive drinking behavior; the functional and structural loci in brain that are influenced by ethanol in a heritable manner; the study of the neurotransmitter systems implicated in abnormal alcohol-seeking behavior in rats; the influence of specific genes (e.g., alcohol dehydrogenase) on responses to ethanol in transgenic mice; and selective breeding of mice for high and low alcohol preference. Core laboratory functions will include the production of selective bred high and low alcohol-preferring rats for research, development of polymorphic markers associated with candidate genes for study in experimental animals and humans, and support of research into the structure-function relationships of molecules important to alcohol metabolism and the actions of ethanol. The pilot projects component will promote new research directions and recruit new investigators.	{ "pile_set_name": "NIH ExPorter" }
The grant is designed to uncover new secondary metabolites from higher plants which have novel chemical structures and which have potential for clinical use as antibiotics in 1 the treatment of investions caused by microorganisms against which current fermentation based antibiotics are unsatisfactory in some significant way. Isolation, structural characterization biological evaluation and synthesis of novel agents is involved. The diseases under attack by these methods include tuberculosis, infections due to Gram-negative bacteria -- particularly those carrying R-factors--and due to fungi.	{ "pile_set_name": "NIH ExPorter" }

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