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This study is designed to further investigations of adaptive changes in skeletal and osseous tissue associated with alterations in muscle length. Alterations in muscle length occur normally during growth but may also be induced in the craniofacial regions by modifications of the intermaxillary space and by orthognathic surgical procedures. Our primary objective is to broaden our understanding of the structure and function of the muscles of the craniofacial regions and to determine the adaptations that occur in the craniofacial musculature and the associated hard tissue resulting from alterations in muscle length. The emphasis in this proposed continuation period is in skeletal and muscular alterations following decreases in muscle length. Several experimental models will be used including partial and complete removal of the dentition. Adaptations will be studied by cephalometric, electromyographic, histochemical, biochemical and physiological techniques that we have used previously in studies of craniofacial adaptations. Data and interpretations gained from this study will be combined with data gained from our previous experimental investigations, leading to a better understanding of adaptations in the complex musculature of the craniofacial region.
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{
"pile_set_name": "NIH ExPorter"
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This project has focussed on studying the ability of human immunodeficiency virus (HIV) to infect various human and mouse cell lines. The CD4 gene which encodes an important receptor for HIV infection was expressed in human and mouse cells and susceptibility of these cells to HIV infection was followed using a sensitive focal immunoassay (FIA) technique. The results indicated that HIV infection in certain human cells (HeLa) was related to the level of CD4 expression in different clones. However, in other human lines such as U87 astroglioma and SCL1 squamous cell carcinoma HIV infection was only rarely successful in spite of very high CD4 expression. Further studies indicated that HIV could bind to the CD4 on these cells, but HIV did not undergo successful fusion and entry of the cells. These results suggested that other cellular molecules in addition to CD4 were required for HIV entry of cells. This possibility was confirmed by showing that HIV could infect these human cells when the usual HIV entry mechanisms were bypassed by using viral DNA to tranfect the cells or by infecting with HIV genomes pseudotyped by packaging in envelope proteins of mouse retroviruses which use receptors other than CD4 for entry. Similar experiments were also carried out using mouse cell lines expressing human CD4. In these mouse cells, there were at least two levels of resistance to HIV infection. The first was a block in entry similar to U87 and SCL1 human cells, but even when this block was bypassed by transfection or viral pseudotyping, mouse cells still had a markedly reduced level of HIV infection. Present data suggests that the reduced HIV expression in transfected mouse cells may be due to poor functioning of HIV promotor and enhancer sequences in these mouse cell cultures.
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{
"pile_set_name": "NIH ExPorter"
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We propose continuation of a basic investigation using gel microdroplets (GMDs; e.g. 10 to 100 mu agarose particles) to analyze biochemical activity in very small volumes (e.g. 10-10 to 10-6 ml). We seek rapid: (1) determination of activity of individual enzyme molecules, cells (or microcolonies), (2) sorting of GMDs with cells of unusual activity, and (3) determination of growth of individual cells (or microcolonies) by using flow cytometry with either aqueous or non-aqueous fluidics. The GMDs are created rapidly and statistically occupied, such that an individual GMD has a high probability of initially containing 0 or 1 active entity (enzyme molecule or cell), and a low probability of initially containing 2 or more entities (a multiply occupied GMD). By providing suitable nutrients and reagents for a extracellular biochemical assay, fluorescent product is accumulated within individual GMDs. Alternatively, initial cells grow to form microcolonies of two or more cells, which are fluorescence labeled in proportion to the amount of specific cellular constituents. Many biochemical systems of importance to fundamental biological and medical problems appear amenable to this method. We propose continued investigation of the basic properties of this small volume method, and its use in the study of several fundamental problems. An important problem in enzymology relates to the mechanisms of changes in enzymatic activity, particularly inactivation. By measuring the activity of a large number (e.g. 104) of individual enzyme molecules, it will be possible to readily study whether inactivation is an "all or nothing" or a gradual process for individual enzyme molecules, using several different enzymes. Microanalysis within the small volumes of GMDs is also applicable to biochemical activity measurements of individual microorganisms and mammalian cells, and is applicable to partially automated strain improvement through rapid sorting of GMDs with individual cells or microcolonies which have unusual activity. by rapidly obtaining many individual measurements of the biochemical activity within GMDs, it is possible to readily compute a population distribution based on specific biochemical activity. Such a distribution can form the basis for population studies, or for screening for sorting cells for specific biochemical activities. Rapid determinations of microbial activities and growth based on individual cells (or microcolonies) are potentially applicable to important clinical tests and environmental health problems.
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{
"pile_set_name": "NIH ExPorter"
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The overall objectives of this project are to perform in vitro and in vivo studies in dogs with spontaneously occurring malignancies in order to design improved immunotherapeutic approaches to the treatment of tumors. More specifically, the proposed work will include: a) Development of a treatment protocol for dogs with lymphoma involving chemotherapy, total body irradiation and adoptive immunothrapy using allogenic cells, b) Studies in dogs with naturally occurring osteogeneic sarcoma involving treatment with BCG, c) Detection of tumor associated antigens in dogs with lymphoma or osteogeneic sarcoma using serological and cell mediated techniques and, d) Determination of platelet and fibrinogen survivals in dogs with lymphoma or solid tumors and their modification by drugs which inhibit platelet function.
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{
"pile_set_name": "NIH ExPorter"
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The Mxi1 protein (and other members of the Mad/Mxi1 family) functions as a Myc antagonist and a growth/tumor suppressor, in part by effecting transcriptional repression of key target genes. This repression has been shown to occur through Mxi1's recruitment (via its amino terminal domain) of the Sin3 proteins and their associated transcriptional repressors including histone deacetylase. Our preliminary data and a recent report suggest that Mxi1 may also bring about a different mode of repression via its carboxyl terminal domain. The objective of this proposal is to pursue this new function assigned to Mxi1's carboxyl terminal domain by defining associated factors that mediate its activities as well as gene targets that are subject to its repression. To accomplish this objective, we will first focus upon novel interactions that we have identified between the Mxi1 carboxyl terminus and (i) members of the RYBP/Yaf-2 adapter protein family and (ii) members of the Polycomb group of transcriptional repressors. Since the RYBP (and Yaf-2) adapter(s) also can recruit Polycomb group proteins, we hypothesize that Mxi1 may be a potential targeting factor for this type of transcriptional repressor that functions in multi-protein complexes to generate repressive chromatin structures. Here we will continue to characterize and expand upon these interactions and will assess their functional ramifications with respect to known properties of Mxi1. A priority will be to ascertain whether these proteins indeed mediate the transcriptional repression activity assigned to the Mxi1 carboxyl terminus to which they bind. Concomitantly, the basis for Mxi1 's harboring two distinct repression domains/mechanisms will be probed through the identification and classification of genes repressed by each or both of these domains. Taken together, our findings will forge further links between factors involved in transcriptional regulation and factors involved in the genesis/suppression of neoplastic disease. Moreover, a better understanding of the molecular functions of the Myc antagonists (e.g., Mxi1) may suggest new therapeutic approaches for the plethora of cancer types in which MYC is disregulated.
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{
"pile_set_name": "NIH ExPorter"
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This Core will support the six research projects of the P01 by providing data and computing resources. Because of the use of common data sets across the projects, the Core can achieve substantial economies of scale. The Core will pool data management and programming tasks where there is overlap. This is particularly cost effective at RAND because of the very substantial investment RAND has made in the development of the HRS data. Working in coordination with project by Gustman, the Data Core will assist the other projects in the selection and use of pension data, particularly making use of the HRS Pension Calculator. Resulting data products that are of interest to the broad research community will be siddeminated. Moreover, the Core will provide a platform for efficient communication and information sharing among project members. This will involve an Internet forum for discussion and posting of information. It will collect, update, and disseminate a library of contextual data relevant to research on aging. A number of the projects in this P01 use restricted data, and the Core will supply expertise in forming and administering data protection plans. Many of the researchers in the P01 are active in primary data collection as well as in analysis. The Data Core will facilitate the flow of information from data collectors to researchers and from researchers back to data collectors. It will assist project by Kapteyn in the collection of experimental subjective data. Overall the Core will contribute to maintaining an efficient, cost-effective computing environment.
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{
"pile_set_name": "NIH ExPorter"
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We are using a range of MS techniques to characterize natural and synthetic b-amyloid peptides associated with Alzheimer's disease.
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{
"pile_set_name": "NIH ExPorter"
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The continuing objective of the research proposed in this renewal application remains the elucidation of the biochemical mechanisms that lead to irreversible cell injury in ischemia. Based on the progress reported below during the first 4 years of support from this grant, a working hypothesis of the sequence of events mediating the loss of cell viability in, at least, liver ischemia has been derived. The specific aims of the studies proposed here are 1) to further assess the validity of this hypothesis and 2) to evaluate its relevance to the mechanisms of ischemic cell injury in general and to ischemic myocardial cell injury in particular. The first aim will be addressed by consideration of a) the role of the release of intracellular, sequestered calcium stores in the genesis of membrane injury; b) the respective roles of an accelerated deacylation versus inhibited reacylation in the net loss of phospholipid; c) the relationship between such a mechanism for lipid depletion and the redistribution of intracellular Ca++ ion stores; d) the further analysis of the consequences of phospholipid loss with respect to the structure of cellular membranes; and 3) the changes in the structure and function of the plasma membrane. Two established experimental models are to be used: 1) in situ ischemia of the intact rat liver and 2) the anoxic death of cultures rat hepatocytes. The second aim will concentrate initially on the sarcolemma of ischemic myocardium with particular attention to the role of phospholipid degradation and consequent changes in membrane structure and function.
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{
"pile_set_name": "NIH ExPorter"
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Obesity and type 2 diabetes are two of the major causes of morbidity and mortality in the Western world. Obese patients have a high incidence of type 2 diabetes and manifest both hyperinsulinemia and hyperleptinemia. Receptors for insulin and IGF-I and for leptin are expressed in the pancreatic islets, and recent reports have suggested that leptin can act through some components of the insulin-signaling cascade in different tissues. Although studies have examined the interactions between insulin and leptin in muscle and hepatocytes, the role(s) of these hormones in modulating islet function is not fully explored. We and others have shown that genetically engineered defects in insulin/IGF-I signaling, at the level of the beta-cell, can contribute to alterations in glucose homeostasis. Thus, mice with beta-cell specific knockouts of the insulin or IGF-1 receptors manifest defects in glucose-stimulated insulin secretion and develop glucose intolerance, but surprisingly do not show alterations in the growth and development of the beta-cell/islets. In contrast, IRS-2 knockouts show insulin resistance and beta-cell hypoplasia leading to early onset diabetes. On the other hand, leptin has been demonstrated to suppress insulin secretion both in vitro and in vivo in mice and humans and to promote beta-cell growth in insulinoma cells. The broad goals of this proposal are to identify the cellular mechanisms by which the proteins transmitting the insulin/IGF-I signal, interface with the leptin signaling pathway, in regulating insulin secretion, hormone biosynthesis and beta-cell proliferation, to understand the cross talk between two relatively novel signaling pathways in the islets, and to understand the cause(s) of (-cell failure in type 2 diabetes in the context of obesity. The specific aims of this proposal are to: 1) investigate the ability of leptin to activate proteins in the insulin/IGF-I signaling pathway in beta-cell lines and islets, 2) examine the effects of leptin on islet hormone secretion, islet metabolism and alterations in Ca++ flux in islets and 13-cell lines isolated from mice lacking the insulin or IGF-I receptors or IRS proteins, and 3) study the effects of leptin on beta-cell proliferation. My past experience in investigating the effects of leptin action in islet physiology and the experience I have gained in cellular signaling techniques by interacting with Dr. Ron Kahn's group, as I move to independence, provides me with a unique perspective to explore the interactions between the leptin and insulin/IGF-I signaling pathways in the islets. The additional funding for studies, during the last two years of the KO8 Award, will be useful to perform pilot experiments to obtain preliminary data to apply for a RO1 application and will allow the applicant to gain independence from the mentor's field.
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{
"pile_set_name": "NIH ExPorter"
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Age-related macular degeneration (AMD) is the most common cause of acquired visual impairment in people over the age 60. AMD is a multifactorial, complex disorder associated both with environmental and genetic factors. All currently available treatment options, i.e., photodynamic therapy, laser photocoagulation, anti-VEGF compounds, are directed towards a temporary relief of some of the symptoms, mainly neovascularization. Despite concerted efforts over the last several years, the general knowledge of genetic determinants of AMD has not advanced substantially. The underlying hypothesis of this proposal is that increased susceptibility to AMD in individual cases results from a combination of subtle defects in many genes, i.e., from specific genotype(s). This proposal suggests a continuation of our current program directed towards deciphering the genetic cause of AMD by a combination of several approaches. These include: 1) Completing large, clinically and genetically well-characterized, cohorts of AMD patients and rigorously matched controls until reaching 2000 samples each; 2) Utilizing high-throughput screening methods, including recently introduced in our laboratory genotyping microarrays, to obtain data on genetic heterogeneity in these populations; 3) Correlating the large numbers (>4 million) of derived genotypes with specific (endo-)phenotypes in AMD by statistical analyses. Identification of genes, alleles, haplotypes, and genotypes underlying the AMD complex trait and understanding how these defects contribute to the development of macular degeneration has the potential to improve the quality of life of the affected individuals. Furthermore, it will enable the accurate identification of at-risk individuals before they develop the disorder, and has the potential to modify or prevent the devastating visual consequences of this disorder in future generations.
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{
"pile_set_name": "NIH ExPorter"
}
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Clozapine is a unique, atypical antipsychotic dibenzoxazebine with unusual efficacy in treating patients with refractory schizophrenia. Randomized studies have shown it to be superior to conventional treatment of this disorder. The use of clozapine, however, has been limited by a 1.6% incidence of life-threatening agranulocytosis. Many times patients who develop agranulocytosis have had meaningful responses and once withdrawn from clozapine experience dramatic psychiatric relapses. Its use in the United States requires weekly blood monitoring, because there exists no predictive tests for the development of agranulocytosis. This proposal will evaluate potential mechanisms of clozapine induced agranulocytosis in patients with schizophrenia. Both toxic and immunologic mechanisms will be explored. Specific emphasis will be placed on the role of clozapine metabolites both as agents which directly suppress the bone marrow and as immunogenic compounds. Due to the prolonged duration of agranulocytosis and the severe marrow suppression there may be either a direct toxic or immune hematopoietic precursors. The specific aims for this small grant proposal are: 1. To determine whether clozapine or its metabolites are cytotoxic to normal human bone precursors. 2. To search for a cytotoxic antibody in the serum which attacks hematopoietic precursors and identify whether the antibody is dependent on clozapine, a metabolite or complement. 3. To measure the toxicity of the proposed toxic metabolite and/or antibody against hematopoietic precursors taken from affected patients after recovery from clozapine-induced agranulocytosis. 4. To measure plasma levels of potential toxic metabolites to determine whether high levels are associated with bone marrow suppression. This data may help unravel the mechanism of clozapine induced agranulocytosis, may identify a predictive laboratory test to identify high risk patients, and may allow the drug to be more successfully administered to patients with schizophrenia.
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{
"pile_set_name": "NIH ExPorter"
}
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Flavonoids are major components of dietary supplements and herbal medicines with reported protective activities against microbial infections in humans. However, the precise mechanisms by which these plant derived natural products attenuate microbial infections are unclear. Using a high-throughput screen for type III protein secretion in Gram-negative bacterial pathogens, we discovered specific flavonoids from medicinal plants that do not interfere with bacterial growth can effectively antagonize this key bacterial virulence pathway and prevent Salmonella typhimurium invasion of host cells. To determine how specific flavonoids can antagonize type III secretion systems (T3SSs) and attenuate bacterial virulence, this application will 1) develop more potent flavonoid analogs, 2) determine flavonoid T3SS inhibitor mechanism of action and 3) analyze more active flavonoids in cellular and animals of S. typhimurium infection. As many Gram-negative bacterial pathogens use type III protein secretion systems to infection host cells, elucidating the mechanisms by which flavonoids inhibit bacterial virulence should reveal new targets and lead compounds that could be used to selectively target pathogenic bacteria and preserve beneficial host microbiota. Given the emergence of new and antibiotic-resistant bacterial pathogens, our studies should afford new anti-infective agents combat bacterial infections in humans. PUBLIC HEALTH RELEVANCE: The potential beneficial effects of flavonoids on human health has been extensively explored in the context of antioxidants, anti-inflammatory and anti-cancer agents and resulted in their distribution as dietary supplements. Our analysis of TCMEs that have been historically used as anti-infective remedies has revealed the unexpected and exciting discovery that specific flavonoids can antagonize T3SSs in bacterial pathogens and attenuate their virulence. The multi-disciplinary studies described in this application should help elucidate the flavonoid mechanisms of action on T3SSs and afford new lead compounds to attenuate bacterial pathogens in vivo. This approach should preserve the beneficial members of the host microbiota and minimize the development of antibiotic resistance to new anti-infective agents.
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{
"pile_set_name": "NIH ExPorter"
}
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Huntington's Disease (HD) is a fatal neurodegenerative disorder caused by an expanded polyglutamine (polyQ) tract in the protein huntingtin (htt). Sadly, a decade of research into the mechanisms of polyQ- dependent neurodegeneration has failed to produce even a single effective therapy for HD. Although small molecules have been identified that inhibit the aggregation of a mutant htt fragment in vitro, many bind to secondary structures shared by other proteins, and it is not known if any of these molecules will be effective and specific in more complex models of HD. We recently completed a loss-of-function (LOF) genomic screen in S. cerevisiae with single gene deletion strains that identified kynurenine 3-monooxygenase (KMO), an enzyme in the KP of tryptophan degradation, as a potent suppressor of mutant htt toxicity. The brain levels of two neurotoxic metabolites in the KP, quinolinic acid (QUIN) and 3-hydroxykynurenine (3-HK), are increased in the striatum and neocortex in early grade HD; similar increases in QUIN and/or 3-HK are present in three mouse models of HD. QUIN and 3-HK have long been hypothetically linked to the pathophysiology of HD. Indeed, intrastriatal injection of QUIN together with 3-HK causes striatal lesions that may be mediated by the combination of N-methyl D-aspartate (NMDA) receptor over-stimulation (excitotoxicity) and free radical formation. In our proposal, we present data showing that Ro 61-8048, a high-affinity, orally bioavailable small molecule inhibitor of KMO, decreases QUIN, 3-HK and mutant htt toxicity in yeast, and significantly improved neurological index, rotarod performance, locomotor activity and ambulatory distance in a small pilot study using a mouse model of HD. Remarkably, we show KMO is expressed exclusively in microglia. Microglial activation has been documented in postmortem brains of early grade HD patients and in HD mouse models. However, little is known about the role of microglia in HD pathophysiology. We show that primary microglia isolated from HD mice have significantly increased levels of 3-HK. We hypothesize that mutant htt induces a transcriptional defect that activates the KP in microglia, and that inhibiting the KP via pharmacological and genetic approaches will improve behavioral and pathological outcome measures in HD mouse models. We propose to test these hypotheses by studying the role of mutant htt and the KP in cultured microglia and in vivo in mouse models of HD. These experiments will establish whether KMO inhibitors such as Ro 61-4048, which showed promising results in a small pilot study, deserves further consideration for pre-clinical development as a HD therapy. More broadly, our genetic experiments in mice will determine whether pharmacological inhibition of KMO is a bona fide therapeutic approach to treating HD. PUBLIC HEALTH RELEVANCE In this project we will use genetic and pharmacological approaches to determine if blocking a metabolic pathway implicated in Huntington's disease (the kynurenine pathway) confers protection in mouse models of this disorder. The kynurenine pathway is found predominantly in microglia, the macrophages of the brain, which are activated abnormally in pre-symptomatic Huntington's disease patients. If successful, our studies may lead to clinical tests of small molecule inhibitors of the pathway in patients with Huntington's disease.
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"pile_set_name": "NIH ExPorter"
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DESCRIPTION (Applicant's Abstract):This grant has focused on the molecular biology and functions of the eosinophil's granule-associated cationic and other proteins to elucidate key structure-function relationships for their potent inflammatory actions in allergic responses, host-parasite interactions, tissue damage and fibrosis. This renewal focuses on the Charcot-Leyden crystal (CLC) protein, which forms the distinctive bipyramidal crystals which are hallmarks of eosinophil (or basophil) participation in allergic and related inflammatory reactions. CLC protein was originally identified as eosinophil lysophospholipase (LPLase), but we have now shown it to be a member of the galectin superfamily of animal lectins based on amino acid sequence, 3D protein structure, lack of LPLase activity and gene structure. The physiologic/effector role this highly abundant eosinophil constituent is therefore unresolved. Quantitative considerations alone (CLC is 10 percent of eosinophil protein), but also its secretion by activated eosinophils, elevated blood levels in patients with eosinophilia, and increased levels in sputum and BAL of asthmatics, all argue for an important role in eosinophil function. Our cloning of the CLC cDNA and analyses of CLC amino acid sequence identified similarities to b-galactoside-binding animal galectins, but not to any LPLases or other lipolytic enzymes. Our X-ray crystal structure showed CLC protein to be nearly identical to human galectins-1, -2, -3 and -7, and to possess a carbohydrate recognition domain capable of binding mannose, but not standard b-galactoside sugars. We demonstrated that CLC protein is not eosinophil LPLase, which is likely identical to human pancreatic LPLase. The goal of this renewal is to investigate the mechanisms by which eosinophils, through their considerable LPLase activity or the lectin activities of CLC protein, function in allergic pulmonary inflammation. Three questions are addressed with regard to the structure and functions of CLC protein and eosinophil LPLase in terms of their pathophysiologic roles in acute and chronic allergic airways inflammation: (1) What is the biologically relevant glycoconjugate ligand for CLC protein and what does it tell us about CLC's role in eosinophil (or basophil) biology and function. We will identify the physiologically relevant ligand(s) for CLC protein; (2) What are the structure-function relationships for CLC protein's carbohydrate-binding "galectin-like" activities and ligand specificity? Site-specific mutagenesis will be used to characterize CLC protein's carbohydrate recognition domain and its mechanism of binding for the glycoconjugate ligands we define; (3) What is the role of eosinophil LPLase in allergic pulmonary inflammation involving eosinophils, and in asthma pathophysiology? We will determine whether eosinophil LPLase alters pulmonary surfactant function, resulting in decreased patency of distal airways in asthma, or modulates allergic pulmonary inflammation. We will analyze the effects of eosinophil LPLase on pulmonary surfactant function in vitro, and in vivo by determining the relationships between eosinophil recruitment and secretion of LPLase in the lung and its localization in distal airways in human asthma and murine allergic asthma models. We will analyze the effects of targeting overexpression of LPLase in the lung using transgenic approaches, and determine the effects of knocking out the eosinophil LPLase gene in murine models of eosinophilic airways inflammation. The proposed work should elucidate unique aspects of the structural biology and functions of CLC protein in eosinophils (and basophils), and the pathophysiologic actions of eosinophil LPLase in disease processes associated with eosinophilic inflammation in the lung and other tissues.
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{
"pile_set_name": "NIH ExPorter"
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General anesthesia is a reversible, drug-induced behavioral state comprised of unconsciousness, amnesia, analgesia and immobility with stability and control of vital physiological systems. This fundamental tool of modern medicine is crucial for allowing thousands of patients daily to safely undergo most surgical and many non-surgical procedures. Today this state is induced and maintained by administering multiple drugs that act at multiple sites in the brain and central nervous system. Emergence from general anesthesia is a passive process whereby anesthetic drugs are merely discontinued at the end of surgery and no drugs are administered to actively reverse their effects. Allowing multiple drugs to act at multiple sites without specific mechanisms to terminate their effects most likely explains a significant component of anesthesia-related morbidity; drug side effects (nausea, hypotension, respiratory depression, hypothermia) are due to actions at sites other than their intended targets whereas persistent effects (delirium, cognitive dysfunction) are due to actions at intended targets for periods longer than desired. Hence, general anesthesia, as presently produced, is highly non-specific and inefficient. Despite the central role of anesthesiology in modern healthcare, research in this field is overly focused on deciphering the anesthetic and toxic mechanisms of current drugs with little to no attention being paid to developing new approaches. The paradigm-shifting question whose answer will revolutionize anesthesiology is not, how do current anesthetics work?, but rather, how should the state of general anesthesia be designed? We hypothesize that the answer is by developing strategies to control directly the brain's natural inhibitory pathways and arousal centers. We propose to redesign general anesthesia by combining optogenetic, electrical and pharmacological manipulations in rodent models to create this behavioral state through precisely timed control of the brain's natural inhibitory pathways and its arousal centers. If successful this research will provide a new fundamental understanding of brain arousal control, and eventually, new anesthesiology practices including: neurophysiologically-designed approaches to creating general anesthesia; reduction in morbidity; improved brain function monitoring; safer anesthesia care by non-anesthesiologists; and possibly novel therapies for arousal disorders such as depression, insomnia, pain and coma.
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{
"pile_set_name": "NIH ExPorter"
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There is a need for a comprehensive, standardized, and engaging curriculum to train addiction treatment counselors to become credentialed clinical supervisors. Danya International, Inc., proposes to conceptualize, develop, and evaluate a multicomponent, multimedia clinical supervision training curriculum for addiction treatment counselors. The Clinical Supervision Training Program will consist of an 18-hour classroom training curriculum; a 12-hour online training curriculum; a PowerPoint presentation; a participant workbook; a 20-minute video; a CD-ROM-based, computer-simulated interactive training tool; and a train-the-trainer curriculum. The Clinical Supervision Training Program will be designed to: increase clinical supervision skills and knowledge among senior addiction treatment staff; help counselors become credentialed Certified Clinical Supervisors of the International Certificate and Reciprocity Consortium, and obtain continuing education units; increase learning, skills acquisition, and adoption of new knowledge regarding clinical supervision using visual, auditory, and kinesthetic learning styles; develop the national standard of clinical supervision training in addiction counseling; and increase opportunities for professional mentorship, clinical competency, and professional growth. During this Phase I SBIR grant, project staff will conduct a needs assessment with clinical supervisors; write the text for the 18-hour classroom training; develop the PowerPoint presentation; write the text for the 12-hour online training; write the text for the participant workbook; conduct an expert peer-group evaluation of the curriculum products text; and conduct a pilot feasibility evaluation of the classroom training modules, PowerPoint presentation, and hard-copy mockup of the online course with clinical supervisors and counselors. Hazelden Publishing and Educational Services has expressed an interest in this product for its Clinical Innovators Series and professional development materials product line. [unreadable] [unreadable] The project has the potential to impact public health by helping qualified addiction counselors become credentialed clinical supervisors and by increasing clinical supervision skills and knowledge among senior addiction treatment staff. These skills will serve to strengthen the supervisory relationship and improve the delivery of clinical supervision directly impacting the services provided to drug abusing clients. [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Signals mediated by the T cell antigen receptor (TCR) complex regulate both T cell development and the balance between activation and tolerance in both early T cells (thymocytes) and mature T cells. Understanding how TCR signals regulate these processes will be essential for understanding autoimmune diseases and designing effective vaccines to pathogens. The antigen recognition ability of T cells is supplied by clonotypic chains (TCRa,b), but the signaling potential is conferred by the CD3 subunits (CD3e, CD3g,and CD3d) and members of the zeta (z)-chain family (z, eta, and Fcg). The ability of z-chain and CD3 components to transduce signals derives from a consensus sequence (ITAM) present within their intracytoplasmic domains. Each CD3 chain contains a single ITAM, whereas full length z-chain contains three ITAMs. To understand better the role of specific and multiple signaling motifs in immune processes, we have studied genetically altered mice lacking TCR-z chain and/or CD3e. We have reconstituted these mice with transgenes encoding natural or genetically engineered variants of z-chain and/or CD3e. We have used these animals to assess the role of TCR signaling in T cell development and activation. Using a breeding process in which we introduced clonotypic TCRab genes with defined specificities into signaling altered mice, we have demonstrated several key findings. 1)Thymocytes expressing TCRs with low affinity for ligand were dependent on multiple ITAM motifs for development. Importantly, TCR's with higher affinity for ligand were less dependent on multiple motifs for their development, revealing an important relationship between TCR affinity and signaling potential for development of the TCR repertoire. 2) The phenotype of cytokine responses (Th1 vs Th2) correlated with the signaling potential of the TCR. 3)The biochemical signals mediated by z chain and CD3 components are at least partially redundant. 4)The ITAM present in the CD3e chain does not have a specific function in development but rather contributes to TCR function through signal amplification. We are currently constructing mice that lack both CD3e and z-chain and will reconstitute these animals with CD3e and z-chain variant molecules to further investigate the multiple signaling structure of the TCR. Despite the dramatic effects alteration in TCR ITAMs have on T cell development, those T cells that populate the peripheral lymphoid organs in ITAM-deficient mice appear functional and proliferate in response to anti-TCR antibodies, mitogens and specific peptides. We think that expression of costimulatory molecules serve to dampen effects of deficiencies in TCR ITAM-mediated signals. Hence, we will test if costimulatory moleucles are particularly important for responses by T cells from ITAM deficient mice. We have begun a series of experiments, using purified MHC molecules loaded with specific antigen to look at the role of such costimulatory molecules in situations in which TCR signals are impaired. Finally, during the last year, we have used our model to examine how alterations in TCR signaling potential affect the development of autoimmune disease. We are using the multi-dose streptozoticin model of diabetes to determine how specific alterations in TCR signaling affect the development and pathology of diabetes. Early data suggest the signaling potential (number of ITAMs) affects the development of diabetes. Current studies are directed at understanding the inflammatory process that underlies diabetes in these mice.
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{
"pile_set_name": "NIH ExPorter"
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This Administrative Core (Core A) will provide the necessary scientific, organizational, and fiscal oversight of the Consortium for Development of Immunotherapeutics Against Viral Hemorrhagic Fevers. This consortium includes multiple investigators and a vast array of antibodies with in vitro or in vivo efficacy against the filoviruses and arenaviruses. An essential feature ofthe consortium is coordination of projects and studies on these antibodies, so that effort is not unduly duplicated, the separate multidisciplinary analyses integrate effectively and seamlessly, and the group achieves maximum synergy. The Administrative Core will develop and implement a management plan to ensure the success of this program. This Core will continuously monitor the scientific progress of each component of the program and ensure that results, the meaning of the results, and next steps are effectively communicated to the rest of the group at each stage. This Core will also facilitate communications with the NIH, the larger research community, and our external advisors so that these comprehensive studies are definitive and defined by consensus in the fields, and so that the resulting products can be effectively translated toward clinical use. The Core will manage financial resources and ensure that the select agent, vertebrate animal, human subject, and intellectual property issues are respected. The Core will also initiate all external collaborations and oversee invitation, selection, and progress of supplemental research activities as directed by the NIH.
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{
"pile_set_name": "NIH ExPorter"
}
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Despite improvements in cancer therapy, the prognosis of metastatic lung and non-colorectal gastrointestinal cancers remains poor, with median survival rates of less than one year. During the course of illness, these patients often experience marked physical suffering, psychological distress and resource-intensive care at the end of life. While cancer treatment can extend survival, its impact on quality of life and distress is minimal for these patients. Moreover, the trend toward increased administration of anti-cancer therapy near the end of life negatively impacts quality of death, family caregiver bereavement, and the cost of health care delivery. Comprehensive cancer care ideally would include interventions to diminish suffering as well as enhance the quality and delivery of care for patients with limited life expectancy and high symptom burden. We hypothesize that a highly integrated model of early palliative care will improve quality of life and mood in both patients and their family caregivers as well as facilitate the delivery of quality end-of-life care. Palliative care clinicians focus on multiple aspects of care including support and education to assist with treatment decision-making, physical and psychological symptoms, as well as spiritual and existential concerns. Our previous randomized study of early, integrated palliative and oncology care versus standard oncology care alone in patients with newly diagnosed metastatic non-small cell lung cancer demonstrated marked improvements in patient-reported outcomes, quality of end-of-life care and resource use. Despite receiving less chemotherapy at the end of life and utilizing more hospice services, patients assigned to early palliative care had prolonged survival compared with patients receiving standard care. We now propose a larger, randomized controlled trial using mixed quantitative and qualitative methods to expand our findings in patients with newly diagnosed metastatic lung and non-colorectal gastrointestinal malignancies. The goals of this study are to: 1) demonstrate the efficacy and generalization of the early palliative care model to improve quality of life, mood, quality of end of life care, resource utilization and survival in a more diverse sample of patients with poor prognosis, metastatic cancers; 2) examine potential mediators of the relationship between receiving early palliative care and patient outcomes, including prolonged survival; and 3) operationalize national guidelines for quality ambulatory palliative care into an evidence-based manual of preferred practices that can be disseminated broadly for clinical and research purposes. This single-institution study will serve as the foundation for a multisite trial utilizing our evidence-based manual of preferred palliative care practices to ensure a measurable and reproducible intervention. The long-term goals of this research are to improve patient outcomes and transform oncology practice by enhancing the quality of end-of-life care and optimizing resource utilization through timely integration of palliative and cancer care in patients with metastatic disease. Achieving these critical goals without negatively impacting patient survival would undeniably challenge existing paradigms in cancer care. PUBLIC HEALTH RELEVANCE: The goal of this project is to enhance the care of patients with poor prognosis, metastatic cancers by involving palliative care soon after diagnosis. Although palliative care teams have traditionally seen patients late in the course of disease, this study seeks to demonstrate that their early involvement aids with treatment decision- making and leads to improved quality of life and mood in patients with advanced cancer. We hope that combining palliative care with traditional cancer treatment will not only prove beneficial for patients and their families but also transform comprehensive cancer care.
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{
"pile_set_name": "NIH ExPorter"
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The ultimate goal of our proposal is to develop a novel therapeutic approach that will save lives of aged septic patients. Sepsis is the most common cause of death in the non-cardiac intensive care units (ICU). It is particularly a serious problem in the geriatric population. The elderly (e 65 years of age) accounts for 12% of the US population but 65% of sepsis cases. Nearly 80% of septic deaths occur in elderly patients. Although this problem is increasingly recognized, current treatment options for aged septic patients are very limited. Ghrelin is an endogenous ligand for the growth hormone (GH) secretagogue receptor 1a (GHSR1a, i.e., ghrelin receptor). We have shown that administration of ghrelin inhibits inflammatory responses, attenuates organ injury, and reduces mortality in young septic animals. Ghrelin's beneficial effects are attributed to the activation of the vagus nerve through ghrelin receptors in the brain. We have also shown a greater production of proinflammatory cytokines in aged than young rats under endotoxemia, and ghrelin treatment fails to rescue aged septic animals. The age-related hyperinflammation is associated with central hyporesponsiveness to ghrelin, which results in reduction of parasympathostimulatory neuronal activity in the dorsal vagal complex (DVC) of the brain. Down regulation of ghrelin receptor is responsible for central hyporesponsiveness. Recently, we have discovered that a low dose treatment of rat GH up regulates the ghrelin receptor in aged rats. In humans, GH levels decline ~15% per decade after age 25. We have demonstrated that treatment with rat ghrelin in combination with rat GH reduces inflammation and attenuates tissue injury in aged septic animals. However, in consideration of the potential immunogenicity of applying rat proteins in humans, development of ghrelin/GH derived from human as therapeutic agents is needed for septic patients. Thus, human ghrelin/GH will be tested in this proposal. Moreover, a more clinically relevant model of sepsis induced by cecal ligation and puncture (CLP) will be used to determine the effect of human ghrelin/GH in aged animals. The primary objective of this project is targeted towards demonstrating the feasibility of the further development and commercialization of human ghrelin in combination with human GH as a novel therapy for sepsis in the geriatric population. We plan to first determine the optimal dosage of human GH as a ghrelin sensitizing agent in aged rats, and then assess the dose-dependent effect of human ghrelin/GH on inflammatory responses and tissue injury in aged septic rats, and finally determine the effect of human ghrelin in combination with human GH on sepsis- induced mortality in aged rats. Our future goal (SBIR Phase II and beyond) is to obtain commercial utilization of human ghrelin/GH as a safe and effective therapy for aged patients suffering from sepsis.
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{
"pile_set_name": "NIH ExPorter"
}
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Thousands of chemicals in wide commercial use have not been tested for adverse effects on humans, but are present in the environment. Accordingly, there is a need to improve chemical prioritization for in vivo toxicity testing and, ultimately, to find cell-based alternatives for evaluating the large inventory of potentially harmful substances. Quantitative high throughput screening (qHTS) assays are multiple-concentration experiments with an important role in the efforts of the National Toxicology Program to meet these testing challenges and advance toxicology from a predominantly observational science to a predominantly predictive science. qHTS can simultaneously assay thousands of chemicals over a wide chemical space with reduced cost per substance. Previous approaches for making activity calls from qHTS data were based on pharmaceutical applications seeking to minimize false positives and usually relied on heuristics rather than statistical tests to make activity calls. For that reason, we developed a three-stage algorithm to classify substances from qHTS data into statistically supported activity categories relevant to toxicological evaluation, seeking to improve sensitivity while minimizing Type I error rate (Shockley, 2012). The first stage of our approach fits a four-parameter Hill equation to find active substances with a robust concentration-response profile within the tested concentration range. The robust criterion specifies that response profiles are statistically significant using both unweighted and weighted non-linear least squares (NLS and WNLS) regression. NLS weights all data points equally and, consequently, may not discriminate between a profile with data along both asymptotes and a profile supported by a single point. WNLS weights each response point based on 1/s2, where s is the sample standard deviation estimated from all response data within a defined concentration range containing the response point of interest, so that more influence is given to neighboring data points with similar response levels than neighboring data points with very different responses. The second stage finds relatively potent substances with substantial activity at the lowest tested concentration, substances not captured in the first stage. The third and final stage separates statistically significant profiles from responses that lack statistically compelling support, or inactives. This framework accommodates large volumes of qHTS data, tolerates missing data, and does not require replicate measurements. We evaluated this three-stage classification algorithm via extensive simulations (Shockley, 2012). The area under receiver operating characteristic curves (AUC) was used to assess performance. Using AUC statistics, our algorithm outperformed overall F-tests comparing the fit of the Hill equation to a horizontal line (no response) when the concentration for half maximal response (AC50) was less than 0.1 micro molar. It also outperformed t-test approaches in detecting known actives when the AC50 was greater than 0.001 micro molar. The three-stage decision strategy yielded good (AUC >= 0.75) to high (AUC >= 0.9) performance for 14 point concentration-response curves when the response was in the detectable region of the simulated assay (>25% of the positive control). Our approach was able to detect relatively potent substances (e.g., AC50 = 0.001 micro molar) with as few as 4 data points when the tested response was at least 50% of the positive control response. The three-stage algorithm described above is based on the Hill equation model. However, concentration-response data can be complex and it may be more informative to find alternative patterns in the data not based on fits to sigmoidal curves. We are currently developing approaches to find complex patterns in qHTS data based on principles of order restricted inference. Multiple hypothesis testing is used in order to compare the significance of all searched patterns and identify the most appropriate pattern describing a response profile. It is often unclear how to prioritize chemicals for follow-up studies due to the large uncertainties that accompany parameter estimates derived from nonlinear regression model fits to data generated in qHTS experiments. Therefore, we have also used a weighted entropy score (WES) as a measure of average activity level in order to rank chemical in qHTS experiments. WES scores can be used to rank all chemicals in a tested library without a pre-specified model structure, or WES can be used to complement existing approaches by ranking returned hits. The performance of WES has been evaluated using data simulated from a Hill model. WES outperforms rankings based on AC50 (estimated concentration of half-maximal response) across the full range of conditions that are typical of qHTS studies.
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{
"pile_set_name": "NIH ExPorter"
}
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A Molecular theory for the rate of nonadiabatic electron transfer was developed and its relation to classical Marcus theory was established. The rate was found to be determined by the probability density of the energy gap, which is defined as the instantaneous change in solvation energy upon moving an electron from the donor to the acceptor. It was shown how this probability density can be obtained from the free energies of transferring varying amounts of charge between the donor and acceptor (as specified by a charging parameter). A simple algorithm was proposed for calculating these free-energy changes (and hence the energy gap probability density) from computer simulations on just three states; the reactant,t he product, and an "anti" product formed by transferring a positive unit charge from the donor to the acceptor. The Marcus relation (i.e., the activation energy as a parabolic function of the free-energy change of reaction) was derived in a way that clearly shows that it is a good approximation in the normal region even when the solvent response is significantly nonlinear. A simple generalization of this relation, in which the activation energy is given by parabolic functions with different curvatures in the normal and inverted regions, was proposed. A novel method has been developed to analyze NMR relaxation experiments on unfolded proteins. It was shown how the spectral density function describing the dynamics of amide bond vectors can be determined at specific frequencies using N15 relaxation parameters alone. Experimental data on the folded and unfolded forms of a small protein was used to establish the validity of this procedure A continuum model of electrostatic interactions in protein has been used to analyze the energetics of a positionally disordered water molecule within an apparently hydrophobic cavity of a protein, as has recently been bound experimentally using NMR.
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{
"pile_set_name": "NIH ExPorter"
}
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The repeated use of psychostimulants produces long-term changes in brain chemistry and behavior that are thought to contribute to persistent and compulsive drug use. Long-term adaptations are also observed in response to other drugs of abuse including opiates and alcohol. Our studies seek to understand the mechanisms, both at the systems and cellular level, which underlie these changes. Included in this goal is the identification of biological factors that may predispose certain individuals to abuse psychoactive drugs and pharmacotherapies that may be effective in the treatment of drug and alcohol addiction. Our studies have suggested an important role of endogenous opioid peptides in modulating dopamine neurons in the brain upon which drugs of abuse act to affect and subsequently control behavior. We have shown that the release of the opioid peptide, dynorphin, the endogenous ligand for the kappa opioid receptor, is increased in response to the use of amphetamine and cocaine and that this increase opposes alterations in behavior and presynaptic dopamine neurotransmission that occur as a consequence of psychostimulant use. Using biochemical and immunocytochemical approaches we have shown that the activation of the kappa opioid receptor causes marked changes in the trafficking of the dopamine transporter, a membrane-bound protein that is a key substrate upon which psychostimulants act. Our studies suggest that the activation of the kappa opioid receptor increases the trafficking of the transporter from the cytosol to the plasma membrane and that this action may be one mechanism underlying the ?cocaine-antagonist-like? effects of drugs that activate this opioid receptor subtype. Data suggesting that kappa agonists may exert their effects via the activation of two kinases (mitogen activated protein kinase and phosphoinositol ?3 kinase) that regulate cytoskeletal dynamics has also been obtained. We have also shown that pharmacological inactivation or genetic ablation of dynorphin or its receptor results in increased vulnerability to the behavioral and neurochemical effects of cocaine. Our more recent studies have shown that this vulnerability extends to another drug of abuse, morphine and that tolerance to the analgesic effects of morphine, can be reversed by administration of a peptide that binds to and complexes with an N-terminal sequence of the dynorphin molecule. Using pharmacological and genetic approaches to selectively inactivate other opioid receptor types (e.g., mu ? and delta-), we have begun to examine whether dysregulation of specific opioid receptor systems affects basal and drug-evoked neurotransmission in two dopamine- rich brain regions implicated in mediating the behavioral effects of drugs of abuse (n. accumbens and ventral pallidum). In parallel studies, intravenous drug self-administration is being used in conjunction with in vivo techniques for the quantification of peptide and neurotransmitter release in the same animal to determine the role of opioid / dopamine system interactions in mediating individual differences in vulnerability to acquire, maintain and reinstate compulsive drug-seeking behavior.
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{
"pile_set_name": "NIH ExPorter"
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The proposed P30 Phase III Pilot Project Program will primarily act as a mechanism to fund pilot studies that allow junior and senior investigators to generate preliminary data for submission of grant applications using the unique technologies available in the cores. Through this program we expect to incentivize use of our core technologies enabling all investigators but particularly past and present COBRE target faculty to gather preliminary data to test and develop new hypotheses. As a result, we expect to greatly increase the competitiveness of grant applications that when funded may also require future use of core resources. Since the projects will vary widely in scope and needs, we have decided to create three different routes to fund pilot applications. SC BioCRAFT TIGER CUB grant (up to $10K): These grants will be used to obtain data critical to submission of a larger (R-type) grant. Investigators who have already submitted a grant application to external agencies and are looking to revise their application with new data will be given priority for such an award. SC BioCRAFT TIGER grant ($10k-50K): These grants will be reserved for High Risk- High Reward type applications that propose groundbreaking research in the area of regenerative medicine. These can be single PI or multiple PI proposals. SC BioCRAFT TIGER STREAK grant: ($50k-75K). The purpose of the TIGER STREAK grants program is to provide financial support for the establishment or expansion of multidisciplinary collaborations, which will lead to the development of more competitive, center-type proposals and significant collaborative proposals. Total money available for pilot projects will be $200K/year. Vouchers are to be used to cover the costs of services available through the cores. The annual budget for the voucher program will be $40K /yr. with limitations to the number of vouchers per PI. The Administrative Core through its Program Coordinator will have responsibility for implementing and managing this program. Program guidelines and application instructions will be posted on the COBRE website and applications will be submitted to the Program Coordinator using web-based forms. Applications will be solicited four times annually and will be advertised on our website as well as locally and regionally by emails via statewide networks. Applications will be reviewed by the Internal Advisory Committee. Criteria will include, 1) scientific merit, 2) alignment to the scientific themes of our center, and 3) the perceived benefit (e.g. manuscripts, grants) to be derived from the expected data.
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{
"pile_set_name": "NIH ExPorter"
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Project Summary. Since HIV and HCV share common risk factors for infection, co-infections with HIV and HCV are frequently found in Injection drug users (IDUs). These two pathogens are also likely to be responsible for the highest infectious disease morbidity and mortality rates among IDUs. IDUs are the single largest risk group for HCV infection in the United States. However, we know little about direct opioids-virus (HIV and/or HCV) as well as virus-virus (HIV-HCV) interactions, which is a major barrier to a fundamental understanding of the immunopathogenesis of HCV disease and HCV-related morbidity in HIV/HCV- coinfected IDUs. A major barrier to address this key issue in vitro is the lack of a HCV infectious cell model system. Recently, three independent research groups have reported the development of a robust HCV infectious system in vitro. This newly established cell model provides an excellent opportunity for this proposed project. We have generated infectious HCV from this cell model in our laboratory. The goal of this study is to address our overarching hypothesis that the interactions between opioids (e.g., morphine), HIV, and HCV are a key determinant of the outcome of HCV/HCV infection. We will address previously un- recognized mechanisms by which morphine and/or HIV/HCV compromise the host cell innate immunity, leading to the persistence of viral infection and replication. Specifically, we will investigate the effects of morphine and the HIV proteins (Tat and gp120) on HCV infection, intracellular innate immunity, and the anti- HCV effects of IFN-a/Ribavirin in the human hepatic cells. We will utilize innovative cocultures of human hepatic cells with Kupffer cells to study the interactions of HIV with HCV in the presence or absence of morphine. Since Kupffer cells are the resident macrophages of liver, and are the target for HIV infections, the inclusion of Kupffer cells in this study is highly significant. Data arising from this study will be critical for a better understanding of opioids as a cofactor in the immunopathogenesis of HIV/HCV infection. The long term goal of this proposal is to develop host innate immunity-based treatment and prevention strategies for HIV/HCV-infected opioid abusers. Relevance to Public Health. There is little direct evidence available about the interactions between morphine, HIV and HCV in different cell systems, which is a major barrier to a fundamental understanding of HCV-related morbidity and mortality in HIV/HCV-infected IDUs. This research will contribute not only to our basic understanding of host cell innate immunity against HIV/HCV, but also to the design and development of innate immunity-based treatment and prevention strategies for HIV/HCV-infected opioid abusers.
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{
"pile_set_name": "NIH ExPorter"
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The Southwest Oncology Group is an adult multi-disease, multi-modality clinical research organization with 33 Member Institutions, 23 Community Clinical Oncology Program (CCOP) institutions, including 5 Minority- Based CCOPs, 27 Urologic Cancer Outreach Program (UCOP) members, 22 High Priority program members and a network of 1,441 Cooperative Group Outreach Program (CGOP) investigators at 272 affiliate hospitals, and 15 institutions participating in the CTEP Minority program. Special emphasis areas for the Group with respect to the inclusion of women and minorities have come under the responsibility of the new Committee on Women and Special Populations. This group will focus on detailed analyses of women in the various disease categories. There have been gender differences identified in melanoma and lung cancer trials. The special populations include a commitment to optimize the enrollment of minorities in Group trials. Analyses of race and ethnicity impacts on the outcome of patients in our clinical trials are in progress. Additional special populations include the elderly patient advocates, and survivors of cancer. The Committee on Women and Special Populations portion of this application details the heavy current and future involvement of patient advocates in the committee activities of the Group. The Group will involve cancer survivors in our clinical trial committee structure. The Chairman of the Group has created an ethics training program which is delivered between 2-3 times to separate audiences, at every Group Meeting in order to help members understand the moral and ethical principles of the conduct of clinical trials. Tied to this, is the fully implemented Conflict of Interest Policy which identifies potential conflicts from all investigators, who then agree to recuse themselves from decision making related to their identified potential conflict(s) of interest. We have an Affirmation of Integrity statement on file for all members of the Group that come in contact with clinical trial data. The Group application includes correlative biologic studies from banked tumor specimens of homogeneously treated patients in breast, gastrointestinal, head and neck, leukemia, lymphoma, and multiple myeloma cancers. There are two new initiatives involved in this application. A proposal in lung cancer biology proposes a 2-year study of the molecular composition of lung cancer specimens obtained from patients on completed Group clinical trials. The second new initiative deals with a 5-year plan for upgrading Informatics at the Statistical Center and at Group institutions to become ready for all electronic data and information transfer into the future.
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{
"pile_set_name": "NIH ExPorter"
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The insulin-like growth factors (IGFs) are clearly involved in maintaining and stimulating breast tumor cell growth, and are therefore potential targets for therapeutic attach in the treatment of breast cancer. The IGF system comprises several elements: two growth factors (IGF-I and IGF-II), two receptors (IGFR-I and IGFRI-II) which both may mediate the growth-stimulating actions of IGF-I and IGF-II, and at least six IGF binding proteins (IGFBPs), which may either enhance or interfere with tumor growth stimulation. In order to target the IGF system effectively, it will be necessary to define the key elements of the system involved in growth regulation, as well as to demonstrate that specifically attacking these elements can inhibit breast cancer growth. We therefore propose three specific aims: (1) To inhibit IGF-stimulated breast cancer cell growth by blocking the expression of the IGFR-I using antisense strategies and by overexpressing a non-functional mutant receptor, (2) To define the role of IGFR-II in breast cancer cell growth by over-expressing it (using inducible promoters), and to determine the effects of the manipulation on basal and IGF-stimulated growth; (3) To investigate the -expression and function of the IGF binding proteins (IGFBPs) in breast cancer cells and clinical breast tumor specimens. It is well established that polypeptide growth factors are important regulators in cancer cell growth in vitro and that interference with growth factor pathways can lead to inhibition of tumor proliferation in many different model systems. This proposal therefore seeks to fully characterize the cellular effectors (receptors and binding proteins) of the IGF system, and to use these observations to develop feasible therapeutic strategies for inhibiting IGF-Medicated breast cancer proliferation.
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{
"pile_set_name": "NIH ExPorter"
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Members of the PSL provide consulting services to scientists and physicians at NIH in different areas of applied mathematics and the physical sciences. A large segment of this effort is devoted to a study of the effects of head injury in veterans of Vietnam. We have continued our study of interpolation error in computerized tomography. We have examined experimental data and suggested further experiments with affinity chromatography to discover the causes of discrepancies from theoretical predictions. We have worked on combinatorial problems that arise from a method of fractionating DNA by alkaline elution.
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{
"pile_set_name": "NIH ExPorter"
}
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The long-term goal of this research is a better understanding of the molecular mechanism and regulatory processes involved in the maintenance of the constant pool of plasma iron that is essential for the biosynthesis of hemoglobin and other important iron- containing systems. It is hoped that studies of this nature will provide insight into the molecular bases of metabolic disorders such as iron deficiency anemia, one of the most common deficiency disorders in humans, and hemochromatosis, the excess storage of iron. The specific objective of this study is to determine, on a molecular level, how the mucosal cell rapidly adjusts to decreases in the iron content of the diet and to decreases in the iron stores in the body to permit the absorption of a greater fraction of the iron present in the diet. Both important phases of iron absorption, uptake into the mucosal cell and processing within the cell, have been implicated as playing a role in this adjustment. Upon transferring animals from a diet of normal iron content to one of low iron content, detailed analyses will be made of the molecular changes that occur both in the uptake of iron into the mucosal cell and in the processing of iron within the mucosal cell. A comparison of these changes as well as a comparison of how well these changes correlate with alterations in the actual absorption of a test dose of iron and in the iron status of these animals should provide insight into the relative regulatory importance of the two important phases of iron absorption.
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{
"pile_set_name": "NIH ExPorter"
}
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There is a fundamental gap in our understanding of how HIV evades innate and adaptive immune responses to establish a persistent infection. Our laboratory's long-range goal is to provide better therapies for HIV- infected people and to move closer to a cure for this infection. We expect that a greater understanding of HIV persistence will inform the design of therapeutic strategies that will result in prevention of infection and/or long term remission or cure. As the next step towards this goal, the objective of this application is to identify cellular intrinsic antiviral mechanisms and to understand how HIV evades them. Our central hypothesis is that APOBEC 3G, an intrinsic antiviral factor that inactivates the virus by converting cytosine residues to uracils, also alerts natural killer cells to the infection. HIV counteracts this cellular response via the concerted activities of Vif, Vpr and Nef. The rationale of the proposed work is that identifying antiviral mechanisms and understanding how HIV evades them will yield mechanistic insights into the establishment of persistent infection and will hasten the discovery of disease-modifying therapies. We plan to test our central hypothesis and accomplish the objective of this application by pursuing the following three specific aims: (1) Determine whether Vif and Vpr counteract deleterious effects of uracil in HIV-infected macrophages. It has been well established that HIV-1 Vpr is necessary for efficient infection of macrophages whereas a potential role for Vpr in T lymphocytes has been ambiguous. Based on our preliminary data, we hypothesize that Vpr is needed in macrophages to counteract the deleterious effects of high concentrations of cytoplasmic dUTP found in non- dividing cells such as macrophages and/or to counteract macrophage-specific expression of APOBEC family members that are resistant to Vif. We will test this hypothesis by measuring uridine incorporation and HIV DNA stability in HIV-infected macrophages plus or minus Vpr. We will also use multiple approaches to assess the role of APOBEC family members. (2) Determine whether UNG2 is both a necessary host co-factor for Vpr-dependent repair as well as a target of Vpr-mediated degradation. Our working hypothesis, based on preliminary data, is that Vpr counteracts APOBEC-mediated cytidine deamination and incorporation of uridine by activating the uracil glycosylase, UNG2, to promote repair of viral DNA. We will use molecular approaches to determine the extent to which UNG2 is responsible for Vpr-dependent removal of uridine residues incorporated in the HIV genome in primary macrophages and T cells. The proposed experiments will help explain apparently conflicting published data regarding a role for UNG2 in HIV infection. (3) Determine whether baseline and/or induced A3G is associated with reservoir size in HIV-infected people. Our preliminary data indicate that APOBEC3G (A3G) expression is variable amongst uninfected donors and furthermore that A3G expression and NK-cell activation and lysis are induced to varying degrees upon HIV infection. Based on these preliminary data we hypothesize that a strong intrinsic antiviral immune response mediated by A3G-dependent activation of NK cells will limit reservoir size. This hypothesis will be tested using samples from HIV-infected donors analyzed ex vivo. If our hypotheses are proven correct, this work will reveal APOBEC and uracilation as key antiviral factors that promote antiviral clearance through natural killer cell activation and lysis of virally infected cells. Moreover, antiviral compounds that disrpt the activity of Nef, Vif and Vpr will limit viral infectivity and promote immune cell clearance of infected cells. These results have particular relevance for cure research as intact immune surveillance systems may be needed to identify reactivated latent reservoirs of HIV and to eradicate the infected cells.
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{
"pile_set_name": "NIH ExPorter"
}
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Electron Transfer (ET) reactions play a central role in chemical and biological processes. Early experimental and theoretical studies have identified the key factors in ET reactions and reach a qualitative understanding using a continuum model for the solvent. However, recent experiments are providing detailed information on the level where the microscopic nature of the actual environment of the donor and acceptor might be very important. The basic objective of this proposal is to contribute toward a more microscopic description of ET reactions in solution and proteins using actual molecular simulation. It is proposed to simulate ET reactions in several key experimental systems, focusing on the detailed correlation between the theoretical and experimental results. Our proposed simulation studies will include the following projects: (i) Studies of the microscopic correlation between activation from energies and the corresponding solvent reorganization energies and free energies for ET in solution and in proteins. Significant effort will be dedicated to studies of chemically modified proteins (e.g. Ru-cytochrome c). (ii) Microscopic simulation of dynamical effects in ET reactions, with the goal of correlating rate constants with the dielectric relaxation times of the given microenvironments. (iii) Calculations of the intramolecular Franck-Condon activation barriers in highly exothermic reactions. (iv) A major effort will be dedicated to studies of the detailed energetics and dynamics of ET processes in bacterial photosynthesis. This project will try to use the X-ray structure or the reaction center to evaluate the energies of relevant charge transfer states and to explore the effects of the fluctuations of the protein dipoles. (v) The redox potential of electron transfer proteins will be evaluated by free energy perturbation methods. This study will include calculations of the effects of genetic modifications on redox potentials.
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{
"pile_set_name": "NIH ExPorter"
}
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The neurobiological events that underlie learning and memory likely include activity-dependent modification of synaptic strength in brain structures known to subserve memory. Examples of use-dependent synaptic modification hypothesized to contribute to memory are long-term potentiation (LTP) and long-term depression (LTD) of synaptic transmission at excitatory synapses in the hippozampus. Hippocampal LTD, similar to hippocampal LTP, is persistent, lasting for days in area CA1 in vivo. The persistence of LTD suggests that the underlying molecular mechanisms involve altered gene expression, although some of the changes in expression must differ from those involved in LTP. The proposed work addresses this issue by examining the role of the serine/threonine protein phosphatases PP1 and PP2A and that of the extracellular signal-regulated kinase (ERK) cascade in LTD and LTP in area CA1 in vivo. Motivated by previous work from our laboratory and work by others, we propose to pursue the following Specific Aims: (1) To determine whether PP1 or PP2A is responsible for decreased activation of the transcription factor CREB during LTD and plays a role in the persistence of LTD; (2) To determine whether LTD requires de novo transcription and translation and is associated with an increase in ERK/Elk-l-mediated gene expression but not in CREB-mediated expression; and (3) To delineate the role of ERK in the regulation of CREB and Elk-1 function in LTD vs. LTP, and test whether reduced phosphatase action on CREB during LTP contributes to the persistence of LTP. We will address these questions with a combination of in vivo electrophysiological techniques, enzyme activity assays, Western blot and immunohistochemical analyses, and in situ hybridization. Collectively, the studies will elucidate mechanisms of negative and positive regulation of transcriptional signals in response to plasticity-inducing synaptic activation in the adult in vivo hippocampus, and will yield models of how interactions among different signaling events combine in the regulation of bidirectional synaptic plasticity. The findings from these studies will provide the foundation for future work directed at relating signaling events involved in bidirectional synaptic plasticity to signaling events involved in different types of memory operations performed by behaving animals.
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{
"pile_set_name": "NIH ExPorter"
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Histone N-terminal tails undergo multiple post-translational modifications including methylation, which significantly impact chromatin structure and transcription. Our main interest is to understand histone methylation regulation, a modification that was considered irreversible until our discovery of the first histone demethylase, LSD1. Recent studies implicated LSD1 in differentiation, heterochromatin biology and tumorigenesis. However, the LSD1 biology and in vivo mechanism of action remain incompletely understood. Loss of the C. elegans LSD1 homolog SPR-5 has recently been reported to result in progressive sterility (W. Kelly, per. comm.). In the past funding period, we pursued this further and uncovered an important link between LSD1 and C. elegans germline maintenance based on the findings that 1) spr-5 mutants display increased, p53-dependent germ cell apoptosis, suggesting activation of a DNA damage checkpoint; 2) RAD-51 foci levels are elevated during meiotic prophase in spr-5 mutants suggesting that progression of double-strand break repair is impaired. Thus, a focus of this application is to understand the mechanisms by which SPR-5 [unreadable] regulates germline maintenance. We will determine the demethylase activity of SPR-5 and its related proteins T08D10.2 and AMX-1 in vitro and in vivo, and assess the role of the enzymatic activity in germline maintenance (Aim 1). We will explore the mechanisms by which SPR-5 regulates germline maintenance by considering two alternative but not mutually exclusive models (Aim 2): 1) SPR-5 exerts a direct effect on chromatin structure impacting DNA damage susceptibility and repair. 2) SPR-5 exerts its function indirectly via controlling transcription of DNA damage response genes. We will also investigate the progressive infertility observed in spr-5 mutants, which is reminiscent of defects in telomere maintenance (Aim 2). Lastly, we will identify protein networks that regulate SPR-5 germline functions through protein complex purification and investigate the genetic relationship between SPR-5 and other histone modifying enzymes by genetic suppressor/enhancer screens (Aim 3). Collectively, this application will provide significant new insights into the poorly understood role of chromatin and chromatin modifying enzymes in germ cell maintenance. Findings will also shed new light on molecular mechanisms that control germ cell functions and will impact on our understanding of analogous processes in higher eukaryotes. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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This study is designed to assess changes in cardiovascular risk factors in women using Norplant as compared to women using a low dose oral contraceptive containing estrogen.
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{
"pile_set_name": "NIH ExPorter"
}
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The yeast spindle plaque and associated microtubules mediate the fusion of nuclei during mating. By identifying the genes and gene products responsible for nuclear fusion we aim to identify the components of the spindle plaque and the mechanism of its function. The spindle plaque is of wide interest as it is the sole organizer of microtubules in the yeast cell. Little is known about the mechanism of function of any microtubule organizer. Therefore understanding of the mechanism of nuclear fusion is likely to be important to the understanding of a broad set of phenomena common to all eukaryotic cells such as organelle movement, membrane and organelle fusion, and the regulation and function of microtubules in mitosis and meiosis. Yeast is an organism which is easily manipulated genetically, which should allow rapid analysis of these very complex processes. We will address the following specific experimental aims: 1) The full spectrum of functions on one nuclear fusion gene, KARI, in mitosis, nuclear fusion and sporulation, will be determined by a combination of the genetic analysis of mutations in the gene and the subcellular localization of the gene product. Genes that interact with the KARI gene will be identified by mutations that suppress different KARI mutations. 2) The mechanism of nuclear fusion will be examined by the development of an in vitro system for nuclear fusion. The proteins that are required for nuclear fusion will be characterized in the in vitro nuclear fusion assay. 3) The pathway of nuclear fusion will be dissected by the isolation of mutations in the other genes that are required for nuclear fusion. The functions of the different genes will be ordered by reciprocal shift experiments and cytology of the different mutants. Nuclear fusion genes will be isolated and analyzed by in vitro mutagenesis and gene replacement to determine their role in the mitotic functions of the spindle plaque.
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{
"pile_set_name": "NIH ExPorter"
}
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The overall objective of this proposal is to analyze the mechanisms regulating cell lineage determination and cell commitment during mammalian development. Specifically, we propose to explore the role of imprinted genes and the role of cell surface molecules in initiation and maintenance of specific embryonic lineages. Genomic imprinting, which occurs during gametogenesis in mammals, imposes different functional states upon maternally and paternally derived genomes. Accordingly, completion of normal embryogenesis requires the presence of both a maternally derived and paternally derived nucleus. Zygotic transfer of pronuclei shows that the trophectodermal lineages fail to develop in gynogenetic embryos, which lack the male genome, while the inner cell mass derived lineages fail to develop in androgenetic embryos, which only contain the male genome. Analysis of gynogenetic <-> normal and androgenetic <-> normal chimeras has demonstrate the cell autonomous nature of lineage failure. In addition, analysis of parthenogenetic <-> normal chimeras has demonstrated an apparent need for the male genome in formation of skeletal muscle. Specific cell surface molecules, which appear in a distinct place and time during embryogenesis, serve as unique cell lineage markers and, very likely, also mediate cellula interactions necessary for lineage formation. In order to examine the role of imprinting in lineage formation we propose to isolate embryonic stem (ES) cells from androgenetic, parthenogenetic, an gynogenetic blastocysts, test the ability of these cells to contribute to chimeras, and compare this with the developmental capacity of androgenetic and gynogenetic embryos. In this way we will address the stability and extent of imprinting in cultured stem cell lines and determine whether changes occur in imprinting which alter the capacity for lineage allocation Chimeras between gynogenetic and normal embryos will be analyzed at progressive developmental stages using in situ hybridization in order to determine at which stage of muscle lineage development cells from parthenogenetic embryos fail to participate. Myoblast cultures from chimeric embryos will be initiated and clones derived from cells of parthenogenetic and normal embryonic origin isolated. Comparison of gene expression in normal v. parthenogenetic myoblasts will pinpoint the genes which are differentially expressed (i.e. imprinted). In order to approach the role of cell surface molecules in early lineage allocation we will analyze the control of synthesis and the function of early embryonic antigens present in embryonic ectoderm and endoderm. We propose to clone the gene for the specific fucosyl transferase involved in synthesis of stag specific embryonic antigen 1 (SSEA-1). This will be achieved by expression cloning using mammalian expression vectors and a monoclonal antibody. The cloned gene will be characterized and its function in development determine using gain and loss of function induced mutations.
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{
"pile_set_name": "NIH ExPorter"
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For 6 years, the UCLA OAIC Recruitment and Retention Core (RRC) has provided invaluable assistance to academic investigators seeking to enhance the reach and impact of their funded science by enrolling lower income and minority seniors. Under the leadership of Dr. Mangione until 2011 and Dr. Sarkisian since 2011, the OAIC RRC has successfully facilitated recruitment of lower income and minority senior subjects for 30 different projects. With the RRC's track record and the increased national focus on translational research, the OAIC RRC has evolved into a highly sought-after resource. In this renewal application we seek to continue the highly successful model of the RRC's past 6 years while at the same time expand the level of the Core's collaborations to include not only recruitment and retention but also other critical components of community-partnered research: bi-directional project development, evaluation and dissemination of findings, and creating new research capabilities of our community partners. The RRC will facilitate new partnerships between community partners and affiliated scientists, work closely with our Community Action Board (CAB) to disseminate findings in a manner designed to maximize community impact, and work in partnership with the UCLA Clinical and Translational Science Institute (CTSI) to develop and implement a research training program for senior center staff .across the Los Angeles aging services network. The OAIC RRC challenges the current inefficient project to project community-partnered research model by utilizing an innovative single point of entry for scientists and community partners to come together to deploy equitably partnered research. This infrastructure fundamentally changes the way research is conducted, will enhance the quality, relevance and scientific impact of aging research, and will greatly accelerate the rate of translation of research into community-based programs. Dr. Sarkisian will continue as Core Leader, bringing her expertise as a geriatrician and Director of the NIA-funded Los Angeles Community Academic Partnership for Research in Aging (L.A. CAPRA) Center, with its influential and well-connected CAB and vast network of community partners. Dr. Mangione will continue as Core Co-leader; as Director of the UCLA Robert Wood Johnson Clinical Scholars Program and the NIA-funded UCLA/Drew Resource Center for Minority Aging Research, she brings expertise and access to large numbers of investigators in training who can use RRC resources to become the next generation of researchers in aging.
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{
"pile_set_name": "NIH ExPorter"
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To determine the influecne of virus load, CD4+ cell counts, biologic phenotype, and the presence of symptomatic HIV disease on the response to a new therapeutic regimen.
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{
"pile_set_name": "NIH ExPorter"
}
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The myxobacteria are a group of bacteria ubiquitous in nature. Studies in the last 25 years have demonstrated these organisms produce a host of novel secondary metabolites. Current estimates suggest approximately 50% of all myxobacterial strains produce a bioactive molecule. For some myxobacteria, such as the cellulose-degrading Sorangium cellulosum, this estimate nears 100%. S. cellulosum is best known for its production of epothilones, which are secondary metabolites that possess a mode of action similar to Taxol. Since this organism's genome is approximately 12 million base pairs, the genetic potential for secondary metabolite production is significant. Unfortunately, it's slow growth, copious production of slime, and variable appearance make identification and propagation difficult, resulting in few laboratories involved in new strain isolation. Although several S. cellulosum metabolites have been described in the literature, the vast potential of this organism remains largely unexplored. The broad, long-term goal of this project is to access the capacity of S. cellulosum to produce biologically significant molecules. Our working hypothesis is that since this organism is largely understudied as compared to other secondary metabolite producing organisms, many new biologically active metabolites remain to be discovered. To address this hypothesis, aim 1) of this project will be to isolate new S. cellulosum strains and propagate a historical collection of S. cellulosum known as the Peterson collection. Once strains are isolated, the goal of aim 2) will be to determine the biological activity of S. cellulosum extracts using the National Cancer Institute Yeast Anticancer Drug Screen and to generate a high performance liquid chromatography profile of each extract. Once a strain is identified which produces a metabolite of interest, the goal of aim 3) is to optimize fermentation conditions for production of the molecule of interest followed by its purification. Students pursuing both B.S. and M.S. degrees will be involved in all aspects of this project. Sorangium cellulosum is a bacterium that produces a wide array of molecules that may benefit human health. This project will attempt to identify S. cellulosum isolates that may be producing these molecules followed by their purification. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
}
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This proposal will examine the relationship of immunoregulatory immune cell phenotype and function to the development, clinical activity and disease sequellae of hypersensitivity pneumonitis in human and in a nonhuman primate model of the disease. One aspect of these studies will involve a comparison of in vitro immunologic reactivity (mitogen and antigen responsiveness) as well as immunoregulatory cell phenotype (FCGamma, T4, T8) and function using autologous cocultures in the peripheral blood and bronchial lavage fluids of symptomatic pigeon breeders, asymptomatic but equally exposed breeders and controls. In addition, after development of the Rhesus model, we will perform similar in vitro comparative studies in both symptomatic and asymptomatic but exposed monkeys. A second part of this proposal will examine the relationship of immunoegulatory cell imbalances in vitro and in vivo to disease development and activity using a combination of natural infection and abrogation of T (TBI and/or cimetidine) and prostaglandin mediated monocyte (indomethacin) suppressor cell function. In addition, we will examine the relationship of immunoregulatory imbalances (in vitro and in vivo) and the development of fibrosis (in vivo) using long-term abrogation of suppressor cell function (cimetidine and/or indocin) in conjunction with prolonged daily antigen expsoure. These studies, then, will provide correlative (human and monkey) and direct (monkey) evidence of the relationship between altered immunoregulation andn disease development, activity and sequellae of hypersensitivity pneumonitis in general and may have import for our understanding of other pulmonary inflammatory diseases.
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{
"pile_set_name": "NIH ExPorter"
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The educational aim of the proposed K01 proposal is to allow the applicant to train in developmental affective neuroscience and pediatric bipolar disorder and acquire the skills necessary to characterize neurodevelopmental abnormalities in neural systems of emotion regulation in young adolescents at high genetic risk of bipolar disorder. Bipolar disorder is a chronic and debilitating psychiatric disorder in adults and even more so in children and adolescents. It is characterized by significant impairments in emotion regulation, which have been associated with functional abnormalities in prefrontal and subcortical neural regions. Bipolar disorder may be mediated by neurodevelopmental abnormalities in these neural regions. The onset of bipolar disorder increases dramatically in adolescence, which is a key period for the development of prefrontal systems involved in modulating and regulating emotions. Studies in adolescent offspring of parents with bipolar disorder, who are significantly at risk of developing the illness, promise to help elucidate some of the neural mechanisms involved in the development of bipolar disorder. This proposal will use functional magnetic resonance imaging (fMRI) to investigate differences in neural activity associated with implicit emotional information processing and emotional cognitive control in young healthy adolescents at high genetic risk of bipolar disorder compared to low-risk controls. The secondary goal is to explore to what extent these differences in neural activity are associated with pubertal maturation. The central tenet of the proposed project is that reduced modulation of emotional information processing by higher-order cognitive control processes represents a key risk factor for bipolar disorder. This project complements the proposed career development plan that will involve obtaining training in: 1) developmental affective neuroscience approaches to the study of emotion regulation in adolescence, 2) clinical issues associated with pediatric bipolar disorder, 3) methodology used to examine neural systems of emotion regulation, including increased knowledge and skills in fMRI of neural systems at the interface of cognitive control and emotion and ethical issues associated with pediatric neuroimaging, 4) design and methods used in longitudinal research and advanced statistical techniques for modeling neurodevelopmental trajectories. Findings from this study will contribute to identifying potential neurodevelopmental risk markers for bipolar disorder and establishing endophenotypes of emotion dysregulation specific to bipolar disorder, which will facilitate the development of early intervention strategies.
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{
"pile_set_name": "NIH ExPorter"
}
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Activation of T-cells depends on numerous interactions of surface receptors and cytoplasmic proteins. Cell adhesion and recognition of foreign peptides presented by MHC molecules is mediated by extracellular portions of T-cell receptors, such as the adhesion domains of CD2 and the Fv fragment of the alpha-beta heterodimer (Ti) of the T-cell receptor of antigens (TCR). During the previous granting period we have characterized structures of several extracellular portions of such receptors, such as the adhesion domains of CD2, CD58 and a Fv fragment of the class II-specific alpha-beta-TCR Dl0, and we have elucidated the role of glycans in the adhesion domains of CD2 and CD58. Finally, we determined the structures of the CD2/CD58 complex and a complex between the Dl0 Fv fragment and a peptide/MHC Class II complex. The proposed continuation of this research is focused on structural and functional characterization of CD3 components of the TCR, the signaling components of the TCR, and on proteins that mediate signaling via the cytoplasmic tail of CD2. We will pursued via three specific aims: 1. Structure and interactions of the heterodimer: We will solve the structure of a recently engineered correctly folded single chain epsilon-gamma-CD3 construct using NMR spectroscopy. Based on the structure we will investigate interactions between the epsilon-gamma-CD3 heterodimer and the alpha-beta-TCR. 2. Structure and interactions of the epsilon-delta-CD3 heterodimer: Using similar strategies we will engineer single-chain constructs of the epsilon-delta-CD3 heterodimer and study its interaction with the alpha-beta-TCR and CD4. 3. Structural and functional studies of CD2-BP2: We will solve the structure of the complex of the GYF domain of CD2-BP2 with a fragment of the CD2 tail. We will also study larger fragments of or full-length CD2-BP2 and investigate their interactions with CD2 or potential other factors. This research will provide new insights into the mechanism of T-cell activation at the receptor-cytoplasm border.
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{
"pile_set_name": "NIH ExPorter"
}
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Our work focused on the use of live imaging techniques to examine the behavior of retinal microglia in living tissue. We employed ex vivo time-lapse confocal imaging techniques to visualize fluorescence-labeled microglia from transgenic CX3CR1+/GFP mice and follow dynamic microglia behavior in intact retinal explants in real time. Previous studies have demonstrated that microglia in the cerebral cortex demonstrate structural dynamism in their ramified processes, but whether this behavior extends to areas outside the brain has not been previously examined. Our imaging system enables us to follow detailed changes in the structure of individual microglial processes and to quantitate process and migration velocities in intact retinal explants. We have also examined dynamic microglial responses to focal laser treatment using parameters similar to those used in the grid laser treatment of diabetic retinopathy. Using live imaging, we found that under normal conditions, resting retinal microglia are not static in structure but instead exhibit extensive structural dynamism in their cellular processes, changing their structure at remarkably rapid rates on the scale of micrometers per minute. These processes extended in all directions, and appeared to sample the surrounding extracellular space in a random fashion. Despite marked dynamism, the overall area of the cell remained relatively constant;processes did not cross into a neighboring cells territory, and the position of cell bodies remain relatively fixed, and did not display any overt cellular migration. We found that this phenomenon was present in the retina both neonatal and adult animals and is a property of both developing and mature systems. We also examined how retinal microglia behavior changes in response to focal retinal injury. Focal photocoagulative injury using an argon laser was applied, and microglial behavior in the vicinity was recorded and analyzed. After injury, we found that the rate of microglia process movement increased significantly (67% increase) over baseline rates. Also, microglia in the injury vicinity directed processes preferentially toward the injury site, while withdrawing processes on the other side of the cell, hence adopting a polarized cellular phenotype. In addition, while resting microglia have stationary cell body positions, microglia post-laser injury acquire a migratory capacity, and are capable of translocating through tissue in a ramified state. The migratory speed of polarized microglia was on the scale of 0.5 microns/minute. In summary, retinal microglia normally occupying uninjured tissue display a continuous, dynamic behavior that suggests functions of tissue surveillance and intercellular communication. Microglial behavior is highly regulated by, and immediately responsive to, focal tissue injury and may constitute a therapeutic response to focal laser photocoagulation. Future work will focus on elucidating the cellular signals modulating and directing the dynamic behavior of microglia in terms of process movement and migratory dynamics.
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{
"pile_set_name": "NIH ExPorter"
}
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The prime goal of this core is to provide expert histopathology services, facilities, and resources for investigations involving cell and tissue microscopic analyses. Our Histopathology Core was established in 2004 with funding from the COBRE in Stem Cell Biology and Regenerative Medicine (8P20GM103465), with Dr. Volkhard Lindner as the Core Director. The success of this core has engaged all Center investigators, attracted many investigators from our Institute?s COBRE in Vascular Biology and Maine Medical Center, and a number of regional clients. Current services provided include the fixation and processing of specimens for frozen, paraffin and plastic embedded sectioning. A comprehensive spectrum of histological stains and specialty stains are offered. Detection of specific proteins by immunohistochemistry (IHC) is a key service, and our staff expertly develops and optimizes antigen retrieval, analysis, and versatile detection methods. This core facility has also developed and characterized highly specific rabbit monoclonal antibodies for the detection of several antigens where quality antibodies do not exist. In transitioning to Phase III, we outline several innovative features. First, we recently have developed the capacity to prepare custom tissue microarrays, and this is being applied to genetically engineered mouse models, human specimens, and mice generated from the Knockout Mutant Mouse Project (KOMP) at The Jackson Laboratory. This core will collaborate with the Mouse Transgenic Core of our sister COBRE in Vascular Biology to generate unique tissue arrays from this mutant mouse resource. In addition, Histopathology will collaborate with our Molecular Phenotyping Core (Oxburgh) to facilitate quantitative analysis of immunostained antigens. The ability to quantify both histological and immunostained specimens will significantly sharpen cellular and pathological phenotypic analysis of disease models and human pathological specimens. Thus, for Phase-Ill, our Histopathology Core is well-positioned to provide, and expand, comprehensive expert analyses of a broad spectrum of cell and tissue specimens.
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{
"pile_set_name": "NIH ExPorter"
}
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Project Summary: The Infectious Diseases Training Program at the University of Virginia is in its 29th year. The objective is to provide a rich interdisciplinary experience in infectious diseases research in order to prepare our trainees for careers as independent investigators. The cornerstone is the side-by-side education of predoctoral, M.D. and Ph.D. postdoctoral fellows. The rationale is that integration of clinical and basic science provides the varied perspectives and techniques required for creative research to combat infections. The design includes a Program Director reporting to an Executive Committee that selects predoctoral trainees from a pool of rising 2nd year graduate students, and selects postdoctoral applicants based on nominations from both mentors and the Infectious Diseases Division. Training is enriched by the monthly "ID Breakfast" for trainees chaired by the Director, graduate courses taught by Program faculty, an M.S. degree in clinical investigation (taken by all 9 fellows doing clinical investigation), seminars, research in progress and journal clubs. The 26 faculty are 100% NIH supported (median annual direct costs $648,000), interactive (65% co-published), senior (19/26 full Professors, although select "up and coming" junior faculty are included), from 7 Departments, with 20 PhDs, 10 MDs, and 1 DVM. The average mentor graduated 8 trainees in the last 10 years. Evaluation of the Program includes an annual faculty retreat and trainee questionnaire. Institutional support includes funding of 1st year predoctoral fellows, the seminar series and the Director's effort, and construction of a new research building. NIH support is requested for 6 predoctoral, 5 M.D. and 2 Ph.D. postdoctoral fellows (an increase of 1 predoctoral position). Progress since the last funding period includes increases in applicants (95% predoctoral and 43% postdoctoral), average GRE V/Q of 600/703 and GPA of 3.50, 100% retention of trainees in the Program, and a 44% increase in mentor NIH support. Peer-reviewed original publications averaged 4 for postdoctoral and 3 for predoctoral fellows. Academic and research positions are held by 91% (20/22) of postdoctoral and 100% (16/16) of predoctoral graduates, and 8 K08 and 4 foundation fellowships have been awarded. Women constitute 50% of trainees and 23% of mentors;13% of trainees are underrepresented minorities. Relevance: The Program prepares fellows to conduct research on infectious diseases, a paramount threat to international health in the 21st century.
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{
"pile_set_name": "NIH ExPorter"
}
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This application requests an Applied Biosystems model 477A/120A automated Protein Sequencer and a free-standing microbore HPLC (ABI model 130A) for interdepartmental use by investigators in the School of Medicine and Biomedical Sciences of the State University of New York at Buffalo. 'Mere is no protein sequencer in the School of Medicine. Nor is sequencing available routinely on a fee-for-service basis in the University at Buffalo, that can satisfy the needs of this user group. When samples are accepted, they are accepted on a "second" priority basis. Four applicants are having sequencing done outside of the University, and others are planning to do the same. The model 477A is requested, rather than the less costly model 473A, because of its 2-fold advantage in sensitivity and its greater versatility. The microbore HPLC will expedite optimum use of the sequencer and will enhance the sequencing facility's utility. The sequencer will be operated and maintained by a skilled technician with previous experience in operating an automated protein sequencer. The HPLC will be operated by individual users under the supervision of the PI. The technician's salary will be derived from state funds through the Department of Biochemistry, major repairs will be offset by the School of Medicine, other repairs and some operating costs will be provided by the Center for Applied Molecular Biology and Immunology, and the remaining routine operating costs will be obtained through user fees. A three-person internal advisory committee, consisting of the PI and two of the Co-PIs who have the most pressing need for this instrument, will govern the care and use of the instrument. The instrument will serve a core user group of 14 investigators, representing 18 projects, all supported by the National Institutes of Health, with the exception of one, who is supported by the NSF. The instruments will be in use about 90% of the time. The projects can be grouped into two categories, with several fitting into both. The first type concentrates on protein chemistry and enzymology. These users require protein/peptide sequencing routinely to derive functional information, such as the nature of active sites, precursor product relationships, or confirmation of sequences in site-directed mutagenized proteins. Six of the applicants have these needs (projects 1 through 5 and 13). The second type will use sequence information from tryptic digests of partially-purified enzymes/proteins to plan syntheses of oligopeptides and oligonucleotide probes for raising antibodies and cloning the cDNA of the genes of the target proteins. Ten of the applicants have this need (projects 1 and 6 through 18). A protein sequencer and dedicated microbore HPLC will enhance funded research by providing the capacity and accessibility required for optimum progress. An additional advantage is that the cost to the individual will be lower than other alternatives.
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{
"pile_set_name": "NIH ExPorter"
}
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The goal of this project is to construct an OLCR system utilizing a high powered low coherence source that provides a photon flux into the highly scattering biological tissues sufficient for acquisition of high resolution images. Results from OLCR are analogous to ultrasound B-scan except the imaging is performed optically instead of acoustically. OLCR is far superior to ultrasound in that the technique is non-contact, has high sensitivity, and exceptional spatial resolution in both the axial and radial directions. Other features of OLCR also make this methodology particularly attractive for industrial applications: (1) the high spatial resolution in the recorded tomographic images; and (2) potential for flow diagnostics of fluids. OLCR is ideally suited for industrial applications such as characterization of fibrous materials and various resin composites where rapid and non-contact tomographic imaging of constituents embedded in highly scattering material is required.
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{
"pile_set_name": "NIH ExPorter"
}
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The research deals with the structure and functions of leukocytes in various types of reactions. Studies are continuing on intracellular mechanisms that permit certain parasites to survive and multiply within macrophages, and on modifications in these mechanisms in cells from the immunized animal. Detailed observations are being made at the subcellular morphologic and biochemical level on the mechanisms involved in incorporation in macrophages of iron into ferritin, and in storage and mobilization of ferritin in these cells. Megakaryocytes have been isolated from guinea pig bone marrow. We are attempting to obtain information on morphology of the maturation of these cells, and on the mechanisms that underlie platelet formation. Studies are to be made on eosinophils, comparing their locomotion and chemotactic responses to those of neutrophils, in the hope of obtaining clues to eosinophil function.
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{
"pile_set_name": "NIH ExPorter"
}
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We have recently demonstrated that both white and black adolescents in two rural Kentucky counties have higher blood pressures than we had previously measured in either white or black adolescents in Washington, D.C. consequently, these Kentucky adolescents provide a unique opportunity to: a) identify demographic, physiologic, and psychosocial correlates of elevated blood pressure in young people, b) determine if young people with relatively high blood pressure have clinical evidence of cardiovascular disease, c) determine if blood pressure in adolescence is predictive of blood pressure at a later age, d) identify factors in young adolescents predictive of the development of elevated blood pressure in late adolescence, and e) determine if factors associated with relatively high blood pressure in a population are also associated with clinical hypertension. We propose to re-evaluate, at 5 and 7 years people who at age 16-19 years had blood pressure above the 90th percentile or below the 10th percentile for their sex-race group. Correlates and consequences of elevated blood pressure will be compared in these two groups. We have previously measured blood pressure in a population of 14-15 year old adolescents, and we propose to re-evaluate these young people to identify distinguishing characteristics of young persons who develop elevated blood pressure during adolescence and to determine if young people maintain the same relative blood pressure, compared to their peers, over time. We also propose to evaluate young people with clinical hypertension, to determine if distinguishing factors associated with high blood pressure in a population are also associated with hypertension in patients referred for clinical evaluation. We have previousy studied a group of pregnant adolescents homogenous in age and parity. This group provides the opportunity to determine if blood pressure during adolescent pregnancy correlates with blood pressure 3-6 years later. In addition, we will evaluate the factors associated with blood pressure of their 3-6 year old children.
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{
"pile_set_name": "NIH ExPorter"
}
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Integrin ?v?3 is expressed in the most aggressive and metastatic cancers. Here, new experimental data suggests that ?v?3 also drives a novel pathway of resistance to EGFR-targeted therapies. In fact, different epithelial carcinoma cell lines exposed to the EGFR inhibitors erlotinib or lapatinib for several weeks show enhanced expression of the integrin ?3 subunit in the subpopulation of surviving cells. Mechanistically, ?v?3 integrin promotes the plasma membrane clustering of oncogenic K-Ras to drive its signaling to RalB and its downstream effectors. This ?v?3/K-Ras/RalB oncogenic unit not only increases anchorage-independence in vitro and tumor growth in vivo, but it also renders tumors resistant to EGFR inhibition. These findings suggest that disabling components of this novel pathway may enhance the sensitivity to EGFR-targeted therapies. Experiments outlined in this proposal will examine the significance of the ?v?3/K-Ras interaction in terms of EGFR inhibitor resistance for a number of epithelial cancers in vitro and in vivo. The goal of Aim 1 is o define the structural basis for the association between integrin ?v?3 and K-Ras, which appears to drive RalB- mediated signaling in epithelial carcinoma cells. Studies in Aim 2 are designed to determine which ?3/K- Ras/RalB effectors drive anchorage-independence and erlotinib resistance, and to test both genetic and pharmacological strategies to block this pathway in vitro. Finally, Aim 3 will evaluate strategies to overcome EGFR inhibitor resistance for subcutaneous and orthotopic mouse cancer models. If successful, these studies will support the continued use of EGFR-targeted therapies in the clinic, and offer novel strategies to improve response to such therapies once tumors acquire resistance.
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{
"pile_set_name": "NIH ExPorter"
}
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Project 1: Finding pathogenetic steps. Accelerated plasma cell tumor formation occurs in BALB/c-Bcl-xL transgenic mice given a single intraperitoneal injection of pristane. This generates a chronic inflammatory tissue (oil granuloma) in which the tumors exclusively develop. 90+ percent of the mice develop a plasma cell tumor by day 100 in contrast to 60% at day 300 when three doses of pristane is given to BALB/c mice. The oil granuloma is invaded by large numbers of B-lymphocytes that form aggregates. Some of the lymphocytes that escape the aggregate become proliferating plasma cells. This is a new step or stage in plasmacytomagenesis. Fluorescence In Situ Hybridization of the plasma cells associated with these aggregates shows that IgH/C-Myc chromosomal translocations can be found by day 14 post pristane. Analysis of the B cells in the aggregates for Activation Induced Cytidine Deaminase is in progress. The basic question is whether the oncogenic chromosomal translocation takes place at this new late stage of development. Project 2: Inhibition of Plasma Cell Tumor Development The high predictable incidence of plasma cell tumors in pristane induced models makes it possible to test potential inhibitors. The nonsteroidal anti inflammatory agents indomethacin or sulindac administered continuously for 200-300 days in the diet are powerful inhibitors of pristane induced plasma cell tumor formation. In collaborative experiments with Jim Mitchell we have found the free radical scavenger Tempol substantially but not completely inhibits non-accelerated plasma cell tumor formation. PolyIC an interferon gamma stimulator in a first experiment with Sam Baron partially inhibits plasma cell tumor development. To test these effects in the accelerated model current experiments are designed to find the minimal amount of pristane required to initiate tumor formation and to maximize the inhibitory effects of Tempol and polyIC . Other shorter-acting agents such as squalene or alum that might be alternatives to pristane are being tested. The inhibition of IgH/C-Myc chromosomal translocations is also being tested by inducing plasma cell tumors in mice that are genetically defective for Activation Induced Cytidine Deaminase and Deoxy uracil glycosylase has shown former mice were partially but not completely resistant to developing IgH/C-Myc chromosomal translocations while the latter are strongly resistant. Thus, preventing chromosomal translocations can inhibit plasma cell tumorigenesis.
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{
"pile_set_name": "NIH ExPorter"
}
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The overall objectives of the proposed research are to define in molecular terms the initial biochemical and physiological responses of the neutrophil to chemotactic agents and to investigate the nature and role of activatable esterases (proteases) in the neutrophil and other cells. To this end we propose to study the structure activity relationships of formylmethionyl peptide for neutrophils, isolate the formylmethionyl peptide receptor and prepare antibodies to it. The calcium requirements for chemotactic peptide induced granule enzyme secretion and stimulated orientation and locomotion of neutrophils will be further studied. The role and nature of arachidonic acid metabolites involved in granule enzyme secretion in neutrophils will be studied using 14C-arachidonic acid followed by identification and isolation of metabolites. The role of neutrophil esterase-activation in phospholipase activation and production of arachidonic acid is to be investigated in neutrophils using the same approach employed for platelets. Antibodies to the purified chymotrypsin-like esterase of rabbit peritoneal neutrophils will be prepared and used to localize the enzyme in the cell. The substrate specificity of the esterase activated during the antigen-antibody induced locomotion of B lymphocytes will be studied using radioactive benzoyl L arginine methyl ester and protein substrates. Additionally, we shall seek evidence that the stimulus specific esterases activited during platelet serotonin release are the same as those that aid phospholipase activation in the same cell.
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{
"pile_set_name": "NIH ExPorter"
}
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Thymidylate synthase is a well-recognized target for anti-cancer, anti-fungal and anti-parasitic drugs. We have crystallized thymidylate synthase from the species Cryptococcus neoformans, Cryptosporidium parvum, Pneumocystis carinii and Toxoplasma gondii. We plan on doing structure-based drug design studies on all four species using anti-folate and lipophilic drug leads available in the laboratory.
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{
"pile_set_name": "NIH ExPorter"
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The broad goal of this research is to improve the diagnostic accuracy of clinical positron-emission tomography (PET) scans and the quantitative accuracy of research scans by accurately correcting for the effects of Compton scattering. Current methods of correcting for Compton scatter are lacking in three respects: (1) They assume that the scattering medium is homogeneous; (2) They treat scattering as a two-dimensional phenomenon, ignoring scatter from adjacent slices; and (3) They are empirically based, despite the fact that the physics of Compton scatter are well known. These shortcomings are more pronounced in cardiac scans, the scans that are currently of greatest clinical interest, for two reasons: First, scatter is a major source of image degradation in the chest cause of the relatively long path-lengths encountered there; and second, the lungs introduce significant heterogeneities in the scattering medium. The proposed method, termed model-based scatter correction, makes no assumptions about the scattering medium, takes the three-dimensional nature of scatter into account and is grounded in a model of the fundamental physics of Compton scatter. By appropriately formulating the method as a ray-tracing algorithm, the computational complexity is reduced by a factor of n over what might be expected, thereby making the algorithm computationally practical on currently available processors. A preliminary version of this algorithm has been developed based on several restrictive assumptions. Initial phantom studies suggest that a fully developed version of the algorithm will perform significantly better scatter correction than current approaches. The specific aims of this proposal are to: (1) Develop the preliminary version to the point that it can be used clinically; (2) Evaluate the performance of this method; (3) Extend the algorithm for use with fully 3D PET systems; (4) Evaluate the performance of the 3D model; and (5), Formulate the algorithm for use on high-speed processors. Methods include the use of mathematical modeling, phantom studies, and previously acquired clinical PET data. This project will be performed in cooperation with the departments of Neurology, Radiation Sciences, and Internal Medicine.
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{
"pile_set_name": "NIH ExPorter"
}
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Studies were continued on cells derived from the fetal rat lung alveolar epithelial cell line, FRLE, in which neoplastic transformation can be directly induced by quartz particles, without mediation by mesenchymal cells. The patterns of expression of TGF-beta mRNAs and proteins were reported in quartz-transformed cells and derived tumors. Additional tumorigenicity assays in nude mice were performed for quartz-treated and control cells. New studies were undertaken in order to investigate quartz-induced transformation mechanisms in mature alveolar epithelial cells derived from young adult rather than fetal rat lungs. Since rat alveolar type II cells usually cannot be maintained in culture for more than a few passages, they were transfected with plasmids containing immortalizing genes (either SV40 T antigen or adenovirus 2 E1A) and/or a selective marker. Transfected colonies, selected for morphological characteristics of epithelial polygonal cells forming pavement-like colonies, were expanded and cryopreserved at passage 5. These new cell lines have been passaged 15 times so far, and have maintained normal morphology. Characterization of the new cell lines and studies of quartz-induced cytotoxicity and transformation are under way. The SV40Tag-transfected cell line showed a much higher sensitivity to quartz-induced toxicity than the E1A-transfected cell line.
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{
"pile_set_name": "NIH ExPorter"
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The broad, long-term objective of this proposal is to understand the pathophysiology underlying altered pain perception in depressed individuals as a way of understanding the neural dysfunction in depression. The specific aims are to (1) test the effectiveness of our recently developed mood induction paradigm that will effectively induce depressed mood in both pain and fMRI experiments, (2) use phasic and tonic pain stimuli to rest the hypothesis that depressed mood potentiates the perception of pain, and (3) use fMRI to test the hypothesis that mood-altered pain perception is accompanied by activity changes in limbic and prefrontal regions. The health relatedness of the project is that an understanding of the underlying pathophysiology of altered pain in depression could lead to the development of new treatment strategies for both depressive disorders and chronic pain. The research designs are (1) test the effectiveness of a mood induction paradigm to induce depressed moods, (2) measure the ability of depressed mood to alter perceived intensity of both phasic and tonic pain stimuli and to reduce tolerance to tonic pain, and (3) use fMRI in phasic pain experiments to compare activity in limbic and prefrontal brain regions during depressed and non-depressed moods. The methods to be used are psychological mood scales, delivery of phasic electric shock and tonic cold pressor pain, and functional magnetic resonance imaging during delivery of pain stimuli in different mood states. [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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Project Summary/Abstract Over the past five years, my lab has made significant strides in the development of genetic systems capable of driving the rapid mutation and evolution of user-selected genes of interest in vivo. These systems have allowed us and others to quickly and scalably evolve enzymes, proteins, and antibodies to address a range of problems spanning from studying drug resistance to creating affinity reagents on demand. These systems have also begun to allow us to use rapid mutational accumulation as a method for tracing cell lineage in developing animals. One of our key accomplishments has been the invention of an orthogonal DNA replication system (OrthoRep). In OrthoRep, an error-prone orthogonal DNA polymerase (DNAP) exclusively replicates a special cytosolic plasmid encoding only genes of interest (GOIs), driving their continuous evolution fully in vivo. This MIRA will integrate our lab?s work on OrthoRep and support its further development and application in the next five years. In particular, we will grow the core OrthoRep technology in order to accelerate GOI evolution in yeast even more than we currently have, attempt to establish OrthoRep in mammalian cells in order to extend the range of problems OrthoRep can directly address, apply OrthoRep to the engineering of bespoke Cas9s to extend the range and efficacy of targeting, apply OrthoRep to improve the capabilities of a lineage tracing tool developed by my lab, and apply OrthoRep to the generation of mutually orthogonal collections of aminoacyl- tRNA synthetase (aaRS)/tRNA pairs to support genetic code expansion efforts also ongoing in my lab. We hope the set of activities proposed for this MIRA will solidify OrthoRep as an exceptionally powerful genetic system for evolving enzymes and proteins capable of solving high-reward problems in the chemical, biological, and biomedical sciences.
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{
"pile_set_name": "NIH ExPorter"
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Ceramide glycosylation determines the stemness of cancer stem cells Drug resistance with disseminated metastases is a cause of cancer death; in breast cancer, it ultimately claims approximately 40,500 American women per year. Tumors often progress rapidly once treatments fail, and enrichment of these tumors with cancer stem cells generally correlates with poor patient prognosis. Thus, cancer stem cells that initiate tumorigenesis and drive tumor progression might be a root cause of chemotherapy failure. Cancer stem cells are distinguished from normal adult stem cells by their tumorous behaviors. It is well known that anticancer drugs can kill normal stem cells, such as bone marrow stem cells-a source of severe side-effects; in contrast, however, some evidence suggests that anticancer drugs might act to expand cancer stem cell fractions or numbers. Our long-term goal is to understand how cancer stem cells can be specifically manipulated for preventive and therapeutic purposes. The objective of the research proposed herein is to determine how selected anticancer drugs affect breast cancer stem cell (BCSC) populations during the course of chemotherapy. It has been reported that glucosylceramide synthase (GCS) is overexpressed in metastatic breast cancers, and is a risk factor predicting poor chemotherapy response. Embryonic stem cells crucially require ceramide glycosylation catalyzed by GCS in order to retain their pluripotency. Little is known, though, concerning the possible involvement of GCS in regulating cancer stem cells. Our hypothesis is that ceramide glycosylation by GCS is a key step enabling tumor enrichment with BCSCs, and that enrichment with cancer stem cells drives rapid tumor progression and metastasis once chemotherapy fails. This application is formulated from our own preliminary data and previously published work, in which it was found that GCS is over-expressed in metastatic breast cancer, and that silencing of GCS was able to reverse drug resistance as well as prevent BCSC enrichment. We will: (1) identify the effects of anticancer drugs on enrichment of BCSCs and apoptosis of normal stem cells; (2) determine GCS-catalyzed ceramide glycosylation is a key process governing the accumulation of BCSCs and apoptosis of normal stem cells; and if so, (3) determine the signaling pathway(s) by which ceramide glycosylation promotes BCSC formation. Findings from in-vitro studies will be further explored in vivo, in orthotopic tumors exposed to anticancer drugs, employing gene silencing and overexpression strategies, and focusing on proposed role(s) of GCS in increasing the formation of BCSCs vs. contrasting apoptotic effects on bone marrow stem cells and normal mammary epithelial stem cells. Results will be further corroborated via studies of tumor and bone marrow samples from patients undergoing chemotherapy. Lastly, the therapeutic efficacy of disrupting ceramide glycosylation by an antisense oligonucleotide or a small molecule in order to effect reductions in BCSCs in orthotopic tumors will be examined, wherein success will lend validity to this unique approach for selectively eliminating cancer stem cells via deletion of key glycosphingolipids. Completion of the proposed studies will introduce new concepts and advanced technologies in cancer stem cells and drug discovery to students who participate, thus greatly enhancing their biomedical sciences education.
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{
"pile_set_name": "NIH ExPorter"
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PROJECT SUMMARY (See instructions); The mission ofthe Nonhuman Primate (NHP) Core (Core H), located at the Tulane National Primate Research Center (TNPRC), is to provide to Penn CFAR investigators highly integrated clinical and laboratory resources and expertise in using NHP models relevant to AIDS. The Core is lead by Andrew Lackner with Preston Marx, Ronald Veazey, and Marcelo Kuroda as additional key personnel. Core H adds value to the CFAR by enhancing and facilitating the ability of CFAR investigators to perform HIV/SIV-related studies in NHPs, providing mentorship and on-site training to junior investigators, and promoting scientific collaborations between the TNPRC and CFAR investigators. Specific goals are to (1) provide oversight and assistance in all phases of research using NHPs, supporting CFAR investigator projects and proposals and providing resources, assistance with experimental design, and compliance with all animal welfare and biosafety requirements; (2) provide housing, clinical care, animal husbandry, and other routine and necessary services for NHPs assigned to CFAR projects; (3) provide technical expertise and laboratory support for studies utilizing NHPs; (4) support a NHP Pilot Grants Program (separate from but complementary to the Developmental Pilot Program run by Core B) designed to encourage junior and new investigators to use NHP models; (5) provide education, training and mentoring to junior and new investigators who have not previously worked with NHP; and (6) stimulate and facilitate translation of bench-based findings to NHP experimentation. Collaborations between Core H and the CFAR are facilitated by jointly sponsored symposia focused on NHP HIV/SIV research and a seminar series integrated with TNPRC via video-conference links. Core H includes specialized clinical, laboratory and consultative components. The Core performs a variety of specialized services including in situ hybridization, immunohistochemistry, confocal microscopy and image analysis. Core H also participates in the CFAR's mentoring mission by training and educating CFAR members and their students and postdocs on use of NHP models. Core performance is evaluated as part ofthe CFAR's annual strategic planning process. Core H will also play an active role in working with the CFAR's newly formed scientific working groups. In the current cycle Core H has supported 12 NHP Pilot Projects, closely worked with 20 CFAR investigators, provided onsite training for 6 students and/or post docs, and contributed to 20 publications, and 8 NIH grants.
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{
"pile_set_name": "NIH ExPorter"
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The dorsal motor nucleus of the vagus (DMV) provides the parasympathetic motor output to the gastrointestinal (GI) tract and plays an integral role, along with the nucleus of the tractus solitarius (NTS), in the reflex control of gastric motility and compliance. In addition, each of these nuclei receive descending projections from hypocretin (HCRT)- containing neurons in lateral hypothalamus and oxytocin (OT)-containing neurons of the paraventricular nucleus (PVN) that together provide both the means for central control of GI activity and an important route for coordinating digestive processes with feeding behavior. A major objective of the proposed research is to define the cellular and synaptic actions of HCRT peptides and OT in the DMV and ascertain how they relate to the more global influence that these peptidergic inputs have on gastric-related functions. To accomplish this, we will conduct in vivo experiments and obtain patch clamp recordings in brain slices and cell culture from retrogradely labeled DMV neurons with defined GI projections. In Aim 1, we will conduct microinjection experiments in anesthethized rats together with immunohistochemical studies to determine the sites and cellular substrates where HCRTs act in the DMV to stimulate gastric motor function. The pharmacological sensitivity of gastric responses to HCRT will also be examined to determine the nature of the vagal efferent pathways involved. In Aim 2, patch recordings in slices will be used in conjunction with electrical stimulation in NTS to determine the effects of bath application of HCRTs or OT on the membrane properties and synaptic responses of identified GI-projecting DMV neurons. The neurochemistry of each sampled neuron will then be defined using single cell reverse transcription -polymerase chain reaction. Responses of individual cells to HCRT or OT application will be correlated with their specific GI target and chemical phenotype to test the hypothesis that these peptidergic signals exert opposing effects on gastric motility and tone by modulating the excitability of functionally distinct subpopulations of preganglionic vagal motor neurons. In Aim 3, we will obtain whole-cell recordings from identified GI-projecting DMV neurons in culture to characterize the voltage dependent calcium (Ca 2+) conductances in these cells and determine whether HCRTs regulate excitability of neurochemically distinct groups of preganglionic neurons by modulating Ca 2+ influx through specific channels types. The proposed research will increase our understanding of hypothalamic mechanisms that control gastric-related functions, and in so doing may lead to the design of new interventions for effective weight management and treatment of digestive disorders such as gastric stasis or non-ulcer dyspepsia associated with alterations in gastric motility and stomach emptying.
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{
"pile_set_name": "NIH ExPorter"
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The mechanisms governing the transition from a terminally differentiated oocyte to a totipotent zygote remain poorly understood. During the previous funding period, we cloned and characterized a highly-abundant oocyte and embryo-restricted gene, peptidylarginine deiminase 6 (PADI6), from the murine egg proteome and found that it represents a novel member of an enzyme family (PADI) that converts protein arginine residues to citrulline. Our ongoing analysis of the PADI6 -/- phenotype indicates that PADI6 -/- females are infertile due to an arrest in embryogenesis at the two-cell stage, thus demonstrating that PADI6 represents a novel maternal effect gene. At the ultrastructural level, PADI6 localizes to an egg and embryo-restricted structure, the cytoskeletal sheets (CSS), that occupies over 15% of the cytoplasm in many mammals. While the function of this cytokeratin-containing structure is unknown, the CSS form during oocyte growth and undergo dramatic reorganizations at critical developmental time points;suggestive of a role in embryonic reprogramming. Strikingly, preliminary ultrastructural analysis reveals a complete dispersal of the cytoskeletal sheets in PADI6 -/- oocytes and eggs while most other structures appear normal. We also found that transcriptional activity in PADI6 -/- two-cell embryos is severely compromised, suggesting that the PADI6 defect arises due to failure to activate embryonic transcription. Based on our preliminary findings, we hypothesize that, in the oocyte, PADI6 interacts with cytokeratin via its N-terminal domain leading to nucleation of the dispersed CSS components. Following fertilization, we hypothesize that PADI6 becomes activated (most likely by calcium signaling) and citrullinates cytokeratin via its C-terminus, leading to CSS reorganization and a stage-dependent release of associated proteins which are required for embryonic genome activation (EGA). The specific aims of this application are to: 1) Test the hypothesis that PADI6 is required for nucleation of CSS components into the mature 60 nm complex in the oocyte. 2) Test the hypothesis that PADI6 plays a role in cytoplasmic-to-nuclear signaling events in the early embryo prior to genome activation. 3) Test the hypothesis that citrullination of cytoskeletal sheet proteins by PADI6 is required for CSS anastomoses and for early development.
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{
"pile_set_name": "NIH ExPorter"
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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Focal hand dystonia (FHD) is a disorder characterized by involuntary contraction and co-contraction of muscles of the hand. There is growing evidence that a single treatment of rTMS can transiently modify brain excitability and produce temporary symptom improvement in people with FHD. The specific aim of this proposal is to collect pilot data regarding the efficacy of longer-term rTMS treatment vs. sham rTMS in ten subjects with FHD.
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{
"pile_set_name": "NIH ExPorter"
}
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An animal model of human syphilis has been developed, where inbred Syrian hamsters infected with Treponema pallidum develop lesions like "hard chancre" at the site of inoculation (inguinal area) which eventually heal, then animals develop oral ulcers corresponding to mucosal lesions of human secondary syphilis. The proposed research will examine the underlying cellular mechanisms of immunosuppression associated with acute and chronic syphilitic infection. Specific B lymphocyte, T lymphocyte and macrophage cell functions will be assessed in hamsters at various stages of infection. Conclusions will be made concerning the level of immunosuppression (i.e., whether B, T, or macrophage cells are affected), the nature of immunosuppression (i.e., whether it is passive or active) and the cause(s) of immunosuppression (development of treponema specific suppressor T cells or antiidiotypic antibodies and/or non-cytophilic or non-opsonizing antibodies). Elucidation of the specific cellular mechanisms involved in the development of secondary and tertiary syphilis may ultimately enable researchers to correct the deficiencies by immunotherapeutic means.
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{
"pile_set_name": "NIH ExPorter"
}
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Parkinson's disease (PD) and related Lewy body disorders (LBD), including incidental Lewy body disease (ILDB), Alzheimer's disease with Lewy bodies (ADLB) and dementia with Lewy bodies (DLB) may affect up to 40% of the elderly population. Current FDA-approved therapies for PD are still based on neurochemical deficits discovered more than 40 years ago. Experimental model-systems and molecular genetics research since that time, while contributing exponentially to the knowledge base, have not converted this work into new approved agents. Renewed and intensified human tissue-based research may help break this stalemate. Essential to such studies, however, is the availability of high quality diseased and normal control tissue with maximal preservation of molecular entities and detailed clinical characterization. The Brain and Body Donation Program (BBDP) at Banner Sun Health Research Institute is uniquely suited to fill this critical need, as demonstrated by its 20 years of operation and a median 2.8 hour postmortem interval (PMI) for the entire collection of 1,200 brains. Other unique features include a focus on normal aging, comprehensive standardized antemortem cognitive, motor and non-motor assessments and the accompanying donation of bodily organs. The BBDP has the proven capability of serving as a national resource. In the most recent 5 year period it has supported 91 different researchers located in 20 different states, collectively holding 126 NIH grants. There is no other brain bank in the US or world with these unique capabilities. This proposal would enable the BBDP to become a powerful resource for researchers focused on PD and related disorders. The Specific Aims are directed at supporting a major expansion of service to researchers investigating PD and other LBDs. To accomplish this, resource availability will be publicized, additional staff will be hired and a web portal will be developed so that external researchers may search for and request tissue as well as linked clinical data. Steering and Resources Committees with external investigator representation will manage the resource and evaluate requests, allowing the BBDP to supply between 50 and 100 LBD-focused research projects over the five year funding period. PUBLIC HEALTH RELEVANCE: Parkinson's disease, dementia with Lewy bodies and other Lewy body disorders (LBD) may affect up to 40% of the US elderly, causing major movement and mental disabilities for affected individuals as well as economic hardship for families. This proposal would seek to assist between 50 and 100 LBD-focused research projects by supplying them with critically-needed human tissue that is not available elsewhere.
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{
"pile_set_name": "NIH ExPorter"
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The title of this SBIR Phase II research and development contract award is "Thin-Film Peripheral Nerve Electrode." The project objective is to develop a thin-film neural cuff electrode and to demonstrate the efficacy of the electrode for grasp in an in vivo study using a raccoon model. The project will focus developmental research efforts on: i) fabrication of 4-electrode nerve cuffs; 2) sputter deposition of iridium electrode sites; 3) optimization of vacuum deposition processes; 4) optimization of AIROF activation conditions; 5) fabrication of 12-electrode nerve cuffs; 6) evaluation of explanted electrodes; and, 7) acute in vivo studies of implanted 4-electrode nerve cuffs on the raccoon median nerve.
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{
"pile_set_name": "NIH ExPorter"
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The goal is to develop a safe, potent, monoclonal antibody (MAb) product with neutralizing activity against variola (smallpox) virus. This immunotherapeutic product primarily is intended for prevention of infection in the face of an immediate threat of smallpox attack and for treatment to suppress infection and disease in case of exposure to smallpox. The neutralizing MAbs will provide immediate immunity, which might be crucial for providing protection to individuals that are immune-compromised or at risk for smallpox vaccination. The neutralizing MAb product might also be used as a replacement for VIG (vaccinia immune globulin) in the treatment of complications of smallpox vaccination. The product will consist of a cocktail of high affinity MAbs that are directed against critically important antigens located on the surface of EEV (extracellular enveloped virions) or IMV (intracellular mature virions) forms of the virus. IMV are released after cell lysis, are stable in the environment, represent the majority of infectious particles and play a predominant role in host-to-host transmission. By contrast, EEV have an additional outer membrane, are released from infected cells, are unstable in the environment, but appear to be responsible for direct cell-to-cell and long-range virus spread within a host. The studies described in this proposal will specifically test the hypothesis that combinations of MAbs against IMV and EEV antigens will act synergistically and have greater virus neutralizing activity than MAbs directed against single antigens. The specific targets of the MAbs in this proposal are B5R and A33R (EEV-specific proteins) and A27L and L1R (IMV-specific proteins). The first aim is selection of the MAb cocktail that provides the greatest neutralizing activity in mice (vaccinia virus challenge) and in non-human primates (monkeypox virus challenge). The second aim is completion of process development for the selected MAbs. The third aim is cGMP manufacturing of the MAb cocktail drug product. The fourth aim is completion of stability, safety and toxicology studies to support the filing of an IND for a Phase I safety study in human volunteers. The project is relevant to public health in the area of biodefense. The anti-smallpox antibody product is intended to be used to prevent smallpox infection in the face of an immediate threat of smallpox attack. It is also intended to be used to suppress infection and disease in individuals who have been exposed to smallpox as a result of an attack.
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{
"pile_set_name": "NIH ExPorter"
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Description: The Mass Spectrometry Center is equipped with (1) a VG/Micromass 7070E double focusing mass spectrometer with electron ionization (EI), chemical ionization (CI), and fast atom bombardment (FAB) ionization modes that may be operated in either the positive-ion or the negative-ion mode, equipped with a continuous-flow FAB probe, (2) a VG/Micromass gas isotope ratio mass spectrometer, (3,4) two Hewlett-Packard MSD capillary gas chromatograph-mass spectrometer systems with low resolution, electron ionization, positive-ion mode GC/MS operation only, and (5) a Hewlett-Packard 5989A MS engine capable of electron ionization and chemical ionization, equipped with an HP 5890 gas chromatograph and auto injection. The laboratory also performs organic synthesis of stable-isotope-labeled and other specialty chemicals. About 10 percent of the activity of the MS Center serves this MRRD program.
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{
"pile_set_name": "NIH ExPorter"
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DESCRIPTION (Adopted from the Applicant's Abstract): The aryl hydrocarbon receptor (AhR) is a member of the growing family of Per/ARNT /Sim transcription factors which modulate a wide variety of cellular functions. Historically, the function of the AhR has been evaluated in the context of its activation by environmental chemicals such as polycyclic aromatic hydrocarbons (PAH). These studies associate AhR activation with reactive metabolite production, proto-oncogene induction, induction of other transcription factors, and aberrant cell growth. More recent studies strongly suggest that the AhR can be constitutively active in the absence of exogenous ligands. A consequence of this activation is postulated to be dysregulation of cell growth. Among other results, this hypothesis is supported by: 1) nuclear localization and high level expression of the AhR in rodent and human tumors, 2) physical association of the AhR with RelA, an NF-kB family member which regulates proto-oncogene transcription, 3) up-regulation of the c-myc promoter and c-Myc protein after AhR and ReIA gene transfection in mammary tumor cell lines, and 4) inhibition of human mammary tumor cell growth with AhR antagonists and inducible AhR antisense cDNA. Accordingly, the central hypothesis of this project is that high levels of potentially active AhR in mammary tumors influences cell growth. Two specific aims have been defined to test these hypotheses: 1. Generate transgenic mice in which high level AhR expression is directed to mammary tissue. Studies with these mice will define the role of the AhR in epithelial cell transformation in vivo and in normal mammary gland development. Furthermore, the mice produced will facilitate studies defining the molecular mechanisms of transformation in the presence and, potentially, the absence of exogenous AhR ligands. 2. Determine AhR function in transformed human breast cancer cell lines. In addition to extending results to human systems, these studies will elucidate molecular mechanisms through which the AhR is postulated to influence cell growth. Particular emphasis will be place on potential interactions between the AhR signaling pathway and regulation of cell cyclins, Rb, NF-KB, and c-Myc, and on the composition of the AhR complex in mammary tumors.
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{
"pile_set_name": "NIH ExPorter"
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More than 575,000 Americans have end-stage renal disease, which is typically complicated by anemia, and most of these patients receive maintenance dialysis treatment. The proposed work will compare the effectiveness and safety of darbepoetin alfa, a novel injectible drug for the treatment of anemia, with its established cousin, epoetin alfa. Similarly, ferumoxytol, a new intravenous iron supplement for the treatment of anemia will be compared with an established iron supplement, iron sucrose. Both drugs were approved using information from randomized studies that evaluated their efficacy on reducing anemia. Very limited information, however, is available for either drug regarding their longer-term efficacy and safety. We are proposing to study the effectiveness and safety of these two medications when compared to their established cousins. Events of scientific interest include short-term outcomes (anaphylaxis, laboratory parameters including hemoglobin, iron saturation, ferritin) and longer- term outcomes including infection, cardiovascular events, cancer, and mortality. We will use already collected data from Medicare billing claims and electronic medical records of dialysis providers to address these research questions. Since facilities choose among these treatment options based on what is available in their formulary, and not necessarily based on patient characteristics, we can exploit natural experiments that occur when facilities switch their ESA or iron agent for their entire patient population. We will apply modern epidemiological techniques to eliminate bias and aspire to provide valid estimates of relative benefits and risks. Findings from the proposed work have the potential to immediately impact and improve the care that patients with end-stage renal disease receive. Our results may improve the quality of care received and, thus, the outcomes of this vulnerable patient population. The aims and scope of work are in full congruence with the mission of the National Institutes of Diabetes and Digestive and Kidney Diseases, and more specifically the Division of Kidney, Urologic, and Hematologic Diseases, which will consider this application for funding.
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{
"pile_set_name": "NIH ExPorter"
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The mechanism of leukocyte activation by chemotactic factors has been studied using electrophysiology, fluorescent probe, surface charge and ultrastructural techniques. Studies assessing the mechanism of modulating leukocyte locomotion indicate that limited secretion of specific granules, which accompanies chemotaxis, is associated with increased cell adhesiveness and increased availability of chemoattractant receptors. Vigorous exocytosis is associated with depressed chemotaxis, decreased availability of chemoattractant receptors, chemoattractant hydrolysis by secreted products and markedly increased cell adherence and aggregation. Human pyrogen has been shown to be a potent stimulator of neutrophil exocytosis and activation of the hexose monophosphate shunt. Studies of the two populations of neutrophils we had identified previously indicate that during the neutropenia that follows in vivo endotoxin or hemodialysis, a subpopulation of neutrophils with poorly demonstrable Fc receptors is the predominant neutrophil left in the circulation. Clinical studies assessing the effect of pharmacologic agents on the neutrophil subpopulations in normal subjects, patients with recurrent infection and host defense defects are underway.
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{
"pile_set_name": "NIH ExPorter"
}
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Sudden cardiac death annually claims approximately 300,000 American lives. Its very name underscores the urgent need for an accurate methodology to assess an individual's risk of experiencing unstable cardiac propagation that may result in malignant arrhythmias. Our initial work has identified an inexpensive, sensitive diagnostic procedure to perform that assessment. Our proposed research will further explore the biophysical characteristics of unstable cardiac electrical propagation and suggest criteria useful for non-invasively assessing the risk of ventricular arrhythmias. QT and RR interval spatial and temporal variations are known to be intimately related to the stability of cardiac conduction. Our preliminary theoretical and experimental studies demonstrate that the stability can be assessed by measuring QT and RR intervals collected during an ECG stress test employing a proprietary quasi-stationary exercise protocol (i.e., with gradually changing exercise loads). This is a hypothesis that we propose to test by studying a mutual interdependence of the QT and RR interval fluctuations, which accompany the dynamics of the heart rate gradual trends during the quasi-stationary exercise. We will evaluate quantitative criteria for unstable cardiac conduction that are suggested by the physics and may serve as the basis for a new, non-invasive diagnostic tool for evaluating the risk of cardiac arrhythmias.
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{
"pile_set_name": "NIH ExPorter"
}
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Pancreatic cancer is a devastating disease in need of novel therapeutics. To this end, the molecular hallmark of pancreatic cancer is an activating mutation in the small GTPase KRAS. Unfortunately, it has been difficult to pharmacologically inhibit KRas, prompting us to identify druggable proteins required for KRas oncogenesis. To this end, we discovered eNOS, one of three proteins in the Nitric Oxide Synthase family (composed of eNOS, nNOS and iNOS) that generate nitric oxide (NO), mediates oncogenic KRAS tumorigenesis. Specifically, eNOS is activated by oncogenic Ras, and moreover, knockdown of eNOS in human pancreatic cancer cell lines, or genetic ablation of the eNOS gene in mice, greatly retarded oncogenic Ras-driven tumor growth. Most importantly, small molecular weight inhibitors of NOS enzymes have been developed and tested in human clinical trials of septic and cardiogenic shock. Capitalizing on the development of NOS inhibitors for the treatment of other diseases, we tested and found that one such general NOS inhibitor impeded tumor growth of human pancreatic cancer cell lines and extended survival in the most aggressive mouse model of pancreatic cancer. Thus, eNOS is a bona fide target for pancreatic cancer that is druggable. There are no eNOS-specific small molecular weight inhibitors. While general NOS inhibitors are attractive molecules to explore, at least initially since they have already been tested in humans, they nevertheless also inhibit iNOS and nNOS. Two arguments suggest the importance of developing eNOS-specific inhibitors. First, eNOS is the NOS family member involved in pancreatic cancer. Second, as knockout of progressively more NOS family members results in more severe phenotypes, general NOS inhibitors may have undesirable off-target effects compared to inhibitors that specifically target eNOS. Given this, we propose to screen for eNOS inhibitors. We have developed a cell-based assay suitable for high throughput analysis that differentiates between cellular eNOS, iNOS and nNOS enzyme activity. We now propose to utilize this cell-based assay to screen a library of small molecules for their ability to inhibit cellular eNOS preferentially over nNOS and iNOS. Completion of these studies will provide novel eNOS inhibitors as a first step to specifically target this enzyme in pancreatic cancers. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is a devastating disease in need of novel therapeutics. In this regard, our efforts to identify small molecular weight inhibitors of eNOS, an enzyme that is required for pancreatic tumorigenesis, provides a new avenue to treat this fatal disease.
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{
"pile_set_name": "NIH ExPorter"
}
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Regulated membrane fusion is essential to many cell processes and to the life cycle of lipid-sheathed viruses such as HIV, Influenza, and Ebola. Several proteins ("fusion machines") involved in neurotransmitter release and enveloped viral infection have been identified and characterized structurally. Still, how these proteins catalyze fusion is not fully understood. The Lentz lab studies lipid rearrangements associated with fusion between synthetic membranes. The approach is first to define these rearrangements in pure lipid systems and then to ask how fusion proteins might promote them. Fusion requires close contact between membranes, which is induced using the inert polymer poly(ethylene glycol) (PEG). Fusion between lipid vesicles aggregated by PEG is shown to be minimally a three-step process (contacted bilayers D "stalk" intermediate D "diaphragm" intermediate D pore) that mimics biomembrane fusion. The Lentz group calculated, using the mechanical properties of lamellar lipid phases, the free energy reaction profile of this "stalk" fusion mechanism and showed it to be consistent with their unique studies effusion kinetics. Experiments and calculations show that the free energies of bent lipid monolayers and of defects between non-lamellar and lamellar structures (hydrophobic interstices) dominate the fusion process. Most researchers have focused on the bending energy to explain fusion. The Central Hypothesis is that fusion proteins catalyze fusion not just by altering bending energy but also in good measure by stabilizing hydrophobic interstices. To test this hypothesis, the project will address seven Specific Aims: 1] Compare the abilities of different bend-inducing lipids to partition into or promote bent lipid structures, and 2] alter the fusion reaction mechanism; 3] Determine whether an infection-blocking mutation in the membrane spanning domain of HIV alters the structure of a synthetic membrane spanning domain peptide, alters membrane structure, and alters the effect of this peptide on fusion; 4] Determine whether mutations in a key region of Influenza virus (the "fusion" peptide) alter a} the structure of a synthetic fusion peptide, b} membrane structure, and c} the effect of this peptide on fusion; 5] Determine whether neurotransmitter- release-blocking mutations in the membrane-spanning region of a neuronal fusion protein (syntaxin) alter a} the structure of a synthetic membrane spanning domain peptide, b} membrane structure, and c} the effect of this peptide on fusion; 6] Determine by electron microscopy whether fusion pores form at the point of contact between membranes held in contact by neuronal fusion proteins; and 7] Determine the ability of a neuronal calcium-binding protein (synaptotagmin) to perturb, and trigger fusion between, model membranes brought [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Vertebral malformation associates with a large number of distressful human birth defects. Elucidation of the developmental and genetic mechanisms underlying normal vertebral formation will undoubtedly have great impacts on human health. The formation of vertebral system is a complex process involving the morphogenesis of neural tube, notochord, somites and cell [unreadable]cell signaling events among these tissues. In particular, the formation and differentiation of somites play a critical role in vertebral development and vertebral disorders. Each somite has to establish an anterior-posterior (A-P) polarity and disruption of this A-P polarity during development often leads to the abnormal fusions between vertebrae bodies and ribs resulting in serious vertebral and spinal cord disorders. The development of vertebrate tail provides an excellent model system to study somite formation and differentiation due to its simple anatomic structure compared with the trunk vertebral system. The abnormal fusion between vertebrae bodies usually cause tail bending, kinky tail, that can be easily visualized. The A-P differentiation of somites can be examined by a set of well established molecular markers. In this study, we will focus on the regulation of TGF-b signaling pathway in somitogenesis by exploring the functions of two Smads interacting factors: Zfhx1a and TGIF. Zfhx1a, also called dEF1, encodes a zinc finger protein that contains multiple DNA and protein binding domains including homeodomain and Smad binding domain. In vitro studies showed that members of Zfhx protein family can bind to Smads proteins and recruit transcription activator p300 or repressor CtBP. Depending on the types of recruited co-factors, activators or repressors, Zfhx proteins can either up- or down- regulate TGF-b activity. Biochemical studies have revealed that TGIF antagonizes TGF-b signaling by binding to Smad2. Our recent studies with Zfhx1a and TGIF mutant mice uncovered that loss of Zfhx1a or TGIF function leads to a severe kinky tail phenotype, strongly indicating that these genes play important roles the A-P differentiation of somites. We therefore propose here to examine the functional requirements of Zfhx1a and TGIF during mouse somitogenesis and vertebral development. In addition, we will employ Zebrafish system to determine the specificity of Zfhx1a and TGIF mediated TGF-b suppression in vivo, namely which TGF-b member(s) activity is regulated by Zfhx1a and TGIF in vertebrate somitogenesis. Results from the proposed research will be significant as it address the regulation of an important pathway, TGF-b signaling, in a fundamental biological process, vertebrate body segmentation. In addition, the project features several advantages that fit the purpose of this COBRE application suitably: 1) It is a suitable project for the investigator to enter a new area of birth defect research, vertebral disorders;2) An additional model system, Zebrafish, will allow the investigator to conduct more mechanistic studies as Zebrafish is more accessible to embryonic and genetic manuscript;3) Since Zebrafish system is a popular and powerful model system in developmental biology, having a zebrafish model system in the laboratory will improve the research strength to a great extend.
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"pile_set_name": "NIH ExPorter"
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This is a renewal of a K24 Career Investigator Award on sleep disturbances in alcohol-dependent patients. Insomnia is common, frequently persistent in newly recovering patients, and associated with relapse. In the first 5 years, the investigator acquired knowledge about sleep medicine, sleep research methods, traditional sleep stage scoring, and pharmacotherapy trial methods. He then designed and conducted a randomized, controlled pilot trial to study night-time administration of the sedating anticonvulsant drug, gabapentin, for its effects on both sleep and relapse to alcohol. This work produced the first evidence from a randomized controlled trial that gabapentin was superior to placebo for reducing time to relapse. Although robust effects on traditional measures of subjective and objective sleep were not found, analysis of sleep microarchitecture revealed that gabapentin increased delta (slow frequency) activity during rapid eye movement sleep, the period of sleep during which the cortex is paradoxically activated. Thus, gabapentin may have exerted its effect on daytime abstinence by slowing cortical hyperactivity during sleep. The current proposal builds on this work by investigating potential mechanisms for sleep disturbance in alcohol-dependent patients that could be targeted by medications. A novel sleep laboratory paradigm is described (and will be refined) that can inform the field about mechanisms of alcohol-related sleep disturbances as well as screen for medications that improve sleep. To demonstrate proof-of-concept, two medications with known potential for treating either alcohol dependence or insomnia (gabapentin, trazodone) will be studied for their short-term effects on disregulated sleep in alcohol-dependent patients. Training will comprise 10-15% effort for 5 years including mostly time in the sleep lab, but also local and national conferences on the neurobiology of sleep and alcoholism, and selected readings. Another 5-10% effort is for training new investigators supported by T32 and other fellowships. With Drs. Armitage and Hoffmann, the candidate will acquire new knowledge and skills about quantitative sleep EEG analysis and interpretation, as well as methods for measuring and analyzing homeostatic sleep drive and circadian rhythm regulation. He will apply these methods to alcohol dependent patients, an understudied population that may benefit clinically from a more sophisticated understanding of their sleep problems.
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{
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The objective of this application is to develop the career of Dr. Christian Badr to facilitate the transition to a stable independent phase and attain his ultimate goal to become an independent scientist and establish his own research program. The career development plan created by the candidate will contribute substantially to his scientific development and further his training in proteomics, RNAi and cancer stem cells biology. The proposed opportunity in the K22 will provide the PI protected time while starting his independent Faculty position and direct him towards a successful career in developing therapeutics for brain tumors. The proposed research will also serve as the foundation for an R01 proposal prior to the end of the second year of this award. Glioma stem cells (GSC) represent a subset population within malignant gliomas, and are highly tumorigenic, intrinsically resistant to current therapies, and capable of self-renewal and differentiation into mature glioma cells (GCs). There are two major subtypes of GSCs: Proneural (PN) and Mesenchymal (MES), the latter being more aggressive and more resistant to conventional treatment. We have identified a subpopulation of GSCs, termed floating cells (FC) that could be isolated from in vitro-cultured glioma cells. These cells possess a stem-like signature with prominent mesenchymal features. They tend to be more aggressive when implanted into the brain of nude mice as compared to their parental GSCs line and have a superior resistance to radiation and therapy. Due to such properties, FC represents an ideal model to identify and test new GBM therapeutics. In Aim 1 of this proposal we will use quantitative mass spectrometry to perform a proteomic analysis on different models of PN and MES GSCs including the FC population. This work will provide a deeper characterization of the FCs and identify differentially expressed proteins/pathways in PN and MES GSCs, which could be exploited to develop new therapeutics. In Aim 2, we will use imaging-based reporters and perform an RNAi screen to identify GSCs modulators which either push GSCs to a more differentiated state, making them susceptible to conventional therapy, or simply kill these cells. Finally we will test a novel gene delivery approach based on extracellular vesicles packaged in adeno-associated virus vectors (vAAV) to deliver our therapeutic RNAi to brain tumors in mice. This work can add valuable information for understanding the biology of this elusive tumor population and plasticity between GSCs subtypes. It also has the potential to identify new vulnerabilities and therapeutic avenues for PN and MES gliomas.
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{
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Central anatomical correlates of aging and age-related hearing loss (presbycusis) will be studied in two mouse models: the C57BL/6J, which demonstrates progressive sensorineural hearing loss (SNHL) with onset during young adulthood to middle age; and the CBA/J, which maintains good auditory sensitivity until late in life. Several age groups will be studied, ranging from young (2-months old) to very old (over 30-months). Two groups of CBA mice with induced chronic SNHL (similar to that occurring in C57 mice) will also be employed. Quantitative morphometric evaluations will be made in subdivisions of the cochlear nucleus (CN) and inferior colliculus (IC) using Nissl-stained, fiber-stained, and Golgi- impregnated tissue. A variety of cellular features will be assessed including cell number (neurons and glia), neuronal soma size and shape, dendritic dimensions and orientation, presence of spines, signs of degeneration, etc. Data will be statistically analyzed as a function of age, brainstem subnucleus, and when appropriate, cell type or location with regard to tonotopic organization. Peripheral input to the brainstem will be evaluated using quantitative counts of spiral ganglion cells. The final product will be a comprehensive, detailed anatomical description of two major auditory brainstem regions in two mouse models of presbycusis. In addition, by comparisons among data from the various subject groups the contributions of aging per se, the occurrence and chronicity of SNHL, and genotype will be elucidated.
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{
"pile_set_name": "NIH ExPorter"
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The modified-oddball paradigm has been used to measure ERP components associated with attention and recognition memory in infancy. Infants are familiarized with 2 stimuli and then exposed to brief presentations of three types of memory stimuli: frequent familiar, infrequent familiar, and infrequent novel. A middle latency negative ERP component over central leads labeled Negative Central (Nc) is assumed to reflect a general orienting response associated with attention. The Nc has been found to be greater in amplitude following novel stimulus presentations. A commonly used behavioral measure of recognition memory is the visual paired-comparison choice trial comparing two visual stimuli. No study to date has measured ERPs during paired-comparison trials because of the eye-movement artifacts produced during shifts between stimuli. One goal of the present study is to examine infant ERPs during paired-comparison trials by utilizing independent component analysis to identify and remove eye-movement components from the EEG data. A second goal is to examine the consistency between ERP components and behavioral correlates of attention and recognition memory by embedding paired-comparison trials within the modified-oddball paradigm. Infants 20, 26, and 32 weeks of age will serve as participants. Participants will be exposed to alternating blocks of paired-comparison trials and brief stimulus presentations in order to measure the infants' visual preferences as the study progresses. Electroencephalographic recordings will be made with a 126-channel system and ERP averages will be made from -50 ms to 2000 ms around stimulus onset for brief stimulus exposures, and for the duration of the paired-comparison trials. Results may clarify brain-behavior relationships associated with infant attention and recognition memory, and shed light on the typical course of the development of attention and recognition memory across the first postnatal year. This is a first step toward understanding the development of attention and recogntion memory for atypical populations. [unreadable] [unreadable] [unreadable]
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{
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The objective of this research is to obtain detailed information about the structure of several membrane transport systems in order to understand the molecular mechanism by which ions are moved across the cell membrane. In the current year we have concentrated on probes which give us information about the transport sites of the human red cell anion exchange system and the (Na,K)-ATPase from dog kidneys. In particular, we have addressed the question of whether sites on different subunits interact during the transport process.
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{
"pile_set_name": "NIH ExPorter"
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Annually in the U.S., more than 1.7 million people sustain a traumatic brain injury (TBI), and approximately $76.5 billion is spent on TBI treatment and rehabilitation. Over the last 10 years, the prevalence of women with TBI has risen by 49%. Research shows that women with TBI have worse outcomes than men, including reporting more headaches and dizziness, as well as loss of confidence, perceived need for supervision, and problems setting realistic goals. Pre-injury, women generally have different social roles and expectations than men in their personal and work lives. After TBI, women struggle to return to pre-injury roles, evidenced by fewer women returning to work and school and having more challenges with parenting than men. However, due to limited research on women with TBI, knowledge on how to meet women's needs and maximize opportunities for successful outcomes after TBI is unavailable. Family members play an essential role in supporting women with TBI during the inpatient rehabilitation process, including involvement in decision- making and goal-planning for post-rehabilitation discharge to home. To prepare family members to best support women with TBI following discharge, it is essential to understand how family members are prepared during the inpatient rehabilitation phase. Yet, after discharge, family members report unmet needs, including insufficient education and feeling unprepared to support the person with TBI and manage their care after discharge. To date, no study has focused on the immediate needs of women with TBI and their families during inpatient rehabilitation, limiting knowledge on their experiences and whether and how current interventions could best meet their post-rehabilitation needs. Congruent with the strategic plan of the National Institute of Nursing Research, which seeks to conduct research on the health of individuals and families to improve self- and family-management of care after injury, this study aims to fill a knowledge gap concerning the inpatient rehabilitation phase. The purpose of this study is to investigate the perspectives and expectations of women with TBI's and their families during inpatient rehabilitation regarding preparation to return home from the hospital. This study will use grounded theory, a qualitative methodology, which will allow the researcher to develop a conceptual model explaining the inpatient rehabilitation process, including the development of expectations for discharge. The sample will be composed of 15 to 20 women with TBI and 15 to 20 of their family members. The long-term goal of this study is to inform future research that will develop and test interventions to improve inpatient rehabilitation for women with TBI.
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{
"pile_set_name": "NIH ExPorter"
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Human umbilical cord blood is a rich source of hematopoietic stem cells that can be used to re-populate the blood and immune systems. In preliminary studies we have identified a novel population of non-hematopoietic cells that exhibit extensive self-renewal and markers characteristic of stem cells. We postulate that these cells can be used to repair the brain following ischemic injury. In specific aim 1 we will conduct studies to compare high vs. low passage stem cells to determine whether high passage cells can repair the brain and restore function in a rodent model of experimental stroke. In specific aim 2 we will conduct cell dose-response experiments to determine an optimal range in the number of stem cells that minimizes neurological and neuropathological deficits. Behavioral tests and novel magnetic resonance protocols will be used to assess neurological function. In specific aim 3, the ability of the transplanted cells to differentiate into neural cells will be assessed by immunohistochemistry. The ability of the grafted cells to induce neurogenesis and vascular neogenesis will also be assessed. The results from this study will provide information on whether non- hematopoietic cord blood stem cells can be used as a therapy for treating ischemic brain injury. PUBLIC HEALTH RELEVANCE: Stroke resulting from ischemic brain injury is a major cause of disability and death in this country. At the present time drugs that are used to treat ischemic stroke must be administered within 3 hours after the ischemic event in order to be effective. We have discovered a unique population of umbilical cord stem cells that appear to reduce the size of lesions in the brain that result from experimental ischemia and minimize the extent of neurological deficits even when administered 48 hours after the stroke. Our proposed project intends to determine whether these cells can be developed as a therapy for treating patients who have suffered ischemic injuries to the brain. [unreadable] [unreadable] [unreadable]
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As a continuation of previous projects on clinical, pathological, and epidemiologic surveillance of Guamanian amyotrophic lateral sclerosis (ALS) and Parkinsonism-dementia (PD) in the Mariana Islands, a total of 112 cases, including suspects registered as of January 1, 1983, are to be followed at intervals of six months for detailed clinical descriptions of patterns of progression by a qualified neurologist until all of the patients expire. As the rate of natural death in recent years is at approximately 30 per year, it is expected to complete this project in three to four years. It has been learned that the average duration of ALS is 4.0 to 4.5 years after onset with a range of 2.0 to 25 years. The study of those long surviving cases (over ten years) has been completed. Clinically they showed three patterns: (1) onset with slowly but steady progression at the same pace throughout the course; (2) rapid progression to complete paralysis of the limbs within 1.5 to 3 years and then remaining practically stable for next 5 to 10 years; (3) onset with minimal atrophy and weakness for the first five to six years and then rapid step-wise progression to death. A study of the neuropathology of those long-surviving cases by a guest neuropathologist from Japan showed a burned-out picture: few active neuronal, axonal or myelin destruction with the remaining neurons appearing surprisingly healthy. A Significant number of PD cases were found to show not only lower motor neuron involvement but also severe pelvicrural flexion contractures in the advanced stage of the disease. This observation presents an important question of (1) major neuron involvement as a part of the natural history of chronic diseases of the CNS, or (2) it is an identical process of ALS which occurs in the same patient. If the latter is true, these cases may represent a continuum of ALS and PD, and thus indicate a single etiology of these two diseases.
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{
"pile_set_name": "NIH ExPorter"
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The etiology of breast cancer for the most part, remains unknown. Some risk factors for breast cancer have been elucidated, which are mostly related to hormonal status or family history. Yet, these risk factors explain only a portion of the variability in disease incidence. We have been studying the role of genetic variation and gene-environment interactions for breast cancer. With our collaborators, we have focused both on hormonal and non-hormonal risk factors. We previously reported an association for smoking and breast cancer in women who had a decreased ability to detoxify tobacco smoke constituents through the NAT2 gene, as well as some weaker effects for two other genes. Among the most important findings this year, our data indicates that there is a relationship of breast cancer to genetic variation in the superoxide dismutase gene. This gene plays a role in the maintenance of reactive oxygen species in the mitochondria. By itself, the gene was predictive of breast cancer risk, and we found that the risk decreased in women who ate more foods rich in antioxidants. Thus, women have the option of modifying their susceptibility based upon their dietary behaviors. Separately, we have tested and validate the p53 sequencing methodology for the Affemetrix chip technology. More than 100 human breast cancers were sequenced by the method and by direct sequencing, with excellent concordance. We are now applying the method to several breast cancer case series. - breast cancer, Cigarette smoking, Epidemiology, Genetic Susceptibility, - Human Tissues, Fluids, Cells, etc.
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{
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The specific aims of this pilot study are: 1) to develop a cell culture system of salivary acinar cells; 2) to study some structural and functional characteristics of the cultured cells. To accomplish these objectives we will first isolate acinar cells from the rat submandibular gland by enzymatic digestion of tissue fragments as routinely done in our laboratory. The cells will then be seeded on three types of support matrices, collagen-polycarbonate supports, nucleopore filters coated with collagen and millipore microporous membranes and incubated at 37degreeC in 5% Co2-air in nutrient media with and without hormonal and metabolic supplements. The media will be changed or replenished at timed intervals and proliferation and confluency will be assessed periodically by light microscopy. A major portion of the initial research will be devoted, therefore, to establishing the optimal conditions for the formation of confluent epithelial cell barriers. Monolayers which develop to confluency with one of the procedures outlined above will then be studied morphologically both at the LM and EM levels. Other monolayers will be placed in Ussing-type chambers for the measurement of transepithelial ion fluxes and electrical potential differences (PD). Net and unidirectional fluxes of 36Cl will be measured in the absence and presence of autonomic agents and transport inhibitors. These findings will be correlated with parallel measurements of PD with KCL-agar bridges connected to a high impedance voltmeter by calomel half cells. The development of a cell culture system of salivary acinar cells will set the basis for a number of future studies related to acinar ion transport under basal and stimulated conditions, which would contribute substantially to our understanding of salivary gland function and regulation.
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{
"pile_set_name": "NIH ExPorter"
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This proposal seeks to study how the chick eye adjusts its growth to compensate for defocusing spectacle lenses that impose myopia or hyperopia. This lens-compensation is seen as a model both for emmetropization--the process by which the eye adjusts its refractive state during development-and for how certain visual environments might predispose young human eyes towards myopia. Because brief, frequent episodes of lens-wear cause surprisingly good compensation, but extremely brief episodes are less effective, we propose to study the temporal integration of lens-wearing episodes by the emmetropization control mechanism. Furthermore, because episodes of wearing plus lenses block myopia from negative lenses, we propose to study this interaction by switching lenses over a wide range of frequencies. We also propose to examine chemical signals in the retina and choroid to see if the time course of their response can explain this enduring effect of plus lenses. The chemical signals we propose to study are retinoic acid and glucagon, both of which show opposite changes in levels to plus and minus lenses, suggestive of their involvement in the control eye growth. Finally, we propose to study possible visual signals that the eye might use to discern myopia from hyperopia. By these means we hope to understand the mechanism by which very brief episodes of wearing plus lenses can cancel out an entire day of wearing minus lenses. Such an understanding would strengthen the rationale for arresting myopic progression in children by visual manipulations.
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{
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The objective of this research program is to identify the mechanisms of neoplastic transformation in head and neck squamous cell carcinoma. We have focused on the epidermal growth factor receptor (EGFR) system because EGFR has been found to be constitutively active in high fraction of head and neck squamous cell carcinomas. It has been found that in addition to autophosphorylation, the activated receptor kinase also phosphorylates other specific intracellular proteins in the cell. It is believed that these intracellular substrates of EGFR kinase will then carry out the signal of EGF through a network of interacting proteins (or signal transduction pathways) that ultimately leads to cell proliferation or differentiation. Therefore, studies on these EGFR substrates will provide important information on the mechanisms of neoplastic transformation in head and neck squamous cell carcinoma. In collaboration with Dr. Pier Paolo Di Fiore of the National Cancer Institute, we have been investigating the nature of one of these receptor substrates, eps8. Our works have focused on the role of eps8 in EGFR signaling pathway and the biologic effect it imparts when switched on. We have evidences suggesting that eps8 is likely to play a role in the proliferation pathway of EGFR. We have also identified constitutively phosphorylation of eps8 in human tumor cell lines.
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{
"pile_set_name": "NIH ExPorter"
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Per capita seafood consumption is increasing, not only in the U.5., but also worldwide. This includes the consumption of local and, more recently, imported species of fish and shellfish. Concomitantly, the prevalence and incidence of diseases associated with seafood ingestion (infectious and Marine Seafood Toxin (MST)-associated) appear to be expanding beyond their traditional boundaries of island and coastal communities. Recent evidence links global climate changes with the geographic extent of the algal blooms associated with MST production. All these factors increase the potential risk through seafood consumption for the MST-associated diseases in human communities. Infectious disease surveillance systems utilize diagnostic methodologies and resources to identify infectious disease outbreaks associated with seafood consumption and to implement prevention activities. Such a model does not exist for the MST Diseases, including the most commonly reported disease, Ciguatera. The lack of epidemiological data for these disease has severely hampered investigation and assessment of the extend of the problem. At present, surveillance of these diseases is local and anecdotal, at best. There is a tremendous need for accurate surveillance data, localized not only in time, but in space, to understand and prevent the impact of these diseases on human populations. The multi-dimensional, multi-scaled and multi-variate characteristics of these diseases are not suited to conventional methods of epidemiology. GIS is designed to handle and model complex phenomena, such as the epidemiology of the MST Diseases. This three-year application seeks to establish a model MST Surveillance Network, which focuses on the MST disease Ciguatera in Dade County (Florida) to evaluate its disease burden in a endemic education, outreach and the groundwork for primary prevention. GIS will be used as the main tool for managing, analyzing and presenting the data collected from the Surveillance Network and other sources. GIS will allow the identification of potential spatial and temporal patterns of Ciguatera in human populations, which will promote a more accurate understanding of their impact. Incorporation of additional environmental data such as the geographic local of toxic reefs, as well as the oceanographic and atmospheric data, into GIS will permit predictive capabilities for the future prevention of Ciguatera and the other MST diseases in human populations.
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{
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Project Summary/Abstract Proposed objectives 1. Evaluate the impact of the Utah Clinical Guidelines on Prescribing Opioids. 2. Evaluate the impact of an educational intervention that teaches clinicians how to interpret and use the guidelines. Importance Unintentional fatalities due to prescription medications are an increasing problem in Utah and the United States. In Utah, the annual number of prescription drug-related overdose deaths began to increase substantially in 2001 and the increase has continued through 2007. Since 2003 the leading cause of injury death in Utah has been poisoning from prescription drugs - with opioid-related poisonings as the primary offender. During the years 1997-2007 deaths attributed to non-illicit drug poisoning increased by approximately 290% in Utah from 4 per 100,000 person-years to 12 per 100,000 person-years. The increase was largely due to deaths from prescription opioid pain medications. The number of emergency department encounters stemming from prescription drug overdose has also continued to increase in Utah from 2001-present.4, 5 Evaluation will allow provide valuable information that will help other states adopt or adapt similar opioid pain management recommendations. Objectives Through this evaluation we aim to determine the impact the guidelines have on prescribing patterns and safety monitoring. We also aim to evaluate which tools from the guidelines the physician's are incorporating into their practice. In addition, we plan to evaluate the impact of the educational intervention on following guideline recommendations. Our hypothesis is that the physicians who receive education on how to use the guidelines will be associated with the biggest improvement in pain treatment. Study Design This study proposes to compare pain management practices for the entire state before and after the prescribing guidelines were finalized and supported by Utah Department of Health. We also plan a clinic- randomized design to evaluate the impact of an intensive educational intervention designed to teach providers how to use the guideline and tools for improving the safety of opioid treatment. Comparison will be done using multiple databases state-level databases that include the Utah Controlled Substance Database, the Utah All Payer Data, Vital Records, and Utah Emergency Department Encounter Data. To evaluate the impact of the interventions we will measure change in prescribing patterns, safety monitoring, use of tools, and adverse drug events. Physicians'will also be asked to self-report the change in their practice through a survey. Setting Evaluation of these interventions will be done at the Utah Department of Health (UDOH) using databases that are housed and managed at the UDOH. The educational intervention takes place in clinics and hospitals across the state of Utah. Data is then collected from participating physicians via an online survey. Participants This study proposes to evaluate all opioid prescribers in the state in the pre and post guideline comparison. Approximately 300 providers will be enrolled in the intensive educational intervention and 300 controls. We also propose to conduct chart-review on 88 Utah Community Clinic providers to determine how the educational intervention impacted their use of tools and documentation. Outcome Measures 1. Changes in prescribing patterns of physicians 2. Changes in the number of EKGs and Sleep Studies ordered by physicians 3. Reduction in number of adverse events (opioid-related emergency department visits and deaths). 4. Changes in the tools used by physicians (e.g. treatment plans, dosing guidelines) PUBLIC HEALTH RELEVANCE: Project Narrative: This proposed project is relevant to public health because of the potential it has to provide scientifically based evidence for the use and implementation of prescribing guidelines to reduce morbidity and mortality related to prescription pain medications.
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{
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Four co-principal and two collaborating investigators will utilize core facilities and collaborate and cooperate to investigate the biology, biochemistry, and somatic cell genetics of cells involved in the immune response. Wherever possible, they will use cloned cultured cell lines and derive from them somatic cell variants with defects in specific immunological function. Special efforts will be made to chemically characterize the molecules directly involved in immunological functions. Particular attention will be paid to structural basis of the functions of H-2 antigens; the genetic and molecular mechanisms involved in the somatic generation of antibody diversity and the switching of classes; the structural basis of antibody effector functions; the molecular mechanism responsible for the immunological and phagocytic roles of macrophages; the role of NK cells in resistance to infectious agents and tumors; and the structural basis of idiotype and the role of idiotypes in auto immune disease.
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{
"pile_set_name": "NIH ExPorter"
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Aneuploidy and polyploidy associated with chromosomal instability are hallmarks of many breast cancers and have been implicated as some of the earliest steps in tumor formation. Recent evidence has established that aneuploidy/polyploidy can even predispose to breast cancer. While much is already known about the molecular mechanisms leading to aneuploidy/polyploidy, little is understood at the genetic and epidemiological level. We hypothesize that inherited variation in genes involved in regulating cell division contribute to the development of aneuploidy/polyploidy and subsequently to breast cancer. We propose a comprehensive multi stage approach to the study of genes involved in regulation of cell division using single variants and haplotypes. We will identify candidate variants and haplotypes associated with breast cancer risk in the Mayo Clinic Breast Cancer case-control Study and validate the associations in the Anglican Breast Cancer (ABC) Study. The genes containing the variants will be resequenced and haplotyped tagging variants representing small haplotype blocks will be selected and re-genotyped in an expanded Mayo Clinic Breast Cancer Study to identify the specific variants or haplotype blocks that account for the association with breast cancer risk. These variants and haplotypes will be validated in an expanded ABC study and the influence of these SNPs on gene expression and function will be characterized. Our specific aims are: Aim #1: To identify and confirm associations between Single Nucleotide Polymorphisms (SNPs) and haplotypes in genes that regulate cell division and breast cancer risk. Specific Aim #2: To characterize all genetic variation in the genes displaying association with breast cancer risk in the Mayo Clinic Breast Cancer Study population. Specific Aim #3: To identify the variants in these genes that account for the modified risk of cancer. Specific Aim #4: To characterize the functional significance of inherited variation in mitotic regulation genes that is associated with breast cancer risk. This study will address the contribution of the under-evaluated area of regulation of cell division to breast cancer etiology. The study will go beyond identification of risk markers to the characterization of the variants that actually cause the modification of breast cancer risk. The results will provide information for improved risk assessment and could provide targets for breast cancer prevention and therapeutic agents.
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{
"pile_set_name": "NIH ExPorter"
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OBJECTIVE: This project investigates whether supplementation with vitamin E or beta-carotene reduces incidence of lung, prostate, and other cancers, and tests several hypotheses related to cancer prevention, etiology (including gene- environment interactions), and early detection in a large, prospective cohort. BACKGROUND: Research indicates a role in cancer prevention for certain micronutrients, including vitamin E and beta-carotene, based on antioxidant, antiproliferative, and other properties. METHODS: The ATBC Study is a randomized, controlled intervention trial of daily supplementation with beta-carotene (20 mg) and/or vitamin E (50 mg dl-alpha-tocopheryl acetate) for 5-8 years. The 29,133 participants are 50-69 year old male cigarette smokers in southwestern Finland. Serum, toenails, and questionnaire data were collected at baseline, and serum, whole blood, and RBCs during followup. All cancers are identified annually through the national cancer registry, with central medical record and pathology reviews. PROGRESS: Active intervention concluded in 1993, and continued follow-up of the cohort examines post- trial incidence trends. Cohort and nested case-control investigations are underway, including many aimed at testing environmental-gene (ie, polymorphism) hypotheses. Intervention findings showed reduced incidence in the vitamin E group for prostate (32%) and colorectal (22%) cancers, and modest reductions in major cardiovascular events. Beta-carotene demonstrated no cancer preventive effects and resulted in increased lung cancer incidence (16%) and overall mortality (8%). Organ-specific intervention and cancer etiologic analyses are focused on a wide range of hypotheses that utilize the randomized interventions, baseline risk factor data, and prospectively collected biospecimens (serum, DNA, etc.). The research questions include, for example, effects of vitamin E, carotenoids, folate/homocysteine, vitamin D, selenium, alcohol, and other dietary components, steroid hormones, physical activity, and serum micronutrient concentrations on cancer. Mechanistic investigations of the two trial agents, and the cancer risk relation of several genetic polymorphisms and DNA markers are also being investigated including those for GSTs, CYPs, NQO1, folate and methyl-group metabolism, IGFs, androgen and vitamin D receptors, and DNA damage and repair.
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{
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Adapted from the Applicant?s Abstract) This Core will provide essential technical facilities for microscopic and morphometric studies of muscle and other tissues. Throughout the 25 years of this PPG, the investigators have always taken the stance that structure should be correlated with function. During the first two cycles of the Program Project, extensive use was made of the technique of rapid freezing of lung combined with preparation of tissue for transmission electron microscopy under the direction of Dr. Mazzone. When Dr. Mazzone left, 15 years ago, Dr. Odile Mathieu-Costello replaced him as a faculty member. Her primary interest is in structure-function relationships, especially in skeletal muscle as evidenced by her Project 3 in the 4th cycle , and Project 2 in the previous cycle and in this proposal. She has also closely collaborated with Dr. John West over the past 9 years on the electron microscopy and morphometry of stress failure of pulmonary capillaries. She was trained as a morphologist by one of the leading authorities in both lung and muscle morphology and morphometry, Professor Ewald Weibel from Switzerland. She brings unique expertise to this highly specialized area, and she is the ideal person to continue to head the scientific aspects of this Core. The investigators see the Morphology and Morphometry Core as an essential feature of the Program Project. The Core will also fully support the salary of Li Wu, the histology technician, as in the previous cycle. Mr. Wu has experience in the preparation of sections of muscle and other tissues for light and electron microscopy, and histochemistry. He will work closely with Dr. Mathieu-Costello in the preparation of tissues for both light and electron microscopy, Mr. Wu understands the needs of our group well, and is well trained in the execution of tissue preparation and morphometry. Morphometric analysis requires a large number of sections and fields to be examined, and tissue sectioning, especially for electron microscopy, is extremely time consuming. Based on the output of the Core in the past cycles and the studies proposed in this application, a full time technician is clearly needed to meet the needs of the different projects.
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{
"pile_set_name": "NIH ExPorter"
}
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The purpose of this project is to investigate the mechanisms and effects of the reduction of proline to Delta-NH2 valeric acid in dental plaque from monkeys. Incubation of dental plaque homogenates with a variety of substrates suggests that proline is converted to Delta-NH2 valeric acid by means of Stickland reactions involving other amino acids and also certain end products of glucose metabolism notably pyruvic and lactic acids. The work is being directed towards elucidating the mechanism and products of these reactions and their contribution to the overall metabolism of dental plaque.
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{
"pile_set_name": "NIH ExPorter"
}
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The alpha2 macroglobulins (alpha M) are plasma proteins present in all classes of vertebrates which function as broad-spectrum proteinase inhibitors and as elements in a unique system for clearance of active endopeptidases from the circulation. We have recently identified a proteinase inhibitory activity in the plasma of an arthropod, the American horseshoe crab, Limulus polyphemus (Quigley & Armstrong 1983 a,b,c). The present proposal seeks to investigate the possibility that the Limulus inhibitor is homologous in structure and function to vertebrate alpha M. If such a homology can be demonstrated, then alpha M is presumably of sufficient functional importance to have been preserved over the estimated 0.55 - 0.75 billion years of evolution separating the ancestral stocks of vertebrates and arthropods. Limulus should prove to be a useful model organism to study the function of the alpha M-like inhibitor because the animal is abundant, contains a large quantity of easily obtainable plasma, and the alpha M-like inhibitor is the only proteinase inhibitor in the plasma (in contrast to vertebrates, which possess a variety of plasma proteinase inhibitors). The latter aspect is significant since the presence of the other proteinase inhibitors in mammalian blood has complicated the elucidation of the exact physiological roles and functions of alpha M. We propose to take advantage of this attributed and our recent establishment of a purification scheme for Limulus alpha M (Quigley & Armstrong, 1985) to examine its role in the clearance of endopeptidases from the circulation and the possible existence of specific alpha M- proteinase receptors in Limulus blood cells. In addition we plan to study the reasons for the apparent inability of Limulus alpha M to inhibit the key proteinase in the blood coagulation cascade in Limulus. The Limulus clotting system has been suggested to represent a primitive model for the evolutionary precursors of vertebrate clotting systems. A variety of other invertebrates will be examined for the presence of alpha M homologues in the plasma.
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{
"pile_set_name": "NIH ExPorter"
}
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The H. Lee Moffitt Cancer Center & Research Institute opened in 1986 on the University of South Florida campus with the mission "to contribute to the prevention and cure of cancer." The Cancer Center enjoys close ties at multiple levels of the University structure, and benefits from the clearly articulated authorities of the Center Director and the Center's independent financial status. Over the past five years, the Cancer Center has invested over $18 million in research programs, personnel and infrastructure. In addition, a $14 million renovation of the Research Center and other laboratories during the same time period expanded research space from 18,000 sf in 1993 to 90,000 sf currently. The commitment to research complements the strong clinical operation of the hospital, which admits 4,000 inpatients per year; the clinics, which has more than 80,000 patient visits in 1995-96; and the unique Lifetime Cancer Screening Center, which sees more than 7,000 healthy and high risk individuals each year. The Cancer Center's four established research program include Molecular Oncology, Immunology, Clinical Investigations and Cancer Control. The programs have built on the collective strengths of faculty from multiple departments and colleges and the excellence of a strong clinical care setting. The Cancer Center Support Grant will enable the Cancer Center to expand its research focus to include genetics, to be developed as a joint basic science and cancer control effort. Additionally, the CCSG will support expansion of shared resources to accommodate increased usage from growth in personnel and scope of research. Shared resources includes Flow Cytometry, Pathology, Molecular Biology, Glass Wash, Molecular Imaging, Biostatistics, Data Management, and an animal core in the Cancer Research Center which will add transgenic services in the coming year. Most importantly, the CCSG will provide focused support for enhanced development of translational research through support of senior leaders and program leaders in their roles as they work with cancer center members to promote laboratory-clinical-laboratory interactions and translations.
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{
"pile_set_name": "NIH ExPorter"
}
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Our investigation of signal transduction pathways and their relationship to cancer has grown in many ways over the last year. We have used CAI as a tool to further our studies of key Ca++-mediated pathways. The molecular dissection of CAI demonstrated that CAI inhibition of Ca++- mediated signaling events is concordant with inhibition of proliferation. Novel compounds were identified and characterized and a patent application was filed. Further studies have identified a role for Ca++ influx regulation of tyrosine phosphorylation of phospholipase C-gamma which produces inositol trisphosphate and internal calcium release. Use of CAI helped clarify that receptor-operated Ca++ influx mediated maintenance of the malignant phenotype of CHO cells. Biotinylated CAI (B-CAI) has been produced. Avidin-linked B-CAI and a CAI analog have been used to immunize rabbits. B-CAI and/or antisera will be used for expression cloning of the CAI binding sites and studies at the protein level. Subtraction analysis of gene products amplified in cells resistant to chronic CAI exposure has identified several candidate genes. Transcripts of 1.2 and 2.7 kb were expressed in A2058 resistant to 10, 20, and 30 micromoles CAI and not in wild type A2058. Clones identified using the cDNA subtraction probe are being characterized. CAI has been used as a tool to investigate Ca++-regulated gene expression. CAI inhibits IL-2 gene transcription through several transactivating proteins. It also stimulates and inhibits Ca++ and growth factor-mediated production of a viral enhancer, VL-30. We have shown that CAI is antiangiogenic in vitro and in vivo. Incubation of human umbilical vein endothelial cells with CAI inhibited proliferation, adhesion, motility, MMP-2 production, and tube formation, and CAI inhibited microvascular proliferation in chick chorioallantoic membrane assays. Studies are ongoing to identify the sites of this Ca++- mediated signal transduction inhibition of angiogenesis.
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{
"pile_set_name": "NIH ExPorter"
}
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The purpose of this study is to examine the three-year outcome of relaxation training used as a adjunctive procedure in the treatment of essential hypertension. 260 participants will be randomly allocated to receive relaxation training or no training. All participants will be followed every two months to monitor blood pressure, and prescription of antihypertensive agents will be monitored at six-month intervals. Several other issues concerning the procedure and effectiveness of relaxation training will be examined in randomized experiments with subsamples of the participants. These include an examination of the generalization of blood pressure lowering during the working day; a study of adherence to the relaxation procedure; a test of methods to enhance the effects of relaxation training for nonresponders to initial training; and a study of the need for continued relaxation practice.
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{
"pile_set_name": "NIH ExPorter"
}
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[unreadable] [unreadable] The Environmental Mutagen Society (EMS) is the primary scientific society fostering research on the basic mechanisms of DNA repair and mutagenesis and application of this knowledge to understanding human health effects from exposure to environmental genotoxins. Studies of DNA repair and mutagenesis by EMS scientists are integrated with research on inherited and acquired genetic alterations that predispose individuals to cancer, premature aging, and other diseases. Fundamental understanding of the mechanisms and consequences of cellular, tissue, and whole organism responses to genotoxic agents is crucial to informed regulatory decision-making with respect to environmental health hazards. The integration of multi-disciplinary basic and applied research is essential to this process. Such integration remains the central focus of the EMS, and the annual meetings are key to the dissemination of information and fostering of interactions that are necessary to achieve it. In addition, the meetings provide an important forum for students and postdoctoral investigators to present their research and to interact with leading scientists in their field in a setting that encourages open exchange of ideas. The 39th Annual Meeting of the EMS will be held in Rio Grande, Puerto Rico from October 18 - 22, 2008. Its theme is "Genes and the Environment: From Molecular Mechanisms to Risk". The scientific program, which encompasses the full range of scientific interests within EMS, is comprised of symposia, workshops, topical reviews, plenary lectures, and contributed poster and platform sessions. Throughout its history, the EMS Annual Meetings have brought together academic, industrial and governmental scientists interested in how environmental factors lead to genotoxicity. Awarded funding support for the meeting is critically necessary in order to achieve this mission. The dual objectives of this conference grant application are to request funds for travel awards to students and postdoctoral fellows to attend the 2008 EMS Annual Meeting, and funds to partially support travel expenses for key non-member speakers. EMS believes that its students and postdoctoral investigators are the future of both the Society and the scientific endeavors fostered by its community of scientists. Travel awards assist in offsetting the financial burden that might otherwise prevent trainees from attending. The application also requests funds to partially defray travel costs for invited non-member speakers. EMS member speakers all agree to cover their own expenses for participation in the conference. However, it is necessary to assist with travel expenses for invited non-member speakers, who provide the cross- fertilization of ideas necessary for a world-class meeting. [unreadable] [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
}
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Conceptual links and a nascent literature suggest that in-home caregiving arrangements have a bearing on disabled older adults' use of medical services. Larger caregiving networks, the presence of unpaid help, lower levels of caregiver burden and higher levels of reward, and the absence of unmet need with activities of daily living have previously been linked to desirable health outcomes and a lower likelihood of permanent nursing home entry for recipients. Here we extend this line of inquiry by hypothesizing that these attributes of in-home caregiving arrangements are indicative of the depth of available resources that may be mobilized to assist recipients with personal care as well as necessary medical care. The proposed project examines the relationship between these attributes of in-home care arrangements and hospital outcomes that are thought to be particularly sensitive to the ability of in-home caregivers to facilitate appropriate medical care. This prospective cohort study draws on data from the 1999 National Long Term Care Survey, its accompanying Caregiver Survey, and Medicare claims data for the 12-month period following completion of the 1999 survey. The relationship between in-home caregiving arrangements and hospitalization outcomes (specifically, potentially preventable hospitalizations, repeat hospital admissions, and delayed discharges) will be estimated using multivariate regression models that control for individual and caregiver characteristics known to be related to the outcomes of interest, with particular attention to patient morbidity and severity of illness. The proposed research will provide an understanding of the influence of in-home caregivers in an important area that is beyond the scope of outcomes typically attributed to them. The ultimate goal of this work is to develop strategies that support caregivers and improve the health of older adults. Findings from this pilot project will inform measurement and data collection efforts, as well as interpretation of results from a multi-component caregiver intervention in Baltimore, Maryland that is expected to begin in July 2006. [unreadable] [unreadable] [unreadable]
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{
"pile_set_name": "NIH ExPorter"
}
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The goal of this proposal is to continue to participate as a clinical site in the nation-wide study of neuroprotection in Parkinson's disease (NET-PD). Parkinson's Disease and Movement Disorders Center at Northwestern University (PDMDC-NU) has been a NET PD Clinical Center since 2005. The Center has demonstrated success in subject's recruitment and retention for the LS1 study. The Center is actively working on fulfilling recruitment for the FS-Zone study. We are committed to recruit, retain and follow research participants as a Clinical Center in the NET-PD consortium. The objective of the project will be to: A. Retain participants in the NET-PD LS1 (a phase 3 randomized, double-blind, placebo-controlled trial of creatine treatment for the prevention of Parkinson's Disease progression). B. Recruit and retain appropriate research participants in the NET-PD FS-ZONE (a randomized, double-blind, placebo-controlled futility study evaluating pioglitazone for the prevention of Parkinson's Disease progression). We are committed to collaborate with the NET PD Clinical Trial Coordination Center (CTCC), Statistical Center (SCC), and other Clinical Centers to assure successful study completion, and to participate in the data analysis of the study results. As a Clinical Center, we are committed to continue working cooperatively with the NET PD CTCC, Statistical Center (SCC) by assuring quality and timely data collection. We also are prepared to participate in the data analysis for both studies. Dr. Simuni chairs one of the NET PD LS1 writing groups. Dr. Simuni has assumed the leadership role as the national Principal investigator for the FS Zone study in October 2010 and as such is committed to spearhead the successful recruitment, retention, data acquisition and analysis of the study.
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{
"pile_set_name": "NIH ExPorter"
}
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