Tetrahydropterin forms the basic structure of a series of analogues which serve effectively as cofactors for hydroxylase enzymes. Most analogues are formed by placement of various side chains on the 6-carbon. We have recently acquired a compound which has phenyl group attached at this 6-position. The present investigation compared this compound with tetrahydrobiopterin for effective hydroxylation of phenylanine, tyrosine and tryptophan. The enzymes used were: phenylalanine hydroxylase purified from rat liver and tyrosine and tryptophan hydroxylase from rat brain. In addition we examined the ability of the reductase system to continually regenerate reduced 6-methylpterin. Our kinetic analysis showed that 6-phynylpterin is as effective as the natural cofactor (tetrahydrobiopterin) for the various hydroxylases. Also, 6-phenylpterin is incorporated effectively into the endogenous reductase system.


Cdk5 and nociception: In general nociceptive (painful) neural signals originate from sensory neurons in peripheral tissues and are transmitted to second-order neurons in the spinal cord, which then convey the message to specific nuclei in the brain for perception of pain. Nociception results from the activation of molecular and cellular mechanisms in damaged tissue, sensory neurons, and the spinal cord. Many cellular pathways have been implicated in nociceptive signaling, but their precise molecular mechanisms have not been clearly defined. Abnormalities in molecular pathways underlying nociceptive processes may result in chronic pain conditions. It follows that a detailed characterization of these pathways is necessary for developing effective strategies to treat pain. Signal transduction mechanisms activated by nociceptive stimuli have not been fully characterized. Recent advances, especially the cloning and characterization of the capsaicin receptor (also known as vanilloid receptor 1, VR1, TRPV1) remarkably improved our understanding of signal transduction in nociceptive neurons. TRPV1 is a member of the transient receptor potential (TRP) family. It is a polymodal ligand-gated ion channel that is expressed in small-diameter sensory neurons (C-fibers) and activated by heat, protons, anandamide, and leukotrienes. TRPV1 knockout mice show reduced thermal hyperalgesia following inflammation and reduced nociceptive responsiveness in a model of bone cancer pain. TRPV1 has been connected to a broad spectrum of physiological conditions and diseases such as thermal hyperalgesia, allodynia, inflammatory bowel disease, Crohn's disease, vulvodynia, osteoarthritis, pancreatitis, gastroesophageal reflux disease, bladder disease, cystitis, and asthma. There is mounting evidence that TRPV1 is subject to multiple interacting levels of control. Phosphorylation of TRPV1 is critical for its function in response to nociceptive stimuli. We previously described the expression and specific activity of Cdk5/p35 in TRPV1-positive cells of dorsal root ganglia (DRG) and trigeminal ganglia (TG) and the modulation of Cdk5 activity in response to peripheral inflammatory pain. We also found that p35 knockout mice, which have significantly reduced Cdk5 activity, behaviorally mimic the TRPV1 knockout pain phenotype. [unreadable] [unreadable] Cdk5, a neuron-specific, proline-directed serine/threonine kinase is activated by 2 noncyclin activators, p35 and/or p39. Cdk5 phosphorylates serine and threonine immediately upstream of a proline residue. In addition to an absolute requirement for proline in the +1 position, Cdk5 shows a marked preference for a basic residue in the +3 position and phosphorylates the consensus sequence (S/T)PX(K/H/R). Several neuronal and nonneuronal substrates are phosphorylated by Cdk5, and the list of new substrates is increasing. Earlier studies showed that Cdk5 knockout and p35/p39 double-knockout mice are embryonically lethal with neuronal migration defects, whereas p35 knockout mice show inverted neuronal layering with a concurrent attenuated response to noxious heat; conversely mice overexpressing p35 were hyperalgesic. [unreadable] [unreadable] Subsequently, we analyzed TRPV1 for potential phosphorylation by Cdk5. We report that Cdk5 can directly phosphorylate TRPV1 at threonine 407, and this in turn modulates agonist-induced calcium influx. We initially found that inhibiting Cdk5 activity resulted in attenuation of capsaicin-induced calcium influx in cultured DRG neurons, and this attenuation was reversible. These observations suggest that Cdk5-mediated phosphorylation of TRPV1 is important for capsaicin-mediated calcium influx through this receptor. Since germline Cdk5 knockout mice are embryonically lethal and p35 knockout mice present various neuronal disorders, we generated primary nociceptor-specific Cdk5 conditional knockout (Cdk5-CoKO) mice to identify the precise role of Cdk5 in primary afferent pain signaling. In the basal state, the conditional knockout mice showed significant hypoalgesia, confirming the direct role of normal Cdk5 activity in primary afferent. Collectively, our findings describe a novel molecular mechanism for the functional regulation of TRPV1 by Cdk5 and provide further insights into the role of Cdk5 in the pain pathway.[unreadable] [unreadable] Cdk5 and brain development: Analysis of functional roles of Cdk5 in postnatal brain development has been hampered because of perinatal lethality of Cdk5-/- mice and the compensated phenotype of p35-/- mice. To study the role of Cdk5 in postnatal development of the midbrain-hindbrain (MHB), we generated MHB-specific Cdk5 conditional knockout mice by crossing Cdk5 floxed mice with Wnt1-Cre mice. Wnt1-Cre-mediated Cdk5 conditional knockout (WCOKO) mice have lower body weight than controls, an ataxic gait, and early postnatal lethality. Histological analysis indicated a smaller cerebellum with defective migration of the Purkinje cells. A detailed analysis of WCOKO mice showed a complete lack of inward migration of the granule cells. In addition, we also identified a complete absence of the superior colliculus in Cdk5-/- mice and its abnormal development in WCOKO mice. These results indicate that Cdk5 plays important roles in mouse MHB development.


Protein-protein interactions are essential to almost all cellular processes. The ability to rationally manipulate these interactions would allow for the creation of new tools for studying cell biology and the creation of new protein therapeutics. The objective of this research is to develop and test computer protocols for protein interface design that include full backbone and side chain flexibility. We will test these protocols on two important problems in interface design: enhancing the binding affinity of naturally occurring interactions and changing the binding specificity of signaling proteins. To achieve this objective we will make use of a computer program, Rosetta Design, that we developed for simultaneously optimizing the amino acid sequence and backbone conformation of a target structure. Encouragingly, this protocol has been previously used to design a novel protein structure with atomic level accuracy. Several lines of evidence suggest that the accuracy of protein design simulations increase when more side chain conformations are considered during the simulation. To allow complete flexibility in side chain torsion angles we will use a Monte Carlo minimization procedure in which discrete rotamer substitutions are followed by gradient-based minimization of side chain torsion angles. We will test the flexible side chain model by designing functionally orthogonal binding interactions between two sets of model proteins: proteins from the ubiquitination pathway and proteins involved in G-protein signaling. We will test our model for protein design with a flexible backbone by designing protein extensions that enhance binding affinity by increasing the number of favorable contacts with the partner molecule. Specifically, we will incorporate additional residues off the N-terminus of the ubiquitin conjugating enzyme, UbcH7, that are predicted to enhance its binding affinity for its natural binding partner, the ubiquitin ligase E6AP. Binding affinities will be determined with various biophysical techniques and activity assays will be used to determine if the redesigned proteins bind as an active complex.


Understanding the mechanisms for the breakdown of self-tolerance in autoimmunity is essential for the development of strategies to prevent and treat autoimmune diseases. Abundant evidence has shown that defective apoptosis in the immune system is associated with the development of systemic autoimmune diseases in humans and mice. However, inhibition of apoptosis in lymphocytes alone is not sufficient to break immune tolerance, indicating that impaired apoptosis in other cell types plays a critical role in the breakdown of self-tolerance. Targeted inhibition of apoptosis in dendritic cells (DCs) has been shown to induce systemic autoimmune responses. Experiments are proposed to test the hypothesis that apoptosis in DCs is essential for limiting lymphocyte activation and preventing autoimmunity in the following specific aims: 1) to characterize the apoptosis pathways in DCs. Death receptor-mediated and mitochondrion-dependent apoptosis pathways will be characterized in DCs. Preliminary studies suggested that the lifespan of DC subsets in vivo was correlating to the molecular ratios between anti-apoptotic and pro-apoptotic bcl-2 family members. Experiments will be performed to further characterize the bcl-2-regulated mitochondrial apoptosis pathways in regulating DC apoptosis; 2) to test the hypothesis that defective apoptosis in DCs contributes to dysregulated lymphocyte activation. The lifespan of DCs can potentially influence immune responses by affecting the duration of DCs in stimulating lymphocytes. The potentials for apoptosis-deficient DCs in over- activating lymphocytes will be examined; and 3) to test the hypothesis that defective apoptosis in DCs contributes to the development of autoimmunity. Studies will be performed to examine whether DCs harboring apoptosis deficiency in the mitochondrial apoptosis pathways leads to the development of autoimmunity. Experiments are proposed to test the hypothesis that apoptosis regulates DC homeostasis and immunogenicity, and defective apoptosis in DCs contributes to the onset of autoimmunity. In the long term, the knowledge gained from these studies will be used to develop more specific and effective strategies to prevent the onset of autoimmunity by targeting apoptosis in DCs. [unreadable] [unreadable] [unreadable]


A new electrophoretic method is proposed for efficiently separating membrane proteins while retaining function. Fluid, smooth, electrically neutral lipid bilayers are proposed as the electrophoretic medium. The proposed separation would maintain the membrane proteins in their natural type of environment to produce separated zones of membrane proteins, which are then displayed as functional arrays. The separation would support such applications as membrane proteomics, drug screening and development of cancer diagnostics. Membrane proteins play key roles in human health, since they direct cellular signaling, recognition and transport, yet the steps for isolating membrane proteins for research are expensive, laborintensive and inefficient. Today's methods for separating proteins rely on water solubility. The proposed work will test the feasibility of separating membrane proteins in lipid bilayers by investigating 3 essential properties. First, the fluidity and uniformity of the lipid bilayer will be studied on ultrasmooth polyacrylamide films, grown by atom-transfer radical polymerization. The polyacrylamide film provides a uniform hydrophilic boundary on the underside of the bilayer, an aqueous buffer will be used on the upper side. Second, the electrophoretic mobilities of 5 well characterized receptors will be studied: the 3 opioid receptors and 2 melanocortin receptors. All 5 are medically important G-protein coupled receptors. Their ligands are peptides, which will be singly labeled with rhodamine. Third, the function of these membrane proteins will be studied by single-molecule fluorescence spectroscopy of the rhodamine labeled ligands. The R21 research will critically test the idea of whether the electrophoretic separation of functional membrane proteins is feasible in supported lipid bilayers. If feasible, an R01 proposal would address separations of real mixtures of membrane proteins from cell lysates. The work could have enormous impact on drug discovery, allowing inexpensive screening that encompasses many receptors.


PROJECT SUMMARY Specific compositions of periodontal microbiota are associated with risk for the various forms of periodontitis. Serum antibody levels are considered as surrogate markers for the activity levels of periodontal microbiota, which represent the protein synthesis and inflammatory potential of the microbiota. The chronic trickling of periodontal microbiota into the bloodstream also elicits a low-grade systemic inflammation which has been suggested to increase the risk for several extra-oral diseases, such as atherosclerosis, rheumatoid arthritis, metabolic disorders, and neurodegenerative diseases, including cognitive impairment and Alzheimer's disease. Furthermore, our recent study using data from the Third National Health and Nutrition Examination Survey (NHANES III) indicated that specific compositions of periodontal microbiota, even without inducing clinically significant periodontitis, may have a significant impact on the risk for human disease and mortality, and that Porphyromonas gingivalis (P gingivalis) play a key role in the dysbiotic periodontal microbiota. In this proposal, we will collaborate with experts from related disciplines to test our hypothesis that the activity levels of specific periodontal microbiota/P gingivalis are associated with the occurrence of age-related macular degeneration (AMD). We will first use partial least squares (PLS) regression to identify specific patterns of 21 serum immunoglobulin G (IgG) levels for periodontal microbiota which are associated with the risk for AMD in a case-control study from a representative sample of the US population, the NHANES III. PLS is an important statistical method in bioinformatics and used to discover variables (i.e. specific patterns of IgG levels) that are not directly observed but are rather inferred from other observed variables (i.e. 21 individual IgG levels). Since our hypothesis is novel and no previous study in the topic has been done, it is possible that the observed associations between specific IgG patterns and AMD will be confounded in spite of adjusting for confounding variables. To further verify the associations, we will use complement factor H (CFH) genetic polymorphisms as an instrumental variable in our Mendelian randomization study. CFH genetic polymorphisms are a well-established risk factor for AMD. The Mendelian randomization approach is a novel epidemiologic study design that incorporates genetic information into traditional epidemiologic methods and addresses exposure-outcome relationships without many of the typical biases that impact the validity of traditional epidemiologic approaches. The Mendelian randomization approach is especially useful when a randomized trial is impossible. This project will serve as our first step toward our long-term objectives of understanding how the phenotypes (such as serological markers) and genotypes/strains (microbiome) of oral microbiota, together with the human genome and nutrition and other known risk factors, affect eye health. Since periodontitis is one of the most prevalent diseases in humans and AMD is the leading cause of blindness among elders, understanding the associations between periodontal microbiota and AMD may lead to new therapeutic and preventative strategies for AMD.


We are developing an artificial bone marrow for ex vivo culture of hematopoietic stem cells (HSCs). This platform has significant scientific value for testing hypotheses regarding the cascade of external signals responsible for directing HSC fate decisions within the bone marrow. An artificial marrow would also have significant clinical value for therapeutic expansion of HSCs or for study of the etiology and treatment of hematopoietic pathologies. However complicating this effort is limited information regarding the regulatory role played by the continuum of sub-niches that exist in close spatial order across the marrow responsible for maintaining hematopoietic homeostasis. We have recently described microfabrication approaches to generate an engineered bone marrow (EBM) containing overlapping patterns of marrow-inspired cellular, biophysical, and biomolecular cues to begin to examine the coordinated impact of these signals on HSC quiescence vs. activation. However, the small scale that makes the EBM advantageous introduces concerns regarding the unknown heterogeneity of a stem cell's response to these cues. While not surprising HSCs may exhibit a range of responses to a niche signal, effects may be magnified in multi-cue environments. We therefore propose to demonstrate a label-free approach to temporally track and quantify the heterogeneity of single HSC fate decisions in response to multiplexed EBM niche signals. To do this, we will combine photonic crystal enhanced microscopy (PCEM) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) to create an integrated detection instrument able to trace single HSC fate decisions. We hypothesize that individually profiling these decisions across a continuum of biomaterial sub-niches will allow us to better predict niche regulation of HSC fate than more traditional metrics that report ensemble averages. Aim 1 will integrate SIMS and PCEM to determine the heterogeneity of HSC response to biophysical cues demonstrated to have an effect on populations of HSCs. Aim 2 will employ the PCEM/SIMS-based biosensor to examine HSC fate decisions within multi-cell colonies containing HSCs and supportive niche cells. Combining this novel biosensor with traditional functional assays will allow us, for the firt time, to determine the heterogeneity of HSC response to engineered niche signals. It also offers a framework to rapidly assess the impact of multiple signals using a minimal number of cells in order to identify hierarchies and/or synergies between these cues. Such data will bring new richness to our understanding of how HSCs integrate niche signals as well as identify critical design elements of an engineered bone marrow. By quantifying the level of heterogeneity in these fate decisions, we will also establish where along the continuum between single HSC and ensemble averages future investigations must focus.


Human rotaviruses are an important cause of severe diarrhea in young children throughout the world. Rotavirus infections are especially devastating in developing nations because of the high mortality rate associated with gastroenteritis. By the age of 3, the incidence of rotavirus infections decreases markedly, but outbreaks of rotavirus-induced diarrhea have been reported among the elderly, the acutely ill, and the immunocompromised. Other mammals and birds can develop rotavirus diarrhea as well, making this disease one of veterinary interest and economic concern. Effective means of preventing this disease are obviously needed. The rotavirus genome consists of 11 segments of double-stranded RNA. Most of the gene segments appear to be moncistronic and together they encode at least 12 viral polypeptides (VP). The genome is enclosed in a core which is surrounded by a double- shelled capsid. VP3 and VP7 are the major outer shell proteins. VP3 is activated by protease cleavage and is associated with host range, hemagglutination, virulence, and both homotypic and heterotypic neutralization. The glycoprotein VP7 is the primary determinant of serotype specific neutralization. This proposal focuses on defining and characterizing those genetically-determined features of VP3 that are involved in gastrointestinal infection. To accomplish this, we plan to: 1) determine the nucleotide sequence encoding the homotypic and heterotypic neutralizing domains on VP3, 2) compare the gene 4 sequences of various murine rotavirus stains with human or simian strains in an effort to clarify the genetic basis of host range, 3) design and execute a series of genetic experiments to determine whether rotavirus RNA is "infectious," 4) develop a strategy to clone full length gene 4 cDNA, and 5) use full length cDNA clones of rotavirus gene 4 in a suitable expression vector to produce a biologically active VP3.


Dietitian support only.


The purpose of this project is to add approximately 22,500 person years for the detection of cancer and other major medical illnesses among the 5,464 women who have previously been enrolled in the DESAD Project. This continued surveillance will enable us to accomplish five specific aims: to determine the magnitude of the excess of squamous neoplasia in exposed over unexposed women and to determine if the increased risk continues over the next five years, to determine if the risk of adenocarcinoma increases during the next five years, to determine the rates of serious medical illnesses in the DES exposed women and to compare these rates with the control groups and published age specific incidence rates for the same diseases, to determine the frequency of abdominal and gynecologic surgery among DES exposed women and to compare these rates with the unexposed controls and the rates available from the National Center for Health Statistics, and to maintain contact with the DESAD Project exposed cohort at minimal expense. These aims will be accomplished by soliciting information by the method of mailed questionnaire. Each participant will be questioned annually concerning the diagnosis of medical diseases or the performance of surgical procedures. Previous experience of the DESAD cohort has indicated that in excess of 90 percent of the women previously enrolled in the project can be expected to comply with the requests for information. The sample size considerations are explained in detail in the project and indicate that it will be possible to detect, with a power of .9, a two-fold difference in the rates of many common medical diseases and surgical procedures at the p=.05 level.


The primary objective of this study is to compare the long-term safety of a hydrofluroalbane propellent (non-ozone depleting) with that of the standard chloroflurocarbon based propellant used in metered-dose inhalers. The secondary objective of this study is to evaluate the safety and efficacy of salbutamol sulfate in those subjects who were randomized to receive Ventolin (trade name for salbutamol) in a previous study. This study has been completed.


Ovulation is essential for successful reproduction. The ovulatory luteinizing hormone (LH) surge stimulates periovulatory follicles to produce prostaglandin E2 (PGE2). Blockade of PGE2 production causes ovulation failure. LH-stimulated neovascularization of the follicle is also essential for ovulation as inhibition of follicular angiogenesis prevents ovulation. This proposal puts for the overall hypothesis that PGE2 mediates the ability of the ovulatory LH surge to regulate angiogenesis within the ovulatory follicle. Angiogenesis is regulated via vascular growth factors. In Aim 1, we will determine if LH acts at follicular granulosa cells to regulate production of vascular growth factors in a PGE2-dependent manner. Monkey ovaries with multiple follicles will be obtained before and after administration of an ovulatory dose of the LH-like hormone hCG; some monkeys will also receive concomitant administration of a prostaglandin (PG) synthesis inhibitor. Granulosa cells, follicular fluid, and whole ovaries will be assessed for vascular growth factor mRNAs by qPCR and proteins by western blotting, ELISA, and immunofiuorescent detection on tissue sections. In vitro studies will be performed to determine if PGE2 acts directiy at granulosa cells to regulate vascular growth factor mRNA and proteins. Vascular growth factors act at endothelial cells to promote the formation of new blood vessels. In Aim 2, we will determine if PGE2 and vascular growth factors act at endothelial cells of the follicle to stimulate angiogenesis. Using a novel proliferating population of monkey ovarian endothelial cells, we will determine if PGE2 stimulates endothelial cell proliferation, migration, and formation of new capillaries. Similarly, vascular growth factors produced by granulosa cells, granulosa cell-conditioned media, and coculture with granulosa cells will be used to determine if vascular growth factors stimulate endothelial cell functions necessary for angiogenesis. Finally, monkey ovaries obtained after treatment in vivo with hCG, hCG+PG synthesis inhibitor, or hCG+PG synthesis inhibitor+PGE2 will be assessed histologically for neovascularization of the ovulatory follicle. The proposed studies will likely demonstrate that PGE2 and vascular growth factors act directly at endothelial cells to promote the neovascularization of the primate ovulatory follicle. Modulation of follicular PGE2 levels or PGE2 receptor activity may provide effective treatments for infertility or suggest targets for the development of novel contraceptives.


The effects of illness and nutritional status on various areas of the immune system are important considerations. This is especially true in attempting to define the effects of aging on immune function. Vitamin E apparently has little effect on immune function in vivo or in vitro. Burn trauma results in an overall depletion of circulating T cells, especially CD8+ cells. The receptor for IL-2, an activation marker, is induced not only by antigen and mitogen activation but by endotoxin. This can be seen frequently in older individuals. The T cell antigen receptor is under the control of at least 4 genes, which all are equally transcribed by an activation signal. Studies on genes involved in neurologic disorders have demonstrated that abnormal superoxide dismutase and tubulin gene transcription are not a features of Alzheimer's disease. Also, there are no common viral infections associated with this disorder.


Marrow myeloid cells can be separated into classes which exhibit different self-renewal capabilities carrying morphological features with respect to maturation and differing immunophenotype. The greater the self-renewal potential, the less frequent are these cells. Fluorescent activated cell sorting (FACS) has been used to develop populations of early myeloid cells which are enriched with the phenotype CD34+CD38-Lin-Thy-1 +(lo)Rho(lo). These cells have been shown to have high proliferative potential. Some AML cells show negativity for the Thy-1+lo phenotype. The t(15; 17) AML subset has been shown to be negative for CD34+CD33- cells. This data has suggested to some that the good prognosis AML subsets do not penetrate into the early myeloid compartment, whereas the poor prognostic subsets may contain cells with the immunophenotype CD34+CD38-CD33-Thy-1(lo). We will test this hypothesis by using high speed FACS cell sorting to develop populations of cells from AML patients which have the CD34+CD38- Lin-Thy+1(lo)Rho(lo) phenotype and then test this by FISH and PCR for evidence of AML cells.


The long-term objective of this grant is to analyze the central neural circuits that regulate autonomic functions with an emphasis on the cardiovascular system. Neuroanatomical techniques will be used to identify and chemically characterize specific CNS neural circuits which affect the sympathetic supply to the heart and adrenal gland. Much of the work described here will utilize the viral transneuronal tracing method-a technique which depends upon the use of an attenuated pig herpes virus to produce transneuronal infections that spread within functionally related chains of central neurons. The first project will analyze the descending projections arising from the amygdala-an important brain region thought to be responsible for emotionally triggered cardiovascular changes. The second project deals with the preoptic region and its potential role in sympathetic control. The third project explores the sympathetic control systems represented within the suprachiasmatic hypothalamic nucleus the CNS pacemaker that provides circadian modulation of some sympathetic functions. The fourth project focuses on the descending cell groups. The fifth project investigates the efferent projections of the periaqueductal gray matter in reference to sympathetic control; this area of the brain has been implicated in programming the fight-or-flight response. These studies will be useful for understanding the brain circuits responsible for sustaining and modulating basic life functions, such as blood pressure and cardiac regulation. In addition, they may provide insights into the underlying CNS mechanisms of certain clinical conditions, such as stress-induced cardiovascular changes and essential hypertension.


The overall goal of this research is to evaluate relevant issues confronting the integration of research findings into clinical practice, and to outline the perceived benefits of an automated, multi-use computer program for the assessment and treatment of alcohol abuse, and associated outcome and program evaluation functions. This multi-site, statewide (NM) randomized trial utilizes an automated computer-based assessment, treatment, and follow-up evaluation program to examine two empirical questions. First, does the program effectively disseminate the use of clinical (brief) interventions known to be efficacious for the treatment of alcohol and/or drug abuse across a variety of treatment settings? Second, how useful do treatment providers find an automated intervention and data management program to be, and what, if any, issues arise with respect to the diffusion of such an innovation?


The growing HIV pandemic and its increasing burden on women highlights the urgent need for novel safe and effective preventative strategies such as topical microbicides. We hypothesize that a truly safe and effective microbicide will require sustained, local delivery of a combination of drugs that target different steps in the HIV life cycle and a delivery system that addresses the challenges related to adherence. A safe microbicide should not interfere with host defenses nor increase the risk of selecting for drug resistant viruses. Achieving this goal requires more stringent pre-clinical and clinical evaluation of microbicides, focusing on their interactions with the genital tract mucosal environment. The goal of this iterative Program is to develop a combination microbicide delivered via intravaginal ring (IVR) to prevent the sexual transmission of HIV. This will be accomplished either through IVR formulation of a dual-active single agent, a pyrimidinedione, which is both a potent reverse transcriptase and entry inhibitor, or by combining two distinct antiretroviral (ARV) drugs. We will address the importance of the biological synergy between HIV and HSV by also combining acyclovir with ARV drugs to provide local sustained suppression of HSV. The safety of the microbicides and formulations will be explored using novel assays designed to assess their impact on the epithelial barrier and mucosal immunity. Exploratory clinical studies to evaluate the genital tract mucosal environment of Rwandan women will be conducted as defining the healthy genital tract environment will provide crucial information for establishing clinical biomarkers of microbicide safety. Pilot studies to examine the safety and tolerability of three different polyurethane rings will be conducted to identify the optimal composition for formulation of microbicide rings. Results of this study will guide the IVR formulation of our lead drugs, which will be advanced for further safety and regulatory studies including macaque PK and safety studies. A pre-Phase I clinical study is proposed to evaluate PK and safety of our lead IVR formulated microbicide and to determine if the released drug retains its antiviral activity using a spiking strategy. RELEVANCE: These studies will provide crucial information for advancement of safe and effective IVR combination microbicides to prevent HIV and to suppress HSV. These studies will also address critical gaps in microbicide science focusing on the development of sustained delivery systems and safety biomarkers. PROJECT 1: Efficacy &Safety of Multitargeted Combination Microbicides to Prevent HIV &HSV (Herold, B) PROJECT 1 DESCRIPTION (provided by applicant): Topical microbicides, a female-controlled strategy to prevent HIV, are crucial in stemming the pandemic. An optimal microbicide should protect against infection without disrupting the mucosal environment or its mediators of host defense. This project focuses on the preclinical evaluation of combination microbicides with the goal of identifying safe and effective prevention strategies. Combinations that target at least two steps in the HIV life cycle will be prioritized, thus providing protection against circulating resistant viruses and reducing the risk that the drugs will select for resistant variants. This will be accomplished through the dual activities of a single antiretroviral drug (ARV), a pyrimidinedione, which is a potent reverse transcriptase inhibitor and also blocks HIV entry, or by combining two different ARV microbicides. The biological synergy between HIV and HSV will be addressed by evaluating the possibility of delivering acyclovir in combination with the ARV microbicides to provide local sustained suppression of HSV replication. The rationale for focusing on HSV suppression reflects the pre-existing high prevalence of HSV-2 (60-90%) in the developing world and the overwhelming biological and epidemiological evidence demonstrating that subclinical HSV recurrences, which are quite common, increase the risk for HIV acquisition. Critical gaps in microbicide development are the lack of biomarkers predictive of safety and efficacy. This project will address these gaps by developing and expanding novel assays, which focus on the interactions between microbicides and the vaginal environment, using primary cells, explant cultures, and a dual chamber culture system and murine models. The proposed safety models focus on the impact of sustained drug delivery on epithelial integrity and genital tract mucosal immunity. The biological significance of any observed changes will be assessed by examining changes in the ability of HIV to traverse the epithelial barrier in the dual culture model and changes in the susceptibility to genital herpes in the mouse. Results obtained from these studies will provide crucial information for the advancement of novel safe and effective combination microbicides.


A number of self-management procedures will be systematically implemented in representative middle school classes (5th grade through 8th). The basic self-management procedures to be used in the project are self-evaluation, self-determined work requirements, self-recording, and self-scheduling. In addition, a self-control science course for middle school students is being developed. The effects of the self-management procedures on classroom academic performance will be evaluated by both within group designs and comparisons with peer control subject. Student attitudes towards school will be carefully monitored using multiple administrations of likert scales. Finally, the Peers Harris Children's Self-concept Scale. The Self-esteem Invetory, and the Intellectual Achievement Responsiblity Questionaire will be given to participating students and peer controls pre and post treatment. The scores on these scales will be analyzed to determine if the self-management procedures have any desirable effects on student self-concepts.


Flow disturbances in model vascular conduits and anastomoses will be studied under conditions of steady and pulsatile flow, including clinically encountered pulse configurations. The flow field will be analysed using dye injection, hydrogen bubble generation, and laser doppler anemometry. Emphasis will be placed on determining the formation and behavior of the boundary layer and on the origin and structure of flow separation and secondary flow. The introduction into the separated flow of fluid which has been subject to prolonged contact with the conduit wall will be determined. This will provide information on the role of contact-activated blood in downstream anastomotic hyperplasia. The contribution of fluid which has been subjected to high shear prior to entering the downstream secondary and separated flows will also be determined. This will provide information on the role of rheologic platelet activation in downstream anastomotic hyperplasia. The work will also have important implications with regard to the role of fluid dynamics in atherogenesis. Downstream anastomotic hyperplasia will be studied in paired prosthetic grafts in dog carotid arteries. The structure and progress of hyperplasia with time will be compared using dacron as the control with umbilical vein, PTFE, and autogenous vein as the test materials. An additional group of paired grafts to be studied will involve covering the downstream end of the test dacron graft with an onlay vein patch to provide a source of endothelial cells in an attempt to retard hyperplasia. Quantitative histologic analysis of the area of hyperplasia will be performed using digital computer integration of the projected microscopic image. Light, scanning, and transmission electron microscopy will be used to determine the cellular and subcellular structure of hyperplasia and neointima. These studies will provide a valuable basis for comparing commonly used prosthetic graft materials. Clinically relevant information will be gained concerning the fate of downstream anastomotic hyperplasia, the period of greatest risk of graft failure, the timing of antiplatelet aggregant therapy and of post implant angiography and angiodilation.


The research described in this proposal will provide access to tetrodotoxin, a potent neurotoxin widely used in pharmacological studies as a probe for sodium channels. This highly complex molecule will be assembled in optically pure form in a concise manner using a novel synthetic strategy. Furthermore, the proposed synthetic route is flexible enough to permit preparation of natural congeners of tetrodotoxin as well as its nonnatural analogs. The structure-activity relationship study of these derivatives, which cannot be accessed by chemical modification of tetrodotoxin itself, will make possible a better understanding of the structure and the mechanism of action of the sodium channel.


The programs of the SERCEB have immediate and projected requirements for monoclonal antibodies (mAbs) for studies of pathogenesis, immune response, vaccine development, therapeutics and diagnostics. These reagents and bio-recognition elements will be vital to the success of proposed research projects. The nature and source of the monoclonal antibodies required will depend on the objective of the research program, ranging from murine monoclonal antibodies for immunohistochemistry and detection to human or humanized mouse antibodies for development of therapeutics. Broad expertise exists within the institutions of the SERCEB in mAb development, testing and use. The spectrum of expertise in mAb development includes human, mouse, humanized mouse, rhesus, and phage display human and mouse F abs and ScFv. Finally, the antibodies will be deliverables that may have immediate application in therapeutics and diagnostics. The proposed Core C is a distributed core that uses established labs at several institutions to provide each of the needed services. The Core will provide all of the biorecognition elements needed in the SERCEB and will develop them into new assays and biosensors. The core has four Specific Aims. Aim 1. Generate human mAbs to poxvirus neutralization targets and anthrax protective antigen. Aim 2. Generate high-quality mAb reagents for SERCEB research programs. Aim 3. Develop biosensors for diagnostics or detection of select agents using novel mAbs available in the SERCEB Aim 4. To develop a database and repository for mAbs within the SERCEB that can be shared across institutions, to facilitate the other scientific programs.


We are studying the development and function of mucosal M cells and their role in immune surveillance. Our goal is to define the genes and mechanisms involved in the development and function of mucosal M cells. By identifying the critical steps and mechanisms in M cell biology, we will begin to establish their specific role in the mucosal immune response and its ability to mediate mucosal tolerance and the balance with commensal microbes. Our Working Hypothesis in these studies is that specific TNF Superfamily and TNF Receptor Superfamily genes along with coordinated expression of Jagged-1 mediate cellular interactions that specify M cell development and function. We will study two main steps in M cell development, defined by our studies on CD137- deficient mice. The first step is the commitment of M cell lineage progenitors from stem cells, which is dependent in part on ligands for the lymphotoxin receptors and the TNFa receptors. Expression of Jagged-1 by the established M cells may also inhibit generation of M cells from adjacent enterocytes. The second step, functional maturation of M cells, appears to be dependent on interactions between M cells and basolateral pocket B lymphocytes. Here, CD137 (TNFRSF9) and its ligand CD137L, may be an important signaling pair in this interaction. Two specific aims examine these components of our hypothesis: (1) How is M cell lineage commitment and development regulated by Jagged-1/Notch interactions? (2) What are the specific CD137/CD137L cellular interactions regulating M cell basolateral pocket formation and M cell functional development? Regulation of the steady state numbers of M cells in the intestinal mucosa is a dynamic process, and the process depends on an active interplay between crypt stem cells, intestinal epithelium, and lymphocyte subpopulations. This process works in parallel among Peyer's patch, Isolated Lymphoid Follicles, and Villus M cells, and will be shaped by intestinal infection and inflammation (e.g., in Inflammatory Bowel Disease). Thus, a feedback loop exists where M cell transcytosis of lumenal microbes induces mucosal immune activation, which in turn drives production of new M cells. Our studies will provide important details on both the positive and negative regulators of this process. PUBLIC HEALTH RELEVANCE: M cells are a specialized subset of epithelial cells overlying mucosal lymphoid tissues such as intestinal Peyer's patches; they have a unique selective particle transcytosis capability that enables particles as large as a few microns in diameter to cross the epithelial barrier. Thus, M cells play a central role in host-pathogen interactions. Curiously, while the host mucosal immune system relies on this transcytosis function to detect pathogens and induce protective secretory IgA immunity, many pathogenic viruses and bacteria also hijack M cell transcytosis to invade. This paradoxical dual role of M cells in infection and immunity becomes even more complex as infection- and autoimmune disease- induced inflammatory cytokines affect M cell differentiation. Regulation of the steady state numbers of M cells in the intestinal mucosa is a dynamic process, and the process depends on an active interplay between crypt stem cells, intestinal epithelium, and lymphocyte subpopulations. This process works in parallel among Peyer's patch, Isolated Lymphoid Follicles, and Villus M cells, and will be shaped by intestinal infection and inflammation (e.g., in Inflammatory Bowel Disease). Thus, a feedback loop exists where M cell transcytosis of lumenal microbes induces mucosal immune activation, which in turn drives production of new M cells. Our studies have identified a set of genes that appear to regulate M cell lineage commitment (Jagged-1, Notch) and functional development (CD137, CD137L) and interactions between M cells and associated B lymphocytes. The proposed project will provide important details on the role of these genes in both the positive and negative regulation of M cell development.


Time-resolved absorption and fluorescence spectroscopy are used to study the dynamics of protein structural changes subsequent to rapid mixing or excitation with short laser pulses. Kinetic models are used to fit and interpret the measured data. Our recent efforts have focussed on understanding the folding of the villin subdomain and the assembly of amylin. The 35-residue villin subdomain consists of three helices which interact to form a hydrophobic core. Amylin is an extremely hydrophobic 37-residue peptide which is coexpressed with insulin and fibrilized in the islet cells of diabetic patients.[unreadable] [unreadable] To probe the folding mechanism of villin we previously carried out structural and physical-chemical studies of molecules with point mutations and other sequence modifications. These studies have included determination of the structures of some of the folded states by X-ray diffraction carried out in collaboration with Dr. Thang Chiu and David Davies in the LMB/NIDDK. Conservative mutations had no significant effects on the folding rates, suggesting that the transition state for folding of villin is close to the denatured ensemble of conformations. On the other hand, the K24Nle and K29Nle mutations both stabilize the folded structure and accelerate folding by factors of ~2.5, and the double mutant, with a folding time of ~ 700 ns, ranks as the fastest folding protein studied to this time.[unreadable] [unreadable] To further explore the folding of villin we are engineering a variant which includes a pair of probes which undergo fluorescence resonance energy transfer. We have replaced the N-terminal leucine by a cysteine to which the dye Alexa-350 can be attached and the C-teminal phenylalanine at position 35 with naphthylalanine. In the native state these two residues are only about 1 nm apart so the naphthylalanine (donor) is more than 90% quenched by the Alexa-350. In the unfolded state this distance increases to ~4.0 nm and the donor is only ~40% quenched. This pair thus provides a probe of the global unfolding of villin and of the average end-to-end distance in the unfolded state. At this point we know that the folding of this doubly-labelled variant is very similar to that of the variant which we have previously studied. More detailed equilibrium and kinetic characterization is in progress.[unreadable] [unreadable] We have also begun to study the dynamics of the unfolded states of human and rat amylin. The rat sequence, which contains three prolines at positions 25, 28 and 29, does not assemble into fibrils. To do so, we make use of the triplet quenching method developed in our laboratory (Lapidus et al 2000). The C-terminal tyrosine (Y37)is replaced by tryptophan, the triplet excited state of which is quenched by an internal disulfide formed between residues C2 and C7 in both sequences. By measuring loop formation dynamics as a function of the concentration of guanidinium chloride, which solubilizes the these hydrophobic peptides, and temperature we will be able to observe the effects of intramolecular interactions on their chain dynamics. These studies may provide insight into the mechanism of assembly of human amylin into fibrils.


Population-based surveys estimate that the prevalence of methamphetamine (meth) use is 20 times higher among men who have sex with men (MSM) compared to the general population. Meth-associated sexual risk behavior is also a driving force in the MSM HIV epidemic: meth use is consistently associated with high-risk sexual behavior and sexually transmitted diseases, including HIV. Despite these alarming data, relatively few interventions have been tested among meth-using MSM, and no studies have tested the efficacy of pharmacologic interventions in reducing meth use in this population. Pilot studies demonstrate that aripiprazole (Abilify), an FDA-approved, well-tolerated antipsychotic and partial dopamine agonist, reduces the effects of meth in humans, decreases meth craving, and exhibits an excellent safety profile. Partial agonists - - ligands with target receptor affinity but low intrinsic activity - - have already been shown to be effective in treating opiate and nicotine dependence. In response to the compelling evidence supporting aripiprazole and the partial agonist approach, we propose conducting a randomized, double-blind, placebo- controlled trial of intermediate size (60 participants) and length (90 days of follow-up) to assess the efficacy of aripiprazole in reducing meth use among meth-dependent, sexually active MSM. Our specific aims are: 1) To test the hypothesis that aripiprazole 20 mg daily will reduce meth use significantly more than placebo among meth-dependent MSM, as determined by the proportion of meth-negative urines and by self- report of meth use in the aripiprazole versus placebo group. 2) To measure the acceptability of aripiprazole and placebo among meth-dependent MSM, by determining (via electronic pill caps and self-report) medication adherence to aripiprazole and placebo. 3) To measure the safety and tolerability of aripiprazole and placebo among meth-dependent MSM, as determined by the number of adverse clinical events in the aripiprazole and placebo arms. All participants will receive HIV risk-reduction counseling and brief substance use counseling. If promising, we anticipate that study results will be used to design a phase III study to determine if aripiprazole's effects on reducing meth use lead to significant reductions in meth-associated sexual risk and, if the trial sample size is appropriate, HIV incidence. [unreadable] [unreadable] [unreadable]


This proposal describes a 5 year career development plan for Dr. Pallavi Gopal to serve as a transition to successful independent physician-scientist. Dr. Gopal completed her clinical training in Anatomic Pathology and Neuropathology at the University of Pennsylvania (Penn), and is now developing an independent research and training program that will allow her to gain expertise in spatial and temporal dynamics of mRNA transport under physiological conditions and in disease. This proposal brings together diverse resources in RNA metabolism, molecular neuroscience and neuropathology and will provide superb training for Dr. Gopal to develop into an independent physician-scientist. Research will be performed under the mentorship of Dr. Erika Holzbaur, an internationally recognized expert in microtubule-based motors and real-time axon transport dynamics. This grant will provide protected time for Dr. Gopal to gain expertise in neuronal cytoskeletal, organelle, and RNA-protein dynamics through formal coursework, scientific seminars and meetings. The collaborative environment at Penn will foster utilization of novel techniques to conduct the proposed project and will provide Dr. Gopal with the training required to proceed towards a successful academic career. Amyotrophic lateral sclerosis (ALS) and front temporal lobar degeneration (FTLD) exist on two ends of a clinic pathologic spectrum but share clinical, genetic, and pathologic features. Ubiquitinated cytoplasmic inclusions composed of Tran's active response DNA-binding protein of 43 kDa (TDP-43) are a shared feature of sporadic ALS and the most common form of FTLD; there is concomitant loss of normal nuclear TDP-43 expression. Moreover, the discovery of disease-linked mutations in TDP-43 and other RNA processing proteins highlights altered RNA metabolism as a common pathogenic mechanism of neurodegeneration. However, our knowledge of how TDP-43 mislocalization disrupts its nuclear and cytoplasmic RNA processing functions and/or mediates toxicity in the cytoplasm is still incomplete. The research plan will utilize innovative approaches with real-time imaging techniques in primary neurons to test two main hypotheses: that loss of nuclear TDP-43 results in reduced dynamic flux of TDP-43 target mRNA and proteins in axons and that cytoplasmic redistribution of TDP-43 under pathological conditions results in mislocalization of TDP-43- associated mRNA. The specific aims are to: 1) Determine whether loss of nuclear TDP-43 function reduces axonal trafficking of synaptic proteins and organelle turn over and 2) Determine how (A) loss of cytoplasmic TDP-43 RNA binding function and (B) stress-induced cytoplasmic TDP-43 aggregation affect localization and trafficking of mRNA in axons and dendrites. These studies will provide temporal and spatial resolution of individual RNA transcripts in neurons in order to gain a clearer understanding of altered RNA metabolism in ALS/FTLD pathogenesis.


We have studied the susceptibility of the chicken globin gene in erythrocyte nuclei and the vitellogenin gene in liver nuclei to DNAse I pancreatic digestion. Whereas the globin gene was preferentially digested in erythrocyte nuclei, we were unable to show preferential digestion of the vitellogenin gene in liver nuclei isolated from roosters that were actively synthesizing vitellogenin mRNA.


This modified grant proposal develops methods to address the complexity of modern Pharmacokinetics/Pharmacodynamics (PKPD) population studies focusing on computational PKPD and modeling of complex PKPD systems. AIM 1, Statistical Estimation Methods for Population PK/PD Data, develops improved estimation methods for population PKPD models, and general methods to incorporate patients compliance behavior and uncertain dosing history into PKPD models. AIM 2, Modeling System- PKPD, proposes two specific projects modeling system level PKPD across multiple organs, in particular for Cocaine/Nicotine PKPD and addiction dynamics modeling, and drug absorption physiological modeling. AIM 3, Population Stochastic Differential Equations (SDE), investigates novel approaches to population model building that are based on the use of stochastic differential equation, in particular investigating methodology to include covariates into population PKPD modelsm, and apply SDE to drug-absortion modeling.


The ultimate goal of this proposal is to develop gene therapy for the treatment of hereditary kidney diseases. The carbonic anhydrase (CA) II deficient mouse is an ideal model for kidney-specific gene therapy since it manifests renal tubular acidosis and effects of gene therapy can be easily monitored. CA II is an enzyme present in cells of the kidney and other organs such as the brain and bones, and is important for acid-base homeostasis at both cellular and whole body levels. The investigators have developed a novel approach by infusing the CA II gene, driven by a viral promoter, into the urinary space of the kidney of the CA II deficient mice using liposomes as carriers, and shown a transient (3 week) correction of renal tubular acidosis. During this grant period, new strategies will be tested to improve the efficacy and duration of gene expression using this gene therapy model. Specifically, the goals will be: 1) to add polycations, polymers of positive-charged organic compounds such as basic amino acids, to the liposome delivery system in order to enhance the transfection efficiency in vivo. These polycations may serve as signals for the therapeutic gene to be delivered to the nucleus; 2) to replace the viral promoter of the therapeutic gene with the CA II promoter in order to produce a cell-specific, regulatable, and long-term gene expression in mice; 3) to evaluate the safety of the liposome/polycation-mediated gene therapy in mice; and 4) to determine whether gene therapy in CA II deficient mice will convert the phenotypical changes associated with CA II deficiency, i.e., up-regulation of CA IV, and depletion of intercalated cells, which are responsible for acid secretion in the kidney. It is hoped that results obtained from the proposed study will lead to an optimum gene therapy suitable for future clinical trials on a broad range of renal diseases in humans.


The Gordon Research Conference (GRC) on Cell-Cell Fusion will provide scientists from a wide range of disciplines with the newest research findings on the functions of cell fusion in development in a number of model organisms and in a variety of diseases. The emphasis will be on emerging universal mechanisms and future applications to cancer and stem cell research. This research conference will be the third Cell-Cell Fusion GRC and it is part of over 150 conferences that will be organized in 2011 by the GRC, an organization internationally known for the cutting edge nature of its meetings. Attendance at the meeting has grown steadily since its inception five years ago, and in 2011 the organizers expect to have 140 participants. The 2011 GRC on Cell-Cell Fusion will be held August 7-12, 2011 at the University of New England in Biddeford, ME. The program for the meeting described in this application has been designed around the theme of "Cell Fusion in Sex, Life, Development, and Disease". The organizer will deal mostly on the fundamental process of cell fusion during health and disease as well as the relationship between cell fusion and the larger fields of membrane fusion and fission during exocytosis, endocytosis, viral entry, and cytokinesis. The cell fusion community is entering a particularly fascinating phase as the fundamental principles of fusion emerging from theoretical analyses and studies on viral entry and intracellular trafficking start to be applied to understanding the functions and mechanisms of cell-cell fusion in development as well as disease. Important processes such as membrane fusion and fission, egg-sperm fusion, formation of bones, muscles, eye lens, stem cells, placenta, rare genetic diseases, and the invasion of cells by pathogens will be covered in eight sessions. This progress is being helped by classic and new methodology, including high throughput screening, developmental genetics in model organisms (e.g., fungi, flies, worms, and mice), and single-molecule biophysical methods. Exciting developments will be covered by the invited speakers, along with the latest progress on the basic mechanisms and applications of cell membrane fusion in vitro and in vivo. In addition there will be one round table discussion on myoblast fusion, a hotly debated topic that will encourage wide participation and scholarly organized discussions. The first Gordon-Kenan Research Seminar (GRS) on Cell-Cell fusion will be held August 6-7, 2011. This new GRS is organized by graduate students and postdoctoral fellows, and the activities during the GRS will be oriented to junior investigators and are intended to: (1) provide them with the basic background on mechanisms involved in membrane fusion in health and disease necessary to maximize their understanding of the science which will be discussed in the subsequent conference, (2) receive feedback on their ongoing research projects from experts in the field, and (3) facilitate their interaction with senior members of this scientific community to promote networking between junior and more senior researchers in the field. It is fully anticipate that the scientific discussions, research talks, poster sessions, and other informal interactions among the participants of this conference will contribute to advancing our understanding of novel molecular mechanisms involved in cell fusion during normal development as well as in human diseases, and will set the basis for the development of multidisciplinary projects aimed at discovering new treatments for these disorders. NARRATIVE: The Cell-Cell Fusion Gordon Research Conference (GRC) together with the first sister Gordon-Kenan Research Seminar (GRS) will bring leaders in diverse scientific disciplines together with graduate students and postdoctoral fellows constituting the future generation of researchers in the emerging field of cell fusion. The scientific presentations, discussions, round table discussions, as well as workshops during the GRC and GRS conferences on cell-cell fusion are tailored to expand our knowledge on the mechanisms by which membranes fuse in model organisms, and may contribute to our understanding of the first events in sexual development, the formation of many organs, cancer, stem cell biology, and rare diseases. The exciting, friendly, and multidisciplinary atmosphere that has traditionally characterized the Cell-Cell Fusion GRC will provide the ideal setting for the intellectual development and implementation of groundbreaking approaches to treat some cancers, as well as use stem cells to address fundamental biological questions and diseases.


Physical activity is crucial in maintaining health. Understanding how physical activity impacts pelvic floor disorders is important: half of American women have urinary incontinence or pelvic organ prolapse and one in nine undergoes surgery for these conditions. Studies to date find both protective and harmful effects of physical activity on pelvic floor disorders. The investigators, a multi-disciplinary team of urogynecologists, exercise scientists and statistician, believe this discrepancy is in large part due to inadequate ascertainment of both physical activity data and pelvic floor outcome measures. The primary aims of this study, addressed in two carefully designed case-control studies, are to determine, after adjusting for confounders, whether 1) moderate to severe stress urinary incontinence, defined by a validated questionnaire, and 2) pelvic organ prolapse, defined by structured pelvic examination, are associated with increased or decreased a) current leisure activity and b) total lifetime activity (leisure, household, and occupational). Participants will include 650 women, largely from primary care clinics: 175 with moderate to severe stress urinary incontinence, 175 with pelvic organ prolapse and 300 controls with neither condition frequency matched for age, body mass index and site. A secondary aim of this study is to establish, from the population screened for these studies, a registry of approximately 1,500 women with neither prolapse nor urinary incontinence; this will provide a foundation for future studies on the incidence of these conditions. Physical activity will be measured using validated questionnaires designed to assess current and cumulative lifetime activity in women; activity items will be supplemented with additional loading and impact physical activities. Lifetime physical activity amongst women with and without each condition will be compared using both MET hours/week (the most widely used and traditionally accepted method) and strenuous activity hours/week. PUBLIC HEALTH RELEVANCE: Pelvic floor disorders including urinary leakage and pelvic organ prolapse (dropping of pelvic organs such as the uterus into or out of the vagina) are very common. Limited data to date suggest that physical activity may be both helpful and harmful in terms of its effect on the pelvic floor. We will study the long-term effects of different levels of activity on pelvic floor disorders. Given the health benefits of activity, we believe that women should be encouraged to be active unless there is scientific evidence to the contrary.


Ocular immune privilege is maintained in part by the induction of systemic regulatory T cells mediated by events that are initiated after the introduction of antigen into the Anterior Chamber. This potent (ocular) response specifically preempts the systemic induction of tissue-damaging cell-mediated immunity, some complement fixing antibodies and atopic antibodies. Accordingly, we suggest that regulatory T cells induced by intracameral antigen could be used to selectively regulate systemic autoreactive T cells. Most investigations on the induction of regulatory T cells in autoimmunity do not discriminate between the regulation of the induction of an autoimmune response and the regulation of T cells that effect autoimmunity during on-going disease. Preliminary studies by our laboratories have demonstrated that the injection of myelin basic protein or myelin oligodendrocyte glycoprotein into an anterior chamber of mice immunized to these myelin proteins induces the production of antigen-specific regulatory T cells that suppress delayed- type hypersensitivity and/or experimental allergic encephalomyelitis (EAE) effected by an encephalitogenic T cell clone. Therefore we propose that the injection of myelin proteins into an anterior chamber of mice induces T cells that specifically regulate the induction of EAE by EAE effector T cells and may modulate on-going disease. We will test this hypothesis by raising antigen-specific splenic regulatory T cells by the injection of myelin oligodendrocyte glycoprotein into an anterior chamber and (1) Test the hypothesis that antigen-specific regulatory T cells that effect and/or induce the suppression of an encephalitogenic T cells are produced (but not necessarily activated) during an immune response to the autoantigen. This Aim will also test the hypothesis that inactive regulatory T cells are produced during an immune response to autoantigen and how these cells may be activated in vivo to prevent autoimmunity and/or eliminate an on-going autoimmune response;(2) Determine the mechanism(s) of cell-mediated suppression of EAE induced by encephalitogenic effector T cells. We will test the hypothesis that the suppression of encephalitogenic effector T cells by antigen-specific, regulatory T cells is mediated by cytokines that induce the regulatory T cells or effect suppression. The results of this study could aid our understanding of the systemic basis for the ocular regulation of autoimmunity and develop procedures for the modulation of active autoimmune disease.


The objectives of this undertaking are to originate new epidemiologic approaches to the study of cancer causation in man and animals, to develop sources of data related to specific epidemiologic problems, and to stimulate epidemiologic research in other health agencies. Opportunities for research in cancer epidemiology are based on questions arising from clinical or laboratory observations, on unusual groupings of cancer cases in the population, and on study of the characteristics of groups of persons prior to the onset of a specific type of cancer as compared with similar persons who have not developed the disease.


Dr. Diane Elliot and associates are doing a study entitled "School-Based Drug Use Prevention for Girl Athletes". Subjects will be girls involved in a sports team at school. The purpose of this study is to 1) determine student's knowledge about the effects of weight loss/physique altering drug use, and unhealthy exercise and nutrition habits; 2) measure drug use, 3) gain information about nutrition and exercise habits among female students-athletes; and 4) understand factors that influence drug use and unhealthy eating and exercise habits.


Completed CRC study. Used CDMAS only.


This proposal is to assess the behavioral predictors of weight gain and the development of overweight and obesity in adolescence and early adulthood in the National Longitudinal Study of Adolescent Health (Add Health). In cross-sectional studies researchers have observed a strong association between dieting and being overweight or obese. The cross-sectional association between dieting and weight status may be due to overweight individuals being more likely than their normal weight peers to go on diets to lose weight. Since most dieting efforts are not successful, or at least not successfully maintained, overweight individuals may remain overweight despite, not because of, their dieting attempts. Only prospective analyses can help to determine whether dieting leads to great weight gain or is simply an ineffective strategy to lose and maintain weight. Add Health is a school-based study of adolescents from 80 high schools and 52 middle schools from the U.S. This sample is representative of U.S. schools with respect to region of country, urban/city, school type, ethnicity, and school size. We seek to analyze data from Wave I (1995-1996), Wave II (1996), and Wave III (2001-2002) to assess whether weight change between 1995 and 2001 is predicted by weight change goals (e.g., desire to lose, maintain, or gain weight) or weight control practices in 1995 and 1996. Specifically we will assess whether the development of overweight and obesity is predicted by dieting, using diet pills, self-inducing vomiting, and/or using laxatives to lose weight. In addition, we will investigate whether adolescents who report using exercise to maintain their weight will gain less weight than their peers not using exercise as a weight control behavior. We will also investigate the weight change patterns of adolescents who report that they are trying to gain weight. We will assess whether self-reported use of dieting, exercise, or weight lifting to gain weight or build muscle predicts weight change over a six-year period. We will assess whether these associations exist and whether they vary by gender and/or race/ethnic group. This study will have good power to detect moderate associations (e.g., relative risk >= 1.6). Add Health is the only large prospective study of adolescents with enough males and non-white subjects to assess whether predictors vary by or are modified by gender and/or race/ethnicity, therefore our results should make an important contribution to the field.


The majority of adolescents struggling to avoid substance use during early recovery are also coping with many other issues (e.g., peer relationships) for which they require consistent support. We propose to develop an online relapse prevention program for adolescents called Staying Off Substances (SOS). Specific content in the program will be recommended to counselors based on their client's a) age, b) current motivation level, and, when available, c) reported needs and strengths on the Comprehensive Health Assessment for Teens (CHAT(R)), a Web-enabled interactive adolescent self-report assessment for use in treatment settings. Whereas current online programs for adolescents are geared for primary prevention, CHAT(R): SOS will be an intervention for use in clinical populations. Counselors will assign the client's user name to access privileges for that content and/or other content per their clinical judgment. We expect that CHAT(R): SOS will ultimately have six modules corresponding with the CHAT(R) subscales (i.e., Psychological Health, Family Relationships, Peer Relationships, Tobacco Use, Alcohol Use, Drug Use). CHAT(R): SOS will include opportunities to reinforce the skills and philosophies that adolescents learn in treatment through online interactive exercises, writing activities, and access to a supportive, online community. This resource will be available to them during treatment and post-treatment to help them navigate challenges in early recovery. Adolescents will receive tailored recommendations and an individualized user experience because CHAT(R): SOS will utilize an innovative content management and logic solution called Ektron. CHAT(R): SOS will also provide counselors with guidelines on how to use this program with their clients (e.g., in sessions, during aftercare) and custom user reports. To demonstrate the feasibility of the program concept, the Phase I study focused on the Peer Relationships domain. All feasibility criteria were met and findings indicated: (1) Proposed content was based on highly positive findings from concept mapping, as well as usability and acceptance testing with clients and experts. Ratings of potential effectiveness and appeal were high. (2) Usability testing indicated that the program was usable by the target audience, the skills and outcomes were relevant to the target audience, and the program was regarded as potentially very helpful to the treatment process. We surpassed feasibility benchmarks for both clients and experts. (3) Our technical/design team a) produced a demonstration program that was perceived by key stakeholders as highly usable and engaging multimedia program and b) determined the necessary technologies to produce the complete program in Phase II. In Phase II we will develop the program then test its efficacy in a field trial. The field trial will examine the primary hypotheses that, relative to the control condition (CHAT(R) only), CHAT(R): SOS will be associated with significantly higher motivation, higher self-efficacy, more relapse coping, and lower substance use. PUBLIC HEALTH RELEVANCE: Adolescent substance abuse is a significant public health issue. Unfortunately, the 2005 federal Treatment Episode Data Set indicated that less than 150,000 adolescents received treatment. Typically, 75% of those who receive treatment relapse within the following year. To address the need for cost-effective assessment, Inflexxion, Inc. developed the Comprehensive Health Assessment for Teens (CHAT(R)), a Web-enabled interactive self report assessment for use in treatment settings. We propose developing a supplemental online relapse prevention program called CHAT(R): Staying Off Substances (SOS). CHAT: SOS will use an adolescent's 1) age, 2) motivational level, and, when available, 3) responses to the CHAT(R) assessment to recommend Web-based relapse prevention content to address the adolescent's individual needs. This resource will be available to adolescents during treatment and post-treatment to help them navigate challenges in early recovery.


This project addresses the NHLBI RC2 GO application entitled "Characterizing Differentiated Heart, Lung, and Blood Cells Derived by Reprogramming Human Embryonic and Induced Pluripotent Stem Cells." Emerging technologies to generate induced pluripotent stem cells (iPSCs) by reprogramming human somatic cells promises to revolutionize biomedical research and clinical medicine. Through in vitro culture methods, iPSCs can be differentiated into numerous cells types derived from all three germ layers. This raises the possibility that patient-derived iPSCs can be used to create relevant tissues for the study of many human disorders. In addition, iPSCs may provide starting material to manufacture transplantable cells for transfusion and regenerative therapies. However, the field is in its infancy and many core questions must be solved in order to realize these exciting long-term prospects. This proposal seeks to advance the use of iPSCs for the study of normal and pathological hematopoiesis. Multiple investigators with broad areas of expertise in hematopoiesis, embryonic stem cell/iPSC biology, chromatin biology, clinical hematology, bioinformatics, cell banking and bioethics/regulatory affairs will work together to pursue the following global issues 1) Mechanisms by which hematopoietic developmental potential might vary between different normal iPSC clones;2) The extent to which iPSC-derived hematopoietic precursors resemble normal ones with respect to cellular phenotypes, gene expression and epigenetic signatures;3) Whether hematopoietic disease phenotypes can be recapitulated by in vitro manipulation of patient-derived iPSCs. We will execute these studies using novel methods to create and culture iPSCs and state-of-the art tools to analyze and manipulate their resident genomes. Pursuit of these problems will serve as a framework in which to develop a facile infrastructure where investigators at our large pediatric institution can create, analyze, bank and distribute iPSCs from any patient of interest. If successful, this work will help to accelerate practical applications of iPSCs for the study and treatment of human diseases. This work will be based at Children's Hospital of Philadelphia with subcontracts to The Pennsylvania State University (State College, PA) and The Coriell Institute for Medical Research (Camden, NJ). The project will create 6 new jobs, thereby stimulating the economy in three different regions of the Northeastern United States. PUBLIC HEALTH RELEVANCE: Efforts to better understand blood production from patient-derived induced pluripotent stem cells (iPSCs) will enhance our understanding of blood disorders and generate new therapeutic approaches. Additionally, this work could create new general paradigms for studying the genesis of many normal tissues and their associated diseases.


Legionella pneumophila is a gram-negative bacteria that utilizes a type IV secretion system to alter its intracellular trafficking in host macrophages. Internalized Legionella containing vacuoles (LCV) immediately avoid transport to lysosomes and within 15 minutes of internalization, begin remodeling their phagosomal compartment with ER-derived host cell vesicles. Dot/icm mutants are efficiently transported to and fuse with lysosomes suggesting that the type IV secretion apparatus is required for altered LCV trafficking and remodeling. Identification of factors utilized by Legionella to alter trafficking and remodel its intracellular vacuole may provide new targets for therapeutic intervention. Furthermore, understanding how ER-derived vesicles are recruited to and fuse with a membrane-bound compartment originating from the plasma membrane may provide insight into novel host cell functions. Confocal microscopy, coimmunprecipitation studies, 2D-gel electrophoresis and in vitro budding and fusion reactions will be used to identify host and/or Legionella derived SNARE-like molecules that play a role in ER-derived vesicle fusion with LCV.


General rationale and organization of the Cores The main potential technical barriers to efficient execution of the biology are (1) the quality control and execution of the 3-D culture system, and (2) the requirement for expertise and ready access to state of the art confocal microscopy. To make these essential technologies readily available to everybody, and to coordinate the efforts of the various groups, we propose two core facilities, one to coordinate the 3-D culture methodology (3-D Core), and another to manage the confocal microscope (Confocal Core). Although the 3- D Cell Culture and Microscope Cores will be described separately, they will be located in adjacent labs and are designed to work together to standardize data collection procedures and provide a seamless transition from the 3-D culture phase to the confocal analysis phase of each study.


The focus of this proposal is on the mechanism of host lysis by bacteriophage. Building on recent progress, it is proposed to investigate the biochemical and genetic properties of holins, small phage-encoded membrane proteins that act as the timers of phage infections. Holins have the remarkable ability to accumulate during the phage infective cycle without harming the cell, then suddenly triggering to permeabilize the membrane. This terminates the infection and activates muralytic enzymes called endolysins, or lysozymes, resulting in degradation of the cell wall, leading to bursting of the cell and release of the progeny virions. The work is aimed at determining how these proteins can form holes in membranes, and how the scheduling of the hole-forming event is programmed into the sequence of the holin. Fundamental issues of lipid-protein and protein-protein interactions in membranes will be addressed, including an investigation of how integral membrane domains of some lysis proteins actually are able to exit the membrane upon physiological cues. [unreadable] [unreadable] The holin-endolysin mode is completely general for all phages except those with very small genomes. However, single-stranded DMA and RNA phages, limited to 3 - 10 genes for their entire genome complement, accomplish host lysis by expressing single genes. In two of these cases, recent progress has shown that the phage lysis protein causes lysis by inhibiting different enzymes in the murein precursor biosynthetic pathway. It is proposed to investigate the molecular basis by which these "protein antibiotics" effect inhibition of these conserved enzymes. Other small single-stranded RNA phages effect lysis by an unknown mechanism, the elucidation of which is another goal of this project. [unreadable] [unreadable] Public health implications: These studies are critical to our understanding of how bacterial viruses, or phages, kill their prey and effect dispersal of their progeny. This may have direct practical benefits because there is a growing consensus that phages, as natural antibacterial agents, will become an important tool in combating bacterial pathogens, which are increasingly resistant to available antibiotics. In addition, the research may reveal new modes for design of chemical antibiotics. [unreadable] [unreadable]


Despite advances in resuscitation of trauma victims, a large number of such patients subsequently die of sepsis and multiple organ failure. Improvement in survival of such victims should, however, be possible. Our hypothesis is that the generation of toxic substances and the various metabolic and microcirculatory alterations produced after severe injury are not adequately corrected by resuscitation with conventional fluids alone, and that such alterations eventually lead to organ failure. Although numerous attempts at overcoming these problems have been made, including the use of hypertonic solutions, substrates, inotropes and other agents, there has not been a systematic investigation of the specific needs and interdependance between the heart, liver, and kidney following low flow states. Low flow produces an ischemic environment resulting in depletion of tissue energy stores, cellular electrolyte imbalance, generation of toxic substances, cell swelling and eventually microcirculatory and organ failure. Our plans are to determine the specific needs of the organs which are not met with volume resuscitation alone, and to then correct those abnormalities by pharmacologic means. Organ blood flow and function following injury may be better maintained by providing osmotic and oncotic agents along with metabolic support and agents that improve microcirculatory flow and inhibit the generation of toxic substances. We plan to develop a pharmacologic solution (i.e., composite of pharmacologic agents which will act synergistically) which, together with appropriate volume replacement, will maintain cell and organ function and host defense mechanisms after injury better than conventional solutions alone, and thus prevent subsequent multiple systems failure. Such a solution (osmolarity approximately 600 mOsm) will initially contain dextrose, albumin and electrolytes. To this solution we will systematically add dopamine, diltiazem, imidazole and ATP-MgC12 individually and in various combinations. In this manner, we will evaluate the action of each of the above agents individually and their interaction with each other. Such an approach should yield an optimal combination to produce synergistic beneficial effects on cell and organ function following injury, and produce resistance to a subsequent septic challenge. If these studies uncover a lack of improvement in any cell, organ or system function, the constituents of the pharmacologic solution will be modified to specifically address that problem. To accomplish these goals we will utilize a rat hemorrhage model.


While tremendous advances in pharmacotherapy for adults have been achieved in recent years, expanding the benefits of these new therapies to infants and children remains a significant challenge. Important developmental changes in pharmacokinetics, pharmacodynamics, disease presentation and progression all impede direct translation of adult therapeutics into pediatrics. The discipline of pediatric clinical pharmacolog provides necessary training in developmental physiology to leverage existing knowledge and guide rational therapeutics for infants and children. However, the current pool of pharmacologists and pediatric sub- specialists with formal training in clinical pharmacology is both small and aging. This T32 Fellowship Program led by the Department of Pediatrics at the University of California, San Diego (UCSD), entitled Developing Pediatric Clinical Pharmacologists for the Advancement of Therapeutics (DPCPAT), will provide cutting-edge research training and mentorship in pediatric clinical pharmacology, enabling our trainees to emerge as leaders among the ranks of pediatric academic investigators. The biomedical research community at UCSD has a long- established reputation of excellence, and now ranks in the top 5 in NIH research funding of all US institutions. Notably, the last 5-10 years have witnessed an impressive academic expansion within the UCSD Department of Pediatrics and its research mission, coupled with the solidification of the synergistic relationship among Rady Children's Hospital San Diego (RCHSD) and the UCSD Schools of Medicine and Pharmacy. We will provide ample evidence that the timing for creation of the DPCPAT training program at UCSD has never been better, and that our research training environment and faculty are truly world class by any measure. The overarching goal of the DPCPAT program will be to prepare Pediatric Physician-Scientists for the lifelong challenges of a sustained, independent and highly productive research. Dr. David Brenner, Dean and Vice Chancellor for Health Sciences, Dr. Jack Dixon, Associate Vice Chancellor for Scientific Affairs, Dr. Gabriel Haddad, Chair of the Department of Pediatrics and Dr. James McKerrow, Dean of the School of Pharmacy, have provided letters of UCSD institutional commitment to our proposed DPCPAT, confirming the central importance of this Program to the academic mission of the UCSD School of Medicine. The key objectives of our DPCAT are as follows: (1) To increase the number Pediatric clinical pharmacologists and Clinician-Scientists engaged in clinical pharmacology research as applied to child health; (2) to attract outstanding young pediatric clinicians to UCSD and to expand their translational capacity through clinical pharmacology methods; (3) to facilitate career development of DPCAT trainees under the guidance of world class, established investigator-faculty mentors; and (4) to cultivate the early careers of women and minority investigators in pediatric therapeutics. With our impressive group of mentors with outstanding training track records, UCSD DPCPAT Fellows will be poised to be tomorrow's leaders in pediatric clinical pharmacology.


The main objective of this proposal is to analyze the molecular mechanisms by which retinoic acid inhibits mouse skin tumor promoter 12-)-tetradecanoylphorbol-13-acetate (TPA) induction of epidermal ornithine decarboxylase (ODC), a biochemical change associated with skin tumor promotion by TPA. We first demonstrated that the mechanism by which certain retinoids inhibit skin tumor promotion is due to their ability to inhabit it the induction of ODC by TPA. However, the nature of retinoic acid inhibition of ODC induction by TPA remained undefined. Recently, we found that retinoic acid inhibits TPA-caused synthesis of ODC protein. Retinoic acid may inhibit the synthesis of ODC protein at the transcriptional and/or translational levels. In a preliminary experiment, we found retinoic acid inhibits TPA-increased level of ODC RNA. Thus, we hypothesize that TPA-induce ODC activity (peaks at 6 hr and approximately 200-fold above control) is the result of increased synthesis of ODC protein at the transcriptional level and retinoic acid inhibits ODC induction by inhibiting TPA-increased ODC messenger. Specifically, we plan to investigate: 1) a correlation among TPA-induced epidermal ODC activity, ODC protein, rate of ODC synthesis, and ODC mRNA levels, 2) whether retinoic acid inhibition of TPA-induced ODC activity is proportional to the inhibition of ODC protein and ODC mRNA levels, 3) the mechanism of inhibition by retinoic acid of ODC messenger. We will determine whether TPA causes amplification of ODC gene(s) and retinoic acid inhibits this process. Experiments will be performed with CD-1 mice. Total RNA from skin will be isolated by urea extraction and CsC1 gradient centrifugation. Poly(A)+-contining mRNA will be fractionated by affinity chromatography on an agarose poly(U) column. ODC mRNA will be quantitated by solution hybridization using [32p]-labeled single stranded probe and Dot-blot analysis using radiolabeled cDNA probe. Size of ODC mRNA transcripts will be determined by Northern blotting. Mouse skin genomic DNA will be analyzed by Southern blotting. We will analyze both hybridizable and translatable ODC mRNA; translatable ODC mRNA will be determined by translation in a reticulocyte lysate cell-free syftem. Amount of ODC protein will be analyzed by the radioimmunoassay using [3H] difluoromethylornithine-labeled ODC. These data will permit an evaluation of the role of transcription in the regulation of ODC. These data will permit an evaluation of the role of transcription in the regulation of ODC induction by TPA and whether retinoic acid inhibits TPA induction of ODC at the transcriptional level.


Integrins are critical components of the immune system through their activities in directing lymphocyte development, movement and activation. The function of the b2-integrin CD11c in vivo is largely unexplored. Our data show that CD11c is induced on primary and memory CD8 T cells responding to infection. Moreover, in the absence of CD11c, the CD8 T cell response is severely diminished indicating an integral costimulatory role for CD11c in driving CD8 T cell activation. The studies proposed here will test the hypothesis that CD11c controls CD8 T cell response by promoting T cell- DC interactions early after activation. Three specific aims are proposed to analyze the mechanisms of CD11c costimulation: Specific Aim 1. To determine the role of CD11c in the early CD8 T cell response to infection. Using bacterial and viral infections in conjunction with CD11c-deficient CD8 T cells, the role of CD11c in promoting proliferation, differentiation and survival of CD8 T cells will be tested. Specific Aim 2. To analyze the role of CD11c expressed by CD8 T cells in determining the anatomy of the immune response to infection. Our preliminary results using in situ MHC class I tetramer staining identified a series of stepwise events that drive CD8 T cell activation in response to infection with Listeria monocytogenes. In situ analysis using confocal microscopy will be performed to assess the role of CD11c in activation and movement of responding endogenous and adoptively transferred CD8 T cells. Specific Aim 3. To analyze the role of CD11c in development and function of dendritic cells. The high level expression of CD11c by DC strongly suggests an important role for CD11c in DC function. However, the analysis of CD11c-deficient DC is complicated due to the lack of a reliable marker for DC identification. We have produced transgenic mice expressing a fluorescent reporter protein under control of the CD11c promoter to circumvent this issue. These mice, when rendered CD11c-deficient, will provide the means to track and analyze CD11c-/- DC under conditions of homeostasis or infection. In sum, these studies will provide the first analysis of the role of CD11c in CD8 T cell and DC activation. PUBLIC HEALTH RELEVENCE: This proposal is focused on understanding the role of a specialized adhesion molecule, the integrin CD11c, in CD8 T cell activation in response to a bacterial infection. CD8 T cells are cytotoxic lymphocytes involved in killing of infected cells and play a central role in protection against intracellular bacterial infections. Therefore, this research has substantial impact on understanding immunity to infection and holds implications for design of effective vaccines.


Hospice is a key model of home care for seriously ill individuals and those with dementia represent the fastest growing group of hospice users. Hospices support patients and caregivers through interdisciplinary care with the goal of enabling individuals to remain at home at the end of life. Living in the community in the last phase of life for those with dementia has increased in the last two decades. Although site of death for decedents with dementia in both 2000 and 2015 was predominantly a nursing home, the proportion dying at home in the community almost doubled from 15% in 2000 to 26% in 2016. Community residential alternatives to long term care in nursing homes, such as assisted living facilities, are becoming more prevalent and are considered a positive phenomenon for promoting independence and improving quality of life for those with dementia. Yet research on end of life care for those with dementia is primarily focused on the nursing home setting and little is known about characteristics of those with dementia who remain in the community and the types of supportive services (e.g., personal care, medication assistance, meal preparation, transportation) that they receive. Also unknown is the extent to which hospice is added as an extra layer of support for those with dementia and their families who remain in the community. To address these gaps in knowledge, we propose a supplement to our geriatric palliative care focused Claude D. Pepper Older Adult Independent Center. We will employ the Medicare Current Beneficiary Survey, a nationally representative panel survey of Medicare beneficiaries linked to Medicare administrative data and regional characteristics, which provides a unique opportunity to examine those with dementia living in a range of settings including private residences, assisted living facilities, retirement homes, continuing care communities, and nursing homes, with a range of supportive services (e.g., personal care, medication assistance, meal preparation, transportation). We will determine patient, family, and regional characteristics associated with residential setting and associated with hospice enrollment across residential settings for those with dementia. We will also describe the trajectory of transitions across residential settings at the end of life for those with dementia. The proposed research will lay the foundation for a program of research regarding hospice use in the context of dementia and across residential care settings leading to a more comprehensive picture of the consequences of the increase in home care for those with dementia at the end of life.


This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This event is held on demand during the summer months, if some students cannot attend the NBCR summer institute due to schedule conflict..


This K award will allow me to develop as an independent investigator, focusing on the role that paternal factors play in the risk of schizophrenia in offspring, specifically paternal reproductive health. The proposed program includes research and training in the translation of findings from (1) observational studies into basic experiments through collaborative efforts using animal models and (2) from experimental data into testable hypotheses using epidemiology. The training activities build on my prior experience in psychiatric epidemiology and toxicology, including coursework and research training in neuroscience and experimental animal studies. The skills that I acquire will be directly applicable to the study of many kinds of exposures relevant to schizophrenia. The research program described in this application is based on new findings that prenatal and pre-conceptual exposure that affect paternal reproductive health, including advanced paternal age and exposure to lead (Pb), a known toxin may increase susceptibility to schizophrenia and other psychotic disorders in offspring. Lead exposure results in reductions in fertility in both men and women, and is associated with behavioral abnormalities in animal models of paternal exposure. Two phases of research are proposed that will involve parallel career development activities in both training and research: (I) studies of behavioral phenotypes congruent with schizophrenia using mouse models of paternal lead exposure and advanced paternal age and (II) epidemiologic studies of paternal Pb exposure and SSD. At the conclusion of this award I will design of new collaborative programs, incorporating lifecourse epidemiology and basic research with the goal of submitting an R01 application.