PI: Tang, Guangwen
Title: Retinol Equivalence of Plant Carotenoids in Children
Abstract: This project is to determine the vitamin A value (equivalence) of dietary provitamin A carotenes from spinach, Golden Rice, and pure Beta-carotene (B-C) in oil. These experiments will be conducted in children (ages 6-8) with/without adequate vitamin A nutrition. As plant provitamin A carotenoids are a major and safe vitamin A source for a vast population in the world, it is essential to determine the efficiency of provitamin A carotenoid (mainly B-C) conversion to vitamin A. By introducing B-C into rice endosperm, Golden Rice may directly benefit consumers by providing vitamin A nutrition. Our investigation uses hydroponically grown, decadeuterium labeled spinach and Golden Rice, synthetic Beta-C-d10 and a vitamin A isotope reference, decadeuterated retinyl acetate (RAc-d10), to evaluate the bioavailability and the bioconversion of plant provitamin A carotenes to retinol as compared with B-C in oil capsules in vivo. Our objectives will be to test the following hypotheses and to make the following determinations: (1) The absorption and bio-conversion of provitamin A carotenes taken by children are different between spinach, Golden Rice, and B-C in oil capsules. (2) The absorption of provitamin A carotenes and their bioconversion to vitamin A are different in children with or without adequate vitamin A nutrition. (3) To define the vitamin A equivalence(s) of dietary spinach, Golden Rice, and a B-C in oil dose by using an isotope reference method in children with or without adequate vitamin A nutrition and to compare those values with values derived from model based compartmental analysis. (4) To determine the number and time of blood samples needed for future studies in various field settings on the retinol equivalence of a large number of plant sources. Seventy-two children each will take two meals, lunch and supper, containing equal amounts of B-C in labeled spinach (along with white rice), or Golden Rice (along with light colored vegetables), or B-C oil capsules (along with white rice and light colored vegetables), every day for 7 days. Before the two meals, the volunteers will take a breakfast with a RAc-d10 dose as a reference for 7 days. The enrichment of labeled B-C and labeled retinol in human circulation will be determined using advanced liquid chromatography / mass spectrometry and gas chromatography / mass spectrometry. Through the applications of these novel technologies, we will be able to determine the relative biological activities of endogenous carotenoids; that is, the vitamin A value of spinach, Golden Rice, and B-C in oil capsules for children with/without vitamin A malnutrition. This study will be of importance in planning vitamin A deficiency prevention strategies and also will provide useful information regarding the potential efficacy of a bioengineered crop to provide vitamin A nutrition.

PI: Taylor, Allen
Title: Exploiting Nutrition on Delay Cataract
Abstract: Cataract, or opacification of the lens, is a major cause of visual disability. Cataract extraction is the most frequently performed surgery and costs for cataract-related problems account for the largest line item in the Medicare budget. Delaying cataract by 10 years would halve the number of cataract extractions and save over $1 billion annually. Studies suggest that nutrition may be exploited to diminish the prevalence of or retard the progress of age-related cataract. However, there is little epidemiologic research regarding relations between major components of diet (other than for antioxidants) and risk for prevalence or progress of cataract. This knowledge is important in order to know if people at high risk or with early lens opacities might benefit from proper dietary management. This NEI R03 proposal from the Nutrition and Vision Project (NVP) - a collaboration between Tufts University and the Nurses' Health Study (NHS) at Harvard University and Harvard Medical School - exploits unique existing ophthalmologic and nutritional databases to assess three previously untested hypotheses about the extent to which nutrition affects risk for prevalence and progress of opacification. The NVP ophthalmologic data set includes two sets of graded lens images, gathered five years apart, from 451 women (ages 53-73 y) who are a subset of the NHS. The NHS data set includes long-term nutritional data from 5 food frequency questionnaires and a plethora of data on other personal and environmental factors, all gathered during a 15-year period before the baseline eye exam. Combined, these data will be used to test three hypotheses: that risk for cataract is lower in persons who a) consume higher levels of specific fats (monounsaturated, polyunsaturated, and saturated dietary fats, n-3 fatty acids, etc.), b) consume lower glycemic index foods, and c) eat according to particular dietary patterns.

PI: Taylor, Allen
Title: Ubiquitination Lens Proliferation/Differentiation
Abstract: Lens formation requires a chronologically and spatially executed program of cell division and proliferation, as well as exit from the cell cycle and differentiation into lens fibers. These processes are controlled in most cell types by the timely degradation of cell cycle regulators by Ubiquitin Proteasome Pathways (UPPs). Aberrations, in these processes frequently result in microphthalmia or cataract. Yet there are few published papers that address either the lens cell cycle or its control by- UPPs. During the ongoing grant, we demonstrated that Ub and Ubc3 (a Ub ligase) are required for proliferation and differentiation. However, regulation of the cell cycle by-these moieties occurs at the G2/M transition and not, as predicted, at G1/S. We now seek to determine if the same controls are observed in vivo by directing expression of mutant Ub to the epithelial and differentiating lens cells in transgenic animals. Our recent data beget two new overall hypotheses: 1) proteolysis involving Ub and Ubc3 is required for proliferation, differentiation, and lens formation in vivo; 2) a UPP which involves an undescribed Ubc3-E3 interaction is involved in control of G2/M events in lens. These overall hypotheses are separated into 4 specific aims, to test the hypotheses that: an active UPP is required for proliferation, differentiation and lens formation in vivo; ubiquitination is required for the lens cell cycle, particularly in G2/M; lens Ubc3 cooperates with an E3, which we will identify, to control the G2/M transition; and control of the cell: cycle at G2/M requires ubiquitination of the APC regulators in a Ubc3-dependent process. These studies will address a major objective of the NEI lens and cataract program: to characterize controls of lens cell division and differentiation, and their roles in formation of secondary cataract. The long-term objective is to prolong function of 1) the natural lens by gaining a better understanding of processes involved in control of lens cell proliferation and differentiation, as well as in lens formation, and 2) implanted lenses by limiting secondary cataract due to overgrowth. Our recent papers show that these results will also lead to a better understanding of corneal wound healing and retina responses to stress. This information, and our novel "reagents", will also find use in new ways to limit secondary cataract, prolong function of glaucoma medication, limit cancer and in fighting other proliferative maladies. We are joined in this effort by excellent collaborators, each of whom is a leader in his field.

PI: Thorley-Lawson, David
Title: Computer Simulation of Epstein Barr Virus Infection
Abstract: Epstein-Barr virus persistently infects >95% of the adult human population. It causes infectious mononucleosis and is associated with several important human cancers. Considerable progress has been made recently in understanding how EBV establishes and maintains persistent infection. It is apparent that EBV uses various combinations of latent gene expression patterns to manipulate the biology of normal B lymphocytes. This allows EBV to establish latent persistent infection in resting memory B cells in the blood and replicate in plasma cells in the lymphoepithelium of Waldeyer's ring (tonsils and adenoids). However, these studies have all been static and an understanding of the dynamics of the infection is lacking. In the absence of a suitable animal model, we propose to develop a new generation of computer simulation. The application of supercomputing on distributed clusters of processors will allow us to develop an unprecedented level of sophistication and complexity. We will employ an agent-based approach, which makes it possible to represent the actual anatomy of the relevant tissues and organs and the dynamic changes that occur over time and space. These features are not possible with traditional mathematical models based on ordinary differential equations. A simulation involving approximately 108 agents is within reach and should produce realistic results, based on previous experience with similar simulations of traffic and wireless communication systems. We will use sensitive PCR and immunological techniques to accurately measure levels of virus shedding, virus infected cells, EBV specific CD4 and CD8 cells and neutralizing antibody as acute infection resolves into persistent infection. The data will then be used to inform the computer simulation. This approach will allow us to define the infection parameters necessary to predict the observed kinetics of infection. Ultimately, the simulation will be used to test the effects of perturbations such as lowering the numbers of T cells (immunosuppression) eliminating infectious virus (ant-viral and/or vaccines) and to ask if complete clearance (i.e. cure) of the virus is realistic or even possible.

PI: Thorley-Lawson, David
Title: Epstein Barr Virus Latency
Abstract: Epstein-Barr virus is a herpesvirus that infects >90% of the human population. It has two distinguishing characteristics. First it is able to maintain a life long persistent infection in healthy hosts and second it is associated with several human lymphomas and carcinomas. This proposal will address central issues related to these two properties. Persistence: EBV establishes a persistent latent infection in memory B cells. Much is known about how it does this but less is known about how the latently infected cell produces infectious virus to spread to other hosts. Our preliminary data indicate that the signal for viral replication is the terminal differentiation of the latently infected cell into a plasma cell. We will use standard molecular biological tools to identify the role of plasma cell specific transcription factors in activating the EBV lytic cycle. Since plasma cells replicate the virus only when they are fully differentiated it is likely that they release infectious virus when they migrate to the bone marrow. This would result in early B cell progenitors becoming infected. These cells could also provide a site of life time persistent infection. Therefore, the second aim of this study will be to test the role of the bone marrow as a second site of viral persistence. This will be achieved by fractionating the bone marrow into the known subsets of B cells and testing for the presence of the virus by quantitative DNA and RT PCR techniques that we have developed. Neoplasia: One of the commonest tumors associated with EBV is nasopharyngeal carcinoma. There remains little known about the molecular basis of this tumor and its association with EBV. We will use Affymetrix chip technology to provide a molecular genetic definition of NPC that distinguishes Type II and Type III (poorly and undifferentiated NPC) and tumors negative and positive for the EBV encoded oncogene LMP1. Our preliminary data demonstrates the feasibility of this approach. This analysis will identify candidate marker genes for the different types of NPC and test their roles in in vitro and in vivo models. Specifically this approach will be used to identify genes and signaling pathways activated by LMP1. This will provide a molecular basis for explaining the role of LMP1 in NPC.

PI: Thorley-Lawson, David
Title: Host Immunity to EBV Infection in Vitro and in Vivo
Abstract: The long term objective of this study is to develop a deeper understanding of persistent infection with Epstein-Barr virus (EBV). EBV has the capacity to drive the proliferation of resting B lymphocytes and this makes it a risk factor for human cancers such as Hodgkin's disease, Burkitt's lymphoma, immunoblastic lymphoma and nasopharyngeal carcinoma. However, the virus is able to persist in a quiescent state in vivo where it specifically targets resting memory B cells. By understanding how EBV can persist in most individuals without causing disease we hope to gain insight into what goes wrong when the virus does cause neoplastic disease. This study wilt employ sophisticated cell fractionation techniques and quantitative RealTime DNA and RT PCR assays to address four unresolved issues around EBV persistence. 1. Does acute EBV infection, infectious mononucleosis (AIM), represent a disordered state of EBV infection or simply an amplified version of the stable, long term carrier state? 2. Does EBV, like other herpesviruses, shut off the expression of all protein coding genes when it reaches its final site of persistence - long lived memory B cells in the peripheral blood? 3. What is the nature and origin of the latently infected memory cells proposed as the site of EBV persistence? Are they bona fide memory cells? Does antigen play a role in the production and/or maintenance of these memory cells or do latent proteins perform these functions? How rapidly do the infected cells turn over? 4. Are epithelial cells of the nasopharyngeal lymphoid system e.g. tonsils infected with EBV in vivo or infectable in vitro? Previous studies have analyzed EBV infection of epithelial cell lines and tissues from sites other than the site of persistent infection - the nasopharyngeal lymphoid tissue. However, epithelial tissues are biologically diverse so we will focus our studies on the biologically relevant epithelium from the tonsil.

PI: Thornton, Ronald
Title: Web-Delivered Interactive Lecture Demonstration: Creating an Active Science Learning Environment Over the Internet
Abstract: Educational research has shown that traditional instruction in physics (and other sciences) is rarely effective for teaching conceptual understanding even with academically-talented students and almost totally ineffective for under-prepared students. This project intends to develop further methods and tools that can facilitate effective teaching even in traditional environments. We are proposing a web delivery system for a proven interactive pedagogy, Interactive Lecture Demonstrations (lLDs) which we developed earlier. Prototype WebILDs, like in-class ILDs, have been shown to lead to very significant changes in conceptual understanding and are even easier to implement. Starting with this proven, research-based, pedagogical method and our successful WebILD prototype software we will

• refine, using student learning as a guide, the Motion, Force, and Energy WebILD prototype by testing it with larger and more diverse introductory course audiences and using it for learning at a distance-
• create WebILD sequences in additional physics topic areas.
• create additional WebILD sequences in another science disciplinary area.
• refine the delivery and administration software and make it and the WebILD sequences nationally available.

This project focuses on the learner and uses well established conceptual evaluations to measure learning. We will use the flexibility of internet-delivery to adapt ILDs to increase their already considerable efficacy with under-prepared students including women and minorities.

This project meets a need for learning at a distance, for pedagogical methods that work for internet delivery, and for methods to teach teachers. Even during development we will reach thousands of students per year.

PI: Trimmer, Barry
Title: Central and Peripheral Actions of Nitric Oxide
Abstract: Some neurons signal by producing an unstable soluble gas named nitric oxide (NO). This unusual messenger was discovered only recently and comparatively little is known about its role in the brain. We have identified individual NO-producing and responding neurons in the living nervous system and our goal is to establish how they communicate. We are also studying how NO controls light production in the firefly lantern. This work has potential application in understanding how groups of neurons are coordinated and how the brain is damaged by a stroke or other traumatic injury.

To carry out this research we use a wide range of techniques from molecular biology through biophysics to mechanical engineering. Modern biology is now in a position to cross traditional disciplines and recruit the expertise of physicists, mathematicians and engineers in trying to understand the complexity of living things.

PI: Tzipori, Saul
Title: Specific Human Monoclonal Antibodies for HUS Prophylaxis
Abstract: Our objective is to develop an immunotherapeutic formulation for the treatment and prevention of complications associated with Stx-producing Escherichia coli (STEC) infections. Clinical isolates of STEC are known to predominantly produce Stxl, Stx2, and/or Stx2c. Children are particularly susceptible to development of Stx-mediated HUS. Our hypothesis is that administration of Stx-specific antibodies will prevent or modify the outcome of infection for individuals at risk of developing HUS. In the earlier awards, we have generated a panel of human monoclonal antibodies (Hu-mAbs) specific for Stx 1 or Stx2. Using the gnotobiotic piglet model, we have shown that Stx-specific Hu-mAbs neutralize Stx and prevent development of Stx-mediated complications. We now wish to determine which Hu-mAbs should be included in a formulation suitable for clinical evaluation. Based on superior efficacy, four Hu-mAbs specific for Stx2 (3 against the A subunit and 1 against the B subunit), and 2 for Stxl (both against B subunit) have been selected as candidates. The next step is to determine which combination of Hu-mAbs, is both compatible and highly effective. In this proposal we plan to define the structural and functional characteristics, which facilitate protective efficacy of Stx-1 and Stx2-specific Hu-mAbs (Specific Aim 1). Affinity and efficacy of each HumAb will then be studied against their respective toxin (Specific Aim 2). The efficacy of protection of a given antibody dose will then be determined in terms of time after bacterial infection (Specific Aim 3). Finally, combinations of Hu-mAbs specific for B subunit of Stx 1 and A or B subunits of Stx2, will be examined for relative efficacy and compatibility, to determine which is the most effective and thus suitable for clinical evaluation (Specific Aim 4). At the conclusion of these experiments we will have determined the components, and optimized the formulation of Hu-mAbs which will be recommended for testing in human patients. The Hu-mAbs will first be characterized and ranked according to their efficacy, affinity and compatibility with each other. The optimal amount of each Hu-mAb in the formulation required to provide the longest protection after bacterial infection will also be established. This is not a hypothesis-driven proposal, but an essential segment for the characterization of a promising therapy for HUS, against which currently there is no effective treatment.

PI: Vetter, Douglas
Title: Investigations into the Mouse Olivocochlear System
Abstract: Corticotropin releasing hormone (CRH) receptors, and urocortin, a member of the CRH family of peptides, has recently been discovered expressed in outer hair cells and olivocochlear terminals, respectively. While analysis of the role played by urocorUn, the only known ligand expressed in the inner ear capable of activating the CRH receptors, has been analyzed using urocortin deficient mice, nothing is known of the developmental expression pattern or role of the CRH receptors in the inner ear. The CRH receptors are G protein coupled receptors, and stimulate the cAMP second messenger-signaling cascade. We hypothesize that activation of the CRH receptors may represent one phenomenon underlying protection from noise induced hearing loss. The mechanisms of action may include phosphorylation and inactivation of the sK2 calcium-activated apamin-sensitive potassium channel. In order to further analyze the morphological aspects of the CRH system in the cochlea, its functional role in hearing and protection from noise induced hearing loss, and finally, to assess the cellular mechanisms of action associated with activation of the CRH receptors, three specific aims are proposed. First, we will establish the developmental and adult expression pattern of the CRH receptors in the inner ear. Successful completion of the experiments of this specific aim will establish the precise identity of the cells within the inner ear that express the CRH receptors, and identify the postsynaptic elements of the urocortin immunopositive fibers at the ultra structural level. Second, we will establish the functional roles CRH plays in hearing, and whether they participate in protection of the inner ear from noise induced hearing loss. This aim will be accomplished using mice that lack the gene for either the type 1 or the type 2 CRH receptor, or that lack both. Finally, we will use these mice to establish whether there are alterations in cAMP induced phosphorylation of targets in the outer hair cells following CRH receptors gene ablation. Success in this aim will allow us to identify individual proteins phosphorylated due to activation of the CRH receptors, as well as their role in modulating normal olivocochlear synaptic activity. This will functionally link the urocortin hormone/CRH receptor and classical ACh neurotransmitter systems together in a unified model explaining inner ear based protection from noise induced hearing loss.

PI: Waldor, Matthew
Title: Molecular Biology and Virulence of CTX Phage
Abstract: CTXphi is a filamentous bacteriophage that encodes cholera toxin. This is the principal virulence factor of Vibrio cholerae, the Gram-negative bacterium that causes the severe diarrheal disease cholera. CTXphi is the first filamentous bacteriophage shown to mediate the horizontal transfer of a virulence gene. CTXphi integrates into the Vibrio cholerae chromosome and, in the lysogenic state, most CTXphi genes are not expressed due to the activity of the CTXphi repressor, RstR. Generally, the integrated form of CTXphi is found as part of tandem arrays of prophage DNA interspersed with the related genetic element RS1. RS1 encodes a protein, RstC, that can counter RstR repression and lead to markedly enhanced expression of CTX prophage genes including ctxAB, the genes encoding cholera toxin. The long-term goal of this work is to understand the molecular events in the life cycle of CTXphi and the role that this phage plays in the pathogenesis of cholera. The proposed studies will explore 3 processes central to the phage life cycle: i) the site-specific integration of phage DNA into the bacterial chromosome; ii) the repression of most phage gene expression following integration; and iii) the activation of phage gene expression and virion production by environmental and genetic stimuli. Experiments in Aim 1 to identify the mechanism and factors that mediate the integration of CTXphi DNA into the V. cholerae chromosome will reveal how the chromosome encoded recombinases XerC and XerD interact with phage and chromosome sequences to accomplish CTXphi integration. These studies will elucidate a novel mechanism of phage integration and may shed light on the mechanism of ctxAB amplification as well. Experiments in Aim 2: to characterize the regulation and mode of action of RstR will clarify how CTXphi can be maintained in a quiescent state. rstR autoregulation and modulation of RstR levels by environmental factors will be explored. RstR's binding to its unusual operators will also be studied. Experiments in Aim 3 to determine the mode of action of RstC-will explore how RstC can inactivate RstR-mediated repression. RstC's ability to bind to either RstR and/or RstR's binding sites will be investigated and the expression of rstC during infection will be measured. All of these studies will yield insights into fundamental aspects of phage biology. In addition, they may reveal ways in which changes in phage gene expression or copy number can contribute to the pathogenicity of V. cholerae.

PI: Waldor, Matthew
Title: Role of Hfq in Vibrio cholerae virulence
Abstract: Vibrio cholerae causes the severe diarrheal disease cholera. We found that Hfq, an RNA-binding protein, is essential for Vibrio cholerae virulence. Deletion of hfq abolished V. cholerae colonization of the suckling mouse intestine, but had a minimal effect on growth in vitro and did not influence expression of known colonization factors. Thus, Hfq appears to control previously undescribed pathways essential for cholera pathogenesis. In E. coli, Hfq binds to numerous small untranslated RNAs (sRNAs), modulates their activities, and thereby controls expression of a wide variety of genes. Hfq also binds to some mRNAs in E. coli and alters gene expression directly. Although Hfq in V. cholerae probably acts by similar mechanisms, the distinct phenotypes of hfq V. cholerae and E. coli suggest that the proteins bind different sets of RNAs and control distinct regulons. No RNAs bound by V. cholerae Hfq and no V. cholerae sRNAs have been characterized to date. The goals of this R21 application are to identify pathways controlled by Hfq in V. cholerae, particularly Hfq-regulated genes that contribute to V. cholerae virulence, and to characterize the mechanisms controlling their expression. Experiments in Aim I - to define the Hfq regulon - will generate the first knowledge of Hfq-regulated effectors in V. cholerae. Experiments in Aim II - cloning of sRNAs and mRNAs that interact with Hfq - will utilize a new, unbiased approach for cloning interacting RNAs. Experiments in both Aims I and II will explore which of Hfq's interaction partners and downstream effectors contribute to V. cholerae virulence and thus illuminate currently unknown mediators of pathogenesis. Experiments in Aim III - to match sRNAs to the genes they regulate and characterize processes of Hfq dependent gene regulation - will create the foundation for detailed analyses of the mechanisms by which Hfq controls gene expression in V. cholerae. These studies will also facilitate disruption of Hfq-mediated pathways. As Hfq contributes to the virulence of several other Class B Priority Pathogens in addition to V. cholerae, Hfq or its downstream effectors may prove to be valuable targets for new antimicrobial agents.

PI: Walker, Peter
Title: Humanitarian Response Department
Abstract: Oxfam America will engage the Feinstein International Famine Center (FIFC) at Tufts University to assist their Humanitarian Response Department (HRD) with a range of activities related to development of a strategic planning document. First, FIFC staff will assist HRD with its goal of delivering a document that lays out a multi-tiered strategy. Second is an initial step toward development of an on-going working relationship focused on implementation of strategy in the field. FIFC will participate in Oxfam’s work on the current Ethiopia crisis.

PI: Walker, Peter
Title: Humanitarian Futures Context Mapping: What will humanitarian response look like by 2010?
Abstract: Humanitarian action and the diversity of actors and actions associated with the humanitarian label grew exponentially in the roughly fifteen years since the winding down of the Cold War. That growth brought tremendous opportunities, but also tremendous pressures on the humanitarian system and its inherent linkage of values to action. These strains have become even more apparent and in need of urgent attention, against the backdrop of the rapidly evolving international geopolitical landscape as played out in Afghanistan and Iraq.

Many members of the international humanitarian community have expressed concern over these developments and have sought to finds ways to address them, either as individual agencies or consortia.

This present proposal, in response to a request for assistance from the NGO consortium that World Vision is in the process of helping to constitute, will be driven by four objectives.

1. Research the projected future humanitarian context for 2010.
2. Review and examine the current state of humanitarian NGO practice and business structures.
3. Project the current state of humanitarian NGOs into the predicted humanitarian environment of 2010 in order to develop recommendations regarding appropriate institutional structures and processes structures, NGO programming, preparedness, staffing, and resourcing. (To be projected to 2020 at a later date)
4. Carrying forward the research in an action-research participatory fashion with the intent of building understanding and commitment to change from the major stakeholders as the research progresses and is finally presented and reviewed.
5. Positioning the research and the process creating it as a possible initial building block in a wider and more long lasting humanitarian reform and humanitarian sector-wide process.

PI: Waller, William
Title: New England Space Science Initiative in Education
Abstract: New England contains one of the greatest concentrations of space science professionals in the country. The six-state region also presents a diverse set of educational challenges and opportunities. K-16 schools cover the gamut from rural schools of Aroostook County, Maine, to urban neighborhoods of Boston, and some of the world's leading research universities. The region is also home to innovative curriculum development institutions, educational publishers, public television studios, museums, and science centers.

The New England Space Science Initiative in Education (NESSIE) will weave a rich tapestry between space science researchers on the one hand, and educational institutions on the other. The goals of the project are to:

a) facilitate a wide range of educational and public outreach (E&PO) activities to enhance awareness of space science throughout the New England region; and
b) contribute to our understanding of how space science education can .contribute to scientific and technological literacy .

E&OP activities can be visualized on a grid defined by two dimensions:

1) Levels of interaction. Space science professionals can share their knowledge and passion for science by interacting with people on a variety of levels, ranging from personal contact with small numbers of students, to developing television programs and- school curricula that reach large numbers of people.
2) Educational settings. Educational opportunities for various audiences can be defined along a spectrum from formal education (involving students and teachers in accredited programs) to informal learning (such as visitors to planetariums and science centers).

Our role as a regional clearinghouse of space science research and education will be enabled through a fully-maintained website with linkages to relevant institutions and programs. Special efforts will be made to reach underserved groups first identifying the educational needs and interests of these communities.

PI: Wanke, Christine
Title: Feasibility Study of Probiotics for Growth Faltering
Abstract: Diarrheal disease and growth faltering that occur at the time of weaning are significant public health problems. Malnutrition is associated with more than half of the 10 million deaths per year in children under the age of five in the developing world. Exposure to contaminants in the environment is a risk that increases at the time of weaning and is linked to an increased incidence of diarrheal disease when exclusive breast feeding is supplemented by additional foods. Many of the potential interventions for interrupting this cycle of malnutrition and diarrheal disease are not economically or technically feasible for communities in the developing world. The proposed project consists of a study that will determine the feasibility of conducting a randomized-double blinded controlled study of the probiotic Lactobacillus GG (LGG) given in yogurt to infants at the time of weaning as a culturally and economically appropriate intervention for this cycle of malnutrition and diarrhea. The proposed project will also obtain preliminary, background data on the incidence and severity of both diarrheal disease and growth faltering by weight for height and weight for age z scores before and after the time of weaning in children in a slum community in Pakistan. This study will examine the feasibility of enrolling infants at birth, following them weekly at home until the time of weaning and testing the ability to identify the time of weaning. Further, this study proposes to examine the feasibility of following children daily at home after weaning, to monitor health and nutritional status and to administer LGG in yogurt to a group of infants randomized to this intervention. The primary outcomes for the feasibility study are the ability to collect the required data on nutritional, general health, neuro-developmental status, and on the rate and duration of diarrheal disease in the community in Pakistan. Additional primary outcomes include the determination of the background rates and severity of diarrheal disease and growth faltering by height for age and weight for age z scores and the preliminary ability of LGG to ameloriate both diarrheal disease and growth faltering. These data will be used, with the feasibility data obtained, to prepare a proposal for a full clinical trial of LGG as an intervention for growth faltering and diarrheal disease at the time of weaning in these children.

PI: Yee, Amy
Title: Mechanisms of Transcriptional Repressor HBP1 in Cancer
Abstract: Breast cancer is a major killer of women in whom the etiology of tumor induction is poorly understood. The WNT pathway has emerged as a major oncogenic pathway with a complex interplay of oncogenes and tumor suppressor genes. A key feature is the stability of f3-catenin, which functions as a transcriptional co-activator with the LEF and TCF family of transcription factors. The oncogenic phenotypes are ultimately established by the regulation of promoters for key growth regulatory genes. For example, Cyclin D1 is activated by, the Wnt-betacatenin pathway. Cyclin D1 mRNA is increased in many breast cancers, and its role in breast cell proliferation is well established. While the components and the activation of the pathway have been an area of intense study, the molecular mechanisms that inactivate the Wnt pathway in normal tissues are not well understood. These suppressive mechanisms are excellent candidates for the identification of new tumor suppressor genes. The working hypothesis of this proposal is that HBP1 is a suppressor of WNT-beta-catenin signaling through the transcriptional repression of oncogenes and other gene targets. We had previously identified HBP1 as a transcriptional repressor and cell cycle inhibitor. Like LEF and TCF, HBP1 is an HMG box transcription factor. However, HBP1 remains one of few examples of repressors within this important transcription factor family. Recent work indicates that HBP1 is a transcriptional repressor of the Cyclin Dl promoter, which is activated by WNT-B-catenin signaling. Experiments are specifically designed to test the role of HBP1 and transcriptional repression in breast tumorigenesis. The possible role of HBP1 as a tumor suppressor gene in human breast cancer will be tested directly. HBP1 is located in human Chromosome 7q31--a region that is frequently deleted in breast and other cancers. Deletion in cancer is a hallmark of tumor suppressor genes. The long-term goals are the mechanisms that may govern normal breast cell proliferation and that may become aberrant in tumorigenesis. This is critical to understanding tumor suppression and to how mis-regulation may lead to oncogenesis. Together with other work, the proposed studies may provide insights into new diagnostic and/or therapeutic strategies for breast cancer.

PI: Yee, Amy
Title: Phytonutrient Suppression of WNT Signaling in Cancer
Abstract: The framework of this application will be the mechanisms of early breast cancers that are characterized by increased proliferation and of invasiveness. With the advent of earlier detection and better survival, new clinical challenges are the treatment and prevention strategies for possible recurrent and/or advanced disease. Thus, a better understanding of the mechanisms that regulate the increased proliferation and of invasiveness in early breast cancers will advance knowledge for new molecular targets for prognostic and preventative strategies for early breast cancers. For cancer prevention, the use of natural compounds or dietary strategies is an attractive possibility. Epidemiological studies have identified certain foods associated with reduced cancer risk and have resulted in the isolation of phytonutrients as bioactive food components. However, rational applications for cancer prevention require precise knowledge of the molecular mechanisms in selected cancer contexts. Our long-term goal is to advance the knowledge on the molecular and cellular mechanisms by which phytonutrients may modulate specific signaling pathways that are dysregulated in cancer. Diverse genetic studies have highlighted that constitutive WNT signaling is linked to breast and numerous cancers. However, a gap in knowledge exists on strategies to block constitutive Wnt signaling. Epidemiological studies have linked green tea consumption to a reduced recurrence of early breast cancers. This proposal will bridge molecular and nutritional expertise to discover new strategies that may block constitutive WNT signaling and delay mammary tumorigenesis. The focus of this application is an investigation of the green tea phytonutrient EGCG [(-) epigallo-catechin gallate] in suppressing proliferation and invasiveness in early breast cancer. Our preliminary results indicate that EGCG is efficient at suppressing WNT signaling by inducing the HBP1 transcriptional repressor, which is an inhibitor of WNT signaling. The molecular basis for suppression of WNT signaling will be further defined in cell- and animal-based models of WNT signaling. In cell-based studies, the involvement of WNT signaling in breast cell invasiveness will be addressed. With the EGCG mechanisms as a backdrop, dietary supplementation with EGCG will be used to suppress defined mammary tumorigenesis in MMTV-WNT and xenograft mouse models as a pre-clinical investigation. These studies may also define new biomarkers for future clinical studies and advance knowledge on HBP1 as a possible regulatory factor in breast cell invasiveness. Together, these studies should create a valuable foundation for future clinical studies with EGCG and other phytonutrients for early breast cancer prevention.

 

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