David Artis - US grants

DESCRIPTION (provided by applicant): Soil transmitted helminths remain the most prevalent of all chronic human infections, with an estimated two billion people infected worldwide. Field and experimental studies indicate that immunity in infected individuals is associated with expression of T helper type 2 (Th2) cytokines, while persistent heavy infections can result in overproduction of proinflammatory cytokines and the development of severe intestinal inflammation. The goal of this proposal is to identify the innate immunologic events that occur following infection and interrogate how these responses influence T helper cell differentiation and subsequent resistance or susceptibility to infection. Employing an experimental model of Trichuris infection, our preliminary studies identified a critical role for intestinal epithelial cells (IECs) in the innate response to infection. Manipulation of IEC functions revealed that IECs can regulates multiple aspects of the anti-parasite immune response. First, expression of MHC class II in IECs appears to be critical for the development of Th2 cytokine-dependent immunity. Second, secretion of the cytokine thymic stromal lymphopoietin (TSLP) by IECs appears to be an important early event in influencing dendritic cell and CD4+ T cell responses required for worm expulsion and prevention of intestinal inflammation. Third, TSLP-TSLPR interactions appear to play a critical role in immunity to secondary Trichuris infection, suggesting IEC-derived cytokines may have an important influence on the function of Th2 memory cells. Employing cell lineage- specific deletions in MHC class II, TSLP or TSLPR, three specific aims of this project will determine (i) how IEC-intrinsic MHC class II expression governs the development and regulation Th2 cytokine responses, (ii) how TSLP-TSLPR interactions play a dual role in the development of Th2 cytokine responses and prevention of intestinal inflammation, and (iii) how IEC-derived TSLP regulates the maintenance and function of Trichuris-responsive Th2 memory cells. The results of these studies will provide a framework to test the therapeutic potential of manipulating IEC responses in the promotion of anti- helminth Th2 responses and treatment of infection-induced intestinal inflammation following gastrointestinal nematode infection. In addition, it is hoped that the findings of these studies will have broader implications for understanding the pathophysiology and treatment of multiple inflammatory diseases associated with dysregulated cytokine production including asthma, allergy and inflammatory bowel disease.

Scholarships are being awarded to full-time students majoring in engineering or computer science who demonstrate both academic potential and financial need. Twenty qualifying freshmen students are receiving $3,000 annually for four years and forty upperclassmen are reciving $3510 annually for two years. Special emphasis is being placed on recruiting members of groups underrepresented in engineering and computer science. The scholars are engaged in collaborative learning experiences, have access to tutoring, and receive mentoring and individualized advisement. The opportunity to engage in faculty supervised research is also available to the scholars.

DESCRIPTION (provided by applicant): Soil transmitted helminths remain the most prevalent of all chronic human infections, with an estimated two billion people infected worldwide. Field and experimental studies have shown that immunity in infected individuals is associated with expression of T helper type 2 (TH2) cytokines, while persistent heavy infections can result in overproduction of pro-inflammatory cytokines and the development of severe intestinal inflammation. The long term goals of this proposal are to define the immunological pathways that control T helper cell differentiation and subsequent infection outcome, including intestinal inflammation. Employing experimental Trichuris infection of mice, preliminary studies identified two critical roles for IL-25 (IL-17E) in regulating the immune response following infection: first, IL-25 expression is necessary for the development of protective TH2 responses in resistant mice;second, during persistent infection, IL-25 is an important immunoregulatory cytokine that prevents the development of infection- induced intestinal inflammation. However, the cellular and molecular pathways that orchestrate the effects of IL-25 during Trichuris infection remain unknown. Using cell lineage-specific deletions in transcription factors, cytokines or cytokine receptors, three specific aims of this project will determine (i) how IL-25 promotes TH2 cytokine responses, (ii) how IL-25 limits expression of macrophage-derived pro-inflammatory cytokines and prevents intestinal inflammation, and (iii) how IL-25 regulates the maintenance and function of Trichuris-responsive TH2 memory cells. In addition to testing the prophylactic and therapeutic potential of IL-25 during helminth infection, the findings of these studies will have broader implications for the treatment of multiple TH2 cytokine-associated inflammatory diseases including asthma, allergy and ulcerative colitis. NARRATIVE: An estimated two billion people worldwide are infected with soil transmitted helminth parasites. Although there is strong evidence that T helper type 2 (Th2) cytokines are critical for immunity to infection, the early events that promote protective Th2 cytokine responses are poorly defined. The goals of this proposal are to understand how protective immune responses are initiated and to employ this knowledge in the design of successful new anti-helminth vaccines.

The UCSD MARC U-STAR Fellows Programwill annually prepare 16 ethnically underrepresented undergraduate students for biomedical research careers in academics or private industry. Due to-the growing interrelatedness and interdisciplinary nature of contemporary research activity, a variety of research- associated departments and programs have collaborated to submit the proposal, i.e., Bioengineering, Biological Sciences, Chemistry-Biochemistry, Cognitive Science, Psychology, Center for Language Research, and the Temporal Dynamics of Learning Center. The MARC initiative is further provided direct nstitutional support through the enthusiastic participation of some 90 research faculty from across a wide range of UCSD graduate and undergraduate programs, the School of Medicine, and the School of Pharmacy and Pharmaceutical Sciences. MARC Fellows will participate in a 24-month research regimen under the personalized supervision/mentorship of research faculty, as well as take part in a variety of MARC-specific workshops, research-related writing activities leading to authorship/co-authorship of a paper to be submitted for publication to undergraduate science journals, career exploration activities featuring practicing professionals, ravel for research presentations to regional or national professional meetings, and intensive summer research immersion programs, one at UCSD, one at another R-1 institution. A major UCSD MARC U-STAR goal will be establishment of a strong, underrepresented student research-based support community through which MARC Fellows can easily network and collaborate with peers involved in a wide variety of other undergraduate research programs housed under UCSD's Academic Enrichment Programs in which MARC U-STARS will be located. A second major element of UCSD MARC will involve highly focused academic support and research preparation for underrepresented freshman, sophomore and new community college transfer students through the Pre-MARC Activities. This initiative will annually provide 50-60 students with a wide variety of pre-research activities in order to fuel their growing interest to enter MARC and, subsequently, Ph.D. or MD/Ph.D. programs as a means entering productive, exciting research careers. RELEVANCE (See instructions): MARC's primary mission is to attract, train and prepare highly qualified minority students for entry into health-related research in the biomedical sciences. Through such training, the nation's health/medical research workforce can become dramatically more diversified as ethnic populations previously underrepresented in such disciplines are provided expanded access and support from scientists and health research professionals at UCSD.

DESCRIPTION (provided by applicant): The mucosal surface of the intestinal tract is continuously exposed to innocuous environmental antigens, beneficial commensal microorganisms and potential pathogens. Therefore, maintenance of intestinal immune homeostasis requires a regulated balance between tolerance, immune responsiveness and inflammation. Dysregulation of these processes can result in chronic inflammation or impaired immunity to infection. Intestinal epithelial cells (IECs) provide a physical and biochemical barrier that obstructs entry of commensal and pathogenic bacteria into the underlying lamina propria. In addition, IECs also express pattern recognition receptors and a wide range of immune response genes including cytokines, chemokines and MHC molecules. Therefore, IECs can influence both the composition of commensal communities and host immune responses to them. Recent in vivo studies employing IEC-specific knock-out mice, or the generation of bone marrow chimeric mice in which the non-hematopoietic cell compartment including IECs are specifically targeted, suggest a critical role for IECs in innate recognition of commensal bacteria and regulation of the intestinal immune system. Our preliminary studies suggest two previously unrecognized roles for IECs in the regulation of intestinal immune homeostasis. Metagenomic deep sequence analysis of commensal flora of mice with an IEC-specific deletion of IKK[unreadable] (ikk[unreadable]?IEC) revealed dramatic differences in the composition of commensal bacteria in the intestinal tract of ikk[unreadable]?IEC mice compared to littermate control mice and suggest that IEC-intrinsic NF?B signaling is a critical selective force in influencing the acquisition and/or composition of commensal diversity in health and disease. In addition, deletion of MHC class II exclusively on IECs resulted in dysregulated inflammatory responses following exposure to Citrobacter infection, suggesting an antigen presenting cell function for IECs in immunity to enteric bacterial infection. Based on these preliminary data, two specific aims will (i) test the hypothesis that selective deletion of classical or non-classical NF?B activation in IECs differentially regulates the acquisition and/or stable composition of commensal bacteria in the intestine;and (ii) generate and employ new recombinant Citrobacter strains to test the hypothesis that IEC-intrinsic MHC class II regulates the activation and/or differentiation of bacterial-specific CD4+ T cells in the intestinal microenvironment. Together, the results of these studies will provide new insights into basic mechanisms that govern the relationship between IECs and commensal or pathogenic bacteria, and the consequences of these interactions on homeostasis of immune cell function and host defense in the intestinal microenvironment. It is anticipated that these findings will provide new opportunities to harness the biological functions of IECs in the design of mucosal vaccines and immunotherapeutics. The body's immune system plays a critical role in protecting the exposed surfaces of the body such as the intestine from invading pathogenic bacteria. However, in order to limit chronic inflammatory diseases, the immune system must remain inactive following exposure to beneficial bacteria and environmental antigens. The goals of this proposal are to delineate the mechanisms that balance immune cell responses in the intestine and to employ this knowledge in the design of successful new mucosal vaccines and immune-therapies.

DESCRIPTION (provided by applicant): Food allergies are a significant public health concern and their prevalence in the US is on the rise. The CDC estimates that food allergies affect approximately 6% of children under age three and 4% of the overall population, representing 12 million Americans. Peanut and tree nut allergies develop during childhood and are usually life-long. In the US alone, approximately three million people report allergies to peanuts and tree nuts and a study showed that the number of children with peanut allergy doubled between 1997 and 2002. The public health and economic impact of food allergies is further highlighted by reports indicating that food allergy episodes are the most common ER visit for anaphylaxis and that 50% of food allergy patients will have accidental exposure leading to allergy within a two year period. Despite the continued rise in the prevalence of food allergies in industrialized countries, there have been no new treatments developed. The design and delivery of new prophylactic and therapeutic agents require a better understanding of the basic immunological processes that control allergic reactions in intestine. This proposal will employ new genetic and immunologic tools to gain novel insights into the molecular and cellular events that underlie the pathogenesis of food allergy. Food allergies are thought to be the result of a loss of tolerance to food antigens. Current models suggest that food allergies are caused by the development of MHC CII (MHC CII)-dependent activation of antigen-specific CD4+ Th2 cells that promote the production of IgE and subsequent mast cell-dependent allergic inflammation in the intestine. Our preliminary studies suggest a previously unrecognized role for intestinal epithelial cells (IECs) in the development of food allergy. Specifically, two new preliminary observations form the foundations of this proposal. First, we generated mice bearing an IEC-specific deletion of MHC class II (MHC-CII[unreadable]IEC mice) to examine the influence of IEC-intrinsic MHC CII in the development of Th2 cytokine responses. In preliminary studies, we identified a significant defect in Th2 cytokine-dependent immunity in these mice, suggesting MHC CII expression on IECs may play a critical role in the initiation or propagation of type 2 inflammation in the intestine. Second, we have found that IEC-produced TSLP is required for the establishment of type 2 inflammatory responses in the skin and lung. In a model of food allergy, we also have found that TSLP is required for IgE production, implicating TSLP in the response to food antigens. Based on these preliminary data, the focus of this proposal is to test the role of IECs in the pathogenesis of food allergy. Specifically, we will delineate the functions of IEC-intrinsic MHC CII or IEC-secreted TSLP in a murine model of IgE-dependent food allergy to provide insight for the future design of therapeutic agents to treat and/or prevent food allergies. Food allergies are a significant public health concern and their prevalence in the US is on the rise. The CDC estimates that food allergies affect approximately 6% of children under age three and 4% of the overall population, representing 12 million Americans. Despite their increasing prevalence, the underlying processes that cause food allergy are poorly defined. The goals of this research proposal are to provide new insights into how the body's immune system becomes dysregulated during food allergy and to contribute this knowledge to the design of new drugs for treatment and prevention of these diseases.

DESCRIPTION (provided by applicant): Enteric viral infections, including rotaviruses and noroviruses, remain one of the greatest public health challenges worldwide. In addition, the intestinal mucosa is a critical site for early control of HIV replication and dissemination. Understanding the cellular and molecular mechanisms through which the mucosal immune system recognizes and successfully eradicates enteric viral pathogens is critical for designing better oral vaccines. However, fundamental questions remain unanswered regarding (i) where virus-specific effector and memory T cell responses are primed in the intestinal mucosa; (ii) how the innate immune system recognizes enteric viruses; and (iii) how alterations in intestinal microbiota influence anti-viral immune responses. In preliminary studies, we established mouse Norovirus (MNV) as a model of natural enteric viral infection in humans and generated new tools to track MNV-specific T cell responses. Our preliminary data support the hypothesis that intestinal epithelial cells (lECs) and intestinal commensal bacteria can promote virus-specific T cell responses and host protective immunity following MNV infection. Based on these findings, three fundamental questions will be tested in this proposal (i) Where are MNV- specific T cell responses primed and what regulates MNV-specific memory T cell responses? (ii) How do innate immune cells recognize and respond to MNV infection in the gut? (iii) How do commensal microbial communities influence innate and adaptive immunity to MNV? Collectively, these studies will interrogate the influence of the innate immune response and environmental pathways on the priming and maintenance of MNV-specific T cell responses and host protective immunity in the intestine. Understanding the mechanisms through which the mucosal immune system successfully recognizes enteric viral pathogens and mounts a protective innate and adaptive immune response will be critical for the design of a new generation of successful immuno-modulatory drugs and oral vaccines.

DESCRIPTION (provided by applicant): Enteric bacterial infections remain one of the greatest public health challenges worldwide. An estimated four to six million deaths per year worldwide are caused by bacterial-induced diarrhea and the associated dehydration represents the second most common cause of infant mortality around the world. Enteric bacterial infections and diarrhea are also an important public health concern for travelers and US Forces overseas. The goals of this proposal are to interrogate the influence of innate lymphoid cells (ILCs) in the regulation of innate and adaptive immunity following enteric bacterial infection. Employing Citrobacter rodentium, an experimental model of human enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC or EHEC) infection, our preliminary studies identified a role for IL-23-dependent IL-22 in innate immunity to infection. The dominant cellular source of IL-23-dependent IL-22 in Citrobacter-infected mice was a population of ROR3t-positive ILCs that expressed c-kit and CD90, and in vivo depletion of ILCs in Citrobacter-infected Rag-/- mice was associated with a defect in protective innate immunity to infection. ILCs expressed the thymic stromal lymphopoietin receptor (TSLPR) and deletion of TSLPR resulted in exaggerated ILC responses and enhanced immunity to Citrobacter, indicating that TSLP inhibits ILC-dependent innate immunity in vivo. In addition, a sub-population of ILCs expressed MHC class II and costimulatory molecules and could promote T cell proliferation in vitro, suggesting that these cells also have the capacity to promote T cell-dependent immunity in vivo. Employing selective in vivo depletions, adoptive transfers and cell lineage-specific deletion strategies, three specific aims of this project will determine (i) what signals promote ILC responses in vivo and what is their role in innate immunity to Citrobacter, (ii) how TSLP-TSLPR interactions inhibit ILC responses and innate immunity, and (iii) how ILC-intrinsic MHC class II expression influences CD4pos T cell responses and immunity to Citrobacter. The results of these studies will provide a framework to test the therapeutic potential of manipulating ILC responses in the promotion of anti-bacterial immunity and the treatment of intestinal inflammation. We anticipate that defining the contribution of ILCs to anti-bacterial immunity will direct future clinical efforts to boost innate immunity in the context of human conditions involving immunodeficiency including HIV, cancer and transplantation, to improve the efficacy of oral vaccines, and to dampen ILC responses and reduce chronic inflammation in diseases such as inflammatory bowel disease.

DESCRIPTION (provided by applicant): With an estimated two billion people infected with soil-transmitted helminthes worldwide, this group of parasitic infections represents a significant public health and economic concern. While CD4+ T helper type 2 (TH2) cell responses characterized by IL-4 and IL-13 production are required for protective immunity to intestinal helminth infection, the innate immune responses that promote TH2 cell responses in vivo remain incompletely understood. The goals of this proposal are to interrogate the influence of the epithelial cell-derived cytokine thymic stromal lymphopoietin (TSLP) on regulating basophil responses and the differentiation of TSLP- responsive progenitor cell populations and to assess the role of these cells in influencing TH2 cytokine- dependent host protective immunity following intestinal nematode infection. Employing Trichuris muris and Trichinella spiralis, two well-characterized experimental murine models of human helminth infections, our preliminary studies identified that TSLP selectively promotes basophil responses and basophil-restricted expression was sufficient to partially restore TH2 cell responses and host protective immunity following helminth infection in susceptible mice. In addition, we identified a population of TSLP-elicited progenitor-like cells in the periphery that express the TSLP receptor (TSLPR), exhibit multipotent potential and differentiate to effector cells that produce IL-4 and IL-13. Collectively these data provide insight into two previously unrecognized pathways by which TSLP-dependent basophils and peripheral progenitor-like populations are critical regulators of anti-helminth immunity. Employing a series of novel adoptive transfers and in vivo depletion approaches, Aim 1 of this proposal will utilize experimental Trichuris or Trichinella infection in mice to determine how TSLP regulates innate immune responses required for host protective immunity. Aim 2 of this proposal will utilize similar adoptive transfer studies in mice, in conjunction with cutting edge studies in human patients and humanized mice (hu-mice), to assess the influence of TSLP on progenitor cell responses in the periphery. The results of these studies will provide a framework to test the therapeutic potential of manipulating TSLP-elicited basophils or TSLP-elicited progenitor-like cells in the promotion of anti-helminth immunity. We anticipate that defining the contribution of TSLP-elicited basophils and progenitor-like cells to anti-helminth immunity will direct future efforts to design and improve the efficacy of anti-parasitic oral vaccines and to dampen TH2 cytokine-associated inflammation in the context of asthma and other allergic diseases.

DESCRIPTION (provided by applicant): Respiratory viral infections, including seasonal epidemics of influenza A viral infections, remain a significant global public health challenge. Successful recovery from influenza virus infection requires clearance of the virus, resolution of infection-induced inflammation and effective repair of damaged lung epithelium. While multiple studies have demonstrated an essential role for adaptive immunity in controlling viral replication, the processes that promote repair and remodeling of lung epithelial cells following infection-induced damage remain poorly characterized. This proposal will interrogate the influence of innate lymphoid cells (ILCs) on promoting lung epithelial repair following influenza A virus infection. In preliminary studies, we identified a population of ILCs that is found constitutively n the healthy lung of mice and humans. Lung ILCs in mice constitutively express IL-25R, IL-33R and TSLPR and exhibited a significant population expansion following exposure to IL-25, IL-33 or TSLP or following influenza virus infection. Depletion of lung ILCs following influenza virus infection resulted in severely decreased lung function, impaired airway epithelial repair and increased host mortality, indicating a previously unrecognized role for ILCs as regulators of lung tissue homeostasis. Genome-wide transcriptional profiling of murine lung ILCs revealed a transcriptional signature strongly enriched for genes involved in wound healing and tissue repair including the epidermal growth factor (EGF) family member amphiregulin. Critically, delivery of amphiregulin to ILC-depleted mice restored epithelial repair in influenza virus-infected mice. These data provoke the hypothesis that targeting lung ILC responses could be used therapeutically to promote repair and reduce recovery time in multiple lung diseases including influenza virus infection. Employing bone marrow chimeras, selective in vivo cell depletions and manipulation of the IL-25-IL-25R, IL-33-IL-33R, TSLP-TSLPR or EGFR-dependent signaling pathways, two specific aims of this project will determine (i) how the epithelial cell-derived cytokines IL-25, IL-33 and TSLP regulate lung ILC responses; (ii) how ILC-derived amphiregulin and the EGFR pathway contributes to lung epithelial repair following influenza infection. Collectively, these studies will systematically interrogate the role and regulation of ILCs, amphiregulin and EGFR signaling in promoting lung epithelial repair. We anticipate that defining the contribution of lung ILC and the amphiregulin-EGFR pathway to lung epithelial repair will direct future clinical efforts to promote epithelial barrier function and tissue homeostasis in the context of multiple chronic infectious and inflammatory diseases of the respiratory tract.

Chronic infectious and inflammatory diseases of humans are a significant public health challenge worldwide and greater understanding the human immune system will be essential in the design of new therapies to target these conditions. While significant advances have been made in defining the development and function of the murine immune system, a lack of access to non-diseased human tissue samples has hampered our progress in defining the phenotype and functional potential of immune cells isolated from different human tissue sites. Elucidating a whole body map ofthe human immune system will provide fundamental new insights into the pathways that regulate immunity and chronic inflammation that could aid in the design of new vaccines and immuno-therapeutic approaches. The focus of this proposal is to test whether human lymphoid versus non-lymphoid tissues are populated with phenotypically and functionally distinct innate lymphoid cells (ILCs) and to interrogate the functional significance of ILCs in maintaining tissue homeostasis in the lung and intestine. Employing flow cytometry and genome-wide transcriptional profiling, studies outlined in Aim 1 will comprehensively characterize the phenotypic and functional potential of tissue-resident ILCs isolated from the bone marrow, blood, spleen, lymph nodes, lung and intestine. To complement this analysis, we will also test whether ILCs isolated from distinct tissue sites exhibit differential responsiveness to host-derived cytokines versus microbial products. Using a novel ex vivo 3-dimensional human organoid culture system, studies in Aim 2 will test how ILCs isolated from lymphoid versus lung and intestinal barrier sites can differentially regulate epithelial cell proliferation, differentiation, barrier function and repair. Using antibody-mediated blockade approaches and chemical inhibitors, we will also test the differential contribution of different cytokine-dependent pathways in ILC-mediated regulation of epithelial function in both the lung and intestine organoid systems. RELEVANCE (See instructions): The focus of this proposal is to test whether human lymphoid versus non-lymphoid tissues are populated with phenotypically and functionally distinct innate lymphoid cells (ILCs) and to interrogate the functional significance of ILCs in maintaining tissue homeostasis in the lung and intestine.We anticipate that the outcome of these studies will aid in the design of new therapeutic strategies to target ILC responses in the context of immunity, inflammation and tissue injury.

DESCRIPTION (provided by applicant): Soil-transmitted helminth parasites infect an estimated two billion people worldwide. Helminth-induced type 2 inflammation, which is characterized by the production of the cytokines interleukin (IL)-4, IL-5, IL-9 and IL-13, promotes both acute host-protective immunity and chronic pathologic inflammation. Thus, there is an urgent need to better understand the regulation of protective versus pathologic type 2 inflammation to develop improved therapies to treat helminth infection and chronic inflammation. Group 2 innate lymphoid cells (ILC2s) are innate cells that produce type 2 cytokines that can mediate worm expulsion and contribute to chronic type 2 inflammation in the lung. However, how ILC2 responses are regulated to maintain a balance between helminth-induced protective versus pathologic type 2 inflammation remains unclear. Changes in smooth muscle contractility controlled by the sympathetic nervous system via the ?adrenergic receptor (?AR) and the accumulation and activation of immune cells in response to prostaglandins are also hallmarks of type 2 inflammation. The ?R and prostaglandin pathways have been targeted to treat asthma, allergies and chronic obstructive pulmonary disease (COPD) in patients, but whether these pathways regulate acute protective versus chronic pathologic ILC2 responses is unknown. Here, we demonstrate that human and murine ILC2s express the ?R and the prostaglandin D2 (PGD2) receptor CRTH2 (chemo-attractant receptor- homologous molecule expressed by Th2 cells). Additional preliminary data utilizing in vitro assays and in vivo approaches employing infection with Nippostrongylus brasiliensis, a rodent model for human hookworm infection, led to the central hypotheses that: (1) ?R is a direct negative regulator of acute protective ILC2 responses in the gut and (2) that the PGD2-CRTH2 pathway regulates ILC2 responses and promotes chronic pathologic type 2 inflammation in the lung. To directly test these hypotheses, we propose two Specific Aims. In Specific Aim 1, we will test how the ?R pathway influences human and murine ILC2 phenotype and function in vitro and murine ILC2 responses in vivo during acute protective type 2 immunity to N. brasiliensis in the gut employing genetic and chemical manipulation of the ?R pathway, mutant mouse models and adoptive cell transfer approaches. Studies in Specific Aim 2 will test how the PGD2-CRTH2 pathway regulates human and murine ILC2 phenotype and function and murine ILC2 responses in vivo during chronic pathologic inflammation following N. brasiliensis infection employing cutting-edge bone marrow chimera and adoptive cell transfer approaches. Translational studies will focus on the phenotype and function of CRTH2- expressing ILC2s in the lung of human patients with severe chronic pulmonary inflammation in the context of COPD. We anticipate that a better understanding of the regulation of ILC2 responses during acute protective versus chronic pathologic type 2 cytokine-associated inflammation could lead to the development of new therapies that enhance anti-helminth immunity or limit pathologic inflammation.

Summary The lung is a barrier surface of the mammalian body that is continuously exposed to microbes, allergens and other irritants, and dysregulated immune responses to these stimuli underlies the pathogenesis of multiple airway diseases, including asthma, COPD, IPF, and respiratory viral infections. Basic and translational evidence suggests that populations of innate lymphoid cells (ILCs) are critical orchestrators of inflammation at mucosal barrier sites. While recent research has defined the priming and downstream effector pathways of ILCs in the lung, the cell-intrinsic mechanisms that limit ILCs or influence immune-regulatory responses remain poorly understood. Based upon on our new preliminary data, this renewal will directly test the role of two previously unknown ILC-intrinsic pathways in limiting inflammation in the lung. Group 2 ILCs (ILC2) are a recently described populations of innate immune cells that are enriched in the lung parenchyma that respond to epithelial-derived cytokines IL-25, IL-33 and TSLP, constitutively express the transcription factor GATA3, and can mediate inflammatory processes in the lung through production of effector cytokines IL-4, IL-5, IL-9 and IL-13 or promoting of a Th2 cell response. Despite their recent discovery and the upstream pathways that promote ILC2 responses, it remains unclear what negatively regulates or turns off ILC2 responses in the lung. In new preliminary data, we identify for the first time that ILC2 are enriched in receptors for the Beta-2 adrenergic receptor (?2AR) pathway, a common drug target in asthma. Our new gain- of-function and loss-of-function studies identify that the ?2AR is essential to limit pathogen- and allergen- induced ILC2 responses and inflammation at mucosal sites. Studies outlined in Aim 1 will directly test whether the ?2AR pathway is acting directly on ILC2 in a cell-intrinsic manner and mechanistically how the ?2AR pathway may regulate ILC2 responses. In contrast to type 2 cytokine-dominated inflammation in the lung, recent translational studies suggest that a mixed Th2 and Th17 cell response in airway inflammation results in more severe disease and often patients are refractory to most conventional therapies. Previously we defined that group 3 ILCs (ILC3) directly limit dysregulated Th17 cell responses in the intestine of mice and humans through MHCII-dependent interactions. In new studies, we now demonstrate that while the lung parenchyma is dominantly populated with ILC2, ROR?t+ group 3 ILCs (ILC3) are the dominant ILC group in the lung-draining lymph node of healthy humans and mice. Further, in a model of chronic house dust mite (HDM)-induced lung inflammation, mice with a genetic deletion of ILC3-intrinsic MHCII exhibited increased Th2 and Th17 cell responses, granulocyte recruitment and airway inflammation. Collectively, these data provoke the central hypothesis of Aim 2 that ILC3-intrinsic MHCII critically limits pathologic CD4+ T cells in the context of airway inflammation.

The 4th Annual Meeting of the International Cytokine and Interferon Society (ICIS) will be held in San Francisco, California at the Hyatt Regency Hotel from October 16 - October 19, 2016. This international conference will bring together leading investigators in the fields of cytokine signaling and biology, immunology, cancer research, metabolism, and infectious diseases. The Co-Chairs for this meeting are Dr. David Artis (Cornell University), Dr. John O'Shea (NIH), Dr. Erika Pearce (Max Planck); together, these scientists cover a broad spectrum of scientific expertise relevant to the interests of the ICIS, and have an excellent working relationship as a group. A major goal of the meeting will be to promote the interactions between scientists focusing on cytokine signaling and function in diverse areas of biology, such as host-microbiota interactions, innate immunity, host defense, immune regulation of whole body metabolism, cellular metabolism in immune cells, primary immunodeficiencies, and epigenetics ? and how these translate into emerging therapies. Another goal of this meeting is to facilitate interactions between young investigators and trainees with established researchers in the interferon and cytokine field. Outstanding junior investigators, postdoctoral researchers, and graduate/medical students will be encouraged to participate, and awards will be given to young researchers in each of these categories, as described in the body of this proposal. In addition to approximately 34 invited speakers (some slots remain open for late-breaking speakers) scheduled for Plenary Sessions and Symposia, a number of abstracts submitted by registrants will be selected for oral presentations in Special Topics Sessions. Particular attention will be paid to giving junior investigators an opportunity to present their work during these sessions. Up to 300 participants are welcome to present a poster and will be given the opportunity to discuss their work during their poster session. The program includes scientists from academic institutions, as well as from biotechnology and pharmaceutical companies. We have paid special attention to ensure a program that is well-balanced by gender, ethnicity, seniority and geographic location. A special effort will be made to support the career development of under-represented minorities and women, and an important part of the 2016 meeting is to have 2 workshops and a special reception exclusively for trainees and young (1st year) faculty, designed to foster these interactions. It is the firm belief of the Co-Chairs and Scientific Organizing Committee (composed of 17 scientists from 8 countries including 7 females and two URMs) that, by bringing together leaders, junior investigators, and trainees in these diverse areas of cytokine research, this meeting will inspire important new avenues of investigation and will be of great benefit to the career development of promising young investigators and trainees in the cytokine field.

Project Summary Inflammatory bowel disease (IBD) is often neuropsychiatric disorders, such as anxiety, depression and panic disorders. However, the etiology of IBD-associated neuropsychiatric disorders remains understudied. The focus of this R21 research proposal is to define the molecular and cellular mechanisms through which changes in the intestinal microbiota influence neuronal circuits and brain health that underlie neuropsychiatric disorders associated with IBD. The microbiota is known to regulate mammalian physiology, metabolism and immune homeostasis, and dysbiosis has been linked to the pathogenesis of diverse diseases including IBD. Recently, there is increasing evidence that changes in the intestinal microbiota can have a direct impact on the brain. Studies have reported correlations between gastrointestinal abnormalities and neuropsychiatric disorders, such as autism, cerebral palsy and depression. Animal studies indicate that the absence or modification of the gut microbiota affects neurogenesis, cortical myelination, as well as a variety of behaviors, such as stress, anxiety, cognitive behaviors and panic disorders. Metabolomic studies have shown that metabolites derived from the microbiota and host metabolites altered by the microbiota may influence metabolic, immunologic, and behavioral phenotypes in mice and humans. Collectively, these studies suggest that altered microbiota in IBD patients may contribute to the neuropsychiatric comorbidities associated with IBD. In preliminary analyses, we employed treatment of wild-type mice with a cocktail of broad-spectrum antibiotics to significantly alter the microbiota and examined host behavior in the fear extinction test ? a pre- clinical model of fear-related anxiety disorders. Direct manipulation of the microbiota had a profound effect on fear extinction in these mice. Moreover, GF mice exhibited similar defects in fear extinction that was associated with significantly decreased levels of serum serotonin and dopamine, both of which are reported to play important roles in fear extinction. These findings indicate that the microbiota has an effect on host behavior in the fear extinction model. To dissect the mechanisms by which changes in the intestinal microbiota influence brain function and behavior, in Aim 1, we will employ in vivo two-photon microscopy to directly test whether manipulation of the intestinal microbiota influences neuronal synapse remodeling in mice in the context of fear extinction. In Aim 2, we will employ comparative metabolomics to test how changes in the microbiota influence the metabolite profile in mouse serum and CSF in the context of fear extinction. Using these combined approaches of manipulating the microbiota, coupled with cutting-edge methods of live neuronal imaging and metabolomic profiling, we will generate fundamental new insights into how alterations in the intestinal microbiota are associated with the neuropsychiatric comorbidities associated with IBD.

PROJECT SUMMARY Inflammatory bowel diseases, which include both ulcerative colitis and Crohn's disease, are estimated to affect 3 million Americans, and the number of people living with IBD continues to rise. Currently available medications are costly, ineffective for some patients, and associated with serious risks including opportunistic infections, bone marrow suppression, hepatic inflammation, pancreatitis, and cancer. Thus, there is an urgent need to improve our understanding of modulators of intestinal inflammation and repair in order to identify novel therapeutic targets for the treatment of IBD. Innate lymphoid cells (ILCs) are a relatively-recently characterized family of immune cells that are enriched at barrier surfaces and modulate inflammation in response to cytokine and microbial signals. In particular, group 2 ILCs (ILC2s) sense alarmins and cytokines such as IL-25, IL-33, and TSLP, can be activated by the nervous system, and produce type 2 cytokines that promote anti-helminth immunity and allergic inflammation. Furthermore, our lab has shown that ILC2s also exert tissue-protective functions via secretion of the epidermal growth factor receptor (EGFR) ligand, amphiregulin (AREG), resulting in amelioration of tissue damage following intestinal injury. In new preliminary studies presented here, we show that expression of the neuropeptide, neuromedin U (NMU), is increased during intestinal inflammation in mice, and lack of endogenous NMU results in more severe disease in a model of chemical-induced intestinal damage and inflammation. Conversely, therapeutic administration of NMU results in upregulation of ILC2- derived AREG and ameliorates chemical-induced intestinal damage. Furthermore, similar to in inflamed murine intestines, NMU expression is also elevated in IBD patient biopsies, and the receptor for NMU is detected on human colonic ILCs. Based on our new preliminary data, we hypothesize that enteric neuron-derived NMU activates the tissue-protective functions of ILC2s. We propose to generate a detailed understanding of how NMU mediates tissue protection in both murine models of intestinal inflammation and human IBD. In Aim 1, we will test the hypothesis that during intestinal injury and repair, expression, cellular sources, and spatial pattern of NMU expression are altered. We will also test the role of endogenous enteric-derived NMU in maintaining tissue homeostasis. In Aim 2, we will employ novel reporter mice to directly test the cellular and molecular mechanism by which NMU mediates tissue protection. In Aim 3, we will define the NMU-NMUR1 axis in the healthy human intestine and determine how alterations in NMU-NMUR1 signaling correlate with clinical and endoscopic measures of IBD disease activity. In addition to uncovering fundamental and novel neuropeptide biology and their unique roles in IBD, these studies will provide preclinical justification for development of novel therapeutics to target this pathway.

PROJECT SUMMARY Helminth parasites, including hookworms, infect approximately 2 billion people worldwide and represent a significant public health concern. To combat these parasites, the mammalian immune system has evolved mechanisms to maintain a delicate balance between promoting beneficial inflammation needed to reduce parasitic burdens, but also subsequently restricting that inflammation once the infectious threat is eliminated. When properly regulated, this allows for protective immunity to be achieved without the development of unwanted immunopathology. It is well established that type 2 inflammation, characteristic of helminth-induced immune responses in humans and mice, is initiated via the production of type 2 cytokines by group 2 innate lymphoid cells (ILC2s) and type 2 T helper (TH2) cells. The activation of both innate and adaptive lymphocytes results in the induction of smooth muscle contraction, eosinophilia, mucus production and the population expansion of basophils. Despite our knowledge of the factors that promote type 2 inflammation, the mechanisms that restrict its ability to promote immunopathology remain poorly defined. Our preliminary studies revealed that helminth-induced ILC2 responses, type 2 cytokine production, lung eosinophilia and mucus production are significantly elevated following the depletion of basophils. Moreover, depletion of basophils resulted in dramatic lung pathology and decreased lung function. Strikingly, our new studies also revealed that ILC2s activated in the absence of basophils failed to upregulate expression of the receptor for the neuropeptide neuromedin b (Nmb). Further, delivery of Nmb to helminth-infected mice resulted in reduced ILC2 responses, eosinophilia and mucus production. These data suggest that Nmb is a potent inhibitor of type 2 inflammation. Nmb belongs to the bombesin-like family of neuropeptides consisting of neuromedin B, N, S and U. Importantly, neuromedin U was recently shown to be an important positive regulator of helminth- induced ILC2 responses. Collectively, our studies suggesting that Nmu and Nmb operate as neuropeptide ?rheostat? that properly balances helminth-induced inflammation. Based on our strong preliminary studies and generation of novel Nmbr-floxed and Nmur-Cre mouse models, three specific aims will address the following questions: (i) Do helminth-induced basophils regulate Neuromedin b receptor expression on immune cells, (ii) Does Nmb restrict the activation of multiple immune cells in a manner that properly regulates helminth-induced inflammation, and (iii) Do Nmu and Nmb directly counterbalance each other and operate as a neuropeptide rheostat? Collectively, these studies will interrogate novel mechanisms through which type 2 cytokine- mediated immunity and inflammation are negatively regulated. Defining the mechanisms through which basophils initiate a Nmu/Nmb-mediated rheostat may inform new therapeutic strategies to treat helminth- induced immunopathology.

PROJECT ABSTRACT The immune system at mucosal sites must be tightly regulated to mediate rapid immunity to invading pathogens, while limiting over-reactive responses that drive chronic inflammation. In particular, type 2 immune responses in the airway or gastrointestinal tract are essential to protect from helminth parasites, but if dysregulated, drive asthma and allergic inflammation. Despite this knowledge, we do not yet fully appreciate the complexity of cellular and molecular signals that control these responses, which will be critical for developing the next generation of preventative, therapeutic or curative treatments. The fundamental focus of this renewal application for the Mucosal Immunology Studies Team is to define novel pathways by which the type 2 immune response harnesses signals associated with the nervous system to regulate rapid mucosal immunity and inflammation. In this context, we will define: (i) the pathways that induce and regulate these neuronal signals, (ii) the functional significance of these pathways in type 2 mucosal immunity and inflammation, and (iii) whether it is possible to therapeutically target these signals to boost immunity to helminth infection or reduced chronic allergic inflammation. We will employ innovative approaches and develop new tools to address these fundamental gaps in knowledge, and where possible, translate our findings from mice into human samples. Results from these studies will significantly advance our understanding of the pathways that are essential to mediate rapid type 2 immunity and inflammation at mucosal sites and could provoke the next generation of preventative, therapeutic and curative treatment strategies.

This project will contribute to the national need for well-educated scientists, mathematicians, engineers, and technicians by supporting the retention and graduation of high-achieving, low-income students with demonstrated financial need at the University of California San Diego, Imperial Valley College, and Southwestern College. University of California San Diego is a public research university and an emerging Hispanic Serving Institution (HSI). Imperial Valley College and Southwestern College are 2-year community colleges and fully recognized HSIs. Over its 5 year duration, this project will fund 370 student-years of full-time scholarships to approximately 185 unique students who are pursuing associate’s and bachelor’s degrees in engineering. Transfer-track students at their host institution will receive 2-year scholarships. This project focuses evidence-based high-impact practices on the transition experience endured by transfer students before, during, and after their engagements with 2-year and 4-year institutions. The major significance of this work, beyond financially supporting these students in need, will be implementing, assessing, and ultimately sharing a model educational framework grounded in Schlossberg’s Transition Theory. The transfer students, who undergo frequent and challenging transitions, will be supported with enrichment activities including mentorship, summer programming, research opportunities, and academic year technical and professional workshops.<br/><br/>The overall goal of this project is to increase Engineering degree completion of low-income, high-achieving undergraduates with demonstrated financial need. The challenges faced by transfer students on their journey toward a STEM degree must be better understood. Each transitional step, in, through, and out of their respective institutions requires well-directed support and guidance. This project aims to evaluate the diverse transitions experienced by transfer students and to intervene appropriately, using planned academic year and summer programming as vehicles to deliver the necessary support. Methodologies and results derived from this project will be shared through conference presentations and journal publications. This project is funded by NSF’s Scholarships in Science, Technology, Engineering, and Mathematics program, which seeks to increase the number of low-income academically talented students with demonstrated financial need who earn degrees in STEM fields. It also aims to improve the education of future STEM workers, and to generate knowledge about academic success, retention, transfer, graduation, and academic/career pathways of low-income students.<br/><br/>This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.