This is an amended renewal application for an on-going project, the long-term goal of which has been to understand the development and function of intestinal intraepithelial lymphocytes (IELs) in mice as a means of understanding the human intestinal immune system in health and disease. Chronic intestinal inflammation in humans is a hallmark of inflammatory bowel disease (IBD), an illness that afflicts thousands of people in the U.S. and worldwide. Due to an inability to produce IL-10, a cytokine with strong immune- modulating activity, IL-10-deficient (IL-10-/-) mice develop chronic colonic inflammation with high penetrance beginning in early adulthood. Production of the proinflammatory cytokine, IL-17, is severely dysregulated in IL-10-/- mice, and is one of the major causes of colonic pathology. Additionally, membrane expression of the inducible costimulator (ICOS) molecule, an important T cell activation component, is upregulated on a large proportion of colonic IELs (cIELs) in IL-10-/- mice. A newly discovered RING finger E3 ligase, termed Roquin, has been shown to suppress inflammation by controlling ICOS expression in Roquin-deficient mice. Our Preliminary Studies indicate that Roquin functions as an intermediary effector molecule used by IL-10 to negatively-regulate IL-17;however, the details of that pathway have yet to be fully characterized. The objective of this application is to characterize the molecular events by which IL-10 and Roquin regulate IL-17 expression. The CENTRAL HYPOTHESIS to be tested is that Roquin, under the influence of IL-10, is an intermediary effector molecule involved in negatively regulating IL-17 expression. This hypothesis draws upon preliminary data gathered in our laboratory, and on newly- published work by others. The rationale behind the proposed research is that understanding the role of Roquin in the IL-10 regulatory process will provide new insight into how chronic intestinal inflammation can be controlled, and it may offer a new perspective for therapeutically controlling the inflammatory response in IBD.
Three specific aims will be used to test this hypothesis and to accomplish the objective of this grant.
Specific Aim 1 : To identify IL-10-responsive regulatory elements that control IL-17 and Roquin expression in cIELs.
Specific Aim 2 : To determine the in vitro and in vivo effects of Roquin suppression on IL-17 synthesis and colonic inflammation.
Specific Aim 3 : To determine the capacity of Roquin overexpression to modulate IL-17 synthesis and inflammation in vitro and in vivo. This work will be the first to investigate the interplay between the two anti-inflammatory molecules, IL-10 and Roquin. The expected outcome is that IL- 10 will upregulate Roquin expression and Roquin will then mollify IL-17 expression. These results are expected to have a positive impact by defining the relationship between IL-10 and Roquin;by identifying cognate transcription factors that could be developed into targets for therapeutic intervention of intestinal inflammation;and by assessing the potential for using Roquin for gene therapy in the treatment of IBD.
Inflammatory bowel disease (IBD) is responsible for high morbidity and mortality in the United States and worldwide. At present the etiology of IBD remains unknown and, although some therapies for IBD exist, there is an urgent need for effective long-term treatment. Studies in this renewal application will be done that will use IL-10-/- mice as a means of understanding the immunosuppressive effects of IL-10 and Roquin on IL-17-mediated intestinal inflammation. Roquin is a newly identified molecule that has been shown to be involved in suppressing chronic inflammatory responses;however, its role in controlling IL-17 has not been explored. These studies will use in vitro approaches and in vivo gene therapy strategies to explore the involvement of Roquin, under the influence of IL-10, in the control of colonic IL-17 expression.
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