The goal of this proposal is to advance as a surgeon-scientist to become an independent investigator through completion of the proposed research project and career development under the oversight of an outstanding mentoring team of gastrointestinal and FXR signaling experts. The career development described will provide foundational insight as I design experimental approaches and assess study outcomes. This award will also support the protected time needed to establish a strong publication record in the field of intestinal physiology and develop preliminary data for a competitive R level submission. The farnesoid X receptor (FXR) is a key bile acid receptor that influences the intestinal epithelial barrier. While activation of FXR seems to be beneficial in chronic injury models of the intestine, our preliminary data show that in acute, inflammatory injury, FXR activation is deleterious. FXR knock-out (KO) animals, in contrast, are protected from acute injury. In this proposal, we will define the role of FXR in intestinal barrier function during acute injury. We hypothesize that FXR activation compromises intestinal barrier function by upregulating small heterodimer protein (SHP), while interfering with EGFR signaling, leading to tight junction disruption. We will test this hypothesis with the following specific aims: 1) Determine the mechanisms involved in FXR regulating intestinal epithelial barrier function, and 2) Test the effect of tissue-specific FXR inhibition on the intestinal bar- rier using acute injury models in vivo. We will determine the role of FXR and its downstream effector SHP in intestinal barrier function in vitro and in vivo. To explore this, we will use enteroid-derived monolayers from wild type, FXR KO, SHO KO, and EGFR dominant-negative ?Velvet? mice to determine the effects on the intestinal barrier and tight junctional proteins with and without injury. We will determine FXR and EGFR signal activation using qPCR and Western blot anal- yses. We will determine the role of macrophages using bone marrow-derived culture from genetically-modified animals. These in vitro findings will be tested in vivo using an LPS-injection model of acute injury, utilizing FXR (whole body, intestinal-specific, and macrophage-specific) and SHP KO mice. Cecal ligation and puncture will be used to test pharmacologic manipulation of the FXR pathway on disease progression. This project will begin to establish the role of FXR regulation of the intestinal epithelial barrier in the setting of acute injury. Long term, these findings will provide needed insight into the development of preventative and therapeutic strategies in gut origin sepsis, inflammatory bowel disease, or other GI pathologies. This research has the potential to improve the outcomes of patients with diseases that affect intestinal integrity through better understanding of the mechanisms of FXR-mediated intestinal barrier function.
Numerous intestinal diseases, including necrotizing enterocolitis and inflammatory bowel dis- ease, have disruption of the intestinal barrier. Absence of the bile acid receptor FXR protects animals from acute intestinal injury by maintaining their intestinal barrier. Because FXR may be responsible for gut barrier failure in acute injury, this research proposal will explore the signaling cascades involved in these effects with relevance to human disease.
