Ionizing radiation induces apoptosis in rapidly dividing intestinal epithelial cells and is perpetuated by an inflammatory process mediated by innate immune cells. Commensal bacteria translocating across the damaged mucosal barrier cause sepsis and death due to neutropenia from simultaneous bone marrow failure. In addition, enteric bacterial components activate innate immune responses by ligating membrane-bound and intracellular receptors that activate the NFκB and MAP kinase signaling cascades. Membrane pattern recognition receptors (PRR) include toll-like receptors (TLR) and a prototypic intestinal intracellular receptor, NOD 2/CARD 15, which is a CATERPILLER gene similar to the CIAS gene described in Project 1. NOD 2 ligation by bacterial products inhibits production of proinflammatory molecules, suggesting that it can downregulate inflammatory responses. Studies in animal models of intestinal inflammation demonstrate that ligation of these pattern recognition receptors can have both beneficial and detrimental effects, with a suggestion that activation of NFκB by ligating TLR isoforms in epithelial cells is protective but that NFκB activation in the lamina propria (LP) induces inflammation.

 

Our studies in gnotobiotic mice and rats demonstrate a primary role for commensal bacterial adjuvants and antigens in the induction of acute and chronic intestinal inflammation as well as protective innate and adaptive immune responses. Moreover, these studies demonstrate that the complex intestinal microflora is composed of species with variable biologic effects: some are proinflammatory, some have no effect, and some species are protective. Endogenous IL-10 and prostaglandin E2 (PGE2) are important host protective factors that downregulate innate and acquired immune responses to enteric bacteria and maintain mucosal barrier function. Innate immune cells produce these regulatory molecules through NFκB signaling when activated by commensal bacterial products.

 

We will study 2 hypotheses: 1. Commensal enteric microbial species have variable effects after whole body irradiation – some potentiate gastrointestinal (GI) inflammation and cause sepsis, some have little net effect, and others are protective. Selectively blocking key pathogenic species and increasing protective species will decrease mortality and promote recovery from radiation injury. 2. Commensal bacterial products activate innate immune responses that have variable biologic effects after exposure to radiation: activated epithelial cells prevent injury, while activated LP innate immune responses potentiate intestinal inflammation and induce molecules that both stimulate and inhibit mucosal re-epithelialization.

 

We will explore these clinically relevant but complex hypotheses in rodent models and in vitro co-culture systems that take advantage of our unique gnotobiotic facility, our ability to precisely manipulate the GI microecology in gnotobiotic mice, access to mice deficient in targeted pattern recognition receptors and our extensive experience in studying mechanisms by which commensal enteric bacteria induce beneficial and pathogenic mucosal immune responses. We will use these results to develop optimal therapies to prevent inflammation and death after radiation exposure and to promote mucosal healing. These complex hypotheses require rodent models and in vitro co-culture systems to dissect interacting and sometime competing pathways prior to human application.