Depletion of gut commensal bacteria attenuates intestinal ischemia/reperfusion injury

Abstract

Gut commensal bacteria play important roles in the development and homeostasis of intestinal immunity. However, the role of gut commensals in intestinal ischemia/reperfusion (I/R) injury is unclear. To determine the roles of gut commensal bacteria in intestinal IR injury, we depleted gut microbiota with a broad-spectrum antibiotic cocktail and performed mesenteric I/R (M I/R). First, we confirmed that antibiotic treatment completely depleted gut commensal bacteria and diminished the size of secondary lymphoid tissues such as the Peyer's patches. We next found that antibiotic treatment attenuated intestinal injury following M I/R. Depletion of gut commensal bacteria reduced the expression of Toll-like receptor (TLR)2 and TLR4 in the intestine. Both are well-known receptors for gram-positive and -negative bacteria. Decreased expression of TLR2 and TLR4 led to the reduction of inflammatory mediators, such as TNF, IL-6, and cyclooxygenase-2. Intestinal I/R injury is initiated when natural antibodies recognize neo-antigens that are revealed on ischemic cells and activate the complement pathway. Thus we evaluated complement and immunoglobulin (Ig) deposition in the damaged intestine and found that antibiotic treatment decreased the deposition of both C3 and IgM. Interestingly, we also found that the deposition of IgA also increased in the intestine following M I/R compared with control mice and that antibiotic treatment decreased the deposition of IgA in the damaged intestine. These results suggest that depletion of gut commensal bacteria decreases B cells, Igs, and TLR expression in the intestine, inhibits complement activation, and attenuates intestinal inflammation and injury following M I/R.

324 Epithelial Cell Specific MyD88 Signaling Mediates Ischemia/Reperfusion-Induced Intestinal Injury Independent of Microbial Status

“…Mucosal antibodies have the potential to recognize "neoantigens" revealed on ischemic cells and to activate the complement pathway, leading to exacerbate damage during I/R-induced injury. 10,30,39,40 Increased intestinal IgA and IgM deposition after I/R injury in mice was previously reported. 10 Because Myd88 IEC−/− mice have lower basal levels of luminal IgA, we hypothesized that these mice would display diminished deposition of IgA in injured tissue.…”

“…10,30,39,40 Increased intestinal IgA and IgM deposition after I/R injury in mice was previously reported. 10 Because Myd88 IEC−/− mice have lower basal levels of luminal IgA, we hypothesized that these mice would display diminished deposition of IgA in injured tissue. We focused on IgA because a previous study reported similar levels of IgG and IgM but lower IgA levels in Myd88 −/− mice compared with wild-type mice.…”

“…11,13,30 This signaling pathway mediates host responses to bacterial colonization/infection, and depletion of intestinal commensal bacteria through antibiotic treatment attenuates I/R-induced intestinal injury. 10 Thus, we hypothesized that bacteria use MyD88 signaling to mediate injury responses in intestinal epithelial cells. However, intestinal injury was exacerbated in GF mice compared with conventionalized mice suggesting that bacteria protect against injury.…”

“…10 To stringently evaluate the impact of bacteria on intestinal injury response, we used GF and conventionalized wild-type mice. Conventionalized mice and GF mice (3-5 mice per group) were exposed to either 60-minute ischemia (I) or 60-minute ischemia followed by 180-minute reperfus ion (I/R), and tissue injury was assessed by histological analysis.…”

“…9 Antibiotic use is the mainstay of treatment to prevent sepsis from bacterial translocation after intestinal ischemic events and has even been shown to be protective before the inciting event. 10 Interestingly, the microbiota seems to play a protective role in various experimental injury models including dextran sodium sulfate and radiation. [11][12][13] In contrast, global deletion of myeloid differentiation primary response gene (Myd88), an adaptor protein in innate immune signaling, protects mice from I/R-induced injury.…”