Intestinal Ischemia/Reperfusion Injury Triggers Activation of Innate Toll-Like Receptor 4 and Adaptive Chemokine Programs

Background: Ischemia/reperfusion injury (IRI) is a major problem in intestinal transplantation. Toll-like receptor 4 (TLR4) has been implicated as a possible link between the innate and adaptive immune systems, however little data exists regarding TLR4 in intestinal IRI. The goal of this study is to evaluate the involvement of TLR4 in intestinal IRI and to assess the effect on T cell related chemokine programs. Methods: C57BL6 mice underwent 100 minutes of warm intestinal ischemia by SMA clamping. Control WT mice underwent laparotomy without vascular occlusion. Separate survival and analysis groups were performed, and intestinal tissue was harvested at 1 hour, 2 hours, 4 hours, and 24 hours post-reperfusion. Analysis included histology, CD3 immunostaining, myeloperoxidase activity, Western blot, and PCR. Results: Survival was significantly worse in the IRI group vs control (50% vs. 100%). IRI caused severe histopathological injury including mucosal erosions and villous congestion and hemorrhage. Myeloperoxidase activity increased in a time-dependent manner after IRI (2.71 0.25 at 1 hour, 2.92 0.25 at 2 hours, 4 0.16 at 4 hours, 5.1 0.25 at 24 hours vs 0.47 0.11 controls, P < .05). Protein expression of TLR4 followed by NF-κB was increased after IRI. Additionally, mRNA production of IP-10, MIP-2, MCP-1, and RANTES was increased at all time-points, as was mRNA for ICAM-1 and E-selectin. Conclusion: This study is the first to demonstrate increased expression of TLR4 and NF-κB after warm intestinal IRI. This detrimental cascade may be initiated by TLR4 via NF-κB signaling pathways, implicating TLR4 as a potential therapeutic target for the prevention of intestinal IRI.