Hypoxia/reoxygenation-induced HMGB1 translocation and release promotes islet proinflammatory cytokine production and early islet graft failure through TLRs signaling

Abstract

High-mobility group box 1 (HMGB1) translocation and release, which is involved in several tissue types of ischemia-reperfusion injuries, activate innate immunity by inducing proinflammatory cytokine production through its interaction with toll-like receptors (TLRs). Our objective was to determine the role of HMGB1 and the degree of activation of TLR-related signal transduction pathways in hypoxia/reoxygenation (H/R)-induced proinflammatory cytokine production and intra-islet graft inflammation. After islets are exposed to hypoxia-reoxygenation for 24h, TLR2/4 expression and TLR-mediated signaling was up-regulated in islets, and HMGB1 was translocated from the nucleus to the cytoplasm and released to the extracellular space. With H/R exposure, proinflammatory cytokine production (IL-1β and TNF-α) and islet injury were significantly increased, and these effects depend on TLR2/4 signaling pathways. Exogenous HMGB1 also induces islet inflammation and increases the phosphorylation of STAT3, p38 and IκBα in wild-type islets. TLR2 deficiency in TLR2-KO islets resulted in the inhibition of IL-1β production and STAT3/p38 phosphorylation after HMGB1 exposure. TLR4 deficiency in TLR4-KO islets resulted in the inhibition of TNF-α production and IκBα phosphorylation after HMGB1 exposure. Pre-incubation of the STAT3, p38, or NF-κB inhibitors significantly inhibited HMGB1-induced IL-1β or TNF-α production in islets, but the effect of HMGB1 or H/R-induced islet injury was not counteracted by a separate treatment of the STAT3 inhibitor, p38 inhibitor, or NF-κB inhibitors. HMGB1 inhibition by ethyl pyruvate or blockade by neutralizing antibodies significantly decreased the phosphorylation of STAT3, p38 and IκBα, the production of IL-1β and TNF-α, and the islet injury in wild-type islets after exposure to H/R and significantly improved early islet graft failure. Thus, our results suggest that HMGB1 released from H/R induced islets works in an autocrine manner to up-regulate STAT or p38 and augment IL-1β production via TLR2, and up-regulate NF-κB and augment TNF-α production via TLR4 in intra-islet, which are associated with H/R-induced islet injury and early graft failure.