A preconditioning effect occurs when exposure to a nonharmful quantity of a mediator of injury provides protection against injury upon subsequent reexposure. High-mobility group box 1 (HMGB1) protein, an endogenous ligand for Toll-like receptor (TLR) 4, is a TLR4-dependent mediator of kidney ischemia-reperfusion injury. Here we determined whether preconditioning with recombinant HMGB1 can block kidney ischemia-reperfusion injury, whether this effect is TLR4 dependent and, if so, how preconditioning downregulates TLR signaling. Wild-type mice pretreated with rHMGB1 before ischemia were protected against kidney ischemia-reperfusion injury, indicated by lower serum creatinine, less tubular damage, less tubulointerstitial neutrophil and macrophage infiltration, and less tubular epithelial cell apoptosis versus control mice. Gene expression of TLR-downstream cytokines and chemokines in ischemia-reperfusion injury kidney were also significantly reduced. While TLR4 and TLR2 knockout mice were protected against kidney ischemia-reperfusion injury, HMGB1 preconditioning provided additional protection to TLR2 but not TLR4 knockout mice. The protective effect of rHMGB1 preconditioning involved Siglec-G upregulation, a negative regulator of HMGB1-mediated TLR4 pathway activation. Thus, preconditioning with rHMGB1 affords significant protection from TLR4-dependent kidney ischemia-reperfusion injury, indicating therapeutic potential.

High-mobility group box1 (HMGB1) is an endogenous ligand for TLR4. We have reported that HMGB1 mediates kidney ischemia reperfusion injury (IRI) via TLR4. We aimed to determine: 1) the potential of preconditioning with rHMGB1 to block kidney IRI; 2) whether this effect is dependent upon TLR4; 3) whether this involves down-regulation of TLR signaling. Methods: Wild-type (WT) and TLR4-/- mice were receive rHMGB1 (20 ug/animal) or saline by i.p. injection 2 hours before 21 minute kidney ischemia. Results: WT mice pre-treated with rHMGB1 were protected against kidney IRI, with lower serum creatinine, less tubular damage, less tubulo-interstitial neutrophil, macrophage and CD4+ cell infiltration and less tubular epithelial cell apoptosis versus controls at day 1 and/or 5 post IRI. mRNA expression of cytokines IL6 & TNFa and chemokines CXCL2 & CCL2 in the kidney were significantly reduced by rHMGB1 pre-treatment at day 5. TLR4-/- mice were protected against kidney IRI, with no additional protection afforded by pre-administration of rHMGB1. The protective effect of rHMGB1 preconditioning on WT kidney involved Siglec-10, a negative regulator of HMGB1-TLR4 activation, which is induced through NF-kB signalling (p<0.05 versus control). Conclusion: HMGB1 preconditioning affords significant protection from kidney IRI by negatively regulating TLR4 signaling through Siglec-10 and NF-kB signalling, indicating the therapeutic potential.

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