Up-Regulation of HMGB1 Exacerbates Renal Ischemia-Reperfusion Injury by Stimulating Inflammatory and Immune Responses through the TLR4 Signaling Pathway in Mice

Abstract

Background/aims: The aim of this study was to elucidate how high-mobility group box 1 (HMGB1) exacerbates renal ischemic-reperfusion injury (IRI) by inflammatory and immune responses through the toll-like receptor 4 (TLR4) signaling pathway.

Methods: A total of 30 wild-type (WT) mice and 30 TLR4 knockout (TLR4-/-) mice were selected and then randomly assigned to the Sham, I/R or HMGB1 groups. The serum and kidney tissues of all mice were collected 24 h after the perfusion. The fully automatic biochemical detector and ELISA were applied to determine the blood urea nitrogen (BUN) and serum creatinine (Scr) levels, and TNF-α, IL-1β, IL-6, IFN-γ and IL-10 levels, respectively. HE staining was used to evaluate kidney tissue damage, immunofluorescence and immunohistochemical staining were performed to observe CD68 and MPO cell infiltration, and flow cytometry was applied to detect immune cells. qRT-PCR and Western blotting were used to detect the expressions of TLR signaling pathway-related genes and proteins, respectively.

Results: Compared with the Sham group, the levels of BUN, Scr, TNF-α, IL-1β, IL-6, IFN-γ and IL-10, kidney tissue damage score, CD68 and MPO cell infiltration, the numbers of immune cells, and the expressions of TLR signaling pathway-related genes and proteins in the I/R and HMGB1 groups were significantly up-regulated. In the I/R and HMGB1 groups, the levels of BUN and Scr, TNF-α, IL-1β, IL-6 and IFN-γ, kidney tissue damage score, CD68 and MPO cell infiltration, immune cell numbers, and TLR signaling pathway-related gene and protein expressions in the WT mice were all higher than those in the TLR4-/- mice, but IL-10 level was significantly lower. Similarly, all aforementioned indexes but IL-10 level in the WT and TLR4-/- mice were higher in the HMGB1 group than in the I/R group.

Conclusion: Our study indicated that the up-regulation of HMGB1 could exacerbate renal IRI by stimulating inflammatory and immune responses through the TLR4 signaling pathway.c.

Keywords: High-mobility group box 1; Immune response; Inflammatory response; Renal ischemic-reperfusion injury; Toll-like receptor 4 signaling pathway.