Extracellular Hmgb1 Functions as an Innate Immune-Mediator Implicated in Murine Cardiac Allograft Acute Rejection

Abstract

Hmgb1, an evolutionarily conserved chromosomal protein, was recently re-discovered to be an innate immune-mediator contributing to both innate and adaptive immune responses. Here, we show a pivotal role for Hmgb1 in acute allograft rejection in a murine cardiac transplantation model. Extracellular Hmgb1 was found to be a potent stimulator for adaptive immune responses. Hmgb1 can be either passively released from damaged cells after organ harvest and ischemia/reperfusion insults, or actively secreted by allograft infiltrated immune cells. After transplantation, allografts show a significant temporal upregulation of Hmgb1 expression accompanied by inflammatory infiltration, a consequence of graft destruction. These data suggest the involvement of Hmgb1 in acute allograft rejection. In line with these observations, treatment of recipients with rA-box, a specific blockade for endogenous Hmgb1, significantly prolonged cardiac allograft survival as compared to those recipients treated with either rGST or control vehicle. The enhanced graft survival is associated with reduced allograft expression of TNFa , IFNc and Hmgb1 and impaired Th1 immune response.

High-mobility group box 1 promotes early acute allograft rejection by enhancing IL-6-dependent Th17 alloreactive response

“…40 In line with these reports, we have previously demonstrated that high amount of HMGB1 can be passively released by damaged cells of allografts and actively secreted by allograftinfiltrated immune cells such as DCs and macrophages. 22 In this study, we also detected a time-dependent increase of HMGB1 mRNA in allogeneic cardiac grafts after transplantation. Nevertheless, considering organ collection and ischemia/reperfusion insults result in passive release of HMGB1 by damaged cells of allografts, HMGB1 may contribute the initiation of alloimmune response at a fast kinetics than that observed at the mRNA levels.…”

“…22 We now sought to address the role of HMGB1 in IL-17-producing alloreactive T-cell response during acute allograft rejection. For this purpose, an HMGB1-neutralizing Ab was i.p.…”

“…22 LPS contamination in purified rHMGB1 was less than 4 pg/mg protein as determined by endotoxin-specific chromogenic limulus test (Seikagaku Corporation, Tokyo, Japan). HMGB1 antibodies (Abs) were produced by immunizing New Zealand white rabbits with rHMGB1 and Abs with neutralizing activities were tested as reported.…”

“…Tissue sections were also double stained for CD4 and IL-17 for analysis of in situ allograft-infiltrating CD4 þ IL-17 þ T cells using the established techniques. 22,24 To identify the cell types of infiltrates in the allogeneic or syngeneic cardiac grafts, we stained tissue sections for CD4, CD8, Gr-1, or F4/ 80 by immunohistochemical staining method. Briefly, after deparaffinization and rehydration, the sections were treated with 3% H 2 O 2 followed by blocking with 10% goat serum in PBS.…”

“…[19][20][21] Previously, we demonstrated a role for HMGB1 in the initiation and progression of allograft rejection. 22 In this study, we sought to understand the role of HMGB1 in de novo generation of IL-17 þ T cells during acute allograft rejection. Our study data suggest that IL-17 þ T cells could be essential for the early stage of allograft rejection by inducing an influx of neutrophils into allografts, whereas IFN-g þ -Th1 cells are likely responsible for the late graft destruction by enhancing monocyte infiltrations after Th17 response recedes, and this process is tightly controlled by HMGB1 through regulation of IL-6 production from dendritic cells (DCs).…”