Expression of toll-like receptor 4 and MD-2 and protein in Kupffer cells after ischemia-reperfusion in rat liver graft

Abstract

Aim: To investigate the expression of toll-like receptor 4 (TLR4) and MD-2 gene and protein in Kupffer cells (KCs) and their role in ischemia-reperfusion (IR) injury of rat liver graft. Methods: KCs were isolated at 0 (control group), 2, 12, 24 h (IR group) following IR in rat liver graft, mRNA expression of TLR4 and MD-2 was detected by RT-PCR analysis, protein expression of TLR4/MD-2 was detected by flow cytometric (FCM) analysis, and tumor necrosis factor-α (TNF-α) level in supernatant was measured by ELISA. Then isolated KCs were incubated with anti-TLR4 polyclonal antibody (anti-TLR4 group), and TNF-α level was measured again. Results: The mRNA and protein expression of TLR4/MD2 and the level of TNF-α in IR group increased significantly at 2 h following IR, and reached the maximum at 12 h, and slightly decreased at 24 h, but were still significantly higher than those in the control group (P<0.01). The expression of these factors was markedly decreased after anti-TLR4 antibody treatment as compared with the IR group (P<0.01). Conclusion: Lipopolysaccharide (LPS) following IR can up-regulate TLR4/MD-2 gene and protein expression in KCs, and synthesize cytokine TNF-α. Anti TLR4 antibody can inhibit the production of TNF-α induced by LPS. TLR4 and its partner molecule MD-2 may play an important role in Kupffer cell activation and IR injury. Copyright © 2004 by The WJG Press.