CC chemokine ligand-5 promotes macrophage infiltration in early stage of hepatic ischemia-reperfusion injury

Abstract

Objective To study the mechanism of CC chemokine ligand-5 promoting macrophage infiltration in early stage of hepatic ischemia-reperfusion injury. Methods The adult male C57BL/6 wild type and CC chemokine ligand-5 (CCL5) deficient mice were used for the experiment. The mice were kept in captivity at (23±2) ℃, and were exposed to light and dark cycles for 12 hours to obtain food and water freely. The mice were randomly divided into 4 groups. Sham operation group (sham operation group mice experienced ischemia-reperfusion model scheme, but no vascular occlusion, n=10). In the ischemia-reperfusion model group (mice were anesthetized with isoflurane and underwent midline laparotomy, and a noninvasive clamp was placed on the portal, hepatic artery and bile duct to interrupt the blood supply of the left lateral lobe and middle lobe. After 60 minutes of partial liver ischemia, the clamp was removed to start reperfusion, n=10). CCL5 antagonists + model group (before the start of reperfusion, the CCL5 antagonists + model group received a single intravenous injection of the CCR5 receptor antagonist maladono, n=10); CCL5 defective group (CCL5 defective mice, n=10). Alanine aminotransferase (ALT) content and myeloperoxidase (MPO) activity were measured by vtros dt60 Ⅱ chemical system. The expression of ALT mRNA and MPO mRNA was detected by eal-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). The number of macrophages was detected by flow cytometry. The infiltration of macrophages into the lung was detected by histology and immunohistochemistry. The protein expression of related inflammatory factors was detected by Western blotting. Results Compared with the sham operation group, ALT content [(521.36±20.65) vs. (4 126.55±326.68), F=16.237, P 0.05). Compared with CCL5 deficiency group, the number of CD45+ in ischemia-reperfusion model group increased at 24 hours [(2.91±0.23) vs. (1.65±0.17), F=16.311, P<0.05]. At 1, 2, 8 and 24 hours after reperfusion, the number of CD68+ cells in the ischemia-reperfusion model group was higher than that in the sham operation group [(94.62±8.55) vs. (23.68±3.11), F=16.352, P<0.05]. However, in the early stage of ischemia-reperfusion, CCL5 deficiency group was lower than that of ischemia-reperfusion model group [(76.38±6.77) vs. (94.62±8.55), F=16.352, P<0.05]. Compared with the sham operation group, the protein expression of tumor necrosis factor-α (TNF-α) [(1.12±0.23) vs. (3.24±0.42), F=11.352, P<0.05], interleukin (IL)-1 β [(1.03±0.08) vs. (2.87±0.35), F=12.452, P<0.05], IL-6 [(0.95±0.02) vs. (2.58±0.21), F=15.652, P<0.05] in the ischemia-reperfusion model group increased. However, the expression of TNF - α [(3.24±0.42) vs. (1.35±0.14), F=14.252, P<0.05], IL-1 β [(2.87±0.35) vs. (1.22±0.15), F=13.723, P<0.05], IL-6 [(2.58±0.21) vs. (1.35±0.26), F=14.312, P<0.05] in CCL5 antagonist + model group was lower than that in ischemia-reperfusion model group. Conclusion CCL5 can promote the infiltration of circulating macrophages into the liver during the early stage of liver ischemia-reperfusion, and mediate the subsequent pro-inflammatory injury. Key words: Liver ischemia-reperfusion; Macrophage; CC chemokine ligand-5; Inflammatory response; Antagonist