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目的探讨内皮素1(ET-1)对人肝细胞的损伤作用及降钙素基因相关肽(CGRP)的保护作用与机制。方法外科切除肝组织分为:正常对照组(D-Hanks液灌注);ET-1(10-7M)灌注损伤组;CGRP(10-6M,10-7M,10-8M)干预组。采用胶原酶灌注法分离肝细胞,检测肝组织中肿瘤坏死因子α(TNF-α)及丙二醛(MDA)含量,台盼蓝检测新鲜分离肝细胞成活率,并体外培养肝细胞。用MTT法观察肝细胞活力,收集细胞培养上清检测谷丙转氨酶(ALT)、白(清)蛋白(Alb),尿素(Urea)和乳酸脱氢酶(LDH)水平。结果与对照组比较,ET-1组肝细胞成活百分率及活力,ALB合成及Urea代谢显著减低(P<0.05);肝组织中TNF-α及MDA含量,LDH及ALT释放增加(P<0.05);而CGRP(10-6M,10-7M,10-8M)能显著减轻ET-1诱导的肝细胞损伤,使肝细胞成活率及活力、Alb分泌及Urea代谢均显著增加(P<0.05),使LDH及ALT,TNF-α及MDA显著减少(P<0.05)。结论CGRP对ET-1诱导的肝细胞损伤具有保护作用,其机制与MDA和TNF-α释放减少有关。
Objective To investigate the effects of endothelin-1 (ET-1) on the injury of human hepatocytes and the protective effect and mechanism of calcitonin gene-related peptide (CGRP). Methods Surgical resection of liver tissue was divided into normal control group (D-Hanks fluid perfusion), ET-1 (10-7M) perfusion injury group and CGRP (10-6M, 10-7M, 10-8M) intervention group. Tumor necrosis factor alpha (TNF-α) and malondialdehyde (MDA) were detected by collagenase perfusion method. The survival rate of freshly isolated hepatocytes was detected by trypan blue and hepatocytes were cultured in vitro. The viability of hepatocytes was observed by MTT assay, and the levels of ALT, Alb, Urea and lactate dehydrogenase (LDH) were detected by cell culture supernatants. Results Compared with the control group, the survival percentage, the synthesis of ALB and the metabolism of Urea in ET-1 group were significantly decreased (P <0.05); the levels of TNF-α and MDA, the release of LDH and ALT in liver tissue were increased ; While CGRP (10-6M, 10-7M, 10-8M) could significantly reduce the damage of liver cells induced by ET-1, significantly increase the survival rate and viability of Alb cells, the secretion of Alb and Urea metabolism (P <0.05) LDH and ALT, TNF-α and MDA were significantly decreased (P <0.05). Conclusion CGRP has a protective effect on ET-1-induced hepatocellular injury. The mechanism is related to the decrease of MDA and TNF-α release.