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目的 :探讨肝细胞生长因子对肿瘤坏死因子α(TNF -α)诱导的肝细胞凋亡的保护作用以及可能的机制。方法 :选择第 3代人类肝细胞系 (L0 2细胞 ) ,分成三组 :Ⅰ组 ,正常L0 2细胞 ;Ⅱ组 ,10 0 0 0u/mlTNF -α损伤L0 2细胞 2 4h ;Ⅲ组 ,重组肝细胞生长因子 (r -hHGF ,5ng/ml)与L0 2细胞共同孵育 30min后加入 10 0 0 0u/mlTNF -α ,2 4h后处理细胞。观察“Ladder”条带、细胞DNA断裂百分率和用Westernblot印迹分析法检测HSP70和IκB -α的表达。结果 :Ⅱ组DNA电泳出现典型的“梯状”条带 ,而Ⅲ组则未见明显的“梯状”条带。DNA断裂百分率Ⅱ组比Ⅰ组高 (分别为 17.12± 4 .0 2和 5 .0 5± 1.6 7,P <0 .0 1) ,Ⅲ组比Ⅱ组低 (分别为 8.77± 2 .0 1和 17.12± 4 .0 2 ,P <0 .0 1)。重组肝细胞生长因子预处理后再用TNF -α损伤的L0 2细胞 ,1h即可检测到HSP 70的表达 ,且持续到 4 8h。肝细胞生长因子还抑制了TNF -α引起的L0 2细胞IκB -α量的减少。结论 :重组肝细胞生长因子可减轻TNF -α诱导的L0 2细胞凋亡 ,它通过诱导L0 2细胞HSP 70的表达和抑制IκB-α的降解达到减轻L0 2细胞的凋亡。
Objective: To investigate the protective effect of hepatocyte growth factor on hepatocyte apoptosis induced by tumor necrosis factor α (TNF-α) and its possible mechanism. Methods: The third generation of human hepatocyte line (L0 2 cells) was selected and divided into three groups: group Ⅰ, normal L0 2 cells; group Ⅱ, L0 2 cells were injured by 10 000 u / ml TNF- α for 24 hours; group Ⅲ, recombinant The cells were treated with hepatocyte growth factor (r-hHGF, 5ng / ml) and L0 2 cells for 30min and then added with 10 000u / ml TNF-α for 24 hours. The “Ladder” band was observed, the percentage of cell DNA was broken, and the expression of HSP70 and IκB-α was detected by western blotting. Results: The typical “ladder” bands appeared in group Ⅱ DNA electrophoresis, but no obvious “ladder” bands in group Ⅲ. The percentage of DNA breakage in group Ⅱ was higher than that in group Ⅰ (17.12 ± 4.00 and 5.50 ± 1.67, respectively, P <0.01), while in group Ⅲ was lower than that in group Ⅱ (8.77 ± 2.01 And 17.12 ± 4 .0 2, P <0.01). After pretreatment with recombinant human hepatocyte growth factor (TNF-α), the expression of HSP70 was detected in L0 2 cells injured by TNF-α for 1 h and continued for 48 h. Hepatocyte growth factor also inhibits TNF-α-induced loss of IκB-α in L0 2 cells. CONCLUSION: Recombinant human hepatocyte growth factor (HGF) can reduce the apoptosis of L0 2 cells induced by TNF-α, and it can reduce the apoptosis of L0 2 cells by inducing the expression of HSP70 and inhibiting the degradation of IκB-α in L0 2 cells.