硫化氢在SB203580影响肝星状细胞增殖、凋亡中的作用

摘要

目的：探讨H2S在P38丝裂原活化蛋白激酶（P38MAPK）阻断剂SB203580影响大鼠肝星状细胞（HSC）-T6增殖、凋亡中的作用及对Ⅰ、Ⅲ型胶原蛋白mRNA表达的影响。方法设对照组（HSC加含10%胎牛血清的DMEM培养液）、二甲基亚砜（DMSO）组（对照组基础上加DMSO）、NaHS组（对照组基础上加NaHS至最适浓度）、SB组（DMSO组基础上加SB203580至最适浓度）、SB+NaHS组。采用四甲基偶氮唑盐（MTT）法测定SB203580及H2S对HSC的增殖抑制率（IR）；Annexin V-FITC/PI双染法流式细胞术（FCM）检测HSC凋亡率；逆转录PCR法检测Ⅰ、Ⅲ型胶原蛋白mRNA的表达。结果 SB组（7.64±2.46）、SB+NaHS组（12.71±2.73）细胞凋亡率高于对照组（1.70±0.88），且SB+NaHS组高于SB组（均P＜0.05）；DMSO组（2.07±1.18）、NaHS组（2.56±1.23）与对照组差异无统计学意义。Ⅰ、Ⅲ型胶原蛋白mRNA在各组细胞中均表达，其中NaHS组表达高于对照组（P＜0.05），且表达量最多，条带最亮、最宽， SB组与SB+NaHS组表达低于对照组和NaHS组（P＜0.05），表达量较少，条带较暗。结论 H2S激活P38MAPK信号通路，且SB203580特异性阻断P38MAPK信号通路后，H2S刺激的HSC增殖受到抑制，凋亡得到促进。%Objective To study the role of hydrogen sulfide (H2S) in the effect of SB203580 on proliferation and apoptosis of hepatic stellate cells and the effects of H2S on expressions of collagenⅠand collagenⅢmRNA in hepatic stel-late cells. Methods There were five groups of HSC-T6 cells in this study including control group (DMEM medium contain-ing10%fetal bovine serum), dimethyl sulfoxide (DMSO) group, sodium hydrosulfide (NaHS)group,SB203580 (SB)group and SB+NaHS group. MTT method was used to detect the cell proliferation and inhibition rate of HSC-T6 cells treated by SB203580 and H2S. The apoptotic rate of HSC-T6 cells was detected by flow cytometry with annexin V-FITC/PI double staining. RT-PCR was used to detect the expressions of collagenⅠand collagenⅢmRNA in HSC-T6. Results The apop-totic rate of HSC-T6 cells was significantly higher in SB group and SB+NaHS group than that of control group, and the rate was significantly higher in SB+NaHS group than that of SB group (P<0.05). There was no significant difference in the apop-totic rate of HSC-T6 cells between DMSO and NaHS groups than that of control group. The expressions of collagenⅠand col-lagenⅢmRNA were found in five groups of cells. There was a higher expression of collagenⅠand collagenⅢmRNA in NaHS group than that of control group (P<0.05). The expressions of collagenⅠand collagenⅢmRNA were significantly lower in SB group and SB+NaHS group than those of control group and NaHS group (P<0.05). Conclusion H2S activated P38MAPK signal pathway. And P38MAPK was specifically blocked by SB203580 in HSC-T6 cells, which inhibited the cell proliferation stimulated by H2S and promoted the apoptosis.