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目的探讨神经生长因子(NGF)在肝纤维化大鼠肝星状细胞(HSC)增殖、凋亡中的作用。方法HSC株分别在实验组(分别以NGF 50、100和150ng／ml干预HSC)和对照组(单纯HSC培养)中体外培养24、48和72 h。应用四甲基偶氮唑盐(MTT)法检测细胞增殖率;原位杂交凋亡检测(TUNEL)法观察HSC凋亡状况;流式细胞术检测细胞凋亡率;显微镜观察细胞形态学变化。结果①MTT法显示,不同浓度的NGF作用HSC 24 h,实验组HSC增殖率均低于对照组,差异有统计学意义(均P<0．05)。NGF浓度为100 ng／ml时,HSC增殖率低于对照组最显著(0．63±0．02 vs 0．77±0．03,P<0．05);②最适浓度的NGF(100 ng／ml)作用HSC 24、48和72 h,第72小时的HSC增殖率低于对照组最显著(0．48±0．03 vs 0．89±0．01,P<0．05),增殖率的降低呈时间依赖性;③最适浓度的NGF(100 ng／ml)作用HSC 24 h,TUNEL法显示HSC凋亡率显著高于对照组(10．2％±1．2％vs 1．6％±0．1％,P<0．05);同时,流式细胞术显示HSC凋亡率为6．2％±0．2％,而对照组无凋亡;④NGF对HSC形态没有明显影响。结论NGF可诱导体外活化的HSC凋亡。
Objective To investigate the role of nerve growth factor (NGF) in the proliferation and apoptosis of hepatic stellate cells (HSC) in rats with hepatic fibrosis. Methods HSC strains were cultured in vitro for 24, 48 and 72 h in the experimental group (HSC treated with NGF 50, 100 and 150 ng / ml, respectively) and the control group (cultured with HSC alone). The proliferation of HSC was detected by MTT assay. The apoptosis of HSC was observed by TUNEL method. The apoptosis rate of HSC was detected by flow cytometry. The morphological changes were observed by microscope. Results ① The MTT assay showed that the proliferation rate of HSC in experimental group was lower than that in control group at 24 h of NGF treatment (P <0.05). The proliferation rate of HSC was lower than that of the control group at the concentration of 100 ng / ml NGF (0.63 ± 0.02 vs 0.77 ± 0.03, P <0.05). ② The optimal concentration of NGF (100 (0.48 ± 0.03 vs 0.89 ± 0.01, P <0.05) at 24 h, 72 h and 72 h after HSC treatment, (3) The optimal concentration of NGF (100 ng / ml) treated HSC for 24 h, TUNEL method showed that the apoptosis rate of HSC was significantly higher than that of the control group (10.2% ± 1.2% vs 1 .6% ± 0.1%, P <0.05). Meanwhile, the apoptotic rate of HSC was 6.2% ± 0.2% by flow cytometry, but no apoptosis in control group. ④NGF did not have HSC morphology Clearly affected. Conclusion NGF can induce apoptosis of activated HSC in vitro.