肝细胞生长因子对大鼠原代肝星状细胞凋亡的影响及其机制

目的 探讨肝细胞生长因子(HGF)对血小板衍生生长因子BB(PDGF-BB)作用下大鼠原代肝星状细胞(HSC)凋亡的影响及其机制.方法 分离培养大鼠HSC,传代后使用PDGF-BB处理HSC,HGF进行干预,采用AO/EB染色、TUNEL法和流式细胞仪观察HGF对PDGF-BB作用下大鼠原代HSC凋亡的影响,免疫细胞化学分析P65的表达,电泳迁移率分析检测核因子κB DNA-蛋白结合活性. 结果对照组、PDGF-BB+HGF处理组和HGF处理组AO/EB染色法检测细胞凋亡率分别为1.98%±0.42%,5.10%±0.56%和8.43%±0.40%;流式细胞仪观察凋亡率分别为1.64%、3.22%和6.66%;TUNEL法凋亡率分别为2.35%±0.47%、4.89%±0.36%和7.34%±0.51%,各组间比较,差异均有统计学意义(t值分别为3.65、2.46和2.13,P值均＜0.05).对照组、PDGF-BB+HGF处理组和HGF处理组P65的表达依次减少,吸光度值分别为0.250±0.038、0.180±0.025和0.130±0.028,差异有统计学意义(t=2.08,P＜0.05).核因子κB DNA-蛋白结合活性相应减弱,吸光度值分别为283.97±43.20、129.93±15.54和61.56±15.14,差异有统计学意义(t=2.77,P＜0.05).结论 HGF能够诱导大鼠原代HSC和PDGF-BB作用下大鼠原代HSC的凋亡,抑制P65的表达和核因子κ B DNA-蛋白结合活性是HGF促进HSC凋亡的重要机制.

Effects and the mechanism of HGF on the apoptosis of rat primary cultured hepatic stellate cells treated with platelet-derived growth factor

Objective To investigate the inhibitory effect ofhepatocyte growth factor (HGF) on the apoptosis of rat primary cultured hepatic stellate cells treated with platelet-derived growth factor BB (PDGF-BB). Methods Hepatic stellate cells were cultured and treated with PDGF-BB+HGF, HGF or nothing (controls). Apoptosis of these hepatic stellate cells was evaluated by AO/EB staining, TUNEL and flow cytometry. Expression level of P65 was observed with immunocytochemicai staining; DNA-protein binding complex of NF-kappa B was detected by electrophoretic mobility shift assay. Results The cell apoptosis rate of the control group was lower than that of the PDGF-BB+HGF group. The apoptosis rate of the PDGF-BB+HGF group was lower than that of the HGF treated group; the expression of P65 was lower in the PDGF-BB+HGF group and HGF treated group compared to the normal control group; DNA-protein binding activity of NF-kappa B was respectively attenuated in the normal control group, PDGF-BB+HGF treated group and HGF treated group (P＜0.05). Conclusion HGF can induce HSC apoptosis. Its possible mechanism may involve inhibiting DNA-protein binding activity of NF-kappa B and down-regulating the expression level of P65.