毛蕊花糖苷对血小板衍生生长因子BB处理后大鼠肝星状细胞的影响

目的：探讨毛蕊花糖苷对血小板衍生生长因子BB（PDGF-BB）诱导的大鼠肝星状细胞（HSC）活化、迁移、胶原沉积、纤维化作用的影响，并进一步探讨其对ERK1/2、Akt信号通路表达的影响。方法：HSC细胞分为毛蕊花糖苷（1.5、3.0、6.0 mg/L）组[重组大鼠PDGF-BB（rrPDGF-BB）预处理24 h后，不同浓度的毛蕊花糖苷干预]，PDGF-BB组（rrPDGF-BB预处理24 h，无毛蕊花糖苷干预），对照组（无rrPDGF-BB预处理，无毛蕊花糖苷干预）；分别采用划痕实验检测细胞迁移能力，ELISA法检测Ⅰ型胶原（Col-I）含量，Western blot检测纤维化HSC活化标志物α-SMA蛋白、凋亡效应分子caspase-3，及ERK1/2、Akt信号通路相关蛋白的表达水平。结果：与对照组比较，PDGF-BB组能促进HSC的迁移（P < 0.01），毛蕊花糖苷（1.5、3.0、6.0 mg/L）组能明显抑制HSC的迁移（P < 0.01），且毛蕊花糖苷抑制HSC的迁移有明显的剂量依赖性（r=0.894，P=0.038）；随着受试物浓度的增加，Col-I分泌呈现降低趋势，其中毛蕊花糖苷6 mg/L组抑制胶原产生的作用效果最明显（P < 0.01）；毛蕊花糖苷（1.5，3.0，6.0 mg/L）组能够抑制α-SMA蛋白的表达、激活caspase-3的活性，使ERK1/2、P-ERK1/2、Akt、P-Akt蛋白表达明显降低，且测定蛋白的表达在毛蕊花糖苷各剂量组间也呈现一定的剂量-效应关系（0.826 < r < 0.997，P < 0.01）。结论：毛蕊花糖苷可抑制rrPDGF-BB诱导的HSC的活化、迁移和纤维化作用，其机制可能与毛蕊花糖苷激活caspase-3，阻断ERK1/2、Akt信号通路，抑制HSC中细胞外基质过度沉积及胶原形成有关。

Effect of verbascoside on platelet derived growth factor-BB-mediated activities in rat hepatic stellate cells

OBJECTIVE: To investigate effect of verbascoside on recombinant platelet-derived growth factor (rrPDGF)-BB-mediated migration,collagen deposition and fibrosis in rat hepatic stellate cells (HSC). METHODS: HSC cell cultures were divided into:rrPDGF-BB+ verbascoside (1.5,3,6 mg/L) group[recombinant rat PDGF-BB (rrPDGF-BB) pretreatment for 24 h,no verbascoside intervention],rrPDGF-BB group (rrPDGF-BB pretreatment for 24 h,no verbascoside intervention),control group (no rrPDGF-BB pretreatment,no verbascoside intervention). Scratch test was used to detect cell migration ability,ELISA was used to detect content of type I collagen (Col-I),Western blot was used to detect fibrotic HSC activation marker α-SMA protein and apoptosis effector molecule caspase-3,and expression level of ERK1/2 and Akt signaling pathway-related proteins. RESULTS: Compared with the control group,rrPDGF-BB group promoted the migration of HSC (P < 0.01),verbascoside (1.5,3.0,6.0 mg/L) group significantly inhibited the migration of HSC (P < 0.01). Moreover,inhibition of HSC migration by verbascoside was dose-dependent (r=0.894,P=0.038). With increasing test substance concentrations,Col-I secretion showed a decreasing trend. Among them,the effect of inhibiting collagen production was the most obvious in the 6 mg/L verbascoside group (P < 0.01). Verbascoside (1.5,3.0,6.0 mg/L) inhibited expression of α-SMA protein,activated activities of caspase-3,and significantly reduced expression of ERK1/2,P-ERK1/2,Akt and P-Akt proteins. Moreover, expression of the determination protein was also shown to have a dose-effect relationship among the various dose groups of verbascoside (0.826 < r < 0.997,P < 0.01). CONCLUSION: Verbascoside inhibited the activation,migration and fibrosis of HSC which was mediated by rrPDGF-BB. The evidence suggests that activation of caspase-3 by verbascoside blocked ERK1/2 and the Akt signaling pathway,and inhibited ECM excessive deposition and collagen formation in HSC.