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研究莪术含药血清抑制活化的大鼠肝星状细胞(hepatic stellate cells,HSCs)中Hedgehog(Hh)信号通路关键因子Shh(sonic hedgehog),Gli1(glioma-associated oncogene homolog-1)表达的机制。清洁级SD大鼠均分2组,分别给予莪术水煎剂、生理盐水灌胃取血制备含药血清。体外培养大鼠HSCs,分为7组:空白组、模型组、Hh通路抑制剂组、莪术组、Hh通路抑制剂+莪术组、Hh通路激动剂组、Hh通路激动剂+莪术组。除空白组外,其余各组均给予100μg·L-1瘦素诱导后,各组再予以相应药物干预24 h,经MTT法检测HSCs增殖情况,RT-PCR法检测Shh,Gli1的mRNA表达,Western blot法检测Shh,Gli1蛋白表达及免疫荧光法检测Gli1蛋白表达。经瘦素活化的大鼠HSCs中Shh,Gli1的mRNA和蛋白表达均显著上调(与空白组比较P<0.01)。Hh通路抑制剂、莪术含药血清分别干预后,Shh,Gli1的mRNA和蛋白表达显著下调(与模型组比较P<0.01);莪术含药血清与Hh通路抑制剂协同干预后,Shh,Gli1 mRNA和蛋白表达显著下调(与模型组比较P<0.01);Hh通路激动剂干预后,Shh,Gli1 mRNA和蛋白表达显著上调(与模型组比较P<0.01)。用莪术含药血清干预Hh通路激动剂组后,Shh,Gli1mRNA和蛋白表达显著下调(与Hh通路激动剂组比较P<0.01)。莪术含药血清可通过抑制瘦素诱导活化的HSCs中Shh,Gli1的表达,参与Hh信号通路抑制HSCs的活化,发挥抗肝纤维化的作用。
To study the mechanism of curcumin-containing serum on the expression of Shh (sonic hedgehog) and Gli1 (glioma-associated oncogene homolog-1) in activated hepatic stellate cells (HSCs). Clean-grade SD rats were divided into two groups, respectively, to give Curcuma decoction, nasal irrigation with saline to prepare serum. HSCs were cultured in vitro and divided into 7 groups: blank group, model group, Hh pathway inhibitor group, Curcuma group, Hh pathway inhibitor + Curcuma group, Hh pathway agonist group, Hh pathway agonist + Ezhu group. Except for the blank group, the other groups were given 100μg · L-1 leptin induction, each group and then given the appropriate drug intervention 24 h, MTT assay HSCs proliferation, RT-PCR method Shh, Gli1 mRNA expression, Western blot was used to detect the expression of Shh and Gli1 protein and Gli1 protein expression by immunofluorescence. Shh and Gli1 mRNA and protein expressions were significantly up-regulated in leptin-activated HSCs (P <0.01 compared with the blank group). Hh pathway inhibitors, curcumol containing drug serum, Shh, Gli1 mRNA and protein expression was significantly reduced (compared with the model group P <0.01); Curcuma drug serum and Hh pathway inhibitor after co-intervention, Shh, Gli1 mRNA (P <0.01). The expression of Shh and Gli1 mRNA and protein were significantly up-regulated after Hh pathway agonist intervention (P <0.01 compared with model group). The mRNA and protein expression of Shh and Gli1 were down-regulated significantly (P <0.01 compared with Hh pathway agonist group) after intervention of HZ pathway agonist with Ezhu serum. Curcumin-containing serum can inhibit the expression of Shh and Gli1 in HSCs stimulated by leptin, and participate in the activation of HSCs by Hh signaling pathway, and play an anti-hepatic fibrosis role.