肝宁方含药血清对人肝星状细胞增殖及凋亡的影响

Effect of Ganning Formula on Proliferation and Apoptosis of Human Hepatic Stellate Cells

目的研究肝宁方含药血清对人肝星状细胞(HSC-LX2)增殖及凋亡的影响。方法40只SD大鼠随机分为4组,每组10只,空白对照组与TGF-β1组SD大鼠给予生理盐水灌胃,TGF-β1+肝宁方组及肝宁方组SD大鼠给予肝宁方灌胃给药(7mL·kg^-1·d^-1),连续灌胃10d后取血离心,制备干燥血清粉末备用。将HSC-LX2分为空白对照组、TGF-β1组、TGF-β1+肝宁方组及肝宁方组。根据分组加入相应的20%大鼠血清对HSC-LX2进行培养造模。CCK8检测各组HSC-LX2细胞增殖率;流式细胞术检测各组HSC-LX2细胞的周期和细胞凋亡;qRT-PCR以及Western blotting分别检测各组α-SMA、Samd2、Samd3、Samd4、Samd7 mRNA及蛋白的表达。结果CCK8结果显示,TGF-β1+肝宁方组及肝宁方组HSC-LX2增殖受到抑制;流式细胞术结果显示,TGF-β1+肝宁方组及肝宁方组HSC-LX2周期停滞在G0/G1期并能促进细胞凋亡;qRT-PCR及Western blotting结果显示,TGF-β1+肝宁方组及肝宁方组α-SMA、Samd2、Samd3、Samd4 mRNA及蛋白的表达受到抑制,Samd7 mRNA及蛋白的表达增强。结论肝宁方具有抑制HSC-LX2增殖、促进HSC-LX2凋亡的作用,其作用与调控TGF-β1/Samd信号通路相关。

OBJECTIVE To study the effect of Ganning formula on proliferation and apoptosis of human hepatic stellate cells(HSC-LX2).METHODS Forty SD rats were randomly divided into 4 groups,10 in each group.The blank control group and the TGF-β1 group were given normal saline gavage,and the TGF-β1+Ganning formula group and Ganning formula group were given gavage(7 mL·kg^-1·d^-1).HSC-LX2 was divided into blank control group,TGF-β1 group,TGF-β1+Ganning formula group and Ganning formula group.The corresponding rat serum was added to the corresponding groups according to the data for the culture and modeling of HSC-LX2.The proliferation of HSC-LX2 cells in each group was detected by CCK8.Flow cytometry was used to detect the cell cycle and apoptosis of HSC-LX2 cells in each group.The mRNA and protein expressions ofα-SMA,Samd2,Samd3,Samd4 and Samd7 were detected by qRT-PCR and Western blotting.RESULTS The results of CCK8 showed that the proliferation of HSC-LX2 was inhibited in the serum group containing Ganning formula.The results of flow cytometry showed that the cycle of HSC-LX2 in the G0/G1 phase of the serum group containing Ganning formula was stagnated and cell apoptosis was promoted.qRT-PCR and Western blotting results showed that the mRNA and protein expressions ofα-SMA,Samd2,Samd3 and Samd4 were inhibited and the mRNA and protein expressions of Samd7 were enhanced.CONCLUSION Ganning formula can inhibit the proliferation of HSC-LX2 and promote the apoptosis of HSC-LX2,which may be related to the regulation of TGF-β1/Samd signaling pathway.