截断逆挽方含药血清对D-氨基半乳糖诱导的人正常肝细胞LO2细胞增殖影响

Effect of serum containing drugs in Jieduan Niwan formula on human normal liver cell LO2 cells proliferation induced by D-galactosamine

目的研究截断逆挽方含药血清通过调控E2F1介导的细胞增殖途径减轻D-氨基半乳糖(D-Galactosamine,D-GlaN)诱导的人正常肝细胞LO2的损伤。方法体外培养LO2细胞,以D-GlaN进行造模,将细胞分为正常组、模型组、2.5%截断逆挽方含药血清组(2.5%JDNWF)、5%截断逆挽方含药血清组(5%JDNWF)、10%截断逆挽方含药血清组(10%JDNWF)。采用Cell Counting Kit-8(CCK8)法检测细胞活力;Real-time PCR检测转录因子E2F1、细胞周期蛋白E(cyclin E)、细胞周期蛋白A(cyclin A)、细胞周期蛋白依赖性激酶2(cyclin-dependent kinase 2,CDK2)、细胞分裂周期因子25A(cell division cyclin 25A,CDC25A)mRNA的表达;流式细胞术检测细胞周期;Western Blot检测分化调控因子DP1蛋白表达。结果截断逆挽方含药血清能显著促进LO2细胞进入S期(P<0.01),上调E2F1、cyclin E、cyclin A、CDK2、CDC25A mRNA的表达(P<0.01),并且能增加DP1蛋白的表达(P<0.05,P<0.01)。结论截断逆挽方减轻肝细胞损伤的作用机制可能与其促进肝细胞增殖有关。

Objective To study how to reduce D-Galactosamine by serum containing in Jieduan Niwan formula by regulating E2F1-mediated cell proliferation path.Methods LO2 cells were cultured in vitro and modeled with D-galactosamine(D-GlaN).The cells were divided into normal group,model group,2.5%serum containing drugs in Jieduan Niwan formula group(2.5%JDNWF),5%serum containing in Jieduan Niwan formula group(5%JDNWF),and 10%serum containing in Jieduan Niwan formula group(10%JDNWF).Cell activity was detected by Cell Counting Kit-8(cck8)assay.The mRNA expressions of transcription factor E2F1,cyclin E,cyclin A,cyclin-dependent kinase 2(CDK2)and cell division cycle factor 25A(CDC25A)were detected by real-time PCR.Cell cycle was detected by flow cytometry.DP1 protein expression was detected by Western Blot.Results Serum containing Jieduan Niwan formula could significantly promote the entry of LO2 cells into S phase(P<0.01),increase the expression of E2F1,cyclin E,cyclin A,CDK2 and CDC25A mRNA(P<0.01),and increase the expression of DP1 protein(P<0.05,P<0.01).Conclusion Jieduan Niwan formula in reducing hepatocyte injury may be related to its promotion of hepatocyte proliferation.