活血解毒方对HSG细胞凋亡及自噬表达的影响

Effects of the Huoxue Jiedu Decoction on the apoptosis and autophagy expressions of HSG cells

目的观察活血解毒方对HSG细胞凋亡及自噬表达的影响。方法活血解毒方中药冻干粉干预IFN-γ+TRAIL联合诱导的HSG细胞凋亡,以白芍总苷(total glucosides of paeony,TGP)作为阳性对照组,实时荧光定量(quantitative real-time fluorescence,PCR)及蛋白印迹法(Western blotting)检测细胞凋亡内源性途径中及自噬相关蛋白与m RNA的表达、激光共聚焦检测自噬情况。结果与空白组比较,模型组Bax、LC3B蛋白相对表达量明显升高(P<0.01),而Bcl-2蛋白相对表达量明显降低(P<0.01);与模型组比较,两治疗组Bax、LC3B蛋白相对表达量明显降低(P<0.01),Bcl-2蛋白相对表达量明显升高(P<0.01);与TGP组比较,中药组LC3B蛋白相对表达量降低(P<0.05);与空白组比较,模型组Bax mRNA相对表达量明显升高(P<0.01),Bcl-2m RNA相对表达量明显降低(P<0.01);与模型组比较,两治疗组Bax mRNA、LC3B mRNA相对表达量明显降低(P<0.01),Bcl-2 mRNA相对表达量均有不同程度的升高(PTGP组<0.05,P中药组<0.01);与TGP组比较,中药组Bcl-2 mRNA相对表达量明显升高(P<0.01);空白组呈现正常的自噬过程,模型组表现为大量的自噬溶酶体和自噬体聚集物,两治疗组表现为相对较少的自噬体和的自噬溶酶体聚集物,且中药组自噬程度更低。结论活血解毒方可以通过调节内源性途径延缓细胞凋亡及自噬的发生。

Objective To observe the effects of the Huoxue Jiedu Decoction(HXJDD)on the apoptosis and autophagy expressions of HSG cells.Methods The optimal concentration of lyophilized powder of HXJDD was selected by CCK-8 to intervene the HSG cell apoptosis induced by IFN-γand TRAIL,and the total glucosides of paeony(TGP)was used as a positive control,and the apoptosis was detected by a flow cytometry.The quantitative real-time fluorescence PCR and Western blotting were used to detect the related protein and mRNA expressions of apoptosis and autophagy,and a laser confocal microscopy was used to detect its autophagy.Results Compared with the blank group,the relative protein expression levels of Bax and LC3B in the model group were increased significantly(P<0.01),while the relative protein expression level of Bcl-2 was decreased significantly(P<0.01).Compared with the model group,the relative protein expression levels of Bax and LC3B in the two treatment groups were significantly decreased(P<0.01),while the relative protein expression level of Bcl-2 was significantly increased(P<0.01).Compared with the TGP group,the relative protein expression level of LC3B in the HXJDD group was decreased(P<0.01).Compared with the blank group,the relative mRNA expression level of Bax in the model group was significantly increased(P<0.01),while the relative mRNA expression level of Bcl-2 was significantly decreased(P<0.01).Compared with the model group,the relative mRNA expression levels of Bax and LC3B in the two treatment groups were significantly decreased(P<0.01),while the relative mRNA expression level of Bcl-2 was increased in different degrees in the two treatment groups(P<0.05 in the TGP group,P<0.01 in the HXJDD group).Compared with the TGP group,the relative mRNA expression level of Bcl-2 in the HXJDD group was significantly increased(P<0.01).The blank group showed a normal process of autophagy,while the model group showed a large number of autophagic lysosomes and autophagic aggregates,and the two treatment groups showed relatively fewer autophagy and autophagic lysosomal aggregates,and the degree of autophagy in the HXJDD group was lower.Conclusion The Huoxue Jiedu Decoction can delay the apoptosis and autophagy by regulating the endogenous pathway.