[Gyl14]-Humanin对Aβ_(25-35)诱导PC12细胞凋亡的影响

Effect of [Gly14]-Humanin on Aβ_(25-35)-induced PC12 cell apoptosis

目的:探讨神经保护肽[Gly14]-Humanin过表达对Aβ25-35诱导的PC12细胞凋亡的影响。方法:采用脂质体法将重组真核表达载体pcDNA3.1(-)/HNG-FLAG转入PC12细胞,G418筛选获得抗性亚克隆细胞株,免疫细胞化学法分析[Gly14]-Humanin基因的表达。用25μmol/LAβ25-35作用细胞24h,MTT法分析细胞存活率,流式细胞术(FCM)监测细胞凋亡,Hoechst33258染色观察凋亡细胞核形态改变。结果:建立了稳定过表达[Gly14]-Humanin的PC12细胞系。经25μmol/LAβ25-35作用后,稳定表达[Gly14]-Humanin的PC12细胞存活率较空质粒转染组明显提高(P<0.05),凋亡率显著降低(P<0.05)。Hoechst33258染色结果表明,空质粒转染组细胞核出现浓缩及断裂等凋亡形态,而过表达[Gly14]-Humanin组细胞核形态正常。结论:过表达[Gly14]-Humanin能够抑制Aβ25-35诱导的PC12细胞凋亡。

AIM：To investigate the effect of [Gly14]-Humanin overexpression on Aβ25-35-induced PC12 cell apoptosis.METHODS：Recombinant plasmid pcDNA3.1（-）/HNG-FLAG was transfected into PC12 cells by liposome method.The subclone cell lines were obtained by persistent G418 selection.The HNG gene expression of PC12 cells was detected by immunocytochemistry.After being treated with 25 μmol/L Aβ25-35 for 24 h,cell viability was determined by MTT assay,and apoptosis rate was detected by using flow cytometric analysis.Hochest33258 staining was used to observe the morphological changes of cellular nuclei.RESULTS：PC12 cell lines stably expressing HNG gene was successfully selected.After being treated with 25 μmol/L Aβ25-35 for 24 h,cell viability of PC12 cells overexpression HNG was significant elevated compared with empty plasmid transfected cells （P0.05）,and the apoptosis rate was lower significantly （P0.05）.By Hoechst 33258 staining,the nuclei of empty plasmid transfected PC12 cells exhibited highly condensed and fragmented nuclei morphology which was the typical characteristics of apoptosis,and the nuclei of PC12 cells overexpression HNG were round and homogeneously stained.CONCLUSION：Overexpression of HNG prevented the cell apoptosis induced by Aβ25-35 in PC12 cells.