蛋白激酶Cα反义核酸诱导肺癌A549细胞凋亡

Antisense oligonucleotides targeted protein kinase C α induces apoptosis of A549 cells

目的探讨聚乙烯亚胺(PEI)介导的人蛋白激酶Cα(PKCα)基因反义寡核苷酸(ASODN)对人肺癌A549细胞凋亡的影响。方法以PEI介导PKCαASODN转染A549细胞,设空白对照组、PEI组、PEI-ASODN组(PKCαASODN终浓度分别为1.25、1.50μmol/L),通过Hoechst33258染色和电镜检测,观察细胞凋亡的形态学改变;设空白对照组、PEI组、PEI-随机寡核苷酸(RODN)组(RODN终浓度1.50μmol/L)、PEI-ASODN组(PKCαASODN终浓度分别为1.00,1.25,1.50μmol/L),采用PI单染和AnnexinⅤ/PI双染流式细胞术,检测细胞亚二倍体峰和早期凋亡率。结果Hochest33258染色和电镜检测均可见PKCαASODN转染组细胞有核固缩、边集和裂解等凋亡形态学变化。流式细胞术示:PKCαASODN转染组细胞出现了明显的亚二倍体峰和早期凋亡细胞群,与空白对照组、PEI或PEI介导的RODN转染组相比,差异有统计学意义(P<0.05),并呈浓度依赖性效应。结论PEI介导的PKCα反义核酸能诱导A549细胞凋亡。

Objective To investigate the effects of antisense oligonucleotides（ASODN） targeted protein kinase Cot（PKCot） on apoptosis of A549 ceils. Methods PKCot ASODN and random oligonucleotides（RODN） were transfected into A549 cells mediated by polyethyleneimine （PEI）. ①A549 cells were divided into control group, PEI group, PEI-ASODN （ 1.25,1.50 αmol/L） groups, and the morphology was examined by fluorescence microscope and transmission electron microscope. ②A549 cells were divided into control group, PEI group, PEI-RODN （ 1.50 μmol/L） group, PEI-ASODN （ 1.00,1.25,1.50 μmol/L） groups. Then the percentage of hypodiploid cells of A549 cells treated with PKCα ASODN for 48 h was determined with propidium iodide（PI） staining by flow cytometry, and the early apoptosis rate was detected with Annexin V-FITC and PI dual parameter by flow cytometry. Results Nuclear fragmentation and condensation were observed in A549 cells treated with PKC ASODN. The percentages of hypodiploid cells and early apoptosis rates increased dose-dependently in PEI-ASODN groups, significantly higher than in control group, PEI group and PEI-RODN group（P 〈 0. 05 ）. Conclusion The PKCα ASODN mediated by PEI could induce the apeptosis of A549 cells.