摘要
目的:探讨死亡相关蛋白激酶(deathassociatedproteinkinase,DAPK)诱导PGCl3细胞凋亡的可能机制及其对半胱氨酸蛋白酶(Caspases)表达的影响。方法:真核表达载体pcDNA3·1-DAPK导入PGCl3细胞中。荧光染色法观察细胞凋亡,流式细胞仪检测凋亡峰、细胞周期和线粒体膜电位的变化。RT-PCR检测Caspase-3、Caspase-6和Caspase-8表达变化。结果:pcDNA3·1-DAPK转染导致PGCl3细胞发生凋亡。pcDNA3·1-DAPK不影响PGCl3细胞周期和线粒体膜电位,t=1·816,P=0·328。DAPK诱导PGCl3细胞凋亡时,Caspase-3和Caspase-8基因表达上调,Caspase-6基因表达无明显变化。Caspase的抑制剂crmA(100μmol/L)不能完全抑制细胞凋亡,t=0·969,P=0·423。结论:过表达的DAPK诱导PGCl3细胞凋亡与Caspase-8和Caspase-3途径有关,Caspase-6相关途径可能不是主要路径,同时可能存在不依赖于Caspase激活途径。
OBJECTIVE： To investigate the effect on Caspases expression in PGCl3 cells for studying mechanism of apoptosis of PGCl3 cells induced by death associated protein kinase （DAPK）. METHODS： Eukaryotic express vector pcDNA3.1-DAPK transfected into high-metastasis non-small lung cancer cell PGC13. Morphologic assessment of apoptosis was performed with fluorescence microscope. Subdiploid peak of DNA, cell cycle and mitochondrial membrane potential were analyzed by flow cytometry. Changes of Caspase-3, Caspase-6 and Caspase-8 expressions were detected by RT-PCR. RESULTS： The pcDNA3. 1-DAPK-transfected could induce apoptosis of PGCl3 cells, but it didn not affect on PGCl3 cells cycle and mitochondrial membrane potential, t= 1. 816, P = 0. 328. Caspase-3 and Caspase-8 expressions were upregulated, and meanwhile Caspase-6 did not change in apoptosis of PGCl3 cells induced by DAPK. CrmA （100 μmol/L）, a Caspases inhibitor, didn＇t suppress cell apoptosis, t=0.969,P=0.423. CONCLUSIONS： Apoptosis of PGCl3 cells induced by over-expression of DAPK may be associated with Caspase-8 and Caspase-3 pathway. Caspase-6 may not be an important pathway. Additionally, apoptosis of PGCl3 cells induced by DAPK may have another independent pathway at the same time.
出处
《中华肿瘤防治杂志》
CAS
2006年第16期1222-1225,共4页
Chinese Journal of Cancer Prevention and Treatment
