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目的探讨转染人乙酰胆碱酯酶(AChE)基因对全反式维甲酸诱导胰腺癌细胞凋亡的影响。方法应用含人AChE基因的真核表达质粒pcDNA3/AChE和绿色荧光蛋白(GFP)真核表达质粒pcDNA3/GFP,以脂质体介导上述两种质粒DNA对人胰腺癌细胞株Patu8988进行瞬时转染,转染后的细胞经全反式维甲酸(ATRA,1μmol/L)分别处理24h及48h,流式细胞仪分别检测AnnexinV染色后早期细胞凋亡率及转染效率,以细胞化学染色法(KarnovskyRoots染色)检测AChE表达。结果流式细胞仪测定GFP显示转染效率达(45.50±3.84)%,人AChE基因转染后联合ATRA治疗凋亡比例增加,转染前后分别为(14.26±1.16)%/(42.84±7.21)%(24h,P<0.01)和(18.08±0.75)%/(56.29±2.54)%(48h,P<0.01)。细胞化学染色法可见AChE阳性细胞,染色聚集于胞核部位。结论胰腺癌细胞AChE基因的高表达可增加细胞对ATRA诱导凋亡的敏感性。
Objective To investigate the effect of transfected human acetylcholinesterase (AChE) gene on the apoptosis of pancreatic cancer cells induced by all-trans retinoic acid. Methods The pcDNA3 / AChE and green fluorescent protein (GFP) eukaryotic expression plasmids pcDNA3 / GFP containing human AChE gene were used to mediate the transient transfection of human pancreatic cancer cell line Patu8988 by liposome The transfected cells were treated with all-trans retinoic acid (ATRA, 1μmol / L) for 24 h and 48 h, respectively. The apoptosis rate and transfection efficiency of early apoptotic cells were detected by flow cytometry. The cytochemical staining (Karnovsky Roots stain) for AChE expression. Results The flow cytometry showed that the transfection efficiency of GFP was (45.50 ± 3.84)%, and the percentage of apoptotic cells was increased after transfection of human AChE gene combined with ATRA. The transfection efficiency was (14.26 ± 1.16)% / (42.84 ± 7.21) % (24h, P <0.01) and (18.08 ± 0.75)% (56.29 ± 2.54)% (48h, P <0.01). Cytochemical staining showed AChE positive cells, staining gathered in the nucleus. Conclusion The high expression of AChE gene in pancreatic cancer cells can increase the sensitivity of cells to ATRA-induced apoptosis.