论文部分内容阅读
目的研究人端粒酶逆转录酶(hTERT)干扰对肝癌．HepG2、SMMC-7221细胞生物学形为的影响和对肿瘤坏死因子相关的凋亡诱导配体(TRAIL)诱导凋亡的影响。方法将HepG2细胞和SMMC-7721细胞分为转染组 (转染重组质粒真核表达载体)、对照组(转染空载体质粒)和未转染组。采用聚合酶链反应方法检测hTERT干扰序列, 逆转录聚合酶链反应方法检测hTERT表达,HE染色、生长曲线和流式细胞术方法分别检测细胞形态、增殖情况和细胞周期,β-半乳糖苷酶染色方法检测细胞状态,Armexin V／PI染色流式细胞术检测细胞凋亡。结果转染组细胞内均存在hTERT干扰序列,HepG2和SMMC 7221细胞hTERT干扰率分别为100％和43．3％;与未转染组细胞相比, 转染细胞核质比明显缩小,增殖率下降差异有统计学意义(P<0．05),老化细胞和G2-M期细胞明显增加(P<0．05)。细胞老化率分别由未转染组的0增加到转染组的20．4％,由3．60%,增加到10．O％;G2-M期分别由未转染组的7．1％、6．9％增加到转染组的10．6％、7．9％。hTERT干扰显著增加肝癌细胞凋亡和TRAIL诱导凋亡敏感性(P<0．05)。两株肝癌细胞凋亡率分别由未转染组的3．5％、4．8％增至转染组的5．2%、7．9％;100 ng／ml TRAIL作用24 h后两株肝癌细胞凋亡率分别由未转染组的5．3％、13．9％增加到转染组的10．4％、77．2％,而对照组细胞各指标均无显著变化。结论 hTERT干扰明显影响肝癌细胞的生物学行为,显著增加细胞凋亡和TRAIL诱导凋亡的敏感性。
Objective To study the interference of human telomerase reverse transcriptase (hTERT) on liver cancer. HepG2 and SMMC-7221 cells, and the effects of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on apoptosis. Methods HepG2 cells and SMMC-7721 cells were divided into transfection group (transfected with recombinant plasmid eukaryotic expression vector), control group (transfected with empty vector plasmid) and untransfected group. The expression of hTERT was detected by polymerase chain reaction (RT-PCR), hTERT expression was detected by reverse transcription-polymerase chain reaction (RT-PCR), cell morphology, proliferation and cell cycle were detected by HE staining, growth curve and flow cytometry. Cell morphology was detected by staining and apoptosis was detected by Armexin V / PI staining. Results The hTERT interference sequences were found in the transfected cells. The interference rates of hTERT in HepG2 and SMMC 7221 cells were 100% and 43.3%, respectively. Compared with untransfected cells, the ratio of nuclear to cytoplasm was significantly reduced and the proliferation rate decreased The difference was statistically significant (P <0.05), aging cells and G2-M phase cells increased significantly (P <0.05). The cell aging rate increased from 0 in untransfected group to 20.4% in transfected group, from 3.60% to 10. O and G2-M phases increased from 7.1% and 6.9% respectively in the untransfected group to 10.6% and 7.9% in the transfected group. hTERT interference significantly increased apoptosis of hepatocellular carcinoma cells and TRAIL-induced apoptosis (P <0.05). The apoptosis rates of two hepatocellular carcinoma cells increased from 3.5% and 4.8% respectively in the untransfected group to 5.2% and 7.9% in the transfected group, respectively. Two strains of TRAIL treated with 100 ng / ml for 24 h The apoptosis rate of hepatocellular carcinoma cells was increased from 5.3% and 13.9% respectively in the untransfected group to 10.4% and 77.2% in the transfected group, but there was no significant change in the control group. Conclusion The hTERT interference significantly affects the biological behavior of hepatocellular carcinoma cells and significantly increases the sensitivity of apoptosis and TRAIL-induced apoptosis.