摘要
目的 探讨长春瑞滨（NVB）对人肺腺癌Anip973细胞的凋亡、端粒酶活性以及人端粒酶逆转录酶mRNA（hTERT mRNA）表达的影响.方法 以不同浓度的NVB作用于Anip973细胞,在不同时间内收集细胞,采用四甲基偶氮唑蓝（MTT）法观察NVB对Anip973细胞的生长抑制作用;通过流式细胞术观察Anip973细胞凋亡率;用倒置显微镜和电镜观察细胞形态学变化;应用以聚合酶链反应（PCR）为基础的端粒重复序列扩增（TRAP-PCR）银染法检测端粒酶活性;采用逆转录聚合酶链反应（RT-PCR）检测hTERT mRNA表达.结果 不同浓度的NVB可诱导Anip973细胞凋亡,而且可降低Anip973细胞中端粒酶活性和hTERT mRNA的表达,并呈时间-剂量依赖性.0.08μg/ml NVB作用Anip973细胞24 h,细胞增殖被抑制,细胞凋亡率为（7.37±0.35）%,hTERT mRNA表达为57.01±1.71,与对照组差异均有统计学意义（P＜0.01）;端粒酶活性值为6.36±0.06,与对照组差异无统计学意义（P＞0.05）;hTERT mRNA表达的改变比端粒酶活性更敏感.0.4μg/ml NVB组和2.0μg/mlNVB组各时间点的细胞凋亡率、端粒酶活性和hTERT mRNA表达与对照组比较,差异有统计学意义（P＜0.01）.2.0μg/ml NVB作用Anip973细胞72 h时,端粒酶活性为1.36±0.27,而细胞凋亡率为（74.87±1.88）%,表明细胞凋亡率的增加与细胞hTERT mRNA表达呈负相关（r=-0.96046,P＜0.01）.结论 NVB的作用机制与端粒酶有关,诱导凋亡是其发挥抗癌作用的机制之一.检测端粒酶活性和hTERT mRNA的表达,有助于判定NVB诱导肺癌细胞凋亡的敏感性.
Objective To study the effect of vinorelbine on apoptosis, telomerase activity and expression of hTERT gene in human lung adenocarcinoma cell line Anip973 cells. Methods Anip973 cells were cocultured with Vinorelbine at different concentrations and collected at 24 h, 48 h and 72 h after treatment, respectively. The inhibition rate of cell growth was determined by methyl thiazolyl tetrazolium （MTT） assay to evaluate the effect of vinorelbine. The percentage of apoptosis was detected by flow cytometry. The cellular morphology was observed under inverted microscope and electron microscope.Telomerase activity of Anip973 cells was determined by the method of TRAP-PAGE-silver staining. RT-PCR was used to detect the expression of hTERT mRNA. Results Vinorelbine down-regulated the telomerase activity and expression of hTERT gene mRNA, inhibited the growth of Anip973 cells, and induced cell apoptosis in a time- and concentration-dependent manner. Annexin V-FITC assay showed 0.08 μg/ml NVB group could inhibited cell proliferation in 24 hour, and apotosis rate was （7.37 ±0.35）%, RT-PCR detection of hTERT mRNA expression in this group was （57.01 ± 1.71 ）, and they were very significantly different from that in the control group （ P 〈 0.01 ）, but telomerase activity was （ 6.36 ± 0.06 ）, compared with the control group showed no significant difference（ P 〉0.05 ）. The change of hTERT mRNA expression is more sensitive than telomerase activity. The apotosis rate, telomerase activity and hTERT mRNA expression of 0.4 μg/mlgroup and 2.0 μg/ml group were very significantly different from that in the control group （ P 〈 0. 01 ）. The telomerase activity of 2.0 μg/ml at 72 hour group was （ 1.36 ±0.27）, basically completetly inhibited, while the apotosis rate was （74.87 ± 1.88 ） %, showed the cell apoptosis rate was in a negative correlation with the down-regulation of the hTERT mRNA （ r = - 0. 96046, P 〈 0. 01 ）. Conclusions Vinorelbine can induce apoptosis in Anip973 cells, and its mechanism of action is related to telomerase activity. The detection of telomerase activity and expression of hTERT gene is useful in predicting prognosis of patients with lung adenocarcinoma.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
2010年第10期743-747,共5页
Chinese Journal of Oncology
基金
基金项目：教育部“春晖计划”合作科研项目
关键词
腺癌
肺
端粒酶活性
基因
凋亡
长春瑞滨
Cells, adenocarcinoma, lung
Telomerase activity
Genes
Apoptosis
Vinorelbine
