摘要
目的探讨EV71对人恶性胶质瘤细胞株U251的生长、凋亡和人端粒酶反转录酶（hTERT）表达的影响。方法采用RPMI1640培养U251细胞，培养24h后随机分为实验组和对照组，实验组按感染复数值为1加入EV71，并共培养72h，对照组不加EV71。加入EV71后0h、24h、48h和72h用倒置显微镜观察U251细胞形态和生长情况，流式细胞仪测定U251凋亡率，半定量RT—PCR检测hTERT的相对表达水平。结果倒置显微镜观察发现，与对照组比较，实验组加入EV71后24h，U251细胞生长明显受到抑制，48h和72h时U251细胞明显皱缩、变小、形态不规则，而对照组细胞则大小均匀。流式细胞仪检测发现，实验组加入EV71后24h、48h和72h，U251细胞的凋亡率均明显高于对照组[24h：（12．55±2．38）％比（1．42±0．21）％，48h：（65．60±8．48）％比（1．42±0．17）％，72h：（87．52±3．05）％比（1．41±0．16）％，P均=0．000]，且细胞凋亡率逐日升高，对照组细胞凋亡率无明显变化。实验组加入EV71后24h、48h和72h，U251细胞的hTERT相对表达水平均明显低于对照组[24h：（0．58±0．05）比（0．89±0．05），48h：（0．23±0．04）比（0．89±0．03），72h：（0．10±0．03）比（0．90±0．06），P均=0．000]，且表达水平逐日下降，对照组表达水平无明显变化。结论EV71能有效抑制U251细胞的生长，诱导细胞凋亡，具有潜在的抗瘤作用，其作用机制可能部分与下调U251细胞的hTERT表达有关。
Objective To investigate the effects of enterovims 71 （EV71） on growth, apoptosis and human telomerase reverse transcriptase （hTERT） expression of human malignant glioma cell line U251 in vitro. Methods U251 cells were cultured in RPMI 1640 for 24 hours, then randomly divided into experimental group and control group. In the experimental group, EV71 were added in cell culture holes at n4uhiplicity of infection （MOI） equal to 1, and cultured continuously for 72 hours with U251 cells, but in the control group, EV71 were not put. The morphological features and growth of ceils were observed under inverted microscope at 0 hour,24 hours,48 hours and 72 hours of EV71 treatment, respectively. At the same time, cell apoptosis was measured by flow cytometry using Annexin V/PI double staining method, hTERT expression was detected with semi - quantitative reverse transcriptase - polymerase chain reaction （ RT- PCR）. Results The growth of U251 cells was inhibited significantly at 24 h of EV71 treatment under inverted microscope in experimental group when compared with control group, and at 48 h and 72 h, the morphological features of these cells became significantly shrunken, smaller and irregular shape in experimental group, but which were uniform size in control group. At 24 h,48 h,72 h of EV71 treatment,flow cytometry showed the apoptosis rates were significantly higher in experimental group than that in control group[ 24 h：（ 12.55 ± 2.38 ）% vs （1.42 -± 0.21 ）% ,48 h： （65.60 ± 8.48）% vs （1.42 ±0.17）% ,72 h：（87.52 ±3.05）% vs （1.41 ±0.16）% ,all P =0.000] ,and there was upward trend day by day, but no change in control group. At the same time, hTERT mRNA expression levels of U251 cells were significantly lower in experimental group than control group [ 24 h ： （ 0. 58 ± 0. 05 ） vs （ 0. 89 ± 0.05 ）, 48 h ： （ 0.23 ± 0.04） vs （0.89 ±0.03） ,72 h：（0.10 ±0.03） vs （0.90 ±0.06） ,all P =0.000] ,and the downward trend was observed day by day,but there was no this trend in control group. Conclusions EV71 can effectively inhibit the growth of U251 cells,induce the cell apoptosis, and has the potential anti-tumor effect. Its mechanisms may be partly related to down regulation the hTERT expression of U251 cells.
出处
《中华实用儿科临床杂志》
CAS
CSCD
北大核心
2013年第3期172-174,共3页
Chinese Journal of Applied Clinical Pediatrics
基金
国家自然科学基金（30500544）
四川省科技厅支撑项目（12ZC0909）
