Roles of microRNA-150-5p in inhibiting malignant proliferation of nasopharyngeal carcinoma cells and enhancing radiosensitivity
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摘要:目的
探讨微小RNA-150-5p(miR-150-5p)在鼻咽癌组织中的表达水平及其对癌细胞增殖和放疗敏感性的影响。
方法采用实时荧光定量聚合酶链反应(qRT-PCR)检测鼻咽癌组织和鼻咽癌细胞中miR-150-5p的表达水平。常规培养鼻咽癌细胞CNE2进行miR-150-5p mimic转染, 分为对照组(NC组)和miR-150-5p过表达组(miR-150-5p mimic组)。采用MTT实验检测各组CNE2细胞的增殖能力; 采用流式细胞仪检测各组CNE2细胞的凋亡情况; 采用Western blot检测各组细胞中磷酸化磷脂酰肌醇3-激酶(pPI3K)、磷酸化蛋白激酶B(pAKT)和磷酸化哺乳动物雷帕霉素靶蛋白(pmTOR)的表达。
结果miR-150-5p在鼻咽癌组织中的表达水平为(0.74±0.39), 低于癌旁组织中的(1.44±0.54), 差异有统计学意义(t=8.140, P＜0.001)。转染48 h后, miR-150-5p mimic组CNE2细胞中miR-150-5p的表达水平为(6.31±1.20), 高于NC组中的(1.00±0.08), 差异有统计学意义(t=7.647, P＜0.001)。MTT实验结果显示, 在24、48、72 h时, miR-150-5p mimic组CNE2细胞增殖能力均低于NC组，差异有统计学意义(P＜0.05)。经0.5、1.0、2.0、4.0、8.0、16.0 Gy放射线辐射后, miR-150-5p mimic组细胞增殖率低于NC组，差异有统计学意义(P＜0.05)。采用2 Gy射线剂量照射CNE2细胞后，与NC组相比, miR-150-5p mimic组细胞凋亡率增加, CNE2细胞中pPI3K、pAKT和pmTOR蛋白表达均降低，差异有统计学意义(P＜0.05)。
结论鼻咽癌细胞中miR-150-5p表达下调。过表达miR-150-5p抑制鼻咽癌细胞增殖，促进细胞凋亡，同时可有效增强鼻咽癌细胞的放疗敏感性; 其可能是通过调控PI3K/AKT/mTOR信号通路发挥作用, miR-150-5p可能是增强鼻咽癌放疗敏感性药物的作用靶点。
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关键词:
- 鼻咽癌 /
- 微小RNA-150-5p /
- 增殖 /
- 放疗 /
- 磷脂酰肌醇3-激酶 /
- 蛋白激酶B /
- 哺乳动物雷帕霉素靶蛋白
Abstract:ObjectiveTo explore the expression level of microRNA-150-5p (miR-150-5p) in nasopharyngeal carcinoma tissue and its effects on proliferation of cancer cells and radiosensitivity.
MethodsReal-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression level of miR-150-5p in nasopharyngeal carcinoma tissues and nasopharyngeal carcinoma cells. Nasopharyngeal carcinoma cells CNE2 were routinely cultured and transfected with miR-150-5p mimic, and were divided into control group (NC group) and miR-150-5p overexpression group (miR-150-5p mimic group). The proliferation ability of CNE2 cells in each group was detected by MTT assay; the apoptosis of CNE2 cells in each group was detected by flow cytometry; the Western blot was used to detect the expressions of phosphorylated phosphatidylinositol 3-kinase (pPI3K), phosphorylated protein kinase B (pAKT) and phosphorylated mammalian target of rapamycin (pmTOR) protein in cells in each group.
ResultsThe expression level of miR-150-5p in nasopharyngeal carcinoma tissue was (0.74±0.39), which was significantly lower than (1.44±0.54) in paracancerous tissue (t=8.140, P < 0.001). After transfection with 48 hours, the expression level of miR-150-5p in CNE2 cells in the miR-150-5p mimic group was (6.31±1.20), which was significantly higher than (1.00±0.08)in the NC group (t=7.647, P < 0.001). The result of MTT experiment showed that the proliferation ability of CNE2 cells at 24, 48 and 72 hours in the miR-150-5p mimic group was significantly lower than that in the NC group (P < 0.05). After radiation with 0.5, 1.0, 2.0, 4.0, 8.0 and 16.0 Gy, the cell proliferation rates of the miR-150-5p mimic group were significantly lower than those of the NC group (P < 0.05). After radiation of CNE2 cells with 2 Gy, the apoptosis rate of cells in the miR-150-5p mimic group increased significantly when compared to that in the NC group, while the expression levels of pPI3K, pAKT and pmTOR protein in CNE2 cells decreased significantly when compared to that in the NC group (P < 0.05).
ConclusionThe expression of miR-150-5p in nasopharyngeal carcinoma cells is down-regulated. Overexpression of miR-150-5p can inhibit the proliferation of nasopharyngeal carcinoma cells, promote cell apoptosis, and effectively enhance the radiosensitivity of nasopharyngeal carcinoma cells; its role may be played by regulating the PI3K/AKT/mTOR signal pathway, and miR-150-5p may be the target of drugs to enhance the radiosensitivity of nasopharyngeal carcinoma.
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表 1 miR-150-5p对鼻咽癌细胞增殖能力的影响(x±s)
时点 细胞OD值 NC组 miR-150-5p mimic组 24 h 0.29±0.03 0.16±0.07* 48 h 0.59±0.08 0.31±0.10* 72 h 0.94±0.08 0.42±0.03* 与NC组比较, * P＜0.05。 
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表 2 miR-150-5p对鼻咽癌细胞增殖率的影响(x±s)(n=3)
放射剂量 细胞增殖率/% NC组 miR-150-5p mimic组 0.5 Gy 93.91±4.49 81.98±5.55* 1.0 Gy 79.55±4.44 66.14±4.17* 2.0 Gy 60.83±5.78 41.97±4.30* 4.0 Gy 48.37±6.71 26.90±6.65* 8.0 Gy 26.80±4.11 16.64±3.25* 16.0 Gy 16.39±2.61 7.22±2.65* 与NC组比较, * P＜0.05。
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