医学分子生物学杂志 ›› 2022, Vol. 19 ›› Issue (6): 464-471.doi: 10.3870/j.issn.1672-8009.2022.06.005

• 论著 • 上一篇    下一篇

circ_ 0061140 靶向 miR-6838-5p 促进非小细胞肺癌细胞的增殖、 阻滞细胞周期、 诱导细胞凋亡

  

  1. 1郴州市第一人民医院肿瘤内科一区 湖南省郴州市, 423000  2郴州市第一人民医院肿瘤内科 湖南省郴州市, 423000
  • 出版日期:2022-11-30 发布日期:2023-02-14

circ _ 0061140 Regulate Proliferation, Cell Cycle and Apoptosis of Non-small Cell Lung Cancer Cells by Targeting miR-6838-5p 

  1. 1 First Ward, Department of Oncology, Chenzhou First People’ s Hospital, Chenzhou, Hunan, 423000, China  2 Department of Oncology, Chenzhou First People’s Hospital, Chenzhou, Hunan, 423000, China
  • Online:2022-11-30 Published:2023-02-14

摘要: 目的 通过实验研究确认环状 RNA (circRNA) circ_ 0061140 是否能靶向 miR-6838-5p 调控非小 细胞肺癌细胞的增殖、 细胞周期和凋亡。 方法 采用逆转录-定量聚合酶链反应 (RT-qPCR) 检测 circ_ 0061140 和 miR-6838-5p 在非小细胞肺癌细胞中的表达情况。 将非小细胞肺癌 NCI-H1299 细胞分为 si-circ_ 0061140 组 (转染 si-circ_ 0061140; 低表达 circ_ 0061140)、 si-NC 组 (转染 si-NC; 阴性对照)、 NC 组 (未 转染; 空白对照)、 miR-6838-5p 组 (转染 miR-6838-5p 模拟物; 高表达 miR-6838-5p)、 miR-NC 组 (转染 miR-NC; 高表达 miR-6838-5p 的阴性对照)、 si-circ_ 0061140 + anti-miR-NC 组 (共转染 si-circ_ 0061140 与 anti-miR-NC)、 si-circ_ 0061140 + anti-miR-6838-5p 组 (共转染 si-circ_ 0061140 与 anti-miR-6838-5p)。 采用 Western 印迹检测细胞周期蛋白 D1 ( cyclinD1)、 活化的含半胱氨酸的天冬氨酸蛋白水解酶-3 ( cleaved caspase-3) 蛋白表达, MTT 法检测细胞增殖能力, 流式细胞仪检测细胞周期和凋亡情况。 双荧光素酶活性 检测 circ_ 0061140 和 miR-6838-5p 的靶向结合。 结果 非小细胞肺癌组织、 细胞 NCI-H1299、 NCI-H2170、 NCI-H1975 中的 circ_ 0061140 表达量均比癌旁组织或正常肺上皮细胞 BEAS-2B 高, miR-6838-5p 表达量比 癌旁组织或 BEAS-2B 细胞低 (P均 < 0. 05)。 si-circ_ 0061140 组或 miR-6838-5p 组 NCI-H1299 细胞的 CyclinD1 蛋白表达量、 增殖能力、 S 期细胞比例比 NC 组减少, Cleaved-caspase-3 蛋白表达量、 G0 / G1期细胞比 例、 凋亡率比 si-NC 组或 miR-NC 组增加 (P均 < 0. 05)。 circ_ 0061140 靶向调控 miR-6838-5p 的表达。 共转 染 si-circ_ 0061140、 anti-miR-6838-5p 可减弱转染 si-circ_ 0061140 对细胞生物学行为的作用。 结论 低表达 circ_ 0061140 通过靶向非小细胞肺癌细胞的 miR-6838-5p, 抑制细胞增殖、 阻滞 G0 / G1期细胞周期, 并加速 凋亡。

关键词: circ_ 0061140, 非小细胞肺癌, miR-6838-5p, 细胞增殖, 细胞周期, 细胞凋亡 

Abstract: Objective To investigate the effect of circular RNA (circRNA) circ_ 0061140 on the proliferation, cell cycle and apoptosis of non-small cell lung cancer cells via targeting miR-6838- 5p. Methods Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression levels of circ_ 0061140 and miR-6838-5p in the non-small cell lung cancer cells. The NCI-H1299 non-small cell lung cancer cells were divided into 7 groups: the si-circ _ 0061140 group (transfected with si-circ_ 0061140, circ_ 0061140 lowly expressed), the si-NC group (transfected with si-NC, the negative control), the NC group (blank control with no transfection of miRNA), the miR-6838-5p group ( transfected with miR-6838-5p mimic, miR-6838-5p highly expressed), the miR-NC group ( transfected with miR-NC, the negative control), the si-circ_ 0061140 + anti-miR-NC group (co-transfected with si-circ_ 0061140 and anti-miR-NC), the si-circ_ 0061140 + anti-miR-6838-5p group ( co-transfected with si-circ_ 0061140 and anti-miR-6838-5p). Western blotting was used to detect the expression levels of cyclin D1 and cleaved cysteinyl aspartate specific proteinase-3 ( cleaved-caspase-3). The proliferation of cells was detected by MTT method. The cell cycle and cell apoptosis was detected by the flow cytometry. Dual luciferase reporter assay was used to verify the relationship between circ_ 0061140 and miR-6838-5p. Results The expression level of circ_ 0061140 in the non-small cell lung cancer tissues and the cell lines NCI-H1299, NCI-H2170, NCI-H1975 were all higher than that in the adjacent tissues and the BEAS-2B normal lung epithelial cells, while the expression level of miR-6838-5p in the non-small cell lung cancer tissues and the lung cancer cells was lower than that in the adjacent tissues and BEAS-2B cells (all P < 0. 05). the expression level of cyclin D1 protein, the proliferation rate, and the ratio of S phase cells in cells of the si-circ_ 0061140 group and the miR-6838-5p group were lower than those of the NC group, while the expression level of cleaved-caspase-3 protein, the ratio of G0 / G1 phase cells, the apoptosis rate, and the mortality rate were higher than those of the si-NC group and the miR-NC group when compared with the NC group (all P< 0. 05). circ_ 0061140 targeted regulates the expression of miR-6838-5p. Co-transfection of anti- miR-6838-5p and si-circ_ 0061140 could attenuate the effect of si-circ_ 0061140 on the proliferation, cell cycle and apoptosis in non-small cell lung cancer cells. Conclusion The down-regulation of circ _ 0061140 in non-small cell lung cancer cells can suppress the proliferation of cells, block the cells in the G0 / G1 phase, and increase the apoptosis of cells by targeting miR-6838-5p.

Key words: circ _ 0061140, non-small cell lung cancer, miR-6838-5p, cell proliferation; cell cycle, cell apoptosis 

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