王梦洁，孟碧，李皓，高飞，叶婷，刘阳晨

(1蚌埠医学院研究生院，安徽蚌埠233030；2泰兴市人民医院)

转染miR-124模拟物的宫颈癌细胞株Siha放射敏感性变化及其机制探讨

王梦洁1，2，孟碧1，2，李皓2，高飞2，叶婷2，刘阳晨2

(1蚌埠医学院研究生院，安徽蚌埠233030；2泰兴市人民医院)

目的观察过表达微小RNA124(miR-124)对宫颈癌Siha细胞放射敏感性的影响，并探讨其机制。方法将宫颈癌Siha细胞分为观察组和对照组，观察组转染miR-124 模拟物，对照组不转染。采用实时荧光定量PCR法检测两组细胞miR-124和信号转导和转录激活因子3(STAT3) mRNA表达量。采用克隆形成实验检测两组细胞放射敏感性。采用流式细胞术检测两组细胞周期分布和放射后细胞凋亡率。结果观察组、对照组D0分别为1.062、1.542 Gy，Dq分别为1.605、1.970 Gy,SF2分别为0.527、0.685。观察组相对于对照组的放射增敏比为1.451。转染后24h观察组、对照组miR-124相对表达量分别为89.3±13.6、1.0±0.1，STAT3 mRNA相对表达量分别为0.3±0.03、1.0±0.1，两组相比，P均<0.05。观察组G0/G1期细胞比例为82.49%±1.97%、S期细胞比例11.87%±1.38%、G2/M期细胞比例为5.64%±0.72%；对照组分别为74.58%±1.28%、19.88%±0.26%、5.54%±1.05%。观察组G0/G1期比例高于对照组组，S期比例低于对照组(P均<0.05)，两组G2/M期细胞比例相比P>0.05。观察组、对照组细胞X线照射后细胞凋亡率分别为45.87%±3.16%、37.27%±0.87%，两组相比，P<0.05。结论过表达miR-124可能提高宫颈癌Siha细胞的放射敏感性。其机制可能是过表达miR-124会抑制宫颈癌Siha细胞STAT3表达，进而阻滞细胞周期于G0/G1期，促进细胞凋亡，从而提高宫颈癌Siha细胞的放射敏感性。

宫颈癌细胞；微小RNA124；信号转导和转录激活因子3；放射敏感性；细胞凋亡；细胞周期

Abstract:ObjectiveTo observe the effect of microRNA124 (miR-124) overexpression on the radiosensitivity of cervical cancer Siha cells and its mechanism.MethodsThe cervical cancer Siha cells were divided into the observation group and control group. The miR-124 mimics were transfected into the observation group. The expression levels of miR-124 and signal transducer and activator of transcription 3 (STAT3) in the two groups were detected by real-time fluorescent quantitative PCR. Colony formation assay was used to detect the radiosensitivity of the two groups cells. Clone formation assay was used to detect the cell radiosensitivity. Flow cytometry was used to detect the cell cycle distritution and apoptosis rate after radiation.ResultsThe observation group and the control group had the D0 parameters of 1.062 Gy and 1.542 Gy, the Dq parameters of 1.605Gy and 1.970Gy, the SF2 parameters of 0.527 and 0.685, respectively. Compared with the control group, the radiation sensitization enhancement ratio (SER) was 1.451. At 24 h after transfection, the relative expression levels of miR-124 in the observation group and the control group were 89.3±13.6 and 1.0±0.1, respectively, and the relative expression levels of STAT3 mRNA were 0.3±0.03 and 1.0±0.1, respectively; significant difference was found between these two groups (allP<0.05). In the observation group, the percentage of cells in G0/G1phase was 82.49%±1.97%, was 11.87%±1.38% in S phase, and 5.64%±0.72% in G2/M phase；meanwhile, they were 74.58%±1.28%, 19.88%±0.26%, 5.54%±1.05%, respectively, in the control group. Compared with the control group, the percentage of cells in G0/G1phase was higher, while the percentage of cells in S phase was lower, and significant difference was found between these two groups (allP<0.05). Compared with the control group, the percentage of cells in G2/M phase was not significantly different (P>0.05). The apoptosis rates after X-irradiation were 45.87%±3.16% in observation group, versus 37.27%±0.87% in the control group, and significant difference was found between these two groups (P<0.05).ConclusionsOverexpression of miR-124 may enhance radiosensitivity of cervical cancer Siha cells. The mechanism may be that overexpression of miR-124 can inhibit the expression of STAT3 in cervical cancer Siha cells, and then induce the cell cycle arrest in G0/G1phase, promote the apoptosis, and thereby enhance the radiosensitivity of cervical cancer Siha cells.

Keywords: cervical carcinoma cell; microRNA-124; signal transducer and activator of transcription 3; radiosensitivity; apoptosis; cell cycle

放疗抵抗是导致肿瘤放疗失败的主要原因[1,2]。放疗主要是通过损伤癌细胞的DNA，进而导致癌细胞发生凋亡和不可逆性细胞周期阻滞[3,4]。miRNA在这一过程中起重要作用[5,6]。miR-124是一种高度保守的miRNA，在多种实体肿瘤中低表达，如头颈部鳞状细胞癌、胶质细胞瘤、非小细胞肺癌、乳腺癌、肝癌等[7～11]，miR-124在肿瘤的发生发展过程中起着类似于抑癌基因的作用。过表达miR-124可以提高上述肿瘤组织对放疗的敏感性。但是宫颈癌细胞中miR-124表达情况及过表达miR-124对宫颈癌Siha细胞放射敏感性的影响尚未见文献报道。……