| 李长泳,贾萌,王琪,等.miR-375-3p抑制DNA双链断裂的同源重组修复增强结直肠癌细胞放射敏感性的研究[J].中华放射医学与防护杂志,2022,42(3):168-174.Li Changyong,Jia Meng,Wang qi,et al.MiR-375-3p enhanced radiosensitivity of colorectal cancer cells by inhibiting homologous recombination repair of DSBs[J].Chin J Radiol Med Prot,2022,42(3):168-174 |
| miR-375-3p抑制DNA双链断裂的同源重组修复增强结直肠癌细胞放射敏感性的研究 |
| MiR-375-3p enhanced radiosensitivity of colorectal cancer cells by inhibiting homologous recombination repair of DSBs |
| 投稿时间：2021-10-27 |
| DOI：10.3760/cma.j.cn112271-20211027-00425 |
| 中文关键词: 结直肠癌 miR-375-3p 同源重组修复 RAD51 放射敏感性 |
| 英文关键词:Colorectal cancer miR-375-3p Homologous recombination repair RAD51 Radiosensitivity |
| 基金项目: |
|
| 摘要点击次数: 3815 |
| 全文下载次数: 1355 |
| 中文摘要: |
| 目的 探究miR-375-3p调控结直肠癌细胞放射敏感性的作用和机制。方法 在结直肠癌细胞HCT116及HT29中过表达miR-375-3p，利用CCK-8法检测细胞增殖能力，利用克隆形成实验检测细胞克隆形成能力，利用Annexin V-FITC/PI双染法流式细胞术检测细胞凋亡，利用流式细胞术检测细胞周期分布。过表达miR-375-3p后，检测γ-H2AX foci形成点数量、同源重组（HR）及非同源性末端连接（NHEJ）修复效率，分析miR-375-3p表达对DNA双链断裂（DSBs）以及其修复效率的影响；利用生物信息学预测miR-375-3p在HR修复通路中的下游靶基因，利用双荧光素酶报告基因法进一步验证miR-375-3p表达对靶基因重组蛋白A（RAD51）表达调控作用。最后，利用荧光定量PCR技术检测经60Co γ射线2、6 Gy照射后HCT116细胞miR-375-3p的表达量；抑制miR-375-3p表达，经0、1、2、4、6 Gy不同剂量照射后，分析miR-375-3p表达变化对结直肠癌细胞HCT116放射敏感性的影响。结果 过表达miR-375-3p显著抑制了结直肠癌细胞HCT116及HT29的增殖及克隆形成能力，诱发其细胞凋亡、G1期周期阻滞和DSBs损伤，下调了Rad51表达，显著降低了HR修复效率[miR-375-3p（3.55±0.30）%，miR-nc（1.97±0.15）%；t=10.055，P<0.05]；双荧光素酶报告基因实验显示miR-375-3p能够靶向Rad51 3'UTR区域结合（t=5.013，P<0.05）；电离辐射能诱导miR-375-3p表达，抑制其表达显著降低了结直肠癌细胞放射敏感性（t=6.460、5.619、10.150，P<0.05）。结论 miR-375-3p能够靶向抑制Rad51表达，下调DSBs的HR修复效率，增强结直肠癌细胞的放射敏感性。 |
| 英文摘要: |
| Objective To investigate the effect of miR-375-3p on DNA damage repair and radioresistance of colorectal cancer cells. Methods After overexpression of miR-375-3p in HCT116 and HT29, cell proliferation ability was detected by CCK-8 assay, clone formation ability was detected by clone formation assay, apoptosis was detected by Annexin V-FITC/PI double staining method and cell cycle distribution was detected by flow cytometry, and the formation of γ-H2AX foci were used to analyze homologous recombination (HR) repair efficiency. Bioinformatics was used to predict the downstream target genes of miR-375-3p in the HR repair pathway. A dual luciferase reporter gene assay was used to validate the regulation effect of miR-375-3p on Rad51 gene. The expression of miR-375-3p in HCT116 cells irradiated with 60Co γ-rays at 2 and 6 Gy was measured by RT-qPCR. The inhibition effect of miR-375-3p on the radiosensitivity of HCT116 cells was analyzed after irradiation with different doses of 0, 1, 2, 4 and 6 Gy. Results Overexpression of miR-375-3p inhibited the proliferation and colony formation ability, induced G1 phase cycle arrest and cell apoptosis of colorectal cancer cells, enhanced DSBs formation, inhibited Rad51 expression, and significantly decreased HR repair efficiency (t=10.055, P<0.05). Dual luciferase reporter gene assay demonstrated that miR-375-3p bound to Rad51 3'UTR region (t=5.013, P<0.05). In addition, irradiation increased miR-375-3p expression, and inhibition of miR-375-3p expression reduced radiosensitivity of colorectal cancer cells (t=6.460, 5.619,10.150, P<0.05). miR-375-3p inhibited the homologous recombination repair efficiency of DSBs and enhanced the radiosensitivity of colorectal cancer cells. |
| HTML 查看全文 查看/发表评论 下载PDF阅读器 |
| 关闭 |
|
|
|