靶向RAD18的小干扰RNA对人食管鳞癌ECA-109细胞增殖和化疗敏感性的影响

Effect of RAD18-siRNA on proliferation and chemotherapy sensitivity of human esophageal squamous cell carcinoma ECA-109 cells

目的：研究靶向RAD18基因的小干扰RNA （ siRNA ）对食管鳞癌细胞ECA-109增殖和化疗敏感性的影响。方法：合成针对 RAD18基因的 siRNA （ RAD18-siRNA），通过脂质体将其转染人食管鳞癌ECA-109细胞株，采用荧光定量PCR方法检测ECA-109 RAD18和CyclinD1 mRNA的表达，蛋白质印迹法检测其RAD18和CyclinD1蛋白的表达；应用CCK-8法检测ECA-109细胞的增殖和化疗药物对ECA-109细胞存活率的影响，流式细胞术检测ECA-109细胞周期；采用Pearson检验分析RAD18与CyclinD1基因表达的相关性。结果：与未转染组比较， RAD18-siRNA组RAD18表达明显降低（P＜0．05）。 RAD18-siRNA组细胞增殖抑制（P＜0．05），G1期细胞数增多，G2／M期细胞数减少（P＜0．05）。不同浓度顺铂或5-氟尿嘧啶处理细胞后，两组细胞的存活率均下降（均P＜0．05），但RAD18-siRNA组细胞半数抑制浓度较未转染组减少（ P＜0．05）。在食管鳞癌组织中， RAD18基因mRNA的表达与CyclinD1基因mRNA的表达呈正相关（ r＝0．478，P＜0．01）。结论：下调RAD18表达能够抑制食管鳞癌细胞的增殖，提高其对化疗药物的敏感性；CyclinD1在食管鳞癌中的表达水平可能参与这一过程。

Objective： To investigate the effect of RAD18-siRNA on cell proliferation and chemotherapy sensitivity of esophageal squamous cell carcinoma （ ESCC ） ECA-109 cells. Methods： RAD18-siRNA was transfected into human ECA-109 cells by Lipofectamine 3000. Quantitative PCR and Western blot were performed to detect RAD18 and CyclinD1 expression; CCK-8 assay was used to determine cell proliferation and chemotherapy drug sensitivity;flow cytometry was used to determine cell cycle.Correlation between RAD18 and CyclinD1 mRNA expression was analyzed by Pearson’ s correlation.Results：Compared with non-transfected cells, the expression of RAD18 in RAD18-siRNA group was significantly decreased （ P 〈0.05） .The cell proliferation was inhibited （ P 〈0.05 ） and the cell number of G1 phase was increased, G2/M phase cells decreased （ P〈0.05） in RAD18-siRNA group.After treatment with different concentrations of cisplatin or 5-FU, the survival rate of the two cell groups was reduced （all P〈0.05）, and the IC50 of RAD18-siRNA group was significantly lower than that of non-transfected group （ P 〈0.05 ） .The mRNA expression of RAD18 was positively correlated with CyclinD1 expression in ESCC tissues（ r =0.478, P 〈0.01）.Conclusion： Down-regulated expression of RAD18 can decrease the cell proliferation and increase chemo-sensitivity of ESCC cells, and CyclinD1 may participate in the process.