CO_2气腹对结直肠癌细胞增殖及microRNA-221和CDKN1C/p57表达的影响

Effects of CO_2 pneumoperitoneum on cell proliferation and expression of microRNA-221 and CDKN1C/p57 in colorectal carcinoma cells

目的:通过检测不同压力CO2气腹环境下结直肠癌细胞周期和microRNA-221(miR-221)及CDKN1C/p57表达的变化,探讨CO2气腹对结直肠癌细胞增殖的影响。方法:将人结直肠癌CaCO2细胞系分别置于0、10、12和15mm Hg的CO2气腹环境下培养4h,应用实时荧光定量PCR检测CaCO2细胞中miR-221表达状况,运用Western-blot分析CDKN1C/p57蛋白表达状况,并通过噻唑蓝(MTT)比色法和流式细胞仪观察细胞的增殖状态和分析细胞周期。结果:与对照组相比,15mm Hg的CO2气腹组结直肠癌细胞中miR-221表达量下降[(1.212±0.159)vs.(1.937±0.208)],而CDKN1C/p57蛋白表达增高[(2.017±0.808)vs.(0.972±0.416)],同时细胞增殖抑制率[(32.7±5.1)%vs.(21.8±3.6)%]增加,且G0/G1期细胞比例明显增加,S期细胞比例显著下降,差异均有统计学意义(P<0.01);0、10、12mm Hg的CO2气腹组结直肠癌细胞增殖抑制率及miR-221、CDKN1C/p57表达水平与对照组相比差异均无统计学意义(P>0.05)。结论:临床常用压力的CO2气腹对结直肠癌细胞增殖无明显影响;15mm Hg的CO2气腹可抑制癌细胞增殖,其原因可能与miR-221表达下降,继而CDKN1C/p57蛋白表达增高有关。

Objective：To investigate the effects of CO2 pneumoperitoneum of different pressures on cell proliferation and expression of microRNA-221（miR-221） and CDKN1C/p57 in colorectal carcinoma cells.Methods：Human colorectal carcinoma cell line CaCO2 were exposed to 0,10,12 and 15mm Hg CO2 pneumoperitoneum in vitro for four hours.The miR-221 expression pattern was detected by real-time RT-PCR following the protocols recommended by commercial corporation.The protein level of CDKN1C/p57 expression was detected by Western-blot analysis.Furthermore,cell proliferation and cell cycle analysis were measured by MTT assay and flow cytometer.Results：The cellular proliferation inhibition rate in 15mm Hg CO2 pneumoperitoneum group enhanced significantly compared with control group [（32.7±5.1）% vs.（21.8±3.6）%],while that in other groups did not changed significantly.The expression of miR-221 was significantly down-regulated in 15mm Hg CO2 pneumoperitoneum group compared with control group [（1.212±0.159） vs.（1.937±0.208）],while CDKN1C/p57 protein expression enhanced markedly [（2.017±0.808） vs.（0.972±0.416）].Conclusions：There was no obvious effects of clinical CO2 pneumoperitoneum on colorectal carcinoma cell proliferation;15 mm Hg CO2 pneumoperitoneum could inhibit cell proliferation by inhibiting miR-221 expression and up-regulating CDKN1C/p57 protein expression consequently.