人工气腹对结肠癌细胞增殖的影响

Influence of artificial pneumoperitoneal media on colon carcinoma cell proliferation in vitro

目的探讨不同气体介质对人结肠癌细胞增殖的影响。方法建立体外模拟人工气腹的环境。通过MTT法、软琼脂克隆形成试验、免疫组织化学（免疫组化）、丫啶橙/溴乙啶双荧光染色、流式细胞仪等方法对比不同气腹介质对人结肠癌细胞增殖、细胞因子表达、凋亡及细胞周期的影响并作相关统计学分析。结果体外试验中,由MTT法可以看出,人结肠癌细胞LS-174T经CO2气腹作用后,细胞生存率高于对照组,He组较对照组低[吸光度（A）：0.37±0.02比0.33±0.01和0.30±0.01;P〈0.01]。克隆形成试验结果显示,CO2组细胞的增殖能力高于对照组和He组,差异有统计学意义[形成率：（32.8±3.6）%比（28.4±2.3）%和（23.5±2.7）%;P〈0.05],He组较对照组细胞增殖能力明显降低（P〈0.01）。免疫组化结果显示结肠癌细胞中HIF-1α在CO2组的阳性表达率明显高于He组和对照组,差异有统计学意义（P〈0.01）,He组高于对照组（P〈0.01）。血管内皮生长因子（VEGF）的阳性表达CO2组、He组与对照组比较差异有统计学意义（P〈0.01）,CO2组高于He组差异有统计学意义（P〈0.01）。双荧光染色结果可以看出He组凋亡百分率明显高于CO2组和对照组（P〈0.01）,CO2组和对照组差异无统计学意义（P〉0.05）。流式细胞仪结果显示CO2组G0/G1期细胞比例少于对照组、He组（P〈0.01）,He组G0/G1期细胞比例高于对照组（P〈0.01）。结论体外实验中CO2气腹可促进结肠癌细胞LS-174T的生长,He气腹相对抑制其生长。

Objective To evaluate the influence of different pneumoperitoneal media on colon carcinoma LS-174T cell proliferation in vitro. Methods The artificial pneumoperitonum was established. The proliferation of LS-174T cells was detected by MTT assay and soft agar clone formation assay. Expression of HIF-1α and VEGF was examined by immunohistochemistry. Apoptosis of LS-174T cells was analyzed by AO/EB double fluorescein stain and flow cytometry. Results The growth speed and proliferating capacity of LS-174T cells in CO2 pneumoperitoneum group[A：0.37±0.02,formation （32.8±3.6）%] were significantly higher than those in control group [A：0.33±0.01,formation （28.4±2.3）%] and He group [A：0.30±0.01,formation （23.5±2.7）%], meanwhile the He group was the lowest （P〈0.01）. Positive expression of HIF-1α and VEGF in CO2 and He artificial pneumoperitoneum up-regulated significantly as compared to control group（P〈0.01）, meanwhile the above expression was higher in CO2 group （P〈0.01）. The G0/G1 ratio in CO2 group was the lowest as compared to control group and He group （P〈0.01）, and G0/G1 ratio in He group was higher than that of control group（P〈0.01）. Aapoptosis rate in He group was the highest as compared with the other two groups（P〈0.01）. Conclusion CO2 pneumoperitoneum has stronger effect on the proliferation of colon carcinoma cell LS-174T as compared to He pneumoperitoneum in vitro.