腹腔镜二氧化碳气腹环境对肾透明细胞癌细胞增殖和凋亡的影响

Influence of laparoscopic CO_2 pneumoperitoneum on proliferation and apoptosis of RCC-949 cells in vitro

目的通过模拟腹腔镜二氧化碳(CO2)气腹环境,研究CO2气腹对肾透明细胞癌细胞增殖和凋亡的影响,探讨腹腔镜手术治疗肾脏肿瘤的安全性。方法选用人肾透明细胞癌细胞株RCC-949在99% CO2、1%O2、21℃和15mmHg(1mmHg=0.133kPa)条件下模拟腹腔镜CO2气腹环境离体培养1、2、4h,应用脱氧核苷酸末端转移酶介导的生物素化dUTP缺口末端标记技术(TUNEL)检测RCC-949细胞在CO2气腹环境干预后的凋亡;采用噻唑蓝法(MTT法)检测RCC-949细胞在CO2气腹环境干预后的增殖。结果正常对照组、低氧1h组、低氧2h组和低氧4h组增殖的光密度值分别为0.348±0.051、0.372±0.082、0.389±0.110和0.378±0.114,凋亡细胞百分率分别为(3.2±0.5)%、(3.4±0.3)%、(3.8±0.4)%和(4.3±0.5)%,各组的差异均无统计学意义(P值均>0.05)。结论在模拟腹腔镜CO2气腹环境中培养4h之内,RCC-949细胞的增殖和凋亡水平无显著改变。由此推测,腹腔镜CO2气腹环境并不增加肿瘤细胞种植转移的机会,腹腔镜手术治疗肾脏肿瘤安全、有效。

Objective To investigate the influence of laparoscopic CO2 pneumoperitoneum on the proliferation and apoptosis of renal cell carcinoma （RCC-949） cells in vitro, so as to assess the safety of laparoscopic surgery for renal tumors. Methods Human clear cell renal cell carcinoma （RCC-949） cells were cultured for 1 hour, 2 hours and 4 hours in a simulated laparoscopic CO2 pneumoperitoneum environment at 21 ℃, 99% CO2, 1% O2 and 15 mmHg （1 mmHg=0.133 kPa）. The proliferation and apoptosis of RCC-949 cells cultured in the CO2 pneumoperitoneum environment were examined by terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） assay and methyl thiazolyl tetrazolium （MTT） assay, respectively. Results The optical density （OD） values of cell proliferation in the control group, hypoxia 1 h group, hypoxia 2 h group and hypoxia 4 h group were 0.348±0.051, 0.372±0.082, 0.389±0.110 and 0.378±0.114, respectively. Meanwhile, the apoptosis rates in the control group, hypoxia 1 h group, hypoxia 2 h group and hypoxia 4 h group were （3.2±0.5）%, （3.4±0.3）%, （3.8±0.4）% and （4.3±0.5）%, respectively. There were no significant differences in OD values or the apoptosis rates among all the groups （all P0.05）. Conclusion The proliferation and apoptosis rates of RCC-949 cells are not significantly changed within 4 hours of culture in the simulated laparoscopic CO2 pneumoperitoneum. Thus, we speculate that laparoscopic CO2 pneumoperitoneum does not promote tumor metastasis and implantation of RCC-949 cells, and laparoscopic surgery is safe and effective for renal tumors.