Effects Of Carbon Dioxide Pneumoperitoneum On The Proliferation Of Human Renal Carcinoma Cell Line In Vitro And Its Possible Mechanism

Since the first laparoscopic cholecystectomy was successfully performed in France, minimally invasive surgical techniques continued to advance in capability and popularity. Because of the advantages including less postoperative pain, shorter recovery period, and better cosmetic effects than conventional surgery, it is a feasible technology for biopsy and resection of various malignant tumors nowadays.However, whether CO2 pneumoperitoneum will promote trocar implantation and extensive abdominal metastasis after laparoscopic tumor operation was widely discussed in recent years, but there were still no conclusions. Numerous experimental studies confirmed that CO2 pneumoperitoneum changed the behavior of tumor cells, such as breast cancer, colon carcinoma, and ovarian cancer. The behavior of renal carcinoma cells has not been investigated in this respect as yet.In this study, we aimed to investigate the proliferation of renal carcinoma cells after exposure to simulated carbon dioxide pneumoperitoneum in vitro. And tried to elucidate the possible mechanism for providing initial assessment about the safety of CO2 pneumoperitoneum applied on renal carcinoma.PART ONE Effect of simulated CO2 pneumoperitoneum on cell proliferation of human renal carcinoma 786-O cells lineObjective : To study the effects of mimic carbon dioxide pneumoperitoneum environment on cell proliferation of human renal carcinoma 786-O cells.Methods:The renal carcinoma 786-O cells were exposed to simulated CO2 pneumoperitoneum environment for different time (2h or 4h) under the pressure of 14mmHg and those cultured in normal experiment condition as control, then proliferation viability of 786-O cells was observed by 3-[4, 5Dimethylthiazol-2-yl], 5-diphenyltetrazolium bromide or triazolyl blue (MTT) assay close behind the exposure; cell cycle and was detected by flow cytometry 48h after exposure.Result:Both of the treated groups of the cell proliferation was significantly increased than the control group(P<0.05); and the increasing had a linear relationship with exposure time (R2=0.9840, P<0.01); The proliferative index (PI) of treated groups showed significant difference than that of control group (P<0.01); In cell cycle, the percentages of S-phase and G2/M phase were significantly higher in treated groups than those in control group (P<0.05). Between the two treatment groups, compared with 2h group, the cell proliferation of 4h group was more rapid (P<0.05); the PI and the percentages of S-phase and G2/M phase were obviously higher (P all <0.05).Conclusion:Carbon dioxide could promote the proliferation of human renal carcinoma 786-O cells, and the proliferation would be sped up with the prolongation of exposure time.PART TWO Effect of simulated CO2 pneumoperitoneum on the expression of PCNA, Survivin and Caspase-3 in human renal carcinoma 786-O cell lineObjective : To study the effect of mimic carbon dioxide pneumoperitoneum on the expressions of PCNA, Surviving mRNA and protein as well as Caspase-3 protein in human renal carcinoma 786-O cell line, In order to elucidate the promoting mechanism on tumor growth.Methods:The renal carcinoma 786-O cells were exposed to CO2 environment for different time (4h or 2h) under the pressure of 14mmHg, with the untreated group as control, then the Survivin mRNA were examined by RT-PCR, the PCNA, Surviving and Caspase-3 protein were examined by immunocytochemistry respectively.Results : The carbon dioxide pneumoperitoneums significantly upregulated the expressions of PCNA protein(P<0.01), surviving mRNA and protein(P<0.01), but reduced the expression of Caspase-3 protein in human renal carcinoma 786-O cell line (P<0.01).Conclusion : The promoting effect of carbon dioxide pneumoperitoneum on malignant cells growth is probably through enhance the expression of PCNA and Survivin, and inhibiting the expression of Caspase-3 in tumor cells.