The Influence Of Carbon Dioxide Pneumoperitoneum Models On The Proliferation And The Expression Of CDK4 And Cyclin D1 In Human Cervical Carcinoma Cell Line CASKI

With its advantages of minimaly invasion, less pain and quicker recovery, laparoscopy have been used more and more broadly in gynecologic operations in recent years. The laparoscopy equipments and technique have been improved continuously, even used for gynecologic malignant tumors and made some progress. However, there are increasing reports that the malignant tumors grow faster and recur more easily after pneumoperitoneum with carbon dioxide (CO2), particularly some clinical trials show that the port sites occur tumor metastases. Therefore , the influence of CO2 pneumoperitoneum upon malignant tumors is become one of research focuses in recent years. there are some controversy about the influences of CO2 pneumoperitoneum on malignant tumors . As for the mechanism of CO2 pneumoperitoneum effecting on malignant cells proliferation, related publications are much concentrated on the cells exogenous control pathway such as environment changes, immunity factors etc, but lack of reports about the cells endogenous control pathway.Therefore, this research is aimed to observe the influences of CO2 pneumoperitoneum on human cervical carcinoma cells line (CaSki) growth and proliferation in vitro, cell cycle and cycle related protein CDK4 and Cyclin D1 expression, so that the possible endogenous etiologies of malignant tumor cells'proliferation be explored.PART ONE THE INFLUENCE OF CARBON DIOXIDE PNEUMO- PERITONEUM MODELS ON THE PROLIFERATION IN HUMAN CERVICAL CARCINOMA CELL LINE【Purpose】To observe the influence of different treatment time by CO2 condition which imitating laparoscopic pneumoperitoneum of laparoscopic operation, on the proliferate ability of CaSki line in vitro and explore the relative mechanisms.【Method】CO2 artificial pneumoperitoneum model was established. CaSki cell line was treated 2h, 4h with pure CO2 , pressure 15 mmHg respectively. The CaSki cells without treatment as the control. Cell growth curves were determined by spectrophotometer assay (MTT) 1-5d after CO2 treatment; The cell cyclic rate and proliferation rate were measured with flow cytometry method (FCM) to calculated proliferation index (PI). 【Result】Following culture time prolonged, we found that①The OD value of MTT assay of treatment groups were gradually increased and higher than that of the control group at 2d(P < 0.05), even significant higher at 3d to 5d(P < 0.01) ;②The growth curve of treatment groups were gradually sharp than that of control group at 2d to 5d, however, there were gradually flat with the culture time prolonged to 5d;③The PI of treatment groups were higher than that of control at 1d to 5d(P < 0.05);④The ratio of G0-G1phase of the CaSki cells were gradually declined and significantly lower than that of the control group at the same period (P < 0.05) at 1d to 5d;【Conclusion】After treated in the CO2 pneumoperitoneum,the CaSki cells grow faster and the PI become higher than those CaSki cells without any special treatment. It shows that CO2 pneumoperitoneum promotes the growth and proliferation of cervical carcinoma cell.PART TWO THE INFLUENCE OF CARBON DIOXIDE PNEUMO- PERITONEUM MODELS ON THE EXPRESSION OF CDK4 AND CYCLIN D1 IN HUMAN CERVICAL CARCINOMA CELL LINE CASKI 【Purpose】To explore the mechanisms of the growth and proliferation of CaSki cells line in vitro, from the cells ' endogenous control pathway, by observing the changes on expressions of the cell cycle regulating factors CDK4 and Cyclin D1 after different CO2 treatment time in the imitating laparoscopic pneumoperitoneum situation.【Method】CO2 artificial pneumoperitoneum model was established. CaSki cells were treated 2h, 4h with pure CO2 , pressure 15mmHg respectively. The CaSki cells without treatment as the control. The changes on expressions of CDK4 and the Cyclin D1 in CaSki cells were measured by immunity cell chemistry and Western-blot.【Result】①It was found that there were positive expressions of CDK4 and Cyclin D1 in CaSki cells;②Positive expressions of CDK4 and Cyclin D1 in treatment groups were stronger than that of control group(P < 0.01);③All cultured spots of 1d to 5d, the OD value of CDK4 and Cyclin D1 in treatment groups were higher than that of control group(P < 0.05).【Conclusion】After treated in the CO2 pneumoperitoneum,the expressions of CDK4 and Cyclin D1 in cervical carcinoma CaSki cells were higher than those of without any treatment. It shows that CO2 could premote the proliferation of CaSki cells by the endogenous control pathway of proliferation.