Studies On Effects Of CO₂ Pneumoperitoneum Pressure On Cytological And Molecular Mechanisms Variation Of Endometrioal Lesion In A Rat Model

BackgroundEndometriosis(EMs) is a common gynecological disease which occurs more among in reproductive age women.The clinical symptoms contain menstruation haemorrhage, dysmenorrheal,pelvic pain and infertility.With the wide use of laprascopy recent years, the diagnostic rate of EMs becoming increased.Laparoscopy now is the gold standard in disease final diagnosis,staging and treatment.The recurrence rate of drug treatment is still higher,varying with the clinical stage,follow-up phase and medicine variety,with the highest recurrence rate to be 61.5 percent within three years(reported by Domowshi and Cohen).The effect of therapeutic alliance of surgery and medicine is superior to medicine or surgery application alone.It can preserve the reproductive and endocrine function by drug treatment or conservative and semiconservative operation(laparoscopy or abdominal operation).But the recurrence is still a troublesome problem,which lead us to emphasize the fundamental study,searching for more perfect treatment policies and recurrence prevention.In the clinical treatment of EMs,we find a low recurrence rate of laparoscopic treatment,which is lower than those results reported by other studies.Besides the clinical stage,laparoscopic techniques and treatment degree,there is still no any study on whether the CO₂ pneumoperitoneum(PNP) has an effect on EMs lesion during laparoscopic surgery.Many researches showed that CO₂ pneumoperitoneum stimulated the tumor cells implantation and metastases during laparoscopy,which increased the risk of recurrence. There has been no any correlated report on effect of CO₂ pneumoperitoneum on EMs lesion.Whether CO₂ pneumoperitoneum cause a series of cytological and molecular variation of EMs lesion that may affect the post-operative recurrence.The objective of our study is to set up the EMs rat model and to evaluate the pneumoperitoneum pressure on post-operative recurrence of EMs by determining the cytological and molecular variation of EMs lesion caused by CO₂ pneumoperitoneum.Partâ… Effects of CO2 pneumoperitoneum on achievement rate of foundation of EMs rat modelObjective:To evaluate the effect of CO₂ pneumoperitoneum on EMs model foundation.Methods:To erect the EMs rat model according to the method of induced EMs animal model.To select the female SD rats(mean weight 250～300g) and a intramuscular injection of estradiol benzoate(0.1ml/kg) to make the animals in estruation at the same time.The left uterus was resected and cut open,then endometrial tissue were sutured on both sides of the peritoneum.The rats were divided randomly into four groups(ten per group):control group;group A:pre-operation;group B:4th day post-operation;group C:10th day post-operation of model erection.Then CO₂ pneumoperitoneum(15mmHg,1h) was simulated pre or post operation and the achievement rate of model foundation,cytokine and molecular biological parameter were determined to evaluate the effect of CO₂ pneumoperitoneum on EMs model foundation.Results:After four weeks of CO₂ pneumoperitoneum intervention pre and post operation, the achievement rate of model foundation is 100 percent in control group and group A,70 and percent in group B and C respectively.There is a slight increase or equal level of cytokines and molecular variation in group A compared with control group.And there is a significant decrease more often in group B and C than that of gourp A(P＜0.05).Moreover there is no statistic difference between group B and C(P＞0.05).Conclusion:The achievement rate was decreased by CO₂ pneumoperitoneum via inhibition of aggression,adhesion and angiogenesis.Partâ…¡Effects of CO2 pneumoperitoneum pressure on aggression,adhesion and angiogenesis of EMs lesionObjective:To evaluate the effect of different CO₂ pneumoperitoneum pressure on cytokines level of EMs lesion and serum and the correlation factors with post-operation recurrence.Methods:The rats were divided randomly into three groups(control group,10mmHg and 20mmHg CO₂ pneumoperitoneum,1h) after EMs model being founded successfully.After the CO₂ pneumoperitoneum was over,the right EMs lesion was resected and resutured on the situ peritoneum again.The correlated cytokines of aggression(MMP2/TIMP-1), adhesion(CD44v6,ICAM-1) and angiogenesis(VEGF/ENS,TNF-α) in EMs lesion and serum were detected at 1w,2w and 6w after pneumoperitoneum intervention.Then to evaluate the effect of different CO₂ pneumoperitoneum pressure on cytokines level of EMs lesion and serum and the correlation factors with post-operation recurrence.Results:Compared with control group,the cytokine level correlating with aggression, adhesion and angiogenesis in the two CO₂ pneumoperitoneum groups decreased significantly in early post-operation(P＜0.05).Moreover the cytokines level in 10mmHg group was lower than group with 20mmHg pneumoperitoneum(P＜0.05),and this effect can be continued until two weeks after pneumoperitoneum intervention.The EMs lesion after resection can implant and grow sequently and has the similar cytokines level with EMs lesion on the other side of peritoneum(P＞0.05).The level of cytokines increased gradually with time after CO₂ pneumoperitoneum intervention,but no difference compared with control group(P＞0.05).Conclusion:The low CO₂ pneumoperitoneum pressure group is superior to the high pressure group in inhibition of aggression,adhesion and angiogenesis,which may be one of the factors that correlate with the low post-laparoscopic recurrence.There is a similar invasive capability of EMs lesions with or without resection. Partâ…¢Effects of CO2 pneumoperitoneum pressure on apoptosis and the Ultrastructure of cellular organelle of EMs lesionObjective:To evaluate the effect of CO₂ pneumoperitoneum pressure on EMs lesion and its correlation with post-operative recurrence.Methods:Thirty rats were divided randomly into three groups(five per group),control group,10mmHg and 20mmHg CO₂ pneumoperitoneum groups,with duration lasting for 1h.EMs lesion and peritoneum nearby were sampled for electron microscope examination to determine the apoptosis and ultrasturcture variation of cell organelle of EMs gland cells and peritoneal mesothelium.To evaluate the effect of CO₂ pneumoperitoneum pressure on EMs lesion and its correlation with post-operative recurrence.Results:There is a well being structure of glandular cells with matrix cells and peritoneal mesothelium lining up in order and normal structure of microvilli(MV).While in groups of 10mmHg and 20mmHg,the apoptosis of glandular cells increased with mitochondrial crista decreasing or vanishing and mitochondria swelling and moreover there is no significant difference between the two groups.But compared with the 10mmHg group, there is more severe damage of peritoneal mesothelium with microvilli vanishing, apoptosis increasing,cell junction widening and collapse and matrix cells lining up in disorder.Conclusion:There is a significant promotion of glandular cells apoptosis,which weakens the aggression capability of EMs lesion.But there is a little more worse damage of peritoneal mesothelium and matrix cells of EMs lesion,which reduces the defense capability of peritoneum.Therefore it is suggested that the pneumoperitoneum be selected with appropriate pressure.Summary1.The achievement rate was decreased by CO₂ pneumoperitoneum via inhibition of aggression,adhesion and angiogenesis.2.Both of the CO₂ pneumoperitoneum group with 10 and 20mmHg pressure can inhibit the aggression,adhesion and angiogenesis of EMs lesion and the low pressure are superior to the high pressure group,which may be one of the factors that correlate with the low post-laparoscopic recurrence.3.There is a significant promotion of glandular cells apoptosis and ultrastructure destruction,which weakens the aggression capability of EMs lesion.In conclusion,the aggression,adhesion and angiogenesis of EMs lesion can be inhibited by CO₂ pneumoperitoneum with the lower pressure having the better effects, and a significant apoptosis and ultrastructure destruction changing of glandular cells caused by CO₂ pneumoperitoneum,which may be one of the factors that lead to low post-laparoscopic surgical recurrence.