Study On The Effect And Machanism Of Ceramide In Augmenting Sensitivity Of Cisplatin In Ovarian Carcinoma

ObjectiveThe study intends to observe the effect and machanism of ceramide in augmenting sensitivity of cisplatin in ovarian carcinoma,to screen effective reversal agent of DDP resistance in ovarian cancer and to provide experimental evidence.MethodsMTT assay was used to analyze the curve of growth of SKOV3and SKOV3/DDP cells, IC50value of DDP, the effect of ceramide to augment sensitivity of cisplatin in ovarian carcinoma at different dosage and action time. The effect of ceramide and DDP to expression of Bcl2was detected by RT-PCR and Western Blot.The application of flow cytometry analysed the apoptosis rate of cell. MDC fluorescence staining were used to detect the formation of autophagosomes in DDP’s chemical sensitivity on ovary carcinoma cell line SKOV3and drug-resistance on ovary carcinoma cell line SKOV3/DDP.Results1.SKOV3and SKOV3/DDP cells growth adhsively.The multiplicate time of SKOV3cell was33.36hours and SKOV3/DDP cell was42.60hours; DDP’s IC50is3.384μg/ml on SKOV3,and13.770μg/ml on SKOV3/DDP, the multiple of drug resistance is4.07.2.The ceramide could inhibit SKOV3and SKOV3/DDP cells and the inhibition rate showed time-concentration-dependent effect. Under the same concentration of ceramide, the inhibition rate increased along with the time extension; while under the same reaction time, the inhibition rate increased along with the increase of drug concentration.3. Bcl2mRNA in SKOV3cells were less than that in SKOV3/DDP cells.The cells were divided into ten groups which were Cer5group. Cer10group. Cer20group. Cer40group. Cer5+DDP group. CerlO+DDP group. Cer20+DDP group. Cer40+DDP group, DDP group and control group.Comparing to SKOV3cells, the expression of Bcl2mRNA in SKOV3/DDP cells were increased36.38%、37.33%.43.38%、46.40%.42.05%.44.99%.41.90%.39.17%.43.03%.32.67%separately.In SKOV3cells, the expression of Bcl2mRNA in ten groups were different(F=159.722, P=0.000).000),there were significant difference between ten groups(P<0.05). In SKOV3/DDP cells, the expression of Bcl2mRNA in ten groups were different(F=182.380, P=0.000),there were significant difference between ten groups(P<0.05).4. The expression of Bcl2protein in SKOV3cells were less than that in SKOV3/DDP cells.The cells were divided into ten groups which were Cer5group. Cer10group. Cer20group, Cer40group. Cer5+DDP group. Cer10+DDP group. Cer20+DDP group. Cer40+DDP group, DDP group and control group.Comparing to SKOV3cells, the expression of Bcl2protein in SKOV3/DDP cells were increased40.39%.40.95%.47.63%.44.60%.42.93%.44.73%.43.26%.44.68%.47.48%.35.94%separately.In SKOV3cells, the expression of Bcl2protein in ten groups were different(F=l04.062, jP=0.000),there were significant difference between ten groups(P<0.05). In SKOV3/DDP cells, the expression of Bcl2protein in ten groups were different(F=210.458, P=0.000),there were significant difference between ten groups(P<0.05).5. The rate of apoptosis was more obviously increasing along with increasing drug concentration and action time. Under the same drug concentration, the rate of apoptosis increased along with the prolonged reaction time, and there was significant difference between24h and48h groups(P<0.05). Under the same reaction time, there was significant difference during different drug concentration groups(P<0.05).6. With the increasing concentration and the prolonged reaction time of ceramide, the flourescence intensity is augumented.Conclusions1. Ceramide could enhance the action of growth suppression of SKOV3and SKOV3/DDP cells to cisplatin, the inhibitory action is not influenced by cisplatin resistance in ovarian carcinoma in vitro.2. Influencing the gene and protein’s expression of Bcl2, inducing cell apoptosis and autophagy, ceramide intends to augment sensitivity of cisplatin in ovarian carcinoma.