The Study On Drug Reverse Effect And Reverse Mechanism Of Naringin On Multi-drug Resistantce Human Ovarian Cell Line SKOV3/DDP

Purpose: To culture in vitro human ovarian cancer cell SKOV3 and cisplatin resistant ovarian carcinoma cell SKOV3/DDP. Moreover, to investigate the effect of naringin on the vitro proliferation, study its reverse effect on the drug resistance SKOV3/DDP cell and to explore related mechanisms.Method: 1?Different concentrations of cisplatin were used in SKOV3 and SKOV3/DDP cells respectively. The MTT method was used to measure cell proliferation inhibition rate after 48 hours. Being able to obtain the respective IC50, the resistance index of SKOV3/DDP was calculated. 2?With different concentrations of naringin(5, 10, 20, 40, 80?mol/L) acted on human ovarian carcinoma drug resistant cells SKOV3/DDP. Each concentration was given a different action time(24, 48 and 72 hours). MTT method of calculation was used to determine the inhibition cell rate of SKOV3/DDP for groups with different drug concentrations and action times. 3?Determination of the half inhibitory concentration of naringin in combination with cisplatin on SKOV3/DDP cells under the action of non-cytotoxic naringin concentration was made, to judge whether naringin can increase the sensitivity of the resistant cells to cisplatin. 4?Different groups: sedentary control group, separate group of naringin, separate group of cisplatin, group of naringin in combination with cisplatin, after 48 hours the groups function, determine the expression of MDR1 m RNA and MRP2 m RNA in SKOV3/DDP cells of different experimental groups by adopting RT-PCR method; detect the expression level of P- gp, MRP2 protein in SKOV3/DDP cells of different experimental groups by western blot method.Results: 1?SKOV3/DDP cells resistant index RI: 2.73. After 48 hours of treating SKOV3 and SKOV3/DDP cells with different concentrations of DDP, IC50 of SKOV3 cells was 5.48 ?g/ml, IC50 of SKOV3/DDP cells was 14.93 ?g/ml, and RI was2. 72,according to the results using the MTT method. 2 ? Naringin has certain inhibitory effect on the vitro proliferation of SKOV3/DDP. The result showed that naringin being used alone in certain concentration range can effectively restrain the proliferation of human ovarian cancer resistant cell SKOV3/DDP cells, showing dependency both on time and concentration. The proliferation inhibition rate of SKOV3/DDP after 10?mol/L of naringin act on 24h?48h?72h is 3.24±1.18?6.61±0.19?9.53±0.24,The effect of naringin on SKOV3/DDP cells after 48 hours showed that in order to acquire a 10% growth inhibition an IC10 concentration of 12.66?mol/L was required. To avoid excessive cell death, the concentration of 10umol/L was chosen for this experiment. 3?Naringin in combination with DDP can strengthened the cytotoxic effect of DDP on SKOV3/DDP cells. After,48 hours, 10umol/ml of naringin in combination with DDP acted on SKOV3/DDP cells, the calculated IC50 showed that SKOV3/DDP to DDP decreases from 14.93?g/ml to 8.87?g/ml, The RI decreases from 2.72 to1.62,and the drug resistance reversal fold is 1.68 times, which showed that naringin can partially reverse the drug resistance of SKOV3/DDP cells. 4?The expression of MDR1 m RNA and MRP2 m RNA in SKOV3/DDP cells: separate group of naringin and combination group all have a lower expression of MDR1 m RNA and MRP2 m RNA compared to sedentary control group,the difference is statistically significant(p<0.05), The expression of MDR1 m RNA and MRP2 m RNA decrease significantly in group of naringin in combination with cisplatin compared to DDP group, the difference is statistically significant(p<0.05). 5 ? The expression of P-gp and MRP2 protein in SKOV3/DDP cells: combination group all have a lower P-gp and MRP2 protein expression compared to separate group of cisplatin,the result is significant difference( P < 0. 01).separate group of naringin, separate group of cisplatin and combination group all have a lower expression of P-gp and MRP2 protein compared to sedentary control group,the difference is statistically significant(p<0.05)Conclusion: naringin can significantly inhibit the proliferation of SKOV3 / DDP cell line in vitro, and can enhance the sensitivity of SKOV3 / DDP cell to cisplatin sensitivity, the reversal of drug resistance mechanism may be related to inhibit the expression of MDR1 genes and multidrug resistance associated protein(MRP2) gene. Naringin is expected to become a new drug treatment for ovarian cancer, through conbination use with DDP, to improve the survival rate of patients with ovarian cancer.