The Effect Of Psoralen On Reversing P-gp-mediated Breast Cancer Resistance By Reducing Caveolin-1

Objective:Breast cancer is considered to be one of the malignant tumors affecting women's life and health.Although great progress has been made in treatment,multidrug resistance(MDR)remains the leading cause of death in breast cancer patients.As one of the main extracts of psoralen,psoralen has been reported to reduce the resistance of MCF-7/ADR to doxorubicin after psoralen intervention,and it has a wide range of effects and low natural Poison and other characteristics,but the specific mechanism is not yet clear.In this study,the treatment of breast cancer drug-resistant MCF-7/ADR by psoralen was used to detect the activity and expression of P-gp and the expression of caveolin-1,and the co-localization of the two,and to explore significance.This study will help to improve the anti-tumor theory of psoralen and provide a theoretical basis for the development and utilization of psoralen as a multidrug resistance reversal agent.Methods:1.Culture MCF-7/ADR breast cancer resistant cell lines in RMPI-1640 complete medium containing doxorubicin.2.To detect psoralen against MCF-7 by comparing the 50%inhibition rate(IC50)of doxorubicin to MCF-7/ADR and psoralen-treated MCF-7/ADR cells./ADR reversal effect.3.BALB/c nude mice were used for breast cancer modeling experiments.After group intervention,tumor volume was measured and pathological changes of heart,liver and kidney were detected by immunohistochemistry.4.Detection of P-gp ATPase activity and kinetic analysis.Western Blot and RT-PCR were used to detect the effect of different concentrations of psoralen on the expression of P-gp.Immunohistochemistry was used to detect the effect of psoralen on the expression of P-gp in nude mice.6.The effect of P-gp on the efflux of ADR and Rh123 was examined by microscopic fluorescence technique.7.Western Blot and RT-PCR were used to detect the effect of different concentrations of psoralen on the expression of caveolin-1.Immunohistochemistry was used to detect the effect of psoralen on the expression of caveolin-1 in nude mice.8.Immunofluorescence technique were used to detect the effect of co-localization of caveolin-1 and P-gp.Result:1.Measure the chemosensitivity of MDR cells MCF-7/ADR to chemotherapeutic drugs by comparing IC 50 values.Reversal fold value=IC50 of MC50/MCF-7/ADR of MCF-7/ADR+psoralen.The IC50 of MCF-7/ADR+psoralen and MCF-7/ADR were 24.28±1.53?g/ml and 75.32± 2.52?g/ml,respectively,and the reversal fold was 3.1 times(p<0.05).2.The tumor volume of mice treated with psoralen was significantly lower than that of the control group,and significant damage was found in the hearts,kidneys and liver of mice after psoralen treatment.The results of RT-PCR and Western Blot showed that psoralen had no effect on the expression of P-gp at mRNA and protein levels(p>0.05).The analysis of immunohistochemistry results of tumor samples showed that after psoralen intervention,There was no change in the expression of P-gp(p>0.05).4.Immunofluorescence results showed that the accumulation of ADR and Rh123 in MCF-7/ADR after psoralen treatment was increased.5.Western Blot and RT-PCR results showed that the expression level of caveolin-1 was treated with MCF-7/ADR cells treated with 0,8,12,16 mg/L psoralen compared with the control group.The decrease was significantly decreased(p<0.05).The immunohistochemical results of tumor samples showed that the expression of caveolin-1 was significantly decreased after psoralen intervention(p<0.05).6.Yellow fluorescence was observed in the cell membrane region after superposition of red fluorescent caveolin-1 and green fluorescent P-gp in MCF-7/ADR group,which proved that P-gp shared with caveolin-1.Positioning relationship.However,in the MCF-7/ADR group after psoralen treatment,the distribution of caveolin-1 became dispersed.And after the red fluorescent caveolin-1 and the green fluorescent P-gp were superimposed,the yellow fluorescence of the cell membrane region was significantly reduced.The relative fluorescence intensity of MCF-7/ADR+psoralen was 28.49%± 1.03,which was significantly lower than that of MCF-7/ADR group(51.09%± 1.31).Compared with the two,the relative fluorescence intensity of the cells after psoralen treatment was reduced by 0.56 times(p<0.05).This indicates that psoralen disrupts the colocalization relationship between caveolin-1 and P-gp.Conclusion:1.Psoralen significantly reversed the drug resistance of MCF-7/ADR.2.Psoralen can reduce the growth of MCF-7/ADR xenografted nude mice,and does not cause toxic damage to liver,heart,kidney and other organs at effective concentration.3.Psoralen has no effect on P-gp expression.4.Psoralen reduces the drug efflux transport activity of P-gp.5.Psoralen reduces the expression of caveolin-1.6.Psoralen disrupts the colocalization relationship between caveolin-1 and P-gp.