Protective Effects Of Berberine Against Cardiomyocyte Damage Induced By Adriamycin In AC16 Cells

Objective: Adriamycin(ADR)is a kind of anthracycline antibiotic extracted from the Streptomyces peucetius var.caesius,with broad spectrum and high-effective anti-tumor effect.However,the risk of irreversible and potentially fatal myocardial injury is the main drawback of adriamycin in clinical application.The mechanism of myocardial injury induced by adriamycin is not clear which may be caused by multiple factors.Previous study shows that one of the main factors is that Adriamycin can lead to intracellular ROS content increment,and the latter can cause DNA damage in myocardial cells,then the cell chromosome aberration eventually lead to the apoptosis,or even necrosis.Therefore,in order to improve the effect of cancer treatment,expand the scope of application of Adriamycin,development of cardioprotective agents against myocardial injury caused by adriamycin is the direction of study for many scholars.Berberine,mainly extracted from traditional Chinese medicine Coptis,is an isoquinoline alkaloid,with heat-clearing,detoxification,fire-purging,lowering blood sugar,lowering blood pressure,immune regulation and other effects.Berberine has no toxic effect on normal cells,but can inhibit tumor cell proliferation,also berberine has a protective effect against adriamycininduced myocardial injury.However,the underlying mechanism is unclear.Therefore,to verify the improvement of berberine on adriamycin-induced myocardial injury and analyze the underlying mechanism,the effects of berberine on DNA injury induced by Adriamycin were observed.The study can provide basic research data for the clinical application of berberine.Method:1 In order to establish a adriamycin-induced cardiomyocyte model,the viability of cardiomyocytes caused by different concentrations of adriamycinwas determined by MTT assay,and the cytotoxicity of adriamycin was further determined.Finally,the appropriate concentrations of adriamycin were selected.2 Cardiomyocytes were randomly divided into six groups: blank control group,adriamycin group,berberine group,low-concentration(1?mol/L)berberine + adriamycin group,middle-concentration(5?mol/L)berberine +adriamycin group,high-concentration(10?mol/L)Berberine + adriamycin group;The cells in different groups were cultured at incubate box with 5%CO2 at 37 ? after adding the drug.The effects were observed after 24 hours.The adriamycin was given after berberine administration for half an hour.3 The content of ROS in cells was measured by H2 CDFA dye with flow cytometry.The DNA damage was detected by comet assay and the expression of DNA repair enzyme was detected by RT-PCR.The differences between the two groups were observed and compared.Result:1 The optimal concentration of adriamycin on cardiomyocytes was screened by concentration gradient design.The results showed that 2?mol / L was the appropriate concentration,and the survival rate of cardiomyocytes decreased to about 75%.2 The survival rate of cardiomyocytes in adriamycin group was significantly lower than that in blank control group(P<0.05).There was no significant difference in the viability of cardiomyocytes between berberine group and blank control group(P> 0.05).Compared with adriamycin group,the survival rate of the cells in both low and middle concentration of berberine+ adriamycin group was increased,but the difference was not statistically significant(P>0.05).The survival rate of cells in high-concentration of berberine + adriamycin group was significantly higher than that of the control group(P <0.05).3 The content of ROS in adriamycin group was significantly higher than that in the blank control group(P <0.05).There was no significant difference in the content of ROS between berberine group and blank control group(P>0.05).Compared with adriamycin group,low and medium concentrations of berberine can reduce the increment degree of ROS induced by adriamycin,but the trend was not obvious(P>0.05),ROS content was decreased significantly in high-concentration of berberine + adriamycin group,and the difference was statistically significant(P <0.05).4 Compared with the blank control group,the DNA damage of cardiomyocytes was significantly decreased(P<0.05).There was no significant DNA damage in berberine group compared with blank control group(P>0.05).Compared with adriamycin group,low-concentration of berberine could improve the DNA damage induced by adriamycin in different degrees,but the difference was not statistically significant(P>0.05).The medium and high concentrations of berberine could decrease the damage,and the difference was significant(P <0.05).5 Compared with the blank control group,the expression of DNA repair enzyme in the berberine group was not significantly different(P> 0.05),while the expression in adriamycin group was significantly decreased(P<0.05).Compared with adriamycin group,the expression of DNA repair enzyme in low concentration of berberine group was increased,but the difference was not statistically significant(P>0.05),the expression of DNA repair enzyme in medium and high concentrations of berberine + adriamycin group increased significantly(P <0.05).Conclusion:1 Adriamycin induced AC 16 cardiomyocytes injury,which was caused by the increased oxidative stress and DNA damage.The DNA damage induced by adriamycin may be associated with decreased expressions of DNA repair enzymes XRCC1 and XRCC3.2 Berberine reduced DNA damage induced by adriamycin in AC16 cardiomyocytes through enhancing the expression of DNA repair enzymes.