Effects Of Ursolic Acid On Proliferation, Apoptosis Of SK-BR-3 Cells And Its Mechanism

Objective:The present study was designed to investigate the anti-tumor effect and apoptosis -inducing mechanism of ursolic acid(UA) on human breast cancer SK-BR-3 cells,and to dedect the expression of apoptosis-related proteins in the tissue of breast cancer,and to provide a reliable experimental basis for clinical applicaton of UA.Methods:The effect of UA on the proliferation of SK-BR-3 cells was measured by MTT method;Light microscope,Hoechst 33258 fluorescent dying,electron microscope were used to observe UA induced morphological changes;The biochemical characteristics of apoptosis was detected by DNA agarose gel electrophoresis;The cell cycle and apoptosis rate were detected with flow cytometry.The expressions of COX-2,Bcl-2 and Bax protein in estrogen receptor-negative and human epidermal growth-factor 2 positive human breast cancer were determined by immunohistochemistry.The expressions of COX-2,Bcl-2 and Bax mRNA in SK-BR-3 cells were determined by RT-PCR.Results:UA inhibitied the growth of SK-BR-3 cells in a dose-and time-dependent manner,the IC₅₀ value of UA at 24 h,48 h and 72 h were＞60μmol/l,31.54μmol/l and 24.16μmol/l,respectively;The morphological changes of apoptosis,such as nuclear shrinkage or condensed chromatin were identified after incubation by UA;Characteristic apoptotic ladder was observed by DNA agarose gel electrophoresis;Cell cycle analysis revealed that SK-BR-3 cells treated by 10-30μmol/L UA for 48 h were blocked predominantly in G₀/G₁ phase(P＜0.05);The expressions of COX-2 and Bcl-2 mRNA of SK-BR-3 cells treated by UA for 48 h were decreased(P＜0.05),Bcl-2/Bax was reduced(P＜0.05),whereas the expression of Bax mRNA was unchanged;The positive rate of COX-2,Bcl-2,Bax protein in estrogen receptor-negative and human epidermal growth-factor 2 positive breast cancer were 86.67%(26/30),63.33%(19/30) and56.67%(17/30),respectively.The expression of COX-2 and Bax was negatively correlated(r=-0.37864,P＜0.05),whereas the expression of COX-2 and Bcl-2 was no significant correlation.Conclusion- UA had the effects of inhibition to SK-BR-3 cells proliferation and induction of cells apoptosis in a dose-and time-dependent manner;The role was related to arresting cell cycle,inhibiting COX-2 and Bcl-2 mRNA expression and reducing the ratio of Bcl-2/Bax;There was a higher positive expression of COX-2 in estrogen receptor-negative and human epidermal growth-factor 2 positive human breast cancer;The expression of COX-2 and Bax was negatively correlated,whereas the expression of COX-2 and Bcl-2 was no significant correlation.