| Objective:To evaluate the protective effects of hydroxysafflor yellow A(HSYA)on the injured human coronary artery endothelial cells(HCAECs)induced by High ox-LDL,and related molecular mechanisms.Methods:Four groups of HCAECs were cultured individually: the control group,High ox-LDL group,High ox-LDL+HSYA group,and HSYA group.After 24 hours of injury induction,the survival rate of each group was measured by colorimetric method with MTT,and the optimal dose of High ox-LDL and HSYA in the test conditions were selected.NO and LDH released from HCAECs in each group were detected by nitrate reduction and lactate dehydrogenase colorimetry method,respectively;and cell apoptosis was analyzed by flow cytometry.The expression of LOX-1 and eNOS genes in transcription levels were detected by reverse transcription-polymerase chain reaction(RT-PCR),and Western blot(WB)was employed to detect the expression in protein levels of LOX-1、eNOS and Bcl-2/Bax.Results:Compared with the control group,High ox-LDL(20μg/ml)group behaved lower survival rate(99.39±0.66 vs 67.26±2.68,P<0.05);NO content was decreased,and the expression of eNOS mRNA and protein decreased(P<0.05);conversely,LDH content was increased(P<0.05),either the expression of LOX-1 mRNA and protein levels(P<0.05);the number of apoptotic cells increased significantly(16.28%,P<0.001),the proportion of Bcl-2/Bax protein decreased similarly(P<0.001).Compared with High ox-LDL(20μg/ml)group,High ox-LDL(20μg/ml)+ HSYA(0.2mM)increased the cell viability(67.26±2.68 vs 86.63±3.29,P<0.05),promoted NO release(P<0.05);the mRNA expression of eNOS was restored,and then was upregulated in protein level(P<0.05);besides,suppressed LDH secretion(P<0.05),and the expressions of LOX-1 mRNA and protein were decreased(P<0.05);inhibited the cell apoptosis significantly(12.04%,P<0.001),the Bcl-2/Bax protein expression also increased(P<0.05).Conclusion:Endothelial damage could be induced by the lipid peroxidation,such as addition of High ox-LDL.HSYA displayed a protective effect through up-regulating the expression of eNOS mRNA and protein and then improvement of NO content.In contrast,it could inhibit LDH releasement by down-regulating the expression of LOX-1 mRNA and protein.Treatment of HSYA restored the apoptosis of HCAECs by up-regulating Bcl-2/Bax expression. |
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