丹参酮ⅡA磺酸钠不同给药时段对其内皮保护作用的影响

Influence of Different Administration Timepoints on the Endothelial Protective Effects of Sodium TanshinoneⅡA Sulfonate

目的研究不同给药时段对丹参酮ⅡA磺酸钠(sodium tanshinoneⅡA sulfonate,STS)内皮保护作用的影响。方法将人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVEC)分组刺激,分为5组:DMEM组、脂多糖(lipopolysaccharide,LPS)处理组、STS预处理+LPS组、STS+LPS共处理组、STS后处理+LPS组。其中,LPS终浓度为1μg·mL^(-1),STS终浓度为25μg·mL^(-1),LPS刺激时间为12 h。处理完成后,CCK-8法测定HUVEC活力;ELISA检测细胞培养上清中炎症因子白细胞介素-1β(interleukin-1β,IL^(-1)β)的蛋白浓度;Westernblotting检测细胞裂解液中核因子κB(nuclear factorκB,NF-κB)的蛋白水平;划痕试验检测HUVEC的迁移能力;光镜观察细胞凋亡状态,并用Western blotting检测凋亡相关分子cleaved caspase-3和cleaved caspase-9的蛋白水平。结果与DMEM组相比,LPS处理后HUVEC增殖活力减低;IL^(-1)β和NF-κB蛋白水平增高;细胞迁移能力降低、细胞凋亡增加并伴有cleaved caspase-3和cleaved caspase-9蛋白水平的升高(P<0.05);STS处理能够部分逆转LPS引起的上述现象,其中以STS预处理的效果最为明显,而STS后处理的效果相对较弱。结论STS预处理、共处理和后处理均能部分抑制LPS诱导的HUVEC功能异常和凋亡,且以预处理时的效果最为明显。

OBJECTIVE To investigate the influence of different administration timepoints on the endothelial protective effects of sodium tanshinoneⅡA sulfonate(STS).METHODS Human umbilical vein endothelial cells(HUVEC)were divided into five groups,individually stimulated,including DMEM group,lipopolysaccharide(LPS)group,STS pre-treatment+LPS group,STS co-treatment+LPS group,STS post-treatment+LPS group.The final concentration of LPS in the experiment was 1μg·mL^(-1),and that of STS was 25μg·mL^(-1),LPS stimulation time was 12 h.After that,HUVEC viability was determined by CCK-8 assay.The protein concentration of inflammatory cytokine interleukin-1β(IL^(-1)β)in HUVEC culture supernatant was detected by ELISA.The protein levels of nuclear factorκB(NF-κB)in HUVEC lysate were detected by Western blotting.The migration capacity of HUVEC was detected by wound-healing test.HUVEC apoptosis was observed under an optical microscopy,and the expression of apoptosis-related proteins including cleaved caspase-3 and cleaved caspase-9 were detected by Western blotting.RESULTS Compared with DMEM group,after LPS treatment,HUVEC cell viability was decreased;IL^(-1)βand NF-κB protein expression increased;migration capacity impaired;increased apoptosis with the higher protein expression of cleaved caspase-3 and cleaved caspase-9(P<0.05).STS treatments could partially reverse these LPS-induced phenomena,among which the STS pre-treatment was most effective,while the STS post-treatment was with relative lower efficacy.CONCLUSION STS pre-treatment,co-treatment and post-treatment can partially reverse LPS-induced HUVEC dysfunction and apoptosis,among which the STS pre-treatment is most effective.