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目的:研究丹酚酸B镁盐对缺氧复氧引起的内皮细胞内钙升高和一氧化氮释放增加的影响.方法:培养的人脐静脉内皮细胞(ECV304)暴露在95％ N_2 +5％CO_2条件下缺氧30分钟,后在含5％CO_2的空气中复氧30分钟.内皮细胞的损伤用染料排除实验、SOD的活性和MDA的生成来评价.胞内游离钙浓度用钙荧光探针Fura 2-AM测定.一氧化氮含量用一氧化氮试剂盒测定.内皮型一氧化氮合酶(eNOS)mRNA和诱导型一氧化氮合酶(iNOS)mRNA的表达用半定量逆转录聚合酶链式反应(RT-PCR)检测.结果:缺氧复氧引起内皮细胞的活力由正常条件下(93.1±1.2)％降至(88±3)％(P<0.01),SOD的活性也由(0.24±0.07)kNU/L下降到(0.18±0.03)kNU/L(P>0.05),但其使内皮细胞MDA的生成由(1.12±0.06)mmol/L增至(3.78±0.03)mmol/L(P<0.01).丹酚酸B镁盐2.5,5,10 mg/L能明显降低缺氧复氧引起的内皮细胞MDA生成量的增加,并且显著提高细胞活力和SOD的活性.同时,缺氧复氧还增加内皮细胞的胞内游离钙浓度(F_(340)/F_(380)由 1.65±0.16增至 1.89±0.28)和一氧化氮释放[由(7.5±1.3)μmol/L增至(16±5)μmol/L],并上调其eNOS mRNA的表达,但降低iNOS mRNA的表达(P<0.05).但在无钙的条件下,缺氧复氧对内皮细胞的胞内游离钙浓度、一氧化氮含量、eNOSnRNA和iNOS mRNA表达
AIM: To investigate the effect of salvianolic acid B magnesium salt on the increase of intracellular calcium and nitric oxide release induced by hypoxia-reoxygenation.Methods: Cultured human umbilical vein endothelial cells (ECV304) exposed to 95% N 2 +5 % CO_2 hypoxia for 30 minutes, followed by reoxygenation in air containing 5% CO 2 for 30 minutes.Endothelial cell injury was evaluated by dye exclusion assay, SOD activity and MDA production.The intracellular free calcium concentration was measured by calcium fluorescence Probe Fura 2-AM Assay Nitric oxide was measured by nitric oxide kit Expression of eNOS mRNA and iNOS mRNA was detected by semi-quantitative reverse transcription The results showed that the activity of endothelial cells decreased from (93.1 ± 1.2)% to (88 ± 3)% (P <0.01) under hypoxia and reoxygenation, and the activity of SOD (0.18 +/- 0.03) kNU / L (0.24 +/- 0.07) kNU / L (P> 0.05). However, the production of MDA in endothelial cells increased from (1.12 ± 0.06) mmol / L to (3.78 ± 0.03) mmol / L (P <0.01) .2.5, 5 and 10 mg / L of salvianolic acid B magnesium salt can significantly reduce the increase of endothelial cell MDA production induced by anoxia and reoxygenation, and significantly increase the cell viability and the activity of SOD. Simultaneously, Oxygen reoxygenation also increased the intracellular free calcium concentration (increased from 1.65 ± 0.16 to 1.89 ± 0.28 for F_ (340) / F_ (380)) and nitric oxide release [from 7.5 ± 1.3 μmol / L (16 ± 5) μmol / L], and upregulated the expression of eNOS mRNA but decreased the expression of iNOS mRNA (P <0.05). However, in the absence of calcium, the intracellular free calcium Concentration, nitric oxide content, eNOSnRNA and iNOS mRNA expression