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目的:探讨银杏内酯B(GB)减轻缺氧/复氧(H/R)所致血管内皮细胞损伤的作用和机制。方法:将体外培养的人脐静脉内皮细胞分为5个组,即空白对照组(不加任何处理)、GB组(100μmol/L)、H/R组(缺氧2 h/复氧24 h)、GB+H/R组及GB+H/R+N-硝基-L-精氨酸甲酯(L-NAME,NOS抑制剂)组。用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)比色法测定内皮细胞的存活率(%)。用乳酸脱氢酶(LDH)测试盒和一氧化氮(NO)测试盒分别检测细胞培养上清液中LDH和NO的含量。用Western blot法检测培养的人脐静脉内皮细胞中内皮型一氧化氮合成酶(eNOS)表达的水平。结果:MTT比色法的结果显示,H/R可显著降低内皮细胞的存活率(%),增加LDH的含量,降低NO的生成量(与空白对照组比较,P<0.01);而GB则可逆转上述效应,但GB的作用又可被L-NAME所抑制(与GB+H/R组比较,P<0.01)。GB可以显著提高H/R作用后血管内皮细胞中eNOS表达的水平(与H/R组比较,P<0.01),但是该效应不被L-NAME阻断。结论:GB可显著减轻H/R对内皮细胞的损伤,其作用是通过eNOS-NO途径实现的。
Objective: To investigate the effect and mechanism of ginkgolide B (GB) on reducing vascular endothelial cell injury induced by hypoxia / reoxygenation (H / R). Methods: The human umbilical vein endothelial cells cultured in vitro were divided into five groups: blank control group (without any treatment), GB group (100μmol / L), H / R group ), GB + H / R group and GB + H / R + N-nitro-L-arginine methyl ester (L-NAME) Endothelial cell viability (%) was determined by MTT colorimetry using 3- (4,5-dimethylthiazol-2) -2,5-diphenyltetrazolium bromide salt. The content of LDH and NO in the cell culture supernatant was detected by lactate dehydrogenase (LDH) test kit and nitric oxide (NO) test kit respectively. Western blot was used to detect the expression of endothelial nitric oxide synthase (eNOS) in cultured human umbilical vein endothelial cells. Results: MTT assay showed that H / R could significantly decrease the survival rate of endothelial cells (%), increase the content of LDH and decrease the production of NO (P <0.01 compared with the blank control group) The above effect can be reversed, but the effect of GB can be inhibited by L-NAME (P <0.01 compared with GB + H / R group). GB significantly increased the expression of eNOS in vascular endothelial cells after H / R treatment (P <0.01 compared with H / R group), but this effect was not blocked by L-NAME. Conclusion: GB can significantly reduce the H / R injury of endothelial cells, its role is through the eNOS-NO pathway.