目的：观察外源性硫化氢(H2S)对高糖诱导人脐静脉内皮细胞(HUVECs)氧化应激损伤的保护作用及其可能机制。方法用25 mmol/L D-葡萄糖(高糖)、外源性H2S供体硫化氢钠(NaHS)(5×10-5、1×10-4、5×10-4、1×10-3和5×10-3 mol/L)及沉默信息调节子1(SIRT1)特异性抑制剂尼克酰胺(40 mmol/L)处理HUVECs细胞株24 h。实验分为对照组、高糖组、甘露醇组、NaHS(5×10-5、1×10-4、5×10-4、1×10-3、5×10-3 mol/L)组、高糖+NaHS(5×10-5、1×10-4、5×10-4、1×10-3和5×10-3 mol/L)组和高糖+NaHS(10-3 mol/L)+尼克酰胺组。采用MTT比色法检测细胞活力，流式细胞术检测细胞内的内活性氧(ROS)水平，Western bolt法检测SIRT1的表达。结果与对照组比较，高糖组细胞活力显著降低[OD值分别为(0.76±0.09)和(0.18±0.01)，t=5.34，P<0.05]，细胞内ROS水平显著增加[ROS水平分别为(356.18±42.96)au和(1183.63±84.31)au，t=6.72，P<0.05]。与高糖组比较，高糖+NaHS(5×10-4、1×10-3和5×10-3 mol/L)组细胞活力显著增加[OD值分别为(0.18±0.01)、(0.39±0.05)、(0.68±0.04)和(0.51±0.08)，t1=3.16，t2=3.95，t3=3.86，均P<0.05]，细胞内 ROS水平显著降低[ROS水平分别为(1183.63±84.31)、(874.32±85.36)、(628.65±54.27)和(439.56±53.64)au，t1=3.46，t2=3.97，t3=5.13，均P<0.05]。与对照组比较，高糖组细胞中SIRT1蛋白表达显著下调[蛋白相对表达量分别为(0.48±0.04)和(0.17±0.03)，t=3.94，P<0.05]。与高糖组比较，高糖+NaHS(10-3 mol/L)组细胞中SIRT1表达显著上调[蛋白相对表达量分别为(0.17±0.03)和(0.59±0.08)，t=4.36，P<0.05]。 SIRT1抑制剂尼克酰胺取消了NaHS抑制高糖诱导的HUVECs细胞活力的降低[高糖+NaHS组和高糖+NaHS+尼克酰胺组OD值分别为(0.52±0.04)和(0.23±0.03)，t =2.98，P <0.05]和细胞内ROS水平的增加[高糖+NaHS组和高糖+NaHS+尼克酰胺组ROS水平分别为(628.65±54.27)au和(1052.84±113.42)au，t=3.76，P<0.05]。结论外源性H2S抑制了高糖诱导的HUVECs氧化应激损伤，其机制可能与H2S上调SIRT1的表达有关。%Objective To investigate the protective effect of extrogenous hydrogen sulfide (H2S) on the injury of oxida-tive stress induced by high glucose and explore the possible mechanism in human umbilical vein endothelial cells (HUVECs). Methods HUVECs were incubated by 25 mmol/L D-glucose (high glucose) to induce injury and cells were treated with H2S donor sodium bisulfide (NaHS) (5í10-5, 1í10-4, 5í10-4, 1í10-3 and 5í10-3 mol/L) for 24 h. The in-hibitor of silent information regulator 1 (SIRT1) niacinamide was used in the study. The cells were divided into the control group, high glucose group, mannitol group, NaHS (5í10-5, 1í10-4, 5í10-4, 1í10-3, 5í10-3 mol/L)groups, high glucose +NaHS (5í10-5, 1í10-4, 5í10-4, 1í10-3 and 5í10-3 mol/L) groups and high glucose + NaHS (10-3 mol/L) + niacinamide group. MTT assay was used to detect the cell viability. The level of reactive oxygen species (ROS) was measured by flow cytometry. The expression of SIRT1 was tested by Western blot. Results Compared with the control group, the cell viability was significantly decreased [OD were (0.76±0.09) and (0.18±0.01) respectively, t = 5.34, P < 0.05], and the level of ROS was significantly increased [the levels of ROS were (356.18±42.96) au and (1183.63±84.31) au respective-ly, t = 6.72, P< 0.05] in high glucose group. Compared with high glucose group, the cells viability were signifi-cantly increased [OD were (0.18±0.01), (0.39±0.05), (0.68±0.04) and (0.51±0.08) respectively, t1= 3.16, t2= 3.95, t3=3.86, P< 0.05], and the levels of ROS were significantly decreased [the levels of ROS were (1183.63±84.31), (874.32±85.36), (628.65±54.27) and (439.56±53.64) au respectively, t1=3.46, t2=3.97, t3= 5.13, all P< 0.05] in high glucose+NaHS (5í10-4, 10-3 and 5í10-3 mol/L) groups. Compared with the control, the expression of SIRT1 was significantly down-regulated in high glucose group [relative expression levels were (0.48±0.04) and (0.17±0.03) respectively, t =3.94, P < 0.05]. Compared with high glucose group, the expression of SIRT1 was significantly up-regulated in high glucose+NaHS (10-3 mol/L) group [relative expression levels were (0.17±0.03) and (0.59±0.08) respectively, t= 4.36, P<0.05]. Compared with the high glucose+NaHS (10-3 mol/L) group, the cell viability was significantly decreased [OD were (0.52±0.04) and (0.23±0.03) respectively, t=2.98, P<0.05) and the level of ROS was significantly increased [the level of ROS was (628.65±54.27) au and (1052.84±113.42) au, t = 3.76, P< 0.05) in high glucose+NaHS (10-3 mol/L)+niaci-namide group. Conclusion Extrogenous H2S prevents the injury of oxidative stress induced by high glucose in HUVECs, the mechanisms may be related to the up-regulation of SIRT1 induced by H2S.
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