非瑟酮改善高糖高脂诱导的小鼠脑微血管内皮细胞损伤作用与机制研究

Improving effect and mechanism of Fisetin on on brain microvascular endothelial cell by high glucose and high fat in mice

目的探讨非瑟酮(Fis)对高糖高脂诱导的小鼠脑微血管内皮细胞(BEND3)损伤的改善作用及其机制。方法高糖高脂诱导BEND3损伤,建立体外糖尿病BEND3损伤模型,将细胞分为正常对照组(Con,低糖DMEM完全培养基)、25 mmol/L葡萄糖(HG)+200μmol/L棕榈酸(PA)模型组(HG+PA)、HG+PA+1μmol/L Fis组(HG+PA+1Fis)、HG+PA+5μmol/L Fis组(HG+PA+5Fis)、HG+PA+10μmol/L Fis组(HG+PA+10Fis)、HG+PA+20μmol/L Fis组(HG+PA+20Fis)。CCK-8检测BEND3细胞存活率,筛选最佳Fis浓度;乳酸脱氢酶(LDH)检测细胞毒性;ELISA法检测细胞活性氧(ROS)、一氧化氮(NO)、诱导型一氧化氮合酶(iNOS)、VEGF、基质金属蛋白酶2(MMP-2)、基质金属蛋白酶9(MMP-9)、基质金属蛋白酶抑制剂1(TIMP-1)含量。结果与Con组比较,HG+PA组细胞存活率降低,LDH含量升高(P<0.05);与HG+PA组比较,HG+PA+20Fis组细胞存活率升高,LDH含量降低(P<0.05)。与Con组比较,HG+PA组ROS、NO、iNOS、VEGF、MMP-2、MMP-9、TIMP-1含量升高(P<0.05);与HG+PA组比较,HG+PA+20Fis组ROS、NO、iNOS、VEGF、MMP-2、MMP-9、TIMP-1的表达降低(P<0.05)。结论Fis可改善糖尿病BEND3损伤,可能与抑制细胞内源性ROS及相关氧化因子的产生有关。

ObjectiveTo investigate the ameliorative effect of Fisetin(Fis)on brain microvascular endothelial cell(BEND3)injury induced by high glucose(HG)and high fat in mice and its mechanism.MethodsBEND3 injury was induced by high glucose and high fat. Diabetic brain microvascular cell injury model in vitro was established. Cells were divided into normal control group(Con,low glucose DMEM complete medium),model group(HG+PA,25 mmol/L glucose +200 μmol/L palmitic acid),HG+PA+1μmol/L Fis group(HG+PA+1 Fis),HG+PA+5 μmol/L Fis group(HG+PA+5 Fis),HG+PA+10μmol/L Fis group(HG+PA+10 Fis)and HG+PA+20 μmol/L Fis group(HG+PA+20 Fis). CCK-8 was used to detect the survival rate of BEND3 cells and to select the appropriate concentration for administration. Lactate dehydrogenase was used to detect cytotoxicity. The contents of ROS,NO,iNOS,VEGF,MMP-2,MMP-9,TIMP-1 were detected by ELISA.Results group had lower cell survival rate and higher LDH content(P<0. 05). Compared with HG+PA group,HG+PA+20 Fis group had higher cell activity and lower LDH content(P<0. 05). Compared with Con group,the contents of ROS,NO,i NOS,VEGF,MMP-2,MMP-9,TIMP-1 in HG+PA group increased(P<0. 05). Compared with HG+PA group,the expression of each detection factor in HG+PA+20 Fis group decreased(P<0. 05).ConclusionFisetin can improve the damage of diabetic BEND3,which may be related to the inhibition of the production of endogenous ROS and oxidation factors.