摘要
目的研究对羟基苯甲醛对氧糖剥夺/复氧损伤小鼠脑微血管内皮细胞(bEnd.3)的保护作用。方法对羟基苯甲醛(500、250、125、62.5、31.25、15.63μmol/L)给药36 h,三气培养箱培养复制bEnd.3氧糖剥夺/复氧损伤模型,设置正常组、模型组、高剂量组、低剂量组、抑制剂组。相关试剂盒检测乳酸脱氢酶(LDH)、NO、TNF-α、白细胞介素(IL)-6、IL-1β、ATP水平及线粒体关键ATP酶(Na^(+)/K^(+)-ATPase、Mg^(2+)-ATPase、Ca^(2+)-ATPase、Ca^(2+)/Mg^(2+)-ATPase)活性。Western blot法检测bEnd.3细胞磷脂酰肌醇3激酶/蛋白激酶B/内皮型一氧化氮合酶信号通路相关蛋白。结果细胞存活率筛选出高剂量组250μmol/L、低剂量组125μmol/L作为实验条件。与正常组比较,模型组NO、Na^(+)/K^(+)-ATPase、Mg^(2+)-ATPase、Ca^(2+)-ATPase、Ca^(2+)/Mg^(2+)-ATPase活性、ATP水平明显降低,TNF-α、IL-1β、IL-6、LDH水平明显升高,差异有统计学意义(P<0.05)。与模型组比较,高剂量组和低剂量组NO、Ca^(2+)-ATPase活性明显升高,LDH水平明显降低(P<0.05);高剂量组Na^(+)/K^(+)-ATPase、Mg^(2+)-ATPase、Ca^(2+)/Mg^(2+)-ATPase活性、ATP水平明显升高,TNF-α、IL-1β、IL-6水平明显降低,差异有统计学意义(P<0.05)。与正常组比较,模型组磷酸化Akt、磷酸化内皮型一氧化氮合酶表达明显降低,差异有统计学意义(P<0.05)。与高剂量组比较,抑制剂组磷酸化Akt、磷酸化内皮型一氧化氮合酶表达明显降低(0.80±0.05 vs 0.93±0.08,1.23±0.36 vs 1.72±0.10,P<0.05)。结论对羟基苯甲醛能有效保护氧糖剥夺/复氧损伤的bEnd.3,其药理机制与磷脂酰肌醇3激酶/蛋白激酶B/内皮型一氧化氮合酶信号通路有关。
Objective To investigate the protective effect of p-hydroxybenzaldehyde on mouse cerebral microvascular endothelial cells(bEnd.3)with oxygen-glucose deprivation/reoxygenationinduced injury(OGD/R).Methods After 36 h treatment of p-hydroxybenzaldehyde(at a concentration of 500,250,125,62.5,31.25 or 15.63μmol/L),bEnd.3 cells were cultured in a three-gas incubator in the OGD/R model.The bEnd.3 cells were set up in normal group,model group,high dose group and low dose group,and inhibitor group.Related reagent test kits were used to detect the levels of LDH,NO,TNF-α,IL-6,IL-1β,ATP and the activities of mitochondrial key ATPases(Na^(+)/K^(+)-ATPase,Mg^(2+)-ATPase,Ca^(2+)-ATPase,Ca^(2+)/Mg^(2+)-ATPase).Western blotting was employed to measure the expression of the PI3K/Akt/eNOS signaling pathway-related proteins in the bEnd.3 cells.Results According to the results of cell viability,250μmol/L was identified as the treatment dose of the high-dose group,and 125μmol/L as the low-dose group.Compared with the normal group,the levels of NO and ATP and the activities of Na^(+)/K^(+)-ATPase,Mg^(2+)-ATPase,Ca^(2+)-ATPase and Ca^(2+)/Mg^(2+)-ATPase were significantly decreased,and the levels of TNF-α,IL-1β,IL-6 and LDH were obviously lower in the model group(P<0.05).The NO level and Ca^(2+)-ATPase activity were significantly increased,and the level of LDH was markedly decreased in the high-dose group and the low-dose group than the model group(P<0.05).The Ca^(2+)/Mg^(2+)-ATPase activity and ATP level were significantly increased,while the levels of TNF-α,IL-1βand IL-6 were significantly decreased in the high-dose group(P<0.05).The expression levels of p-Akt and p-eNOS were significantly decreased in the model group than the normal group(P<0.05),and the levels were significantly decreased in the inhibitor group than the highdose group(0.80±0.05 vs 0.93±0.08,1.23±0.36 vs 1.72±0.10,P<0.05).Conclusion p-Hydroxybenzaldehyde can effectively protect the bEnd.3 cells against OGD/R injury,and its pharmacological mechanism is re
作者
肖湉
杨丽萍
陈普
段小花
Xiao Tian;Yang Liping;Chen Pu;Duan Xiaohua(Yunnan Provincial Key Laboratory of Dai and Yi Medicines,College of Ethnic Medicine,Yunnan University of Chinese Medicine,Kunming 650500,Yunnan Province,China)
出处
《中华老年心脑血管病杂志》
CAS
北大核心
2022年第11期1201-1205,共5页
Chinese Journal of Geriatric Heart,Brain and Vessel Diseases
基金
国家自然科学基金(81960733)
云南省科技计划项目(2019FB120)。
关键词
脑缺血
内皮细胞
一氧化氮合酶
肿瘤坏死因子Α
磷酸肌醇3-激酶类
蛋白激酶类
brain ischemia
endothelial cells
nitric oxide synthase
tumor necrosis factor-alpha
phosphatidylinositol 3-kinases
protein kinases
