Wang S, Lü H, Wang L, Tian M, Gao J, Liu Z, Wang J, Yu Y
Department of Physiology, Bengbu Medical College, Bengbu 233000, China.
Key Laboratory of Basic and Clinical Cardiovascular Diseases, Bengbu Medical College, Bengbu 233000, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2022 Dec 20;42(12):1882-1888. doi: 10.12122/j.issn.1673-4254.2022.12.18.
To investigate the effect of aldehyde dehydrogenase 2 (ALDH2) on lipopolysaccharide (LPS)- induced damage of mouse brain microvascular endothelial barrier and explore the role of mitochondrial fusion and fission in maintaining the integrity of endothelial barrier.
Mouse brain microvascular endothelial cells were treated with 1 μg/ mL LPS for 24 h with or without pretreatment with 20 μmol/mL Alda-1 (a ALDH2 agonist) for 1 h. The changes in cell viability were assessed using cell counting Kit-8 (CCK8) assay, and the cell permeability was evaluated using transendothelial cell resistance (TEER) and FITC-Dextran assay. The level of oxidative stress in the cells was assessed by detecting the levels of malondialdehyde (MDA) and superoxide dismutase (SOD), and the content of reactive oxygen species (ROS) was detected using a superoxide anion fluorescent probe (DHE). Western blotting was performed to detect the expressions of ALDH2, tight junction proteins ZO-1 and occludin, and mitochondrial fusion- and division-related proteins Mfn2, OPA1, Drp1 and Fis1.
Compared with the untreated cells, the cells treated with LPS showed significantly decreased TEER, increased FITC-dextran leakage, MDA content and ROS production, decreased SOD activity expressions of ALDH2, ZO-1, occludin, Mfn2 and OPA1, and increased expressions of Drp1 and Fis1 ( < 0.05). Pretreatment with Alda-1 prior to LPS exposure strongly suppressed the increase of endothelial cell membrane permeability, reduced ROS production, increased the expressions of ALDH2, ZO-1, occludin, OPA1 and Mfn2, and lowered the expressions of Drp1 and Fis1 ( < 0.05).
ALDH2 can alleviate LPS-induced damage of brain microvascular endothelial cell barrier by inhibiting the mitochondrial ROS production and promoting mitochondrial fusion and inhibiting mitochondrial fission.
探讨乙醛脱氢酶2（ALDH2）对脂多糖（LPS）诱导的小鼠脑微血管内皮屏障损伤的影响，并探讨线粒体融合与分裂在维持内皮屏障完整性中的作用。
小鼠脑微血管内皮细胞用1μg/mL LPS处理24小时，其中一组在处理前1小时用20μmol/mL Alda-1（一种ALDH2激动剂）预处理。使用细胞计数试剂盒-8（CCK8）检测细胞活力变化，使用跨内皮电阻（TEER）和FITC-葡聚糖检测评估细胞通透性。通过检测丙二醛（MDA）和超氧化物歧化酶（SOD）水平评估细胞内氧化应激水平，使用超氧阴离子荧光探针（DHE）检测活性氧（ROS）含量。采用蛋白质免疫印迹法检测ALDH2、紧密连接蛋白ZO-1和闭合蛋白以及线粒体融合和分裂相关蛋白Mfn2、OPA1、Drp1和Fis1的表达。
与未处理的细胞相比，LPS处理的细胞TEER显著降低，FITC-葡聚糖渗漏增加，MDA含量和ROS生成增加，SOD活性降低，ALDH2、ZO-1、闭合蛋白、Mfn2和OPA1表达降低，Drp1和Fis1表达增加（P<0.05）。在LPS暴露前用Alda-1预处理可强烈抑制内皮细胞膜通透性增加，减少ROS生成，增加ALDH2、ZO-1、闭合蛋白、OPA1和Mfn2的表达，降低Drp1和Fis1的表达（P<0.05）。
ALDH2可通过抑制线粒体ROS生成、促进线粒体融合和抑制线粒体分裂来减轻LPS诱导的脑微血管内皮细胞屏障损伤。
