Abstract
OBJECTIVES: To evaluate the effect of resveratrol (RES) on barrier function of mouse brain microvascular endothelial cell monolayers exposed to oxygen/glucose deprivation/reoxygenation (OGD/R) and PM(2.5) and explore the role of mitochondrial fission and fusion in protecting endothelial barrier function. METHODS: Cultured mouse brain microvascular endothelial cells were exposed to OGD/R, treated with PM(2.5) (100 μg/mL) before OGD/R, or pretreated with RES (40 mg/mL) prior to OGD/R+PM(2.5) exposures. The changes in cell viability were examined with CCK-8 assay, and cell permeability was assessed by measuring transendothelial electrical resistance (TEER) and FITC-dextran permeation. Malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured, and intracellular and mitochondrial ROS levels were detected using fluorescent probes. Mitochondrial morphology in the treated cells was observed using Mito-Tracker Red CMXRos. Western blotting was performed to detect the changes in cellular expressions of the tight junction proteins (ZO-1, occludin, and claudin-5) and mitochondrial dynamics-associated proteins (Drp1, Fis1, Mfn2, and OPA1). RESULTS: Compared with the normal control cells, the cells exposed to OGD/R or both OGD/R and PM(2.5) showed significantly decreased TEER levels, increased permeability, elevated oxidative stress, and increased ROS fluorescence intensities. Obvious mitochondrial fragmentation and morphological changes in the mitochondria were observed in the exposed cells, which also showed decreased expressions of tight junction proteins and mitochondrial fusion proteins with increased expressions of mitochondrial fission proteins. RES pretreatment of the endothelial cells before the exposures significantly reduced membrane permeability, lowered ROS levels, improved mitochondrial morphology, increased expressions of tight junction and fusion proteins, and decreased fission protein expressions. CONCLUSIONS: RES can protect barrier function of mouse brain microvascular endothelial cell monolayers exposed to OGD/R and PM(2.5) by modulating mitochondrial dynamics, potentially through promoting mitochondrial fusion and inhibiting mitochondrial fission.