title: Autophagy protects human brain microvascular endothelial cells against methylglyoxal-induced injuries, reproducible in a cerebral ischemic model in diabetic rats.
abstract: Cerebral microvascular endothelial cells (ECs) are crucial for brain vascular repair and maintenance, but their physiological function may be impaired during ischemic stroke and diabetes. Methylglyoxal (MGO), a reactive dicarbonyl produced during glucose metabolism, could exacerbate ischemia-induced EC injury and dysfunction. We investigated the protective effect of autophagy on cultured human brain microvascular endothelial cells (HBMEC) that underwent MGO treatment. A further study was conducted to explore the underlying mechanisms of the protective effect. Autophagic activity was assessed by evaluating protein levels, using western blot. 3-methyladenine (3-MA), bafilomycin A1, ammonium chloride (AC), Beclin 1 siRNA, and chloroquine (CQ) were used to cause autophagy inhibition. Alarmar blue assay and lactate dehydrogenase release assay were used to evaluate cell viability. Streptozotocin was administered to induce type I diabetes in rats and post-permanent middle cerebral artery occlusion was performed to elicit cerebral ischemia. Blood-brain barrier permeability was also assessed. Our study found that MGO reduced HBMEC cell viability in a concentration- and time-dependent manner, and triggered the responsive autophagy activation. Autophagy inhibitors bafilomycin A1, AC, 3-MA, and BECN1 siRNA exacerbated MGO-induced HBMEC injury. FAK phosphorylation inhibitor PF573228 inhibited MGO-triggered autophagy and enhanced lactate dehydrogenase release. Meanwhile, similar autophagy activation in brain vascular ECs was observed during permanent middle cerebral artery occlusion-induced cerebral ischemia in diabetic rats, while chloroquine-induced autophagy inhibition enhanced blood-brain barrier permeability. Taken together, our study indicates that autophagy triggered by MGO defends HBMEC against injuries.
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Fang
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Xue
authors.initials: X
authors.lastname: Li
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Yinbo
authors.initials: Y
authors.lastname: Zhong
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Yu
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Yu
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Haibin
authors.initials: H
authors.lastname: Dai
authors.affiliation: Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
authors.forename: Min
authors.initials: M
authors.lastname: Yan
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D001812:Blood-Brain Barrier / Q000187:drug effects; D002545:Brain Ischemia / Q000139:chemically induced* / Q000188:drug therapy*; D002196:Capillaries / Q000166:cytology* / Q000187:drug effects*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003921:Diabetes Mellitus, Experimental / Q000473:pathology*; D003922:Diabetes Mellitus, Type 1 / Q000139:chemically induced / Q000473:pathology; D003925:Diabetic Angiopathies / Q000473:pathology*; D042783:Endothelial Cells / Q000187:drug effects*; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism; D006801:Humans; D008297:Male; D020768:Middle Cerebral Artery / Q000473:pathology; D011765:Pyruvaldehyde / Q000633:toxicity*; D034622:RNA Interference / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D011765:Pyruvaldehyde; D051417:Focal Adhesion Kinase 1
keywords: FAK phosphorylation; autophagy; diabetes; human brain microvascular endothelial cells; methylglyoxal; stroke