Affiliations: [a] CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal | [b] Department of Life Sciences – Faculty of Sciences and Technology, University of Coimbra, Coimbra, Portugal | [c] Department of Pharmacology, Tulane University School of Medicine, New Orleans, LA, USA | [d] Neuroscience Program, Tulane University School of Medicine, New Orleans, LA, USA | [e] Laboratory of Physiology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal
Correspondence:
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Correspondence to: Paula I. Moreira, Center for Neuroscience and Cell Biology, University of Coimbra and Laboratory of Physiology, Faculty of Medicine, University of Coimbra, 3000-354 Coimbra, Portugal. Tel.: +351 239480012/+351 239820190; Fax: +351 239480034/+351 239822776; E-mails: venta@ci.uc.pt; pimoreira@fmed.uc.pt.
Abstract: We hypothesized that hyperglycemia-induced mitochondrial dysfunction and oxidative stress are closely associated with amyloid-β peptide (Aβ) toxicity in endothelial cells. Brain microvascular endothelial cells from rat (RBMEC) and mice (MBMEC) were isolated from adult Sprague-Dawley rats and homozygous db/db (Leprdb/Leprdb) and heterozygous (Dock7m/Leprdb) mice, and cultured under normo- and hyperglycemic conditions for 7 d followed by 24 h exposure to Aβ1-40. Some experiments were also performed with two mitochondrial superoxide (O2•−) scavengers, MitoTempo and Peg-SOD. Cell viability was measured by the Alamar blue assay and mitochondrial membrane potential (ΔΨm) by confocal microscopy. Mitochondrial O2•− and hydrogen peroxide (H2O2) production was assessed by fluorescence microscopy and H2O2 production was confirmed by microplate reader. Hyperglycemia or Aβ1-40 alone did not affect cell viability in RBMEC. However, the simultaneous presence of high glucose and Aβ1-40 reduced cell viability and ΔΨm, and enhanced mitochondrial O2•− and H2O2 production. MitoTempo and PEG-SOD prevented Aβ1-40 toxicity. Interestingly, MBMEC presented a similar pattern of alterations with db/db cultures presenting higher susceptibility to Aβ1-40. Overall, our results show that high glucose levels increase the susceptibility of brain microvascular endothelial cells to Aβ toxicity supporting the idea that hyperglycemia is a major risk factor for vascular injury associated with AD.
