title: Aflibercept, bevacizumab and ranibizumab prevent glucose-induced damage in human retinal pericytes in vitro, through a PLA2/COX-2/VEGF-A pathway.
abstract: Diabetic retinopathy, a major cause of vision loss, is currently treated with anti-VEGF agents. Here we tested two hypotheses: (i) high glucose damages retinal pericytes, the cell layer surrounding endothelial cells, via VEGF induction, which may be counteracted by anti-VEGFs and (ii) activation of PLA2/COX-2 pathway by high glucose might be upstream and/or downstream of VEGF in perycites, as previously observed in endothelial cells. Human retinal pericytes were treated with high glucose (25mM) for 48h and/or anti-VEGFs (40μg/ml aflibercept, 25μg/ml bevacizumab, 10μg/ml ranibizumab). All anti-VEGFs significantly prevented high glucose-induced cell damage (assessed by LDH release) and improved cell viability (assessed by MTT and Evans blue). High glucose-induced VEGF-A expression, as detected both at mRNA (qPCR) and protein (ELISA) level, while receptor (VEGFR1 and VEGFR2) expression, detected in control condition, was unaffected by treatments. High glucose induced also activation of PLA2/COX-2 pathway, as revealed by increased phosphorylation of cPLA2, COX-2 expression and PGE2 release. Treatment with cPLA2 (50μM AACOCF3) and COX-2 (5μM NS-392) inhibitors prevented both cell damage and VEGF-A induced by high glucose. Finally, challenge with exogenous VEGF-A (10ng/ml) induced VEGF-A expression, while anti-VEGFs reduced VEGF-A expression induced by either high glucose or exogenous VEGF-A. These data indicate that high glucose directly damages pericytes through activation of PLA2/COX-2/VEGF-A pathway. Furthermore, a kind of feed-forward loop between cPLA2/COX-2/PG axis and VEGF appears to operate in this system. Thus, anti-VEGFs afford protection of pericytes from high glucose by inhibiting this loop.
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Giurdanella
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Carmelina Daniela
authors.initials: CD
authors.lastname: Anfuso
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Melania
authors.initials: M
authors.lastname: Olivieri
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Lupo
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Nunzia
authors.initials: N
authors.lastname: Caporarello
authors.affiliation: Institut de laVision, UMRS_968 Inserm/Université Pierre et Marie Curie, Equipe 14, Paris, France; Department of Surgical Sciences, Eye Clinic, University of Torino, Torino, Italy.
authors.forename: Chiara M
authors.initials: CM
authors.lastname: Eandi
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Filippo
authors.initials: F
authors.lastname: Drago
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Bucolo
authors.affiliation: Department of Biomedical and Biotechnological Sciences, Catania University, Viale Andrea Doria 6, 95125 Catania, Italy. Electronic address: salomone@unict.it.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Salomone
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology*; D001095:Arachidonic Acids / Q000494:pharmacology; D000068258:Bevacizumab; D002452:Cell Count; D002470:Cell Survival / Q000187:drug effects; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D025461:Feedback, Physiological; D005786:Gene Expression Regulation; D005947:Glucose / Q000037:antagonists & inhibitors* / Q000633:toxicity; D006801:Humans; D020286:Pericytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D054467:Phospholipases A2 / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D061251:Primary Cell Culture; D000069579:Ranibizumab; D040262:Receptors, Vascular Endothelial Growth Factor / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology*; D012160:Retina / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism
chemical_list: D020533:Angiogenesis Inhibitors; D061067:Antibodies, Monoclonal, Humanized; D001095:Arachidonic Acids; D004791:Enzyme Inhibitors; D011993:Recombinant Fusion Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C081565:arachidonyltrifluoromethane; C533178:aflibercept; D000068258:Bevacizumab; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; C000588866:KDR protein, human; D040262:Receptors, Vascular Endothelial Growth Factor; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D054467:Phospholipases A2; D005947:Glucose; D000069579:Ranibizumab
keywords: Aflibercept; Bevacizumab; Diabetic retinopathy; Ranibizumab; VEGF