title: Erythropoietin protects retinal pigment epithelial cells against the increase of permeability induced by diabetic conditions: essential role of JAK2/ PI3K signaling.
abstract: The outer blood-retinal barrier is formed by retinal pigment epithelial (RPE) cells and its disruption significantly contributes to the development of diabetic macular edema (DME). The aim of the study was to explore whether erythropoietin (Epo) has beneficial effects on the barrier function of human RPE cells and the main downstream pathways involved. ARPE-19 cells were cultured in standard conditions and under conditions leading to the disruption of the monolayer [25 mmol/L D-glucose plus IL-1β (10 ng/mL)]. Epo (200 mU/mL/day) was added during the last 2 days of the experiment. The experiments were repeated in the presence of an Epo neutralizing antibody and specific inhibitors of JAK2 and PI3K (AG490 and LY294002, respectively). Permeability was evaluated by fluorescein isothiocyanate dextran (70 kDa) movements. Distribution of tight junction proteins was examined by immunofluorescence. Changes in cytosolic Ca(2+) induced by Epo were also measured. Epo treatment was able to prevent but not to restore the increase of permeability induced by high glucose plus IL-1β. The protective effect of Epo on RPE barrier function was completely blocked by AG490 and almost completely abolished by LY294002. In addition, Epo was able to increase cytosolic Ca(2+) with dependence on extracellular calcium influx and this effect was blocked by either JAK2 or PI3K inhibition. We conclude that RPE disruption induced by high glucose plus IL-1β is prevented by Epo through the downstream signaling of JAK2 and PI3K/AKT pathways.
authors.affiliation: Diabetes and Metabolism Research Unit, Vall d'Hebron Research Institute, Universitat Autònoma de Barcelona, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Garcia-Ramírez
authors.forename: Cristina
authors.initials: C
authors.lastname: Hernández
authors.forename: Marisol
authors.initials: M
authors.lastname: Ruiz-Meana
authors.forename: Marta
authors.initials: M
authors.lastname: Villarroel
authors.forename: Lidia
authors.initials: L
authors.lastname: Corraliza
authors.forename: David
authors.initials: D
authors.lastname: García-Dorado
authors.forename: Rafael
authors.initials: R
authors.lastname: Simó
mesh_terms: D002118:Calcium / Q000378:metabolism; D002452:Cell Count / Q000379:methods; D045744:Cell Line, Tumor; D002867:Chromones / Q000494:pharmacology; D003911:Dextrans / Q000378:metabolism; D003930:Diabetic Retinopathy / Q000188:drug therapy; D004921:Erythropoietin / Q000494:pharmacology* / Q000627:therapeutic use; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D053583:Interleukin-1beta / Q000494:pharmacology; D053614:Janus Kinase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D009025:Morpholines / Q000494:pharmacology; D010539:Permeability; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D020011:Protective Agents / Q000494:pharmacology; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction; D020032:Tyrphostins / Q000494:pharmacology
chemical_list: D002118:Calcium; D002867:Chromones; D003911:Dextrans; D004921:Erythropoietin; D016650:Fluorescein-5-isothiocyanate; D005947:Glucose; D053583:Interleukin-1beta; D053614:Janus Kinase 2; D008565:Membrane Proteins; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D020011:Protective Agents; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; C015219:fluorescein isothiocyanate dextran; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C507924:JAK2 protein, human