Protection of blood retinal barrier and systemic vasculature by insulin-like growth factor binding protein-3.
Previously, we showed that insulin growth factor (IGF)-1 binding protein-3 (IGFBP-3), independent of IGF-1, reduces pathological angiogenesis in a mouse model of the oxygen-induced retinopathy (OIR). The current study evaluates novel endothelium-dependent functions of IGFBP-3 including blood retinal barrier (BRB) integrity and vasorelaxation. To evaluate vascular barrier function, either plasmid expressing IGFBP-3 under the regulation of an endothelial-specific promoter or a control plasmid was injected into the vitreous humor of mouse pups (P1) and compared to the non-injected eyes of the same pups undergoing standard OIR protocol. Prior to sacrifice, the mice were given an injection of horseradish peroxidase (HRP). IGFBP-3 plasmid-injected eyes displayed near-normal vessel morphology and enhanced vascular barrier function. Further, in vitro IGFBP-3 protects retinal endothelial cells from VEGF-induced loss of junctional integrity by antagonizing the dissociation of the junctional complexes. To assess the vasodilatory effects of IGFBP-3, rat posterior cerebral arteries were examined in vitro. Intraluminal IGFBP-3 decreased both pressure- and serotonin-induced constrictions by stimulating nitric oxide (NO) release that were blocked by L-NAME or scavenger receptor-B1 neutralizing antibody (SRB1-Ab). Both wild-type and IGF-1-nonbinding mutant IGFBP-3 (IGFBP-3NB) stimulated eNOS activity/NO release to a similar extent in human microvascular endothelial cells (HMVECs). NO release was neither associated with an increase in intracellular calcium nor decreased by Ca(2+)/calmodulin-dependent protein kinase II (CamKII) blockade; however, dephosphorylation of eNOS-Thr(495) was observed. Phosphatidylinositol 3-kinase (PI3K) activity and Akt-Ser(473) phosphorylation were both increased by IGFBP-3 and selectively blocked by the SRB1-Ab or PI3K blocker LY294002. In conclusion, IGFBP-3 mediates protective effects on BRB integrity and mediates robust NO release to stimulate vasorelaxation via activation of SRB1. This response is IGF-1- and calcium-independent, but requires PI3K/Akt activation, suggesting that IGFBP-3 has novel protective effects on retinal and systemic vasculature and may be a therapeutic candidate for ocular complications such as diabetic retinopathy.
Department of Pharmacology and Therapeutics, College of Medicine, University of Florida, Gainesville, Florida, United States of America.
Yagna P R
YP
Jarajapu
Jun
J
Cai
Yuanqing
Y
Yan
Sergio
S
Li Calzi
Jennifer L
JL
Kielczewski
Ping
P
Hu
Lynn C
LC
Shaw
Sue M
SM
Firth
Tailoi
T
Chan-Ling
Michael E
ME
Boulton
Robert C
RC
Baxter
Maria B
MB
Grant
D000818:Animals; D001813:Blood-Retinal Barrier / Q000378:metabolism* / Q000473:pathology; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002536:Cerebral Arteries / Q000187:drug effects / Q000378:metabolism; D062507:Claudin-5 / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D016257:Fura-2; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000235:genetics / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D007365:Intercellular Junctions / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D009389:Neovascularization, Pathologic / Q000235:genetics / Q000378:metabolism; D009569:Nitric Oxide; D052250:Nitric Oxide Synthase Type III / Q000037:antagonists & inhibitors / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011485:Protein Binding; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012164:Retinal Diseases / Q000235:genetics / Q000378:metabolism; D012171:Retinal Vessels / Q000187:drug effects / Q000378:metabolism; D051122:Scavenger Receptors, Class B / Q000378:metabolism; D015398:Signal Transduction; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D015820:Cadherins; D062507:Claudin-5; D018972:Insulin-Like Growth Factor Binding Protein 3; D051122:Scavenger Receptors, Class B; D042461:Vascular Endothelial Growth Factor A; D009569:Nitric Oxide; D007334:Insulin-Like Growth Factor I; D052250:Nitric Oxide Synthase Type III; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D002118:Calcium; D016257:Fura-2