IGFBP-3 Attenuates Vascular Injury by Reducing Activation of Acid Sphingomyelinase and Retinal Vascular Permeability

Abstract

Purpose: : Insulin-like growth factor binding protein-3 (IGFBP-3) has vascular protective effects. However, the molecular mechanisms which mediate this effect have yet to be elucidated. Sphingomyelin is a main lipid component of the plasma membrane. Lysosomal acid hydrolases play an important role in the turnover of membrane lipids of cells. Among them, the key inflammatory enzyme acid sphingomyelinase (ASM), hydrolyzes sphingomyelin to ceramide and phosphocoline. Since, retinal injury activates acid sphingomyelinase (ASMase), we asked (1) if IGFBP-3 reduced acid sphingomyelinase (ASM) and (2) if IGFBP-3 protected from injury-induced increases in retinal vascular permeability.

Methods: : In the first set of studies, healthy mice received an intravitreal injection of VEGF (100 ng/µl) to increase retinal vascular permeability, followed by intravitreal injection of IGFBP-3 (100 ng/µl) at 6 hours (n=6) and 24 hours (n=6) post VEGF injection. As a control, vehicle was injected either 6 or 24 hours after VEGF injection (n=6). Mice then received a tail injection of FITC- albumin 2 hours prior to sacrifice and their retinas were harvested 48 hours following initial VEGF injection. In a second group, mice underwent laser photocoagulation of retinal vessels and intravitreal injection of IGFBP-3 expressing plasmid or control plasmid. Mice were sacrificed 4 days post IGFBP-3 injection, the time of peak IGFBP-3 expression and retinal ASMase changes were determined by qRT-PCR.

Results: : At 24 hours, the combination treatment of VEGF and IGFBP-3 in uninjured mouse eyes resulted in a 1.5 fold reduction in retinal vascular permeability (p<0.05) compared to VEGF alone. In mice receiving laser and control plasmid, ASMase mRNA levels were elevated 4-fold compared to controls (p<0.05). However, in mice receiving laser photocoagulation and IGFBP-3 plasmid, ASMase levels approached baseline (p<0.05) and this was accompanied by a reduction in vascular permeability.

Conclusions: : Our studies support that the beneficial effects of IGFBP-3 on retinal vascular permeability may be due to reduction of the pro-inflammatory enzyme ASMase.

Keywords: diabetic retinopathy • growth factors/growth factor receptors • vascular occlusion/vascular occlusive disease