Abstract
Purpose :
ROP, a retinal neurovasculopathy, is a leading cause of childhood blindness. We reported recently that activation of the molecular chaperone, Sig1R, provides a novel rescue of neuronal deficits in the oxygen-induced retinopathy (OIR), model of ROP (Wang et al, ARVO 2020). Here, we focus on Sig1R and its novel therapeutic potential in vascular injury characteristic of ROP.
Methods :
C57BL/6J pups (P7 with nursing dam) were subjected to OIR or room air (CT). (+)-Pentazocine ((+)-PTZ), a high affinity Sig1R ligand, was administered starting P7 (0.5 mg/kg, IP, every other day). Retinal vasculature was imaged by fluorescein angiography & vascular tortuosity analyzed by MATLAB at P17& P48. At P17, retinal vascular distribution (neovascularization (NV) and avascular area) & vascular barrier function (Evans blue (EB) leakage assay) were evaluated. Oxidative stress (DHE, 3-NT, 4-HNE), reactive gliosis (anti-GFAP), expression of proteins/cytokines (Sig1R, ZO-1, Occludin, p-STAT3 and VEGF, IL-6, TNFα) were assessed by immunohistochemistry (IMH), Dot/Western blotting (WB) & ELISA.
Results :
Vascular tortuosity: Retinal arteries in OIR mice were more tortuous (~1.3) v. CT (1.0), whereas (+)-PTZ significantly reduced arterial tortuosity (~1.1) in OIR mice at P17 & P48. Vascular distribution: Retinal avascular region (OIR-non: ~30% v. OIR+PTZ: ~12%) and NV (OIR-non: ~7.8% v. OIR+PTZ: ~2.5%) were significantly decreased in OIR+PTZ mice. Retinal vascular barrier: EB extravasation was elevated in OIR mice, but decreased significantly in OIR+PTZ mice. ZO-1 & Occludin levels decreased significantly in OIR mice, whereas (+)-PTZ preserved their levels. Oxidative stress and reactive gliosis: Detection of oxidative stress by DHE staining, 4-HNE and 3-NT immunostaining showed that (+)-PTZ attenuated superoxide and lipid peroxide levels in OIR mice. (+)-PTZ treatment diminished reactive gliosis in OIR mice. Exploration of Sig1R-STAT3-Inflammation axis. In OIR mice, we observed a significant decrease of Sig1R, along with increase of p-STAT3, VEGF, IL-6 and TNFα, whereas (+)-PTZ treatment normalized these effects.
Conclusions :
(+)-PTZ treatment attenuated OIR-induced vasculopathy, retinal barrier dysfunction, reactive gliosis, oxidative stress and STAT3-mediated inflammation, suggesting that Sig1R is a promising therapeutic target for vascular injury in ROP.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.