Hypoxia Inducible Factor-1 Upregulates the Expression of Plasminogen Activator Inhibitor-1 in Retinal Endothelial Cells to Promote their Survival in Proliferative Diabetic Retinopathy Patients

Abstract

Purpose: Diabetic retinopathy (DR) is the most common diabetic microvascular complication. Damage to the retinal vasculature from sustained hyperglycemia ultimately leads to retinal ischemia, resulting in stabilization of the transcription factor hypoxia inducible factor (HIF)-1α and upregulation of the expression of angiogenic factors that promote the development of neovascularization (NV). This heralds the progression from non-proliferative to proliferative DR (PDR). The introduction of therapies targeting a HIF-1-regulated angiogenic mediator, vascular endothelial growth factor (VEGF), has transformed the treatment of diabetic macular edema. However, inhibition of this potent endothelial cell (EC) survival factor alone is not adequate for treating patients with PDR. Here we explore additional HIF-1-regulated EC survival factors that may contribute to the development and progression of retinal NV in PDR patients who do not respond adequately to anti-VEGF therapy.

Methods: Johns Hopkins School of Medicine Institutional Review Board approval was obtained to collect aqueous fluid (AF) and vitreous biopsies from patients undergoing intraocular surgery. Levels of VEGF were measured using ELISA. Angiogenic potential of AF was measured using tubule formation assay. A multiplex ELISA angiogenesis array was performed on AF from PDR patients with low VEGF levels but high angiogenic potential. Expression of angiogenic factors was confirmed by qPCR and ELISA in ECs in vitro and in the oxygen induced retinopathy model in vivo. Cell survival was measured using the MTS assay. Expression of angiogenic factors was confirmed in vitreous biopsies from PDR patients.

Results: We observed elevated levels of plasminogen activator inhibitor-1 (PAI-1) in AF from PDR patients with low VEGF levels but high angiogenic potential. PAI-1 mRNA expression was upregulated by HIF-1 in hypoxic ECs in vitro and the ischemic retina in vivo. Secretion of PAI-1 by hypoxic ECs cells was necessary and sufficient to promote EC survival. Increased expression of PAI-1 was confirmed in vitreous biopsies from PDR patients.

Conclusions: PAI-1 is a HIF-1-regulated secreted factor that promotes EC survival in PDR. We propose that PAI-1 is a good therapeutic target for the treatment of retinal NV in diabetic patients.