Plasmin Is the Major Protease Responsible for Processing PDGF-C in the Vitreous of Patients with Proliferative Vitreoretinopathy

Abstract

Proliferative vitreoretinopathy (PVR) is the primary cause of failure of retinal reattachment surgery. Growth factors such as platelet-derived growth factor (PDGF) are strongly associated with PVR. Of the five PDGF family members, PDGF-C predominates in the vitreous of experimental and clinical PVR. PDGF-C is secreted as a latent protein that requires proteolytic processing for activation. Although tissue plasminogen activator (tPA) is primarily responsible for processing PDGF-C in cultured cells, it constitutes a minority of the processing activity in the vitreous of experimental animals and in patients with PVR. Identifying the major PDGF-C processing protease was the purpose of this study. The presence of serum proteins in the vitreous was detected by Coomassie blue staining and Western blotting. PDGF-C processing activity was detected in an in vitro processing assay using either native or recombinant PDGF-C as the substrate. Plasmin activity was blocked using alpha(2)-plasmin inhibitor. Phosphorylation of the PDGF receptor (PDGFR) was monitored by antiphosphotyrosine Western blotting. Vitreous specimens were collected from experimental rabbits or from patients undergoing vitrectomy to repair retinal detachment or for other reasons. A number of prominent serum proteins (albumin and IgG) were detected in the vitreous of all patients undergoing retinal surgery. The level of these proteins markedly increased in the vitreous of rabbits as they developed PVR. These observations suggested that serum-borne proteases are also likely to be present in the vitreous. Indeed, plasmin (a protease capable of processing PDGF-C) was present in the vitreous from PVR rabbits and retinal surgery patients. Plasmin was dramatically more effective than tPA in processing PDGF-C in an in vitro assay. Blocking plasmin activity eliminated most of the processing activity in the vitreous of patients and rabbits with PVR. Plasmin was the major PDGF-C processing protease in the vitreous of PVR rabbits and patients undergoing retinal surgery. Blocking plasmin prevented the generation of active PDGF-C, which is the major PDGF isoform relevant for PVR. These observations are the first report of an in vivo protease responsible for processing PDGF-C. In addition, plasmin was identified as a novel therapeutic target for patients with PVR.