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Purpose: Large trials on anti-VEGF/PDGF (vascular endothelial/platelet-derived growth factor) combination therapy have been established to improve management of neovascular activity in age-related macular degeneration. Targeting pericytes, PDGF is thought to induce vessel regression and reduce fibrovascular scarring. The fate of pericytes exposed to anti-VEGF/PDGF combination therapy is not clear. Therefore, this study was designed to study the influence of anti-VEGF/PDGF on pericyte phenotype and cellular behavior.Methods: Human pericytes from placenta (hPC-PL) were treated with axitinib, a tyrosine kinase inhibitor targeting VEGFR1-3 and PDGFR. Toxic effects were excluded using live/dead staining. Phenotypic changes were evaluated using phalloidin staining for actin cytoskeleton and the expression of stress fibers. MRNA and protein expression levels of α-smooth muscle actin (αSMA) as a marker of proto-myofibroblastic transition were evaluated with real-time PCR and Western blotting. Influences of fibrotic cellular mechanisms were evaluated with a scratch wound migration and a collagen gel contraction assay.Results: Treatment with 0.5, 1, and 2.5 μg/ml axitinib strongly induced a proto-myofibroblast-like actin cytoskeleton with a marked increase in stress fibers. Quantitative real-time PCR and Western blotting revealed these changes to be linked to dose-dependent increases in αSMA mRNA and protein expression. However, fibrotic cellular mechanisms were significantly reduced in the presence of axitinib (scratch wound closure: up to − 78.4%, collagen gel contraction: up to − 37.4%).Conclusions: Combined anti-VEGF/PDGF inhibition seems to induce a proto-myofibroblast-like phenotype in human pericytes in vitro, but reduce profibrotic cellular mechanisms due to prolonged anti-PDGF inhibition. [ABSTRACT FROM AUTHOR]

Mutations in myocilin, a 55–57 kDa secreted glycoprotein, are causative for some forms of primary open-angle glaucoma (POAG). In vitro studies indicate that myocilin can modulate the hydrodynamic outflow resistance in the trabecular meshwork (TM) and that elevated amounts of myocilin can obstruct the TM outflow system in POAG. In this study, we analyzed the localization of myocilin in the trabecular meshwork (TM) of eyes with primary open-angle glaucoma (POAG), and compared it with that of normal eyes. Immunohistochemistry for myocilin was performed in the eyes of human donors (nine normal and 14 with POAG, including one with steroid-induced glaucoma). Staining for myocilin was observed in the extracellular spaces of the juxtacanalicular tissue (JCT) in all normal eyes. Some normal eyes did also show cytoplasmic staining for myocilin in TM cells. In the eyes of six donors with POAG, staining of the JCT was more widespread and intense than in normal eyes. In the other eyes with POAG, immunoreactivity for myocilin in the JCT was not markedly different to that of normal eyes. Staining intensity in the JCT of POAG eyes did not obviously correlate with intraocular pressure or clinical severity. In the eyes of one patient with steroid-induced POAG, cells of the TM, Schlemm’s canal endothelium, and the anterior stroma of the iris showed an immunoreactivity for myocilin which was considerably more intense than in normal eyes, or in the eyes with other forms of POAG. In some cases of POAG, the structural changes in the JCT include an increase in myocilin in the extracellular pathways of aqueous humor. Treatment with steroids appears to increase myocilin synthesis in TM and iris of human eyes in situ. [ABSTRACT FROM AUTHOR]