PURPOSE: To investigate the functional properties and intracellular signaling of hyalocytes under platelet-derived growth factor (PDGF)-rich conditions. METHODS: The hyalocytes were isolated from bovine eyes and identified by immunocytochemistry and electron microscope. The expression of PDGF receptor alpha/beta and its phosphorylation in response to PDGF-BB was analyzed by Western blot analysis. PDGF-BB-induced proliferation and migration were evaluated by thymidine uptake and Boyden's chemotaxis assay. The expression of the urokinase-type plasminogen activator (uPA) gene and the fibrinolytic activity were assessed by Northern blotting and fibrin zymography. An in vitro type I collagen gel contraction assay was performed to determine the effect of PDGF-BB on cellular contraction. RESULTS: The hyalocytes were immunocytochemically positive for S-100 and negative for glial fibrillary acidic protein (GFAP) and cytokeratin, as previously described. The electron microscope demonstrated that hyalocytes possess lysosome-like granules, mitochondria, and micropinocytotic vesicles in their cytoplasm. The hyalocytes expressed PDGF receptor alpha and beta, both of which were immediately phosphorylated in response to PDGF-BB. PDGF-BB also activated p85 PI3-kinase, p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase. PDGF-BB induced thymidine uptake and migration in a concentration-dependent (0-10 ng/mL) manner. Inhibitors of the respective kinases prohibited PDGF-BB-dependent thymidine uptake and migration with the exception of the p44/p42 MAP kinase inhibitor, which displayed no inhibitory effects on migration. PDGF-BB increased uPA gene expression and fibrinolytic activity. Collagen gel contraction observed under PDGF-BB-rich conditions was not prohibited by the respective inhibitors investigated. CONCLUSIONS: The hyalocytes demonstrated macrophage-like characteristics and may have both physiologic and pathologic roles, such as the maintenance of vitreous transparency through fibrinolytic activity and the pathogenesis of proliferative-vitreoretinal diseases through cellular proliferation and vitreous hyper-contraction.
PURPOSE: To investigate the functional properties and intracellular signaling of hyalocytes under platelet-derived growth factor (PDGF)-rich conditions. METHODS: The hyalocytes were isolated from bovine eyes and identified by immunocytochemistry and electron microscope. The expression of PDGF receptor alpha/beta and its phosphorylation in response to PDGF-BB was analyzed by Western blot analysis. PDGF-BB-induced proliferation and migration were evaluated by thymidine uptake and Boyden's chemotaxis assay. The expression of the urokinase-type plasminogen activator (uPA) gene and the fibrinolytic activity were assessed by Northern blotting and fibrin zymography. An in vitro type I collagen gel contraction assay was performed to determine the effect of PDGF-BB on cellular contraction. RESULTS: The hyalocytes were immunocytochemically positive for S-100 and negative for glial fibrillary acidic protein (GFAP) and cytokeratin, as previously described. The electron microscope demonstrated that hyalocytes possess lysosome-like granules, mitochondria, and micropinocytotic vesicles in their cytoplasm. The hyalocytes expressed PDGF receptor alpha and beta, both of which were immediately phosphorylated in response to PDGF-BB. PDGF-BB also activated p85 PI3-kinase, p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase. PDGF-BB induced thymidine uptake and migration in a concentration-dependent (0-10 ng/mL) manner. Inhibitors of the respective kinases prohibited PDGF-BB-dependent thymidine uptake and migration with the exception of the p44/p42 MAP kinase inhibitor, which displayed no inhibitory effects on migration. PDGF-BB increased uPA gene expression and fibrinolytic activity. Collagen gel contraction observed under PDGF-BB-rich conditions was not prohibited by the respective inhibitors investigated. CONCLUSIONS: The hyalocytes demonstrated macrophage-like characteristics and may have both physiologic and pathologic roles, such as the maintenance of vitreous transparency through fibrinolytic activity and the pathogenesis of proliferative-vitreoretinal diseases through cellular proliferation and vitreous hyper-contraction.
Authors: Vivian Capilla-González; Javier López-Beas; Natalia Escacena; Yolanda Aguilera; Antonio de la Cuesta; Rafael Ruiz-Salmerón; Franz Martín; Abdelkrim Hmadcha; Bernat Soria Journal: Mol Ther Date: 2018-08-16 Impact factor: 11.454