G Gao1, C Shao, S X Zhang, A Dudley, J Fant, J-X Ma. 1. Department of Ophthalmology, Medical University of South Carolina, 167 Ashley Ave., Charleston, SC 29425, USA.
Abstract
AIMS/HYPOTHESIS: Kallikrein-binding protein (KBP) is a serine proteinase inhibitor (serpin). It specifically binds to tissue kallikrein and inhibits kallikrein activity. Our study was designed to test its effects on retinal neovascularization and vascular permeability. METHODS: Endothelial cell proliferation was determined by [(3)H] thymidine incorporation assay and apoptosis quantified by Annexin V staining and flow cytometry. Effect on retinal neovascularization was determined by fluorescein angiography and count of pre-retinal vascular cells in an oxygen-induced retinopathy (OIR) model. Vascular permeability was assayed by the Evans blue method. Vascular endothelial growth factor (VEGF) was measured by Western blot analysis and ELISA. RESULTS: Kallikrein-binding protein specifically inhibited proliferation and induced apoptosis in retinal capillary endothelial cells. Intravitreal injection of KBP inhibited retinal neovascularization in an OIR model. Moreover, KBP decreased vascular leakage in the retina, iris and choroid in rats with OIR. Blockade of kinin receptors by specific antagonists showed significantly weaker inhibition of endothelial cells, when compared to that of KBP, suggesting that the anti-angiogenic activity of KBP is not through inhibiting kallikrein activity or kinin production. KBP competed with (125)I-VEGF for binding to endothelial cells and down-regulated VEGF production in endothelial cells and in the retina of the OIR rat model. CONCLUSION/ INTERPRETATION: Kallikrein-binding protein is a multi-functional serpin, and its vascular activities are independent of its interactions with the kallikrein-kinin system. Inhibition of VEGF binding to its receptors and down-regulation of VEGF expression could represent a mechanism for the vascular activities of KBP.
AIMS/HYPOTHESIS: Kallikrein-binding protein (KBP) is a serine proteinase inhibitor (serpin). It specifically binds to tissue kallikrein and inhibits kallikrein activity. Our study was designed to test its effects on retinal neovascularization and vascular permeability. METHODS: Endothelial cell proliferation was determined by [(3)H] thymidine incorporation assay and apoptosis quantified by Annexin V staining and flow cytometry. Effect on retinal neovascularization was determined by fluorescein angiography and count of pre-retinal vascular cells in an oxygen-induced retinopathy (OIR) model. Vascular permeability was assayed by the Evans blue method. Vascular endothelial growth factor (VEGF) was measured by Western blot analysis and ELISA. RESULTS:Kallikrein-binding protein specifically inhibited proliferation and induced apoptosis in retinal capillary endothelial cells. Intravitreal injection of KBP inhibited retinal neovascularization in an OIR model. Moreover, KBP decreased vascular leakage in the retina, iris and choroid in rats with OIR. Blockade of kinin receptors by specific antagonists showed significantly weaker inhibition of endothelial cells, when compared to that of KBP, suggesting that the anti-angiogenic activity of KBP is not through inhibiting kallikrein activity or kinin production. KBP competed with (125)I-VEGF for binding to endothelial cells and down-regulated VEGF production in endothelial cells and in the retina of the OIR rat model. CONCLUSION/ INTERPRETATION:Kallikrein-binding protein is a multi-functional serpin, and its vascular activities are independent of its interactions with the kallikrein-kinin system. Inhibition of VEGF binding to its receptors and down-regulation of VEGF expression could represent a mechanism for the vascular activities of KBP.
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