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M. Nukada, N. Unoki, K. Ogino, T. Murakami, N. Yoshimura; Time-Lapse Imaging of Retinal Angiogenesis Demonstrates That Anecortave Desacetate Attenuates Both the Development and Progression of Neovascular Sprouting. Invest. Ophthalmol. Vis. Sci. 2010;51(13):47.
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© ARVO (1962-2015); The Authors (2016-present)
Neovascularization is a dynamic phenomenon, whereas the kinetics of retinal angiogenesis remains ill-defined. Here we evaluated the dynamics of neovascular sprouting and its molecular mechanisms in the retinal angiogenesis treated with an angiostatic steroid, anecortave desacetate (AD).
Retinas were isolated from 7- to 8- week old C57BL6/J mice and cultured for 96 hours in the presence or absence of AD, followed with each additional experiment. For in vivo experiments, AD was injected intraperitoneally into C57BL6/J mice on postnatal day 2 (P2), and the retinas were isolated on P4. After the fixation and permeation, immunohistochemistry was performed with fluorescent 2nd antibodies. Real time PCR was performed for the measurements of mRNA levels. Time-sequential images were obtained at every 15 minute using confocal microscopy, and the movements of neovascular sprouts were quantified
AD treatment significantly reduced the number of neovascular sprouts in retinal explants in a dose dependent manner. Time-lapse imaging demonstrated that AD suppressed both the development and the elongation of sprouts, and the motility of the leading edges in tip cells. AD treatment further disturbed the filopodial extension and significantly decreased the transcriptional levels of KDR and platelet-derived growth factor-B (PDGFB), which are highly expressed in tip cells, suggesting the tip cell dysfunction. The transcriptional level of stromal derived factor-1 (SDF-1), which regulates tip cell motility, was also reduced by AD, whereas AD did not alter the expression of its receptor, CXCR4. Additionally, immunostaining and electron microscopy demonstrated the immature basement membrane in neovascular sprouts treated with AD. Further in vivo experiments showed that AD inhibited the neovascularization and filopodial extension in tip cells in retinal vascular development. These data suggest that AD suppresses both development and progression of sprouting angiogenesis.
AD attenuated VEGF-induced retinal angiogenesis, mediated via the suppression of the development and progression of neovascular sprouts and tip cell motility at least partially.
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