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Harry Sweigard, Ryoji Yanai, Philipp Gaissert, Magali Saint-Geniez, Keiko Kataoka, Aristomenis Thanos, Kip M Connor; The Alternative Complement Pathway Facilitates Neovessel Regression. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5372.
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© ARVO (1962-2015); The Authors (2016-present)
A characteristic of retinopathy is the development of neovessels (NV). In humans, and in mouse models, the NV often spontaneously regress while the established vasculature is kept intact, returning the retina to a relatively normal state. The mechanisms for the selective removal of NV are not fully understood and would be of great therapeutic value.
C57BL/6 mice and alternative pathway deficient mice (Fb-/-) were used in a mouse model of retinopathy. Retinas were collected from postnatal (P) day 14 to P25 and used for immunohistochemistry (IHC), RNA, and protein evaluation of complement activity, NV development and regression. Retinal flat mounts were used to quantitate the number of NV. Laser capture micro-dissection (LCM) was performed to compare complement regulator expression in isolated NV and normal vessels. HUVEC cells were used to study the effects of hypoxia on complement regulatory gene expression.
Flat mounts show that Fb-/- mice develop more NV along with delayed regression compared to control mice. Real time PCR on diseased retinas isolated from P14 - P17 show a gradual increase in Fb expression, compared to controls, with a maximum difference at P17 (P=0.001). IHC for Fb shows expression on NV, which co-localizes with TUNEL positive cells. LCM of NV versus established vessels analyzed by RTPCR shows that CD55 levels are decreased on NV relative to established vessels (P=0.005). Cell culture using HUVECs show that CD55 is down-regulated when kept in hypoxic conditions.
Our results indicate that the alternative arm of complement is involved in the clearance of NV in a clinically relevant mouse model of retinopathy. Expression levels of CD55, a regulator of the alternative arm, are decreased on NV but maintained on mature vessels. Decreases in CD55 expression are likely a direct consequence of low oxygen. This allows the targeting NV by complement for elimination while leaving the established vasculature intact. Our findings help in the understanding of how the retina maintains such organized vessel architecture and will potentially lead to new disease markers or novel ways for selective removal of NV.
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