C-reactive protein dissociation contributes to choroidal neovascularization in vivo: implications for age-related macular degeneration    

Choroidal neovascularization (CNV) and inflammation processes play an important role in retinal disease development and C-reactive protein (CRP) has been shown to contribute to development diseases like Age-related macular degeneration (AMD). Our aim was to evaluate whether monomeric and pentameric CRP (pCRP, mCRP) isoforms contribute to CNV in vivo and to characterize the mechanism of CRP dissociation in-vivo and in vitro. Both CRP isoforms were intravitreally (IVT) or intravenously (IV) injected in mice, CNV was laser-induced, retinography and fluorescein angiography were performed to evaluate edema. Lectin, mCRP, F4/80 and C5b9 localization were assessed by immunofluorescence and visualized under a confocal microscope. Intensity of fluorescence of mCRP (IF mCRP) was also quantified. To confirm pCRP dissociation in ARPE-19 cells and mice, pCRP was coupled to a fluorochrome and IVT injected. A statistical increase in CNV areas was observed in pCRP IVT injected males (p<0.05) while a statistical decrease was shown in females (p<0.05). After IV injection, pCRP males showed an increase in CNV areas only vs. mCRP injected mice (p<0.05) and in females the injection of pCRP injected mice showed higher CNV areas vs. vehicle (p<0.05) and vs. mCRP injected mice (p<0.05). Retinal edema after IVT CRP injection was observed mainly in mCRP injected mice. In females there was an IF mCRP statistical decrease in pCRP IVT injected mice vs. vehicle and a statistical increase in pCRP IV injected mice vs. vehicle (p<0.05). Mice injected with IVT isoforms showed F4/80 positive cells and C5b-9 deposition around the CNV areas. mCRP labeling was observed in the intercellular space of the endothelial cells in the angiogenic area and detected in pCRP IVT injected animals, demonstrating the dissociation of pCRP into mCRP both in vitro and in vivo in proinflammatory microenvironments. In conclusion, CRP administration increased CNV areas and edema in the retina, suggesting that CRP is activated in the CNV inflammatory environment. Additionally, pCRP dissociates in vivo into mCRP close to CNV damaged tissue, hypothesizing that mCRP is the CRP conformation that contributes to the exacerbation of the CNV process.