Previously related to the formation of CNV in AMD and in animal models. The growth of CNV may be further encouraged by the increased secretion of VEGF by RPE cells following co-culture that can directly stimulate endothelial cell proliferation and migration. In support of the relationship betwen RPE alterations and CNV formation, epidemiological studies have also linked the clinical presence of RPE changes occurring in patients with early and intermediate AMD with an elevated 5-year risk for the development of CNV. Our experiments with the in vivo transplantation of retinal microglia demonstrated that the presence of subretinal microglia exerts a strong pro-angiogenic influence in the formation and growth of CNV. The creation of a pro-angiogenic environment in the subretinal space, while mediated significantly by RPE alterations, may also be contributed towards by the subretinal microglia themselves. In our in vitro co-culture experiments, while mRNA and protein analysis in RPE cell lysates relate directly to RPE gene expression, protein Ginsenoside-F4 analyses involving RPE co-culture supernatants may however also contain contributions secreted from retinal microglia cells. However, in our angiogenesis and Tulathromycin B microglia-migration functional assays, we found that the changes induced by RPE co-culture supernatants were significant when compared not only to unexposed RPE supernatants, but also when compared to supernatants of activated retinal microglia not exposed to RPE cells. As such, despite possible contributions by retinal microglia themselves, changes in RPE gene expression and protein secretion are likely play a prominent role in inducing the structural and functional changes of pathological significance in the subretinal space. While our in vivo model system permits direct cell-cell contact between retinal microglia and RPE cells, the in vitro system used here brings the two cell types in close proximity but however precludes direct cellular surface contact. The similarities in the nature of changes induced in both in vitro and in vivo systems suggest that many of these do not require direct microglial-RPE cell contact, although additional studies may be required to investigate the effects that result only from direct cellular contact. In addition to retinal microglia-to-RPE communication, other forms of intercellular interactions may 
also play a role in AMD pathogenesis.