Despite endogenous proangiogenic properties of MCP-1, we did not observe a reduction in NV on P17 in the MCP-1
−/− mice compared with B6 mice.
46 47 48 In addition, the unaltered NV observed on P17 in MCP-1
−/− mice suggested that infiltrating macrophages are not a major contributor to NV in this model. These observations are consistent with Müller cell-derived VEGF playing the major role in stimulating angiogenesis in this model, as previously described.
49 However, with reduced numbers of infiltrating macrophages, the shift toward tuft regression that normally occurs in B6 mice between P17 and P21 is delayed in MCP-1
−/− mice. Similarly, in a preliminary study in B6 mice, macrophages were systemically depleted with liposomal-clodronate, resulting in a reduction in neovascular tuft-associated F4/80
+ cells with an associated twofold increase in neovascular nuclei on P21O
2, supporting the concept that infiltrating macrophages contribute to tuft regression (Davies MH, et al.
IOVS 2006;47:ARVO E-Abstract 3223). In contrast to the MCP-1
−/− mice, mice deficient in the intracellular apoptosis inhibitor Bcl-2 exhibit significantly less NV than B6 control mice in the OIR model.
50 In vivo studies using capillary tube assays revealed a requirement of only 22% apoptotic ECs for a significant decrease in microvascular density.
51 Hence, it appears that the B6 mice had a level of EC apoptosis that allowed for tuft regression between P17 and P21 in contrast to the delayed regression we observed in the MCP-1
−/− mice, whereas Bcl-2
−/− mice likely had a level of apoptosis that did not allow for a significant level of NV, despite the presence of VEGF. However, the vascular tufts eventually regressed in the MCP-1
−/− mice, suggesting that additional apoptotic mechanisms lead to tuft regression after P21. We did observe an increase in the percentage of tuft cells undergoing apoptosis in the MCP-1
−/− mice on P21. Growth factor withdrawal secondary to waning levels of retinal VEGF likely contributed to the eventual regression of the vascular tufts after P21 in the MCP-1
−/− mice.
49 Retinal Fas was also up-regulated on P21 in the OIR model, whereas EC up-regulated this death receptor under irregular flow conditions (vascular tufts).
5 52 Thus, the tuft ECs in the nascent capillaries were vulnerable to autoregulatory FasL-induced cell death from neighboring ECs.
52 Finally, the resident microglia likely also contributed to eventual regression of the neovascular tufts.