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Brian J. Raisler, Michelle R. Smith, Robinette King; Understanding The Role Of Macrophages In Oxygen Induced Retinopathy. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3118.
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© ARVO (1962-2015); The Authors (2016-present)
To analyze the macrophage response in wild-type in CCR2 -/- mice, deficient in macrophage signaling, in the oxygen induced retinopathy mouse model during the establishment and regression of pre-retinal neovascularization.
Inflammatory signaling was queried by bead-array analysis for cytokines (M-CSF, MCP-1, MIP-1alpha and beta, VEGF) in retinal homogenates. Macrophage infiltration was assessed in paraffin embedded retinal cross-sections with IHC/IF for F4/80 and CD68 and in retinal flat mount staining for F4/80. Macrophage numbers were determined by comparative flow cytometry for F4/80+/CD11c- populations between wild-type and CCR2 -/- mice. Lastly macrophage induced apoptosis was measured by staining for activated caspase-3 in retinal cross-sections.
Macrophages were found to be closely associated with retinal vessels and neovascular tufts in particular. Interestingly, CCR2 -/- mice still exhibit macrophage infiltration similar to that of wild-type mice. CCR2 -/- mice also show a more slightly robust neovascular response in the OIR model than wild-type at postnatal day 17, 42.5 nuclei per section vs. 33.7 (p<0.005, n=3). Flow cytometry of retinal cells from CCR2 -/- mice and WT during the OIR model showed increased levels of macrophages in the period immediately following return to room air. VEGF levels in the retinas of CCR2 -/- mice are markedly increased after the return to room-air, 22.5 ng/ml vs. 9.4 ng/ml for WT mice (p<0.05). Levels for IL-18, M-CSF, and bFGF were also significantly altered over the course of neovascular establishment and regression in the OIR model. Activated caspase-3 was observed in association with macrophage infiltrate during the regression of NV vessels.
This work affirms the Janus-like role of macrophages in neovascularization in the eye. During the establishment phase of NV, macrophages contribute to and participate in inflammatory processes that exacerbate new vessel formation. Later in the model, macrophages also play a role in vascular regression, mediated through phagocytosis and apoptosis. The CCR2 -/- mice seem to exhibit a persistant macrophage response despite a lack of signaling through the MCP-1/CCR2 pathway. This provocative finding hints at the existence of an alternate receptor for MCP-1 that may be signaling macrophage response in these animals and modulating their neovascular response. Disregulation of macrophage related cytokines may contribute to the differential progression observed between WT and CCR2 -/- mice in the OIR model.
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