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. 2015 Sep;185(9):2534–2549. doi: 10.1016/j.ajpath.2015.04.029

Figure 1.

Figure 1

CCR3 and its ligands are expressed in the spontaneous choroidal neovascularization (CNV) lesions of JR5558 mice. A: CCR3 (anti-rabbit Alexa Fluor 594, red) is expressed in the neo-vessels (fluorescein isothiocyanate-IB4, green) of the CNV (arrows) and in nonvascular cells associated the CNV (arrowheads) in eyes of JR5558 mice. B: The CCR3-expressing, nonvascular cells around the CNV are macrophages (F4/80, arrows, anti-rat Alexa Fluor 488, green). C: Some CNV-associated leukocytes (CD45, arrows, streptavidin Alexa Fluor 488, green) also express high levels of CCR3. D: There is no detectable CCR3 staining on the retinal pigment epithelium (RPE65, streptavidin Alexa Fluor 647, pseudo-green) directly under or around the CNV lesion (dotted line), whereas the lesion itself is highly positive for CCR3 (red). Note that the positive CCR3 staining is atop the RPE layer; compare RPE65 with CCR3. E and F: CC chemokine ligand (CCL)-11 (anti-goat Alexa Fluor 594, red) and CCL24 (anti-rabbit Alexa Fluor 594, red) are expressed by macrophages (F4/80, green, arrows) around the CNV (note that some ligands are not associated with cells and likely bind to the extracellular matrix). Animals at postnatal day 30 (P30) were used for A–D and P20 mice were used for the staining in E and F. G: Real-time PCR analyses of the relative expression of CCL11 and CCL24 mRNA show that both ligands are expressed at higher levels in the RPE/choroid complex of JR5558 mice compared with wild-type (WT) controls at P20, during the active phase of spontaneous CNV development. H: Protein levels of CCL11 and CCL24 were measured by enzyme-linked immunosorbent assay in the RPE/choroid complex in both WT and JR5558 mice at P20. Data are expressed as means ± SEM. n = ≥3 eyes per group (G); n = 12 (H, CCL11); n = 8 (H, CCL24).∗P < 0.05, ∗∗P < 0.01 versus levels in wild-type control. Scale bars = 20 μm.