Abstract
Purpose :
To investigate ocular inflammatory cell infiltration in a murine laser-induced CNV model.
Methods :
Laser pulses were applied to mice eyes and CNV area measured by i.v. injection of a vascular label, RPE-Choroid flat mount imaging, and quantification of CNV area by morphometric analysis.
CNV area and cellular infiltrate were assessed 7 days after laser application in mice systemically administered an anti-VEGF Ab (“4G3”) or saline. To study cellular infiltration, labeled antibodies were used to identify microglia (Iba-1+) and macrophages (F4/80+). Images of the ocular flatmounts were captured using AxioVision software (Zeiss). Iba-1+ microglia cells peripheral to the CNV lesion were randomized and counted semi-automatically using MatLab, and the F4/80+ macrophages were counted manually. CNV area and cellular infiltrate were analyzed in sex matched Chemokine (C-C Motif) Ligand 2 (CCL2) knockout (KO) and wild-type (WT) litter-mate controls.
Results :
7 days after laser ocular flat mounts demonstrated microglia cell populations were increased by 61% (N = 16 - 20 eyes per group, p = 0.007) and no changes were seen in retinal macrophages (n = 5 retinas per group, p = 0.36) compared to non-lasered mice. Mice administered an anti-VEGF Ab demonstrated an average of 46% reduction in CNV area (p = < 0.0001), but no difference in sub retinal microglia compared to vehicle treated mice (n = 2 Studies, n ≥ 10 eyes/group, p ≥ 0.22). CNV area was increased in CCL2 KO compared to WT controls (n = 2 studies per sex, males average increase CNV area = 67.8%, females 48.1%, n = 9 - 15 mice per group, p ≤ 0.007 in 3 studies, p > 0.05 in one study). CCL2 KO mice had similar numbers of sub retinal microglia and retinal macrophages compared to WT mice (n = 10 eyes/group p ≥ 0.23).
Conclusions :
Anti-VEGF therapy reduces CNV area, but has no effect on microglia and macrophage infiltrate. CCL2 genetic KO increases CNV area, but shows no alteration in cellular infiltrate numbers. Functional changes in cellular infiltrate rather than number may play a key role in ocular angiogenesis.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.