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Fig. 7 M-CSF stimulates an inflammatory response and initiates Müller glia cell cycle re-entry in the absence of a lesion. (A) Scheme of experimental outline. Wild type or NFκB:GFP reporter animals were injected with M-CSF or PBS into the vitreous of the eye and analyzed at 2 and 4 days post injection (dpi). (B) Quantification of TUNEL+ nuclei in control and M-CSF-injected retinae at two dpi reveals no chance of cell death. See also Supplementary Figure S7 for counts of pyknotic nuclei. (C) Immunohistochemistry for proliferating cell nuclear antigen (PCNA) reveals proliferation in close proximity to Zrf-1 (GFAP) positive Müller cells in M-CSF injected but not in control injected animals at four dpi. n both conditions a slight nonspecific labeling can be seen in the ONL. (D) Quantification of PCNA+ cells in control (PBS) and M-CSF injected retinae at four dpi. (E) L-Plastin staining shows more positive cells in M-CSF injected retinae at four dpi compared to controls. The location in the inner nuclear layer (INL) and morphology of the leukocytes indicates reactivity. (F) Quantification of L-Plastin+ cells in control and M-CSF injected retinae at four dpi, show accumulation of L-Plastin cells in M-CSF injected specimen. (G) In comparison to PBS, injection of M-CSF results in strong activation of the NFκB:GFP reporter at two dpi. (H) In situ hybridization of mmp9 shows transcriptional activation of this gene in M-CSF injected but not in control-injected animals at two dpi. Scale bars = 50µm, Error bars indicate SEM; ns > 0.05; ** = p ≤ 0.01; *** = p < 0.001; N ≥ 5; two-tailed Student’s t-test. ONL = outer nuclear layer; GCL = ganglion cell layer.