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Fig. 2 | Journal of Neuroinflammation

Fig. 2

From: Rapid monocyte infiltration following retinal detachment is dependent on non-canonical IL6 signaling through gp130

Fig. 2

Quantification of recruited monocytes and resident microglia with Flow Cytometry. a Representative dot plots for CD11bhigh IB4 low monocytes (top left gate) and CD11blow IB4high resident microglia (lower right gate) in 5 groups. b After one day, there was significant monocyte infiltration in all RD groups, as well as an increased number of microglia (normal = 8 mice; anti-IL6 Rα group = 16 mice; IL6 KO group = 6 mice). Compared with untreated RD1 retinas (n = 16 mice), only anti-gp130 treated retinas (n = 13 mice) had fewer monocytes recruited (**p < 0.01). c After seven days, anti-gp130 still (n = 8 mice) had fewer monocytes compared with untreated RD7 retinas (n = 14 mice, *p < 0.05). The other three RD7 groups sustained substantial monocyte infiltration (untreated RD7 ***p < 0.001; anti-IL6 Rα =13 mice, ***p < 0.001, IL6 knockout =6 mice ***p < 0.001). Moreover, the number of resident microglia increased significantly in the untreated (***p < 0.001), anti-IL6 Rα(**p < 0.01) and anti-gp130 RD7 retinas (***p < 0.001). Interestingly, anti-gp130 seemed to boost microglia numbers (compared with untreated RD7, ***p < 0.001), while IL6 knockout inhibited this rise in microglia numbers (**p < 0.01). d Total numbers of microglia and monocytes in each group. In the untreated and anti-IL6 Rα RD groups, total immune cells fell after seven days (***p < 0.001 and ** p < 0.01), mainly due to fewer monocytes (***p < 0.001 and * p < 0.05). RD retinas with anti-gp130 treatment had no significant change in total immune cell numbers, but replenished more microglia (***p < 0.001). In IL6 knockout mice, total immunes cells decreased sharply (***p < 0.001), and had the fewest microglia numbers (**p < 0.01). e Fluorescent microscope image of dissociated retinal cells used prior to flow cytometry. Retinas were digested, CD11bhigh monocytes (red round cells) and IB4high microglia (green cells) were isolated, nuclei were labeled with DAPI (bar = 10 μm). f Flat mounts (bar = 100 μm) showed that gp130 blocking antibody was associated with increased numbers of iba1high cells, suggesting it promoted the differentiation of monocytes (CD11bhigh, red) into microglia-like macrophages (green) at seven days after RD as evidence by immunohistochemistry co-labeling

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