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Fig. 4 | Journal of Neuroinflammation

Fig. 4

From: Bovine neutrophil chemotaxis to Listeria monocytogenes in neurolisteriosis depends on microglia-released rather than bacterial factors

Fig. 4

Bovine microglia express IL-8 in vitro and in situ, but IL-8 blocking in supernatant of Lm-infected microglia does not prevent chemotaxis of bovine PMN. A ELISA quantification of IL-8 from supernatants of non-infected (Microglia sup.) and Lm-infected (Microglia Lm sup.) cultured microglia. Cultured microglia exhibit a basal secretion of IL-8 into the supernatant, and infection stimulates IL-8 secretion at concentrations chemotactic for PMN. Data are represented as mean ± SEM (standard error of mean) of 3 independent experiments performed in triplicates. Statistical analysis: Mann–Whitney U tests (****P < 0.0001). B Treatment of Lm-infected microglial supernatant with a monoclonal anti-IL-8 antibody (anti-IL-8 microglia Lm sup.) does not block chemotaxis toward conditioned medium from infected microglia. Data are represented as means on a superimposed scatter dot plot of 2 independent experiments performed in duplicates (anti-IL-8 microglia Lm sup.) or triplicates (all other conditions). Statistical analyses: Mann–Whitney U tests (**P < 0.01). C Bovine IL-8 chemotactic activity is specifically neutralized following incubation with a monoclonal anti-IL-8 antibody (Anti IL-8 + IL-8), while IL-8 chemotactic activity is not abrogated after incubation with mouse IgG1 (MoIgG1 + IL-8). Data are represented as means on a superimposed scatter dot plot of 3 independent experiments performed in duplicates or triplicates. Statistical analyses: Mann–Whitney U tests (**P < 0.01; ***P < 0.001; ns non-significant). D In situ immunohistochemistry for IL-8 expression in microabscesses of cows with neurolisteriosis. a Early infection site, not yet developed into a microabscess, cervical spinal cord. IL-8 is expressed by a mononuclear cell morphologically compatible with microglia (inset). Notice how early recruited PMN, recognizable by their multilobed nuclei (arrows), do not yet express IL-8 at this stage. b Acute microabscess, cervical spinal cord. Predominant IL-8 expression by PMN gathering at the microabscess center (inset). c Chronic microabscess, midbrain. Mononuclear phagocytes (likely infiltrating monocyte-derived macrophages) have replaced PMN and lack IL-8 expression. E, F In situ IF for IL-8 expression in cows with neurolisteriosis. E Acute microabscess in the thalamus, mostly composed of microglia (P2RY12-positive cells, red) interspersed with Lm colonies (green), in the proximity of a perivascular cuff (yellow asterisk). Expression of IL-8 (white) by one microglia containing intracellular Lm (yellow arrow) is visible on higher magnification. F Periphery of an acute microabscess in the medulla oblongata, prevalently composed of ramified Iba1+ cells morphologically compatible with microglia (red). IL-8 expression (white) by one of such cells (yellow arrow) can be seen on higher magnification

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