Fig. 2
From: The pathogenic role of c-Kit+ mast cells in the spinal motor neuron-vascular niche in ALS
Mast cells accumulate in the surrounding of motor neurons in the spinal cord of SOD1G93A mice. A Representative confocal immunohistochemical images showing the co-expression of c-Kit+/chymase+ (left upper panels), and chymase+/Cox-2+ (right upper panels) mast cells in the surroundings of degenerating motor neurons (white dotted lines) in the ventral horn of the symptomatic SOD1G93A lumbar spinal cord. Mast cells display two phenotypes, typically bearing granular morphology with images of explosive degranulation (white arrows) and small rounded cells with few or no granules (white arrowheads). Magenta squares show high magnification 3D reconstructions of both, small non-granular cells, and mast cells showing an irregular shape corresponding to an explosive degranulating state (white arrows). Lower panels to the left show representative confocal images of mast cell markers staining in non-transgenic littermates, where only small cells with non-apparent granules or degranulation were observed (magenta squares). The graphs to the right show the quantitative analysis of chymase+/c-Kit+ and chymase+/Cox-2+ mast cells in the ventral horn of the lumbar spinal cord of SOD1G93A symptomatic mice compared to Non-Tg littermates. B Flow cytometry analysis of the c-Kit+ cell populations of the spinal cord of Non-Tg and SOD1G93A symptomatic mice (150d). The images to the left show the representative density plots of c-Kit expression. The graph to the right shows the quantitative analysis between conditions. Note that a statistically significant increase of c-kit expression is observed in symptomatic ALS mice when compared with controls. C Representative confocal images analyzing cells that co-express c-Kit (red) and CD45 (green) in the surroundings of motor neurons (white dotted lines) in both Non-Tg (left panel) and symptomatic SOD1G93A littermates (right panel). The magenta squares show a high magnification analysis of c-Kit+/CD45+ cells in both conditions. Note the presence of few granules in small cells in the symptomatic condition, while significantly fewer smaller cells with no granules are observed in Non-Tg mice. D Flow cytometry analysis of c-Kit+/CD45+ cell population in the spinal cord of Non-Tg and symptomatic SOD1G93A mice. Upper panels show representative density plots of CD45 expression, where a significant increase is observed in the ALS condition. The lower panels show representative density plots of c-Kit expression within the CD45 population previously analyzed. Graphs to the right show the quantitative analysis for CD45 and CD45/c-Kit respectively. Quantitative data are expressed as mean ± s.e.m. Data were analyzed by Kruskal–Wallis followed by Dunn´s multiple comparison test (A) and two-tailed unpaired t-test (B, D, E), with **p < 0,01, ***p < 0.001 and ****p < 0.0001 considered significant. n = 4 animals/condition. Scale bars: 20 μm and 5 μm in (A) in low and high magnification respectively, 10 μm and 5 μm in (C), in low and high magnification respectively