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Fig. 3 | Acta Neuropathologica Communications

Fig. 3

From: The pathogenic role of c-Kit+ mast cells in the spinal motor neuron-vascular niche in ALS

Fig. 3

Ex vivo generation of c-Kit+ mast cells from the spinal cord of ALS rats. A The scheme shows the procedure followed to obtain primary cultures of mast cells from the adult spinal cord of symptomatic SOD1G93A rats and Non-Tg littermates. Spinal cords were maintained in the presence of IL-3 (20 ng/mL) and SCF (20 ng/mL) for 2, 7, and 14 days in vitro (DIV), and then mast cell markers, c-Kit, chymase, Cox-2, and CD45 were analyzed by flow cytometry and immunocytochemistry. B Flow cytometry analysis of c-Kit+ cultured mast cells after 2 and 7 DIV. Representative density plots show the expression of c-Kit at 2 and 7 days. The graph to the right shows the quantitative analysis of c-Kit expression in cells cultured from Non-Tg and SOD1G93A rats. C Representative cytological and confocal immunohistochemical images of mast cells isolated from symptomatic SOD1G93A spinal cord. Left panels show representative bright-field images of non-adherent cells with showing typical granular mast cells (upper panel) and metachromatic granular mast cells stained with toluidine blue (lower panel). Panels to the right show immunohistochemical phenotyping by confocal microscopy of cultured mast cells expressing typical markers such as c-Kit, CD45, chymase, and Cox-2 after 14 days in culture. Quantitative data are expressed as mean ± s.e.m. Data were analyzed by the Mann–Whitney test with *p < 0,05 considered significant. n = 4 animals/condition. Scale bars: 5 μm

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