Fig. 1
Misfolded SOD1 accumulation precedes cPLA2α upregulation and activation. A A representative double immunofluorescence staining of cPLA2α (red) and misfolded SOD1 (B8H10, green) proteins in the lumbar spinal cord sections of WT and mutant SOD1G93A mice during the development of the disease (3, 6 and 17 weeks). Scale bars = 500 μm (upper panel), 100 μm (lower panel) and inset 20 μm. The means ± SEM fluorescence intensity for both magnifications are presented in the bar graph as arbitrary units. Four mice for each time point and five fields for each mouse were analyzed. ***p < 0.001 compared to control mice (WT). n.s. non-significant. B A representative immunoblot analysis of cPLA2α, mutant SOD1G93A and their corresponding calreticulin protein expression in the spinal cord lysates of WT and mutant SOD1G93A mice during the development of the disease. cPLA2α protein expression was determined by dividing the intensity of each cPLA2α or mutant SOD1G93A (SOD1G93A) by the intensity of the corresponding calreticulin band after quantitation by densitometry and expressed in the bar graph as arbitrary units. The bar graphs are the means ± SE of 4 mice in each group. ***p < 0.001 significance—compared to control mice (WT). n.s. non-significant. C. Misfolded SOD1 was immunoprecipitated using anti-B8H10 antibodies from spinal cord lysates of WT and mutant SOD1G93A mice during the development of the disease. A representative immunoblot analysis of the misfolded SOD1 (IP B8H10, left) and 10% of the unbound (Unbound, right) fractions are shown. nc- negative control, immunoprecipitation without anti-B8H10 antibodies. D A representative immunofluorescence staining of phosphor-cPLA2α (p-cPLA2α) in the lumbar spinal cord sections of WT and mutant SOD1G93A mice during the development of the disease (3, 6 and 17 weeks). Scale bars = 500 μm (upper panel) and 100 μm (lower panel). The means ± SEM fluorescence intensity for both magnifications are presented in the bar graph in arbitrary units. Four mice for each time point and five fields for each mouse were analyzed. ***p < 0.001 significance compared to WT mice. n.s. non-significant