Cross-Linking Mast Cell Specific Gangliosides Stimulates the Release of Newly Formed Lipid Mediators and Newly Synthesized Cytokines

Figure 4

Cross-linking GD1b derived gangliosides with mAbAA4 induced MAP kinase phosphorylation in mast cells. RBL-2H3 cells were either stimulated via FcεRI, where cells were sensitized with IgE anti-TNP and stimulated with DNP₄₈-HSA (50 ng/mL) or incubated with mAbAA4 (1, 2.5, 5, and 10 μg/mL) for 10 min. Total cell lysates were immunoblotted with antibodies against phospho-ERK1/2 (p-ERK1/2), ERK1/2, phospho-JNK1/2 (p-JNK1/2), JNK1/2, phospho-p38 (p-p38), p38, and α/β-tubulin (housekeeping protein) and the mean optical density of the bands was determined. Densitometry of the changes in expression and phosphorylation of proteins were corrected for α/β-tubulin. Data is expressed as the fold of nonstimulated (NS) cells. (a) Ratio of phosphorylated ERK1/2/total ERK1/2; (b) a representative blot from (a); (c) ratio of phosphorylated JNK1/2/total JNK1/2; (d) a representative blot from (c); (e) ratio of phosphorylated p38/total p38; (f) a representative blot from (e). Data is expressed as the mean ± SD of three independent experiments. between experimental samples and the nonstimulated (NS) cells. between experimental samples and FcεRI stimulated cells.