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Dimethylsulfoxide (DMSO), a universal solvent, is frequently used to dissolve various classes of chemicals for the evaluation of their biological activities. In one such evaluation, we noticed that DMSO itself caused cellular proliferation and interfered with high affinity IgE receptor (FcepsilonRI)-mediated degranulation of mast cells. DMSO caused cellular proliferation of RBL-2H3 cells by phosphorylating both extracellular-signal regulated kinase (ERK) and M2-type pyruvate kinase (M2PK) through which the enzymatic activity of M2PK was reduced. Allergenic activation of FcepsilonRI caused the tyrosine phosphorylations of signaling components of FcepsilonRI, such as Syk, PLCgamma1, PLCgamma2, ERK, and M2PK. In these allergenic activated RBL-2H3 cells, DMSO specifically inhibited FcepsilonRI-mediated tyrosine phosphorylation of M2PK, blocked FcepsilonRI-mediated inhibition of the enzymatic activity of M2PK, and then inhibited FcepsilonRI-mediated degranulation. These results suggest that DMSO causes cellular proliferation and mast cell degranulation through differential modulation of M2PK in resting and allergenic activated cells.

Background and Purpose: M2-type pyruvate kinase (M2PK) was found to interact directly with the 'ITAM' region of the gamma chain of the high-affinity IgE receptor (FcvarepsilonRI). Our hypothesis was that mast cell degranulation might require the FcvarepsilonRI-mediated inhibition of M2PK activity., Experimental Approach: In rat basophilic leukaemia (RBL-2H3) cells, the effects of directly inhibiting M2PK or preventing the FcvarepsilonRI-mediated inhibition of M2PK (disinhibition) on degranulation was measured by hexosaminidase release. Effects of blocking the FcvarepsilonRI-mediated inhibition of M2PK was also assessed in vivo in a mouse model of allergen-induced airway hyper-responsiveness., Key Results: Activation of FcvarepsilonRI in RBL-2H3 cells caused the rapid phosphorylation of tyrosine residues in M2PK, associated with a decrease in M2PK enzymatic activity. There was an inverse correlation between M2PK activity and mast cell degranulation. FcvarepsilonRI-mediated inhibition of M2PK involved Src kinase, phosphatidylinositol 3-kinase, PKC and calcium. Direct inhibition of M2PK potentiated FcvarepsilonRI-mediated degranulation and prevention of the FcvarepsilonRI-mediated inhibition of M2PK attenuated mast cell degranulation. Transfection of RBL-2H3 cells with M1PK which prevents FcvarepsilonRI-induced inhibition of M2PK, markedly reduced their degranulation and exogenous M1PK (i.p.) inhibited ovalbumin-induced airway hyper-responsiveness in vivo., Conclusions and Implications: We have identified a new control point and a novel biochemical pathway in the process of mast cell degranulation. Our study suggests that the FcvarepsilonRI-mediated inhibition of M2PK is a crucial step in responses to allergens. Moreover, the manipulation of glycolytic processes and intermediates could provide novel strategies for the treatment of allergic diseases.

The molecular mechanism of how resveratrol inhibits mast cell degranulation was studied by examining its effects on the signaling components of the high affinity IgE receptor (FcepsilonRI) pathway. Resveratrol inhibited mast cell degranulation in a dose-dependent manner and reduced the FcepsilonRI-mediated tyrosine phosphorylation of ERK and PLCgamma1 but not of Syk and PLCgamma2. U-73 122 and PD98059, which are PLC and MEK inhibitors, also had inhibitory effects on mast cell degranulation. These results suggest that FcepsilonRI-mediated tyrosine phosphorylation of PLCgamma1 and ERK could be potential cellular targets of resveratrol for the inhibition of mast cell degranulation.