The corresponding values for EGCg treatment with given or without H2O2 were not significantly different from controls. Glycogen synthase kinase 3B is a key component of multiple signalling pathways involved in the regula tion of cell fate, protein synthesis, glycogen metabolism, cell mobility, proliferation, and survival. By preventing cells from entering the cell cycle, GSK 3B participates in the regulation of the B catenin signalling pathway by modulating cyclin D1 expression levels. For cells treated with 400 uM H2O2, phosphorylation of GSK 3B was increased by 20%, whereas EGCg pre treatment with or without H2O2 attenuated both total and phosphorylated GSK 3B levels in cells. In addition, H2O2 decreased Inhibitors,Modulators,Libraries B catenin and cyclin D1 expression levels, which may cause the subsequent cell cycle arrest at the G1 S phase.

This H2O2 induced inhibition of the B catenin/cyclin D1 signalling pathway could be efficiently prevented Inhibitors,Modulators,Libraries by pre treatment with 20 uM of EGCg or 10 uM of SB 216763, an inhibitor of GSK 3/3B. EGCg induced fluorescence changes in intact Triton X 100 soluble and insoluble fractions of EGFP expressing H9c2 cells To investigate the role of EGCg mediated transmembrane signalling in cardioprotection, EGFP was ectopically expressed in H9c2 cells. Fluorescence spectroscopy indicated that increases in EGCg concentrations from 0 to 100 uM caused dose dependent decreases in EGFP fluorescence. In addition, this experimental approach allowed us to monitor the fluorescence changes as a means to distinguish the effects of Triton X 100 soluble and in soluble compartments on cell membrane.

To identify the protein complexes conjugated to EGFP in EGFP expressing cells, Inhibitors,Modulators,Libraries a co immunoprecipitation assay using a specific antibody against EGFP was used for molecular identification, followed by immunoblotting or MS spectra. The EGFP co immunoprecipitated proteins separated by one dimensional SDS PAGE on cell membranes. Cytoskeletal proteins including B actin, myosin IX, and vimentin may form complexes with other protein in cel lular Triton X 100 resistant microdomains, which may play a role in EGCg transmembrane signalling in cardiac cells. Effects of H2O2 and EGCg on the expression of Cavs in H9c2 cells H9c2 cells expressed mRNA encoding Cav 1, Cav 2 and Cav 3, with a dominant Cav 1 expression.

Exposure Inhibitors,Modulators,Libraries to 400 uM H2O2 with or without 20 uM EGCg pre treatment did not show significant effects on mRNA expression for any isoform as compared to controls. Western blot analysis indicated Inhibitors,Modulators,Libraries that protein levels of Cav 3 in whole cell lysates were not significantly changed by H2O2 and/or EGCg. H2O2 induced a 30% decrease concerning in the levels of Cav 1 concomitant with a 20% decrease in phosphorylated Cav 1. These decreases were abrogated by pre treatment with 10 or 20 uM EGCg for 30 min. For cells with EGCg treatment for 30 min, the levels of Cav 1 and phosphorylated Cav 1 were increased by 12% and 15%, respectively.