Anti-CD3-induced changes in protein kinase C isozymes expression in human CD4+ and CD8+ T lymphocytes

In order to determine whether there is a differential expression and activation of PKC isozymes between CD4+ and CD8+ T cells, peripheral blood mononuclear cells were stimulated with anti-CD3 monoclonal antibody (moAb) for various time intervals and the expression of calcium-dependent PKC isozymes (alpha, beta, gamma) and calcium-independent PKC isozymes (delta, epsilon, zeta) was analyzed with dual color flow cytometry, using anti-PKC isozyme antibodies and anti-CD4 or anti-CD8 antibodies. The basal fluorescence intensity of all PKC isozymes was comparable between CD4+ T cells and CD8+ T cells. Following activation with anti-CD3 moAb a marked increase in the fluorescence intensity of all PKC isozymes in both CD4+ and CD8+ T cells, albeit to a different extent and with different kinetics was observed. Among all PKC isozymes studied, the least striking changes were observed in PKC zeta isozyme and the most striking changes were observed in PKC-epsilon isozyme. Laser-based confocal microscopic studies confirmed that the increase in fluorescence intensity of PKC isozymes following anti-CD3 moAb stimulation, as measured by flow cytometry was accompanied by the translocation of PKC isozymes from cytosol to the plasma membrane. This study demonstrates a differential effect of anti-CD3 moAb on the expression of PKC isozymes between CD4+ and CD8+ T cells and suggests that flow cytometry can be used to study the translocation of PKC isozymes from cytosol to the plasma membrane.