Regulation of leukocyte cytokine production by inhibitors of intracellular signalling pathways

Rapecki, Stephen Edward (2001). Regulation of leukocyte cytokine production by inhibitors of intracellular signalling pathways. PhD thesis The Open University.



The activation of leukocytes during inflammation results in the engagement of many related intracellular signalling pathways. The role of two key proteins families, phosphodiesterase (PDE) and Src kinase, has been assessed in these signalling pathways using specific inhibitors.

Inhibition of phosphodiesterase type 4 (PDE4) in lipopolysaccharide (EPS) stimulated human peripheral blood mononuclear cells (PBMC) resulted in the inhibition of TNF-a by an IL-10-independent mechanism. This is in contrast to other agents that elevate cAMP which in turn inhibit TNF-a synthesis by an IL-10-dependent mechanism. When PBMC were stimulated with EPS plus IFN-y, inhibition of PDE4 resulted in an altered effect on cytokine production, elevating IE-10 while still inhibiting TNF-a.

Inhibition of PDE4 in activated T cells only weakly inhibited cellular proliferation. In contrast, PDE4 inhibitors were potent but non-selective inhibitors of T-cell cytokine production. In contrast to studies in murine cells, inhibition of T-cell activation correlates with inhibition of binding to a high-affinity Rolipram binding site on the PDE4 protein.

Kinases from the Src family (Eck, Fyn and Eyn) have been implicated in signalling via the T-cell receptor (TCR) on lymphocytes and the high affinity IgE receptor (FceRI) on mast cells. Potent inhibitors of Src kinases blocked T-cell proliferation and cytokine generation in response to specific ligation of the TCR. However, activation of T-cells by multiple costimulatory pathways was much more resistant to Src kinase inhibition. Inhibition of Src kinase in human cord blood-derived mast cells inhibited IgE-dependent degranulation of these cells. This indicates that Src kinase inhibitors may be useful in down-regulating allergic responses. Conversely, inhibitors of PDE4 and PDE type 7 (PDE7) did not prevent mast cell degranulation.

The observation that PDE4 inhibitors can regulate monocyte cytokine generation further supports the view that these agents could have therapeutic benefit in a number of inflammatory diseases such as asthma, arthritis and inflammatory bowel disease. The failure of Src kinase inhibitors to modify T cell signalling activated by multiple pathways indicates redundancy in these signalling pathways and therefore Src kinases may not represent good targets for immunomodulation. However, a selective Src kinase inhibitor could represent a good target for an anti-allergic drug.

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