CD40Ligand (CD40L) represents a strong endogenous danger signal associated with chronic inflammatory disease. CD40L induces pro-inflammatory activation of CD40 expressing antigen-presenting cells (APC) such as DCs, monocytes, B-cells and endothelial cells; however, CD40 activation alone is insufficient to induce interleukin (IL)-12p70 secretion. Using quantitative Western blotting and siRNA inhibition of protein expression we demonstrate that cytokine-induced Janus kinase (JAK)1 and/or JAK2 activation mediates a complementary second signal for the production of IL- 12p70, whereas tyrosine kinase (Tyk)2 activation has inhibitory effects. Furthermore, JAK1 activation reciprocally reduced IL-10 production. The mechanism of JAK modulation of CD40 signals in dendritic cells (DCs) involved transcriptional regulation. The Th1 and Th2 cytokines interferon (IFN)-γ and IL-4 both enhance IL-12p70 via activation of the same JAKs. However due to a weaker JAK activity, IL-4 signalling has to persist much longer than IFN-γ signalling in order to induce similar amounts of IL-12p70. This different dependence on signalling persistence may give rise to the distinct biological characteristics of Th1/Th2 cytokines and may explain the pleiotropic in vivo and in vitro effects of IL-4. Complementary CD40 and JAK activity is essential for IL-12p70 secretion by all human APC studied. This strict requirement of two complementary signals opens a new way of interfering with CD40L induced APC activation indirectly by modulating the “second signal” JAK.

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