Ligand stimulation of transfected and endogenous growth factor receptors enhances cytokine production by mast cells.

Abstract

IL-3 dependent mast cell lines produce cytokines in response to Fc receptor cross-linkage or to ionomycin. In this study we have observed that cells pre-cultured in IL-3 produce 10-100 times more cytokine after receptor cross-linkage in comparison with IL-4 pre-cultured cells. Although several hematopoetin receptors, including those for IL-3, IL-4 and EPO, do not contain tyrosine kinase domains, their occupancy with ligand causes tyrosine phosphorylation of specific cellular substrates. Therefore, the contribution of tyrosine kinase activation to the ability of an IL-3 dependent mast cell line, CFTL-15, to produce cytokines was analyzed. The CFTL-15 cells were transfected with growth factor receptors containing ligand-inducible tyrosine kinase domains (EGFR and PDGFR, and CSF-IR) or with the EPOR. All of the transfectants were able to proliferate in response to IL-3 or to their respective growth factor and to produce IL-3 in response to IgE receptor cross-linkage. Stimulation of the EGFR and PDGFR transfectants with their respective ligands resulted in the production of IL-3, IL-6, and GMCSF. Stimulation of the CSF-IR or EPOR transfectants with growth factor alone failed to induce cytokine production. However, in co-stimulation assays each of the growth factors enhanced the amount of cytokine produced in response to FC-epsilon-RI cross-linkage. The ability of these stimuli to induce tyrosine phosphorylation in the transfectants was analyzed. Fc-epsilon-RI cross-linkage in the transfectants routinely induced the tyrosine phosphorylation of 145, 86 and 72 kDa proteins, with occasional phosphorylation of 55, 52, and 40 kDa proteins. Costimulation with either EGF or PDGF of transfectants, whose Fc-epsilon-RI was cross linked, led to the enhanced phosphorylation of the 145, 86 (EGF only) and 72 kDa substrates, while co-stimulation with either EPO or CSF-1 enhanced the phosphorylation of the 145 kDa substrate. These results suggest that growth factor treatment increases the 'strength' of signals resulting from Fc-epsilon-RI cross-linkage leading to enhanced cytokine production.