CYTOKINES MODULATE THE SENSITIVITY OF HUMAN FIBROBLASTS TO STIMULATION WITH INSULIN-LIKE GROWTH FACTOR-I (IGF-I) BY ALTERING ENDOGENOUS IGF-BINDING PROTEIN-PRODUCTION

Human dermal fibroblasts produce a number of insulin-like growth factor-binding proteins (IGFBPs) including the main circulating form, IGFBP-3. It has been suggested that the regulation of IGFBP secretion may play a major role in modulating insulin-like growth factor (IGF) bioactivity. We have quantified the effects of two cytokines, transforming growth factor-beta1 (TGF-beta1) and tumour necrosis factor-alpha (TNF-alpha) which have opposing actions on fibroblast IGFBP-3 production, and examined their subsequent role in IGF-I mitogenesis. TGF-beta1 caused a dose-dependent increase in IGFBP-3 in serum-free fibroblast-conditioned media. TGF-beta1 (I mug/1) resulted in immunoreactive IGFBP-3 levels reaching 286.5 +/-22.4% of control after 20 h, the increase being confirmed by Western ligand blot. TNF-alpha caused a dose-dependent decrease in fibroblast IGFBP-3 secretion, 1 mug TNF-alpha/I reducing IGFBP-3 levels to 32.1 +/-11.% of control. This effect was not due to cytotoxicity and was not cell-density-dependent. Fibroblast proliferation was examined using a 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MT-colorimetric cytochemical bioassay. The addition of IGF-I resulted in dose-dependent growth stimulation after 48 h, the effective range being 20-100 mug/l. The IGF-I analogue Long-R3-IGF-I which has little affinity for the IGFBPs was approximately 20-fold more potent in this assay, and was unaffected by exogenous IGFBP-3. While the addition of 1 mug TGF-beta1/1 to increasing doses of IGF-I resulted in a fourfold decrease in mitogenic sensitivity to the IGF-I, no such effect was seen with Long-R3-IGF-I. Conversely, 1 mug TNF-alpha/1 increased fibroblast IGF-I sensitivity five-fold, an effect not observed with the IGF-I analogue. Such data suggest that endogenous IGFBP-3 inhibits IGF-I bioactivity and that the regulation of IGF mitogenesis by TGF-beta1 and TNF-alpha can occur via local IGFBP modulation. This may represent a mechanism by which complex growth signals are co-ordinated in vivo.