Mannose-6-Phosphate/IGF-II Receptors Mediate the Effects of IGF-1-Induced Latent Transforming Growth Factor β1 on Expression of Type I Collagen and Collagenase in Dermal Fibroblasts

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Abstract
We have previously shown that insulin-like growth factor-1 (IGF-1) induces the expression of latent transforming growth factor β1 (LTGF-β1) through activation of c-fos and c-jun oncogenes. In this study we investigated whether IGF-1 induced latent TGF-β1 has autocrine effects on dermal fibroblasts and described a possible mechanism. Human dermal fibroblasts were treated with either vehicle, IGF-1 alone, or IGF-1 with either anti-TGF-β1 neutralizing antibody or mannose-6-phosphate (M6P) and levels of mRNAs for TGF-β1, collagenase and the pro α1(I) chain of type I collagen were then evaluated by Northern analysis. Conditioned medium was also collected from treated and untreated cells and assayed for TGF-β1 protein by enzyme-linked immunosorbent assay. The results of the Northern analysis revealed a differential effect on the expression of the pro α1(I) chain of type I collagen and collagenase in dermal fibroblasts: mRNA for the former being significantly increased in response to IGF-1 treatment while that for collagenase was markedly suppressed. These effects of IGF-1 were blocked to a significant extent by TGF-β1 neutralizing antibody at a concentration of 0.5–2.0 μg/ml. As the TGF-β1 induced by IGF-1 is inactive in the traditionally used mink lung epithelial cell growth inhibition assay, we explored the possible role of IGF-II/M6P receptors in facilitating these autocrine effects. The results showed that the greater than two-fold increase (201.9 ± 38 vs 81.8 ± 13, p < 0.05) in mRNA for the pro α1(I) chain of type I collagen induced by IGF-1 was at least 60% inhibited by M6P in a time-dependent fashion. A direct correlation between the expression of TGF-β1 and the pro α1(I) chain of type I collagen was found in response to either IGF-1 alone or IGF-1 with M6P. Treatment of cell cultures with TGF-β1 neutralizing antibody mimicked the effect of M6P. In contrast to the effects on expression of type I collagen, the level of collagenase mRNA was markedly reduced by IGF-1 alone and was restored by the administration of M6P. The levels of TGF-β1 in conditioned medium from treated and untreated cells showed a similar pattern to that of the mRNA detected by Northern analysis. These findings suggest that IGF-1 induces latent TGF-β1 and that the matrix-modulating autocrine effects of LTGF-β1 on dermal fibroblasts are facilitated by M6P/IGF-II receptors on these cells.

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