Increased transcriptional activities of transforming growth factor beta receptors in scleroderma fibroblasts.
OBJECTIVE
To investigate the molecular mechanism of the overexpression of transforming growth factor beta receptors (TGF(beta)Rs) in dermal fibroblasts from patients with systemic sclerosis (SSc).


METHODS
Dermal fibroblasts from 7 patients with diffuse SSc of recent onset and from 7 healthy individuals were studied. The expression of TGF(beta)R type I (TGF(beta)RI), TGF(beta)RII, and type I collagen proteins in dermal fibroblasts was determined by immunoblotting. TGF(beta)RI, TGF(beta)RII, and alpha2(I) collagen messenger RNA (mRNA) were evaluated by Northern blot analysis. The transcriptional activities of the TGF(beta)RI and TGF(beta)RII genes were examined by luciferase assay.


RESULTS
SSc fibroblasts expressed increased levels of TGF(beta)RI and TGF(beta)RII protein and mRNA, as well as increased levels of type I collagen protein and alpha2(I) collagen mRNA. Moreover, the half-lives of TGF(beta)RI and TGF(beta)RII mRNA in SSc fibroblasts did not change compared with those in control dermal fibroblasts. The promoter activities of the TGF(beta)RI and TGF(beta)RII genes were both significantly increased in SSc fibroblasts compared with those in control fibroblasts. Calphostin C, a specific inhibitor of protein kinase C (PKC), inhibited TGF(beta)RI promoter activity in SSc fibroblasts, and LY294002, an inhibitor of phosphoinositide 3-kinase (PI 3-kinase), inhibited TGF(beta)RII promoter activity in SSc fibroblasts. Moreover, calphostin C and LY294002 inhibited the up-regulation of TGF(beta)RI and TGF(beta)RII mRNA, respectively, in SSc fibroblasts.


CONCLUSIONS
These results suggest that increased levels of TGF(beta)Rs in SSc fibroblasts play a role in excessive collagen production, and that up-regulation of TGF(beta)R expression might occur at the transcriptional levels. PKC and/or PI 3-kinase might contribute to the up-regulation of TGF(beta)R expression in SSc fibroblasts.
D030201:Activin Receptors, Type I / Q000378:metabolism; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D024042:Collagen Type I / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000502:physiology*; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D009025:Morpholines / Q000494:pharmacology; D009281:Naphthalenes / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D012595:Scleroderma, Systemic / Q000235:genetics* / Q000378:metabolism; D012867:Skin / Q000378:metabolism; D014158:Transcription, Genetic; D015854:Up-Regulation
D002867:Chromones; D024042:Collagen Type I; D004791:Enzyme Inhibitors; D005419:Flavonoids; D009025:Morpholines; D009281:Naphthalenes; D018125:Receptors, Transforming Growth Factor beta; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D017346:Protein Serine-Threonine Kinases; D030201:Activin Receptors, Type I; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II; C058819:calphostin C; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one