Basic fibroblast growth factor inhibits basal and transforming growth factor-beta induced collagen alpha 2(I) gene expression in scleroderma and normal fibroblasts.
OBJECTIVE
Studies have shown that scleroderma (systemic sclerosis, SSc) and normal fibroblasts respond differently to basic fibroblast growth factor (bFGF), SSc fibroblasts being less responsive than normal fibroblasts in mitogenic assays in vitro, bFGF also stimulates the expression of platelet derived growth factor-alpha (PDGF-alpha) receptors in normal fibroblasts, but not in SSc fibroblasts. Conversely, transforming growth factor-beta (TGF-beta) stimulates PDGF-alpha receptor expression in SSc fibroblasts, but not in normal fibroblasts. Since bFGF has been shown to inhibit collagen gene expression in several cell types, we examined responses of SSc and normal fibroblasts to bFGF alone and in combination with TGF-beta with regard to collagen alpha 2(I) (COL1A2) expression.


METHODS
Fibroblasts were obtained by skin biopsy from affected areas of patients with diffuse cutaneous SSc and from healthy donors and propagated in vitro. The effects of bFGF and TGF-beta on the COL1A2 mRNA expression levels in SSc and healthy fibroblasts were analyzed by Northern blot. The effects of bFGF on the COL1A2 promoter activities in both cell types were analyzed by transient transfection assays. The effects of bFGF and TGF-beta on collagen protein synthesis were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography.


RESULTS
While bFGF diminished COL1A2 mRNA in both SSc and normal cells, COL1A2 mRNA quantities in the SSc fibroblasts were not depressed to the levels expressed by normal controls. As anticipated, TGF-beta strongly induced COL1A2 mRNA levels in normal fibroblasts, and to a lesser degree in SSc fibroblasts. When cells were incubated with both TGF-beta and bFGF, the stimulatory effect of TGF-beta was completely suppressed in both cell types. bFGF decreased COL1A2 promoter activity in both cell types, suggesting that COL1A2 inhibition by bFGF occurs at least partially at the transcriptional level. The effects of bFGF and TGF-beta on the collagen protein synthesis correlated well with mRNA data, in that TGF-beta stimulated, while bFGF strongly inhibited, collagen synthesis.


CONCLUSIONS
bFGF is a potent inhibitor of basal and TGF-beta stimulated collagen expression in human fibroblasts, and this effect is not different between SSc and healthy fibroblasts.
Ichiki
Y
Y
Department of Medicine, Medical University of South Carolina, Charleston 29425-2229, USA.
Smith
E A
EA
LeRoy
E C
EC
Trojanowska
M
M
D003094:Collagen / Q000096:biosynthesis / Q000235:genetics*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D005347:Fibroblasts / Q000378:metabolism*; D005353:Fibronectins / Q000096:biosynthesis; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D012595:Scleroderma, Systemic / Q000473:pathology*; D012967:Sodium Dodecyl Sulfate; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors
D005353:Fibronectins; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D016222:Fibroblast Growth Factor 2; D012967:Sodium Dodecyl Sulfate; D003094:Collagen