Ichiki Y, Smith E A, LeRoy E C, Trojanowska M
Department of Medicine, Medical University of South Carolina, Charleston 29425-2229, USA.
J Rheumatol. 1997 Jan;24(1):90-5.
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.
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.
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.
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.
研究表明，硬皮病（系统性硬化症，SSc）成纤维细胞与正常成纤维细胞对碱性成纤维细胞生长因子（bFGF）的反应不同，在体外促有丝分裂试验中，SSc成纤维细胞的反应性低于正常成纤维细胞。bFGF还能刺激正常成纤维细胞中血小板衍生生长因子α（PDGF-α）受体的表达，但对SSc成纤维细胞无此作用。相反，转化生长因子β（TGF-β）能刺激SSc成纤维细胞中PDGF-α受体的表达，但对正常成纤维细胞无此作用。由于bFGF已被证明在几种细胞类型中可抑制胶原蛋白基因的表达，我们研究了SSc成纤维细胞和正常成纤维细胞单独及联合TGF-β对bFGF的反应，观察其胶原蛋白α2（I）（COL1A2）的表达情况。
通过皮肤活检从弥漫性皮肤型SSc患者的病变部位及健康供体获取成纤维细胞，并在体外进行培养。采用Northern印迹法分析bFGF和TGF-β对SSc成纤维细胞和健康成纤维细胞中COL1A2 mRNA表达水平的影响。通过瞬时转染试验分析bFGF对两种细胞类型中COL1A2启动子活性的影响。采用十二烷基硫酸钠聚丙烯酰胺凝胶电泳和荧光自显影法分析bFGF和TGF-β对胶原蛋白合成的影响。
虽然bFGF可使SSc细胞和正常细胞中的COL1A2 mRNA减少，但SSc成纤维细胞中COL1A2 mRNA的量并未降低至正常对照所表达的水平。正如预期的那样，TGF-β可强烈诱导正常成纤维细胞中COL1A2 mRNA水平，对SSc成纤维细胞的诱导作用较弱。当细胞同时与TGF-β和bFGF孵育时，两种细胞类型中TGF-β的刺激作用均被完全抑制。bFGF可降低两种细胞类型中COL1A2启动子的活性，这表明bFGF对COL1A2的抑制作用至少部分发生在转录水平。bFGF和TGF-β对胶原蛋白合成的影响与mRNA数据密切相关，即TGF-β刺激胶原蛋白合成，而bFGF强烈抑制胶原蛋白合成。
bFGF是人类成纤维细胞中基础及TGF-β刺激的胶原蛋白表达的有效抑制剂，且这种作用在SSc成纤维细胞和健康成纤维细胞之间无差异。
