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目的在克隆水平上研究系统性硬皮病(SSc)和正常皮肤成纤维细胞(FB)胶原代谢异质性的特点。方法以皮肤克隆FB作为研究对象,采用实时荧光定量RT-PCR法检测其Ⅰ,Ⅲ型前胶原和基质金属蛋白酶(MMP-1,MMP-3)mRNA水平;采用酶联免疫吸附法检测其细胞培养上清中Ⅰ型胶原蛋白的表达,筛选出具有不同胶原合成能力的异质性克隆,分析其胶原代谢的特点。结果 SSc和正常皮肤克隆FB的胶原代谢均具有异质性,SSc患者克隆FBⅠ,Ⅲ型前胶原mRNA水平的异质性较正常人大(F=20.540,P=0.000;F=6.822,P=0.012),SSc患者Ⅰ型前胶原mRNA高表达克隆的比例高于正常人(48.0%vs14.3%);SSc和正常皮肤克隆FB细胞培养上清中Ⅰ型胶原蛋白水平与其mRNA水平均存在正相关关系(r=0.873,P=0.000;r=0.538,P=0.039)。结论 SSc和正常皮肤克隆FB的胶原代谢均具有异质性。
Objective To investigate the heterogeneity of collagen metabolism in systemic scleroderma (SSc) and normal skin fibroblasts (FB) at the clonal level. Methods Skin clone FB was used as the research object. The mRNA expression of type Ⅰ and Ⅲ procollagen and matrix metalloproteinase (MMP-1) and MMP-3 were detected by real-time fluorescence quantitative RT-PCR. Culture supernatants in the expression of type Ⅰ collagen, screened heterogeneous clones with different collagen synthesis ability, analysis of the characteristics of collagen metabolism. Results The heterogeneity of collagen metabolism was observed in SSc and normal skin clone FB. The heterogeneity of FB Ⅰ and Ⅲ procollagen mRNA expression in SSc patients was higher than that in normal controls (F = 20.540, P = 0.000; F = 6.822, P = 0.012 ). The percentage of type Ⅰ procollagen mRNA expression in SSc patients was higher than that in normal people (48.0% vs 14.3%). There was a positive correlation between the level of collagen Ⅰ and the mRNA level in SSc and normal skin clone FB cell culture supernatant Relationship (r = 0.873, P = 0.000; r = 0.538, P = 0.039). Conclusion The collagen metabolism of SSc and normal skin clone FB are heterogeneous.