Uitto J, Bauer E A, Eisen A Z
J Clin Invest. 1979 Oct;64(4):921-30. doi: 10.1172/JCI109558.
To assess potential abnormalities in collagen metabolism in systemic scleroderma, skin fibroblast lines from patients with this disease were established and compared to control cell lines derived from healthy subjects. For studies on the biosynthesis of procollagen, the cells were incubated with [(14)C]proline in a medium supplemented with ascorbic acid and beta-aminopropionitrile, and the synthesis of nondialyzable [(14)C]hydroxyproline, in relation to DNA or cell protein, was taken as an index of procollagen formation. Five of eight scleroderma fibroblast cell lines demonstrated procollagen biosynthesis rates significantly higher than the controls, and the mean rate of procollagen synthesis by scleroderma fibroblasts was about twice that of the control cells. Control experiments demonstrated that the specific activity of the intracellular free proline was not different in scleroderma and control fibroblasts, and the mean population doubling times of the scleroderma and the control fibroblast cell lines were the same. The relative synthesis of the genetically distinct procollagens was examined by isolating type I and type III procollagens from the cell culture medium using DEAE-cellulose chromatography. The ratios of type I/III procollagens in scleroderma cell lines did not differ from the controls. The helical stability of the collagenous portion of type I and type III procollagens, estimated by the resistance of (14)C-collagen to limited proteolytic digestion with pepsin under nondenaturing conditions, was the same in both scleroderma and control cultures. The capacity of the cells to synthesize enzymatically active and immunologically reacting collagenase was also studied; no marked differences in these parameters could be observed. The results suggest that cultured skin fibroblasts from patients with scleroderma demonstrate a metabolic abnormality expressed as increased synthesis of type I and type III procollagens in a normal ratio. This abnormality may play a role in the excessive accumulation of collagen in the skin and other organs affected in scleroderma.
为评估系统性硬化症中胶原蛋白代谢的潜在异常，建立了该病患者的皮肤成纤维细胞系，并与源自健康受试者的对照细胞系进行比较。为研究前胶原的生物合成，将细胞在补充了抗坏血酸和β-氨基丙腈的培养基中与[¹⁴C]脯氨酸一起孵育，相对于DNA或细胞蛋白的不可透析的[¹⁴C]羟脯氨酸的合成被用作前胶原形成的指标。八个硬皮病成纤维细胞系中的五个显示前胶原生物合成率显著高于对照组，硬皮病成纤维细胞的前胶原合成平均速率约为对照细胞的两倍。对照实验表明，硬皮病和对照成纤维细胞中细胞内游离脯氨酸的比活性没有差异，硬皮病和成纤维细胞系的平均群体倍增时间相同。通过使用DEAE-纤维素色谱法从细胞培养基中分离I型和III型前胶原，研究了遗传上不同的前胶原的相对合成。硬皮病细胞系中I/III型前胶原的比例与对照组没有差异。在非变性条件下，通过¹⁴C-胶原对胃蛋白酶有限蛋白水解的抗性估计的I型和III型前胶原胶原部分的螺旋稳定性在硬皮病和对照培养物中是相同的。还研究了细胞合成具有酶活性和免疫反应性胶原酶的能力；在这些参数中未观察到明显差异。结果表明，硬皮病患者培养的皮肤成纤维细胞表现出一种代谢异常，表现为I型和III型前胶原以正常比例合成增加。这种异常可能在硬皮病中受影响的皮肤和其他器官中胶原蛋白的过度积累中起作用。
