Characterization of the phenotype of high collagen-producing fibroblast clones in systemic sclerosis, using a new modified limiting-dilution method

Clinical and Experimental Dermatology
L ZhuM Li

Abstract

Overproduction of type I collagen in fibroblasts of systemic sclerosis (SSc) is the hallmark of fibrosis. Establishment and characterization of the phenotype of SSc fibroblasts has been hindered by the heterogeneity between fibroblasts and the lack of adequate cloning methods. To establish and investigate the characteristics of the SSc high collagen-producing fibroblast phenotype. Primary cultured fibroblasts from skin biopsies of patients with SSc and normal controls were cloned by a new modified limiting-dilution method. All clones were divided into different subpopulations based on their α1(I) procollagen (COL1A1) mRNA level detected by real-time reverse transcriptase PCR assay. In the different subpopulations, cell growth and cycle distribution were analysed by MTT and flow cytometry, COL1A1 promoter activity was examined by transient transfection, and the binding activity of Sp1 to the COL1A1 proximal promoter was investigated by quantitative chromatin immunoprecipitation. The clonogenicities of SSc and normal control fibroblasts were similar, but the mean COL1A1 mRNA level of clones and the percentage of the subpopulation with a high COL1A1 mRNA level were significantly higher in SSc fibroblasts than in controls. There wa...Continue Reading

References

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Citations

Mar 16, 2019·Non-coding RNA·Mousa G Eissa, Carol M Artlett
Jul 17, 2013·Experimental Biology and Medicine·Asish K GhoshDouglas E Vaughan

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