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20 April 2020 : Clinical Research  

The Effects of the Transforming Growth Factor-β1 (TGF-β1) Signaling Pathway on Cell Proliferation and Cell Migration are Mediated by Ubiquitin Specific Protease 4 (USP4) in Hypertrophic Scar Tissue and Primary Fibroblast Cultures

Yong Huang1AEG, Yuting Wang1AEG, Xueming Wang1BC, Lixin Lin1DF, Peng Wang1CF, Junjun Sun1BE, Lei Jiang1DF*

DOI: 10.12659/MSM.920736

Med Sci Monit 2020; 26:e920736

Abstract

BACKGROUND: Hypertrophic scar results from an abnormal repair response to trauma in the skin and involves fibroblasts proliferation with increased collagen deposition. Transforming growth factor-β1 (TGF-β1) and TGF-β receptor type I (TGF-βR1) are involved in tissue repair and are increased by ubiquitin-specific protease 4 (USP4). This study aimed to investigate the effects of TGF-βR1 and USP4 in human tissue samples of hypertrophic scar and on cell proliferation and cell migration in primary fibroblast cultures in vitro.

MATERIAL AND METHODS: Skin excision tissue samples with adjacent normal skin were obtained from 15 patients with hypertrophic scar, which provided tissue sections and primary fibroblast culture for analysis. Immunohistochemistry detected the expression of USP4 and TGF-βR1 in tissue sections. MicroRNA (miRNAs) expression levels were measured by quantitative real-time polymerase chain reaction (qRT-PCR). Western blot was performed to measure protein expression levels. Cultured skin fibroblasts were investigated using immunofluorescence staining. Fibroblast proliferation, apoptosis, and migration were measured with the Cell Counting Kit-8 (CCK-8) assay, flow cytometry, and a wound-healing assay, respectively.

RESULTS: The expression of USP4 and TGF-βR1 in hypertrophic scar were increased compared with normal skin. Fibroblasts cultured from hypertrophic scar tissue showed increased expression of of USP4 and TGF-βR1. Fibroblast transfection with USP4 short-interfering RNA (siRNA) resulted in reduced fibroblast proliferation and migration, and increased apoptosis. Downregulation of USP4 inhibited the expression of TGF-βR1 protein and increased the expression levels of Smad7 protein.

CONCLUSIONS: USP4 regulated the proliferation, migration, and apoptosis of hypertrophic scar fibroblasts by regulating the TGF-β1 signaling pathway.

Keywords: Cicatrix, Hypertrophic, Protease Inhibitors, Transforming Growth Factor beta, Collagen, Immunohistochemistry, primary cell culture, RNA, Messenger, Receptor, Transforming Growth Factor-beta Type I, Skin, Ubiquitin-Specific Proteases

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750