title: Inhibition of Smad3 expression decreases collagen synthesis in keloid disease fibroblasts.
abstract: BACKGROUND
  Keloids represent a dysregulated response to cutaneous wounding that result in an excessive deposition of extracellular matrix (ECM), especially types I and III collagen. In keloid scars, the ratio of 'type I to type III collagen' varies compared to normal skin. Transforming growth factor beta (TGF-beta) plays a central role in the pathogenesis of fibrosis by inducing and sustaining activation of keloid fibroblasts. However, the underlying mechanisms are poorly understood. RNA interference (RNAi) is an evolutionally conserved mechanism for repressing targeted gene expression. In mammalian cells, RNAi is mediated by small interfering RNA (siRNA). In this paper, we examined the function of Sma and Drosophila mothers against decapentaplegic homolog 3 (Smad3), recently characterized as intracellular effector of TGF-beta signalling, in keloid fibroblasts using siRNA.
  
  
  METHODS
  Dermal fibroblasts obtained from one female patient aged 21 years were maintained in Dulbecco's modified Eagle's medium. Cells (<6 passages) were treated with or without Smad3 siRNA and the expression levels of related genes were examined by Reverse Transcription Polymerase Chain Reaction (RT-PCR) and Western Blot. Statistical analysis was performed using one-way ANOVA (Dunnett correction) and the Excel 7.0 software, with significance set at p<0.05.
  
  
  RESULTS
  The knockdown of Smad3 expression in mRNA and protein levels was confirmed using RT-PCR and Western Blot. Compared to blank, the mRNA levels of types I and III procollagen were also significantly and uniquely decreased following the reduction of Smad3 by siRNA (p<0.05). The results indicate that Smad3 plays an important role in the TGF-beta induced fibrosis in keloid. Downregulation of Smad3 expression in keloid fibroblasts can significantly decrease procollagen gene expression. SiRNA targeting Smad3 was an efficient reagent to reduce ECM deposition and attenuate process of fibrosis. It could be a new promising therapeutic approach to improve skin wound healing and inhibit progression of fibrotic conditions by interrupting the TGF-beta signal pathway.
authors: Department of Orthopaedics, Changhai Hospital, Second Military Medical University, Shanghai 200433, China. wzmsmmu@yahoo.com
authors: Zimin
authors: Z
authors: Wang
authors: Zhongyu
authors: Z
authors: Gao
authors: Yi
authors: Y
authors: Shi
authors: Yi
authors: Y
authors: Sun
authors: Zihao
authors: Z
authors: Lin
authors: Hua
authors: H
authors: Jiang
authors: Tiesheng
authors: T
authors: Hou
authors: Qiugen
authors: Q
authors: Wang
authors: Xiangbin
authors: X
authors: Yuan
authors: Xiaohai
authors: X
authors: Zhu
authors: Hong
authors: H
authors: Wu
authors: Youxin
authors: Y
authors: Jin
mesh_terms: D000328:Adult; D002478:Cells, Cultured; D024042:Collagen Type I / Q000096:biosynthesis*; D024061:Collagen Type III / Q000096:biosynthesis*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D007627:Keloid / Q000235:genetics* / Q000473:pathology; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051900:Smad3 Protein / Q000235:genetics* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000502:physiology
chemical_list: D024042:Collagen Type I; D024061:Collagen Type III; D034741:RNA, Small Interfering; C497790:SMAD3 protein, human; D051900:Smad3 Protein; D016212:Transforming Growth Factor beta