title: RNA interference against platelet-derived growth factor receptor α mRNA inhibits fibroblast transdifferentiation in skin lesions of patients with systemic sclerosis.
abstract: OBJECTIVE
  To down-regulate expression of mRNA for the platelet-derived growth factor receptor (PDGFR)-α, block the signalling pathway of PDGF and its receptor, and study their influence on fibroblast transdifferentiation to myofibroblasts in systemic sclerosis (SSc).
  
  
  METHODS
  Fibroblasts from skin lesions of SSc patients and health adult controls were cultured in vitro, and α-smooth muscle actin (α-SMA) expression was determined by immunocytochemistry. Both groups of fibroblasts were stimulated with PDGF-AA, transforming growth factor β1 (TGF-β1), and costimulated with PDGF-AA and TGF-β1, then PDGFR-α and α-SMA mRNA and protein expression were detected with RT-PCR and WB respectively. Three pairs of siRNAs targeting different PDGFR-α mRNA sequences were synthesized for RNAi. SSc and control fibroblasts were transfected with PDGFR-α siRNA; stimulated with PDGF-AA; and assessed for PDGFR-α and α-SMA mRNA and protein expression.
  
  
  RESULTS
  Although the fibroblasts from both groups had similar morphology, the SSc skin lesions had significantly more myofibroblasts than control skin lesions. PDGF-AA stimulation, TGF-β1 stimulation, and costimulation significantly up-regulated PDGFR-α and α-SMA mRNA and protein expression in SSc fibroblasts compared to control (P<0.05), and costimulation had the strongest effects (P<0.05). All three pairs of siRNAs suppressed PDGFR-α mRNA and protein expression (P<0.05), but siRNA1495 had the highest gene-silencing efficiency (P<0.05). PDGFR-α siRNA attenuated the effects of PDGF-AA through up-regulating PDGFR-α and α-SMA mRNA and protein expression and inhibiting fibroblast transdifferentiation to myofibroblasts in SSc (P<0.05).
  
  
  CONCLUSIONS
  PDGFR-α over-expression in SSc fibroblasts bound PDGF-AA more efficiently and promoted fibroblast transdifferentiation, which was enhanced by TGF-β1. PDGFR-α siRNA down-regulated PDGFR-α expression, blocked binding to PDGF-AA, and inhibited fibroblast transdifferentiation to myofibroblasts.
authors.affiliation: Department of Dermatology, First Affiliated Hospital, College of Medicine, Xi'an Jiaotong University, Xìan City, Shaanxi Province, China. tonyl1220@sina.com
authors.forename: Tong
authors.initials: T
authors.lastname: Liu
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Xin
authors.initials: X
authors.lastname: Mu
authors.forename: Hui
authors.initials: H
authors.lastname: Su
authors.forename: Xiaoding
authors.initials: X
authors.lastname: Hu
authors.forename: Wenli
authors.initials: W
authors.lastname: Liu
authors.forename: Enbing
authors.initials: E
authors.lastname: Zhao
authors.forename: Weimin
authors.initials: W
authors.lastname: Li
mesh_terms: D000199:Actins / Q000235:genetics; D000328:Adult; D054338:Cell Transdifferentiation / Q000187:drug effects / Q000235:genetics*; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D006801:Humans; D008024:Ligands; D008297:Male; D008875:Middle Aged; D058628:Myofibroblasts / Q000187:drug effects / Q000473:pathology; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000172:deficiency* / Q000235:genetics*; D012595:Scleroderma, Systemic / Q000235:genetics* / Q000473:pathology*; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology
chemical_list: C541116:ACTA2 protein, human; D000199:Actins; D008024:Ligands; D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D053773:Transforming Growth Factor beta1; C075278:platelet-derived growth factor A; D020796:Receptor, Platelet-Derived Growth Factor alpha