title: Identification of ETFB as a candidate protein that participates in the mechanoregulation of fibroblast cell number in collagen gel culture.
abstract: BACKGROUND
  Fibroblast activation is strongly influenced by mechanical environment in the wound-healing process, especially in fibrosis. Mechanically stressed three-dimensional collagen embedded culture is a useful model representing fibroblasts in morphological as well as biochemical situations encountered during fibrosis.
  
  
  OBJECTIVE
  To find key proteins involved in reducing the number of fibroblasts during mechanical stress, we performed two-dimensional gel electrophoresis (2DE)-based differential display and siRNA-based functional screening with collagen gel culture focusing on the differences between attached and detached culture environments.
  
  
  METHODS
  Membrane extracts of fibroblasts from 1 day of attached or detached cultures were subjected to 2DE. We compared protein expression levels and identified the attached-culture-dominant proteins by MALDI-TOF-MS. Next, fibroblasts were transfected with siRNA and embedded in collagen gel. Cell number was counted after 3 days in culture.
  
  
  RESULTS
  Eight attached culture dominant proteins were identified with MALDI-TOF-MS. Transfection of siRNA against these proteins demonstrated that electron transfer flavoprotein β subunit (ETFB)-specific siRNA reduced the cell number in the attached culture without a decrease in the detached culture.
  
  
  CONCLUSIONS
  ETFB participates in the mechanoregulation of fibroblast cell number in collagen gel culture.
authors: R&D Laboratories, POLA PHARMA INC., 560 Kashio-cho, Totsuka-ku, Yokohama 244-0812, Japan. s-hirokawa@pola-pharma.co.jp
authors: Shigenari
authors: S
authors: Hirokawa
authors: Tomomasa
authors: T
authors: Shimanuki
authors: Hiroyuki
authors: H
authors: Kitajima
authors: Yasutomo
authors: Y
authors: Nishimori
authors: Makoto
authors: M
authors: Shimosaka
mesh_terms: D002452:Cell Count; D002478:Cells, Cultured; D003094:Collagen; D044927:Electron-Transferring Flavoproteins / Q000235:genetics / Q000502:physiology*; D005347:Fibroblasts / Q000502:physiology*; D005782:Gels; D006801:Humans; D034741:RNA, Small Interfering / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013314:Stress, Mechanical
chemical_list: C500319:ETFB protein, human; D044927:Electron-Transferring Flavoproteins; D005782:Gels; D034741:RNA, Small Interfering; D003094:Collagen