title: [Expression of CCAAT/enhancer-binding protein in cultured rat hepatic stellate cells and its significance].
abstract: OBJECTIVE
  The expression of C/EBPalpha protein and mRNA during automatically activation process in primary cultures of HSCs were observed in order to explore its possible association with the proliferation and activation of HSCs.
  
  
  METHODS
  Immunocytochemistry, Western blot and RT-PCR were used to evaluated the expression of C/EBPalpha protein and mRNA; as well as the expression of alpha-SMA, Desmin, MMP2, type I procollagen (alpha1). The eukaryotic vector harboring the full length cDNA of C/EBPalpha was transfected into activated HSC, then immunocytochemistry was applied to confirm the transfection and evaluate the effect of transfection on the proliferation of HSC by calculating the PCNA-positive cells. The morphological changes of HSC were observed by use of phase-contrast microscope.
  
  
  RESULTS
  Constitutive expression of mRNA and protein of C/EBPalpha were detected in primarily cultured HSCs, and the protein was seen in both nuclei and cytoplasm with the latter being dominant. Their expression levels reached highest at day 2 of the culture, then decreased gradually when continually cultured to the day 4, 7, 10, on the other hand, the expression of alpha-SMA, MMP2 and ColI(alpha1) increased steadily. Transient transfection was verified by the fact that much more and stronger C/EBPalpha stain was observed in transfected HSCs than in void-vector transfected cells. In C/EBPalpha gene transfected HSCs, the number of PCNA-positive cells dramatically decreased compared with the void-vector transfected cells 24h after transfection. In addition, the C/EBPalpha gene transfected HSCs died 36 h after transfection, a few surviving cells became longer and thinner in morphology, however the void-vector transfected cells almost all remained alive.
  
  
  CONCLUSIONS
  C/EBPalpha was likely involved in the HSCs activation, and over-expressed C/EBPalpha by transfection had inhibitory influence on the proliferation of cultured rat HSCs.
authors.affiliation: Department of Pathology, Medical Center of Fudan University, Shanghai 200032, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Huang
authors.forename: Jin-sheng
authors.initials: JS
authors.lastname: Zhang
authors.forename: Guang-cun
authors.initials: GC
authors.lastname: Huang
authors.forename: Qi-qun
authors.initials: QQ
authors.lastname: Tang
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.forename: Xiu-rong
authors.initials: XR
authors.lastname: Zhang
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
mesh_terms: D000818:Animals; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000235:genetics*; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection
chemical_list: D022763:CCAAT-Enhancer-Binding Protein-alpha; D024042:Collagen Type I; D012333:RNA, Messenger; D020778:Matrix Metalloproteinase 2