title: siRNA down-regulation of FGA mRNA in HepG2 cells demonstrated that heterozygous abnormality of the Aα-chain gene does not affect the plasma fibrinogen level.
abstract: BACKGROUND
  We encountered two afibrinogenemia patients with homozygous and compound heterozygous FGA mutation. Of interest, the patients' parents, who are heterozygous, had normal levels of plasma fibrinogen; thus, we hypothesized that liver FGA mRNA levels were higher than those of FGB and/or FGG mRNA.
  
  
  METHODS
  To test the hypothesis, we quantitated mRNA levels of a normal liver and a human hepatocyte cell line, HepG2 cells, and performed siRNA-mediated down-regulation of the fibrinogen gene in HepG2 cells. mRNA levels were determined using real-time quantitative RT- PCR for three normal livers and HepG2 cells. Down-regulation of FGA, FGB, or FGG in HepG2 cells was performed by the addition of siRNA corresponding to each of the three genes, and the mRNA levels determined in the cells and the secreted fibrinogen concentration in media.
  
  
  RESULTS
  The mRNA level of normal human liver was FGA=FGB>FGG and the FGG mRNA level was about 2-fold lower than the others, that of HepG2 cells was FGA>FGG>FGB and FGA mRNA was approximately 2- or 4-fold higher than FGG mRNA and FGB mRNA. When FGA, FGB, or FGG mRNA expression levels were down-regulated by nearby 50%, fibrinogen concentrations in media were 78%, 49%, or 57% of the control, respectively.
  
  
  CONCLUSIONS
  Our results suggest that FGG mRNA levels limit fibrinogen expression in normal liver and HepG2 cells and that 50% reduction of FGA mRNA levels would not limit fibrinogen expression in normal liver and HepG2 cells.
authors: Department of Health and Medical Sciences, Graduate School of Medicine, Shinshu University, Japan.
authors: Yuka
authors: Y
authors: Takezawa
authors: Kazuyuki
authors: K
authors: Matsuda
authors: Fumiko
authors: F
authors: Terasawa
authors: Mitsutoshi
authors: M
authors: Sugano
authors: Takayuki
authors: T
authors: Honda
authors: Nobuo
authors: N
authors: Okumura
mesh_terms: D000347:Afibrinogenemia / Q000097:blood / Q000235:genetics; D015536:Down-Regulation; D005260:Female; D005340:Fibrinogen / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D056945:Hep G2 Cells; D006801:Humans; D008297:Male; D020641:Polymorphism, Single Nucleotide; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D034741:RNA, Small Interfering; C464794:fibrinogen Aalpha; D005340:Fibrinogen