title: Intracrine signalling of activin A in hepatocytes upregulates connective tissue growth factor (CTGF/CCN2) expression.
abstract: OBJECTIVE
  Up to now, the effect of activin A on the expression of the important transforming growth factor (TGF)-beta downstream modulator connective tissue growth factor (CTGF) is not known, but might be of relevance for the functional effects of this cytokine on several liver cell types.
  
  
  METHODS
  In this study, activin A-dependent CTGF expression in hepatocytes (PC) primed by exogenous activin A and in PC maintained under complete activin-free culture conditions was analysed by Western blots, metabolic labelling, gene silencing, reverse transcriptase-polymerase chain reaction (RT-PCR) and CTGF reporter gene assays. This study was supplemented by immunocytochemical staining of activin A and CTGF in PC of injured liver.
  
  
  RESULTS
  Using alkaline phosphatase alpha-alkaline phosphatase staining, it is demonstrated that activin A becomes increasingly detectable during the course of CCl(4)-liver damage. Addition of activin A to cultured PC induced CTGF protein expression via phosphorylation of Smad2 and Smad3. This induction can be inhibited by the antagonist follistatin and alpha-activin A antibody respectively. When PC were cultured under serum(i.e. activin A)-free culture conditions, a time-dependent increase of activin expression during the course of the culture was proven by RT-PCR. Silencing of inhibin beta(A) gene expression under serum-free conditions by small interfering RNAs greatly suppressed CTGF synthesis and the phosphorylations of Smad2 and Smad3. However, both the extracellularly acting follistatin and the alpha-activin A antibody could not inhibit spontaneous CTGF expression, which, however, was achieved by the cell-permeable TGF-beta Alk4/Alk5 receptor-kinase-inhibitor SB431542.
  
  
  CONCLUSIONS
  In conclusion, the results point to activin A as an inducer of CTGF synthesis in PC. Intracellular activin A contributes to spontaneous CTGF expression in PC independent of exogenous activin A, which is proposed to occur via Alk4/Alk5-receptors. The findings might be important for many actions of activin A on the liver.
authors.affiliation: Institute of Clinical Chemistry and Pathobiochemistry, RWTH-University Hospital, Aachen, Germany. ogressner@ukaachen.de
authors.forename: Olav A
authors.initials: OA
authors.lastname: Gressner
authors.forename: Birgit
authors.initials: B
authors.lastname: Lahme
authors.forename: Monika
authors.initials: M
authors.lastname: Siluschek
authors.forename: Katharina
authors.initials: K
authors.lastname: Rehbein
authors.forename: Ralf
authors.initials: R
authors.lastname: Weiskirchen
authors.forename: Axel M
authors.initials: AM
authors.lastname: Gressner
mesh_terms: D028341:Activins / Q000378:metabolism*; D000818:Animals; D019898:Autocrine Communication; D002251:Carbon Tetrachloride / Q000633:toxicity; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury / Q000378:metabolism; D055513:Connective Tissue Growth Factor / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism*; D007150:Immunohistochemistry; D008297:Male; D019899:Paracrine Communication; D051381:Rats; D017207:Rats, Sprague-Dawley; D051899:Smad2 Protein / Q000378:metabolism; D051900:Smad3 Protein / Q000378:metabolism; D015854:Up-Regulation
chemical_list: C526316:CCN2 protein, rat; D051899:Smad2 Protein; C497784:Smad2 protein, rat; D051900:Smad3 Protein; C497787:Smad3 protein, rat; C422405:activin A; D028341:Activins; D055513:Connective Tissue Growth Factor; D002251:Carbon Tetrachloride