title: AG-041R, a novel indoline-2-one derivative, stimulates chondrogenesis in a bipotent chondroprogenitor cell line CL-1.
abstract: OBJECTIVE
  To examine the chondrogenic activity of AG-041R and its mode of action in a bipotent chondroprogenitor cell line CL-1.
  
  
  METHODS
  Chondrogenic activity of AG-041R in CL-1 was examined by histology, alcian blue pH 1.0 intensity and mRNA expression of cartilage matrix proteins (collagen type II, aggrecan). Chondrogenic activities of other CCK2/gastrin receptor antagonists were also examined. Since TGF-beta1 induces dominant chondrogenesis and suppressed adipogenesis in CL-1, induction of TGF-beta by AG-041R was examined by enzyme linked immunosorbent assay. Involvement of MAP kinases in the chondrogenic effect of AG-041R in CL-1 was examined by Western blotting and MAP kinase inhibitors.
  
  
  RESULTS
  AG-041R induced dominant chondrogenesis and marked suppression of adipogenesis in CL-1. Neither of the other CCK2/gastrin receptor antagonists tested showed chondrogenic activity in CL-1. AG-041R increased alcian blue pH 1.0 intensity and mRNA expression of collagen type II and aggrecan. TGF-beta1 and -beta2 proteins were increased by AG-041R. The chondrogenic activity of AG-041R in CL-1 was blocked by TGF-beta neutralizing antibody or inhibitors for activation of latent TGF-beta. AG-041R activated both Erk (p44/42) and p38 MAP kinases in CL-1. Inhibition of Erk (p44/42) by PD98059 canceled the adipogenesis suppression by AG-041R in CL-1. Inhibition of p38 by SB202190 completely canceled the chondrogenic activity of AG-041R in CL-1.
  
  
  CONCLUSIONS
  AG-041R has chondrogenic activity in CL-1 not related to CCK2/gastrin receptor antagonism. It is suggested that TGF-beta induction and the activation of MAP kinases mediate the chondrogenic activity of AG-041R in CL-1.
authors.affiliation: Fuji-Gotemba Research Laboratories, Chugai Pharmaceutical Co., Ltd., 1-135 Komakado, Gotemba, Shizuoka 412-8513, Japan. kitamurahdt@chugaipharm.co.jp
authors.forename: Hidetomo
authors.initials: H
authors.lastname: Kitamura
authors.forename: Makoto
authors.initials: M
authors.lastname: Okazaki
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D020219:Chondrogenesis / Q000187:drug effects* / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology*; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology; D043887:Receptor, Cholecystokinin B / Q000037:antagonists & inhibitors; D015398:Signal Transduction / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000096:biosynthesis
chemical_list: C109755:AG-041R; D004791:Enzyme Inhibitors; D007211:Indoles; D043887:Receptor, Cholecystokinin B; D016212:Transforming Growth Factor beta; D020928:Mitogen-Activated Protein Kinases