title: TGF-ß induces Lysyl hydroxylase 2b in human synovial osteoarthritic fibroblasts through ALK5 signaling.
abstract: Lysyl hydroxylase 2b (LH2b) is known to increase pyridinoline cross-links, making collagen less susceptible to enzymatic degradation. Previously, we observed a relationship between LH2b and osteoarthritis-related fibrosis in murine knee joint. For this study, we investigate if transforming growth factor-beta (TGF-ß) and connective tissue growth factor (CTGF) regulate procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) (gene encoding LH2b) and LH2b expression differently in osteoarthritic human synovial fibroblasts (hSF). Furthermore, we investigate via which TGF-ß route (Smad2/3P or Smad1/5/8P) LH2b is regulated, to explore options to inhibit LH2b during fibrosis. To answer these questions, fibroblasts were isolated from knee joints of osteoarthritis patients. The hSF were stimulated with TGF-ß with or without a kinase inhibitor of ALK4/5/7 (SB-505124) or ALK1/2/3/6 (dorsomorphin). TGF-ß, CTGF, constitutively active (ca)ALK1 and caALK5 were adenovirally overexpressed in hSF. The gene expression levels of PLOD1/2/3, CTGF and COL1A1 were analyzed with Q-PCR. LH2 protein levels were determined with western blot. As expected, TGF-ß induced PLOD2/LH2 expression in hSF, whereas CTGF did not. PLOD1 and PLOD3 were not affected by either TGF-ß or CTGF. SB-505124 prevented the induction of TGF-ß-induced PLOD2, CTGF and COL1A1. Surprisingly, dorsomorphin completely blocked the induction of CTGF and COL1A1, whereas TGF-ß-induced PLOD2 was only slightly reduced. Overexpression of caALK5 in osteoarthritic hSF significantly induced PLOD2/LH2 expression, whereas caALK1 had no effect. We showed, in osteoarthritic hSF, that TGF-ß induced PLOD2/LH2 via ALK5 Smad2/3P. This elevation of LH2b in osteoarthritic hSF makes LH2b an interesting target to interfere with osteoarthritis-related persistent fibrosis.
authors.affiliation: Rheumatology Research and Advanced Therapeutics, Radboud University Nijmegen Medical Centre, Geert Grooteplein Zuid 26-28, 6525 GA, Nijmegen, The Netherlands.
authors.forename: Dennis F G
authors.initials: DF
authors.lastname: Remst
authors.forename: Esmeralda N
authors.initials: EN
authors.lastname: Blaney Davidson
authors.forename: Elly L
authors.initials: EL
authors.lastname: Vitters
authors.forename: Ruud A
authors.initials: RA
authors.lastname: Bank
authors.forename: Wim B
authors.initials: WB
authors.lastname: van den Berg
authors.forename: Peter M
authors.initials: PM
authors.lastname: van der Kraan
mesh_terms: D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D005355:Fibrosis / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D006801:Humans; D010003:Osteoarthritis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008240:Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase / Q000235:genetics* / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D051898:Smad1 Protein / Q000378:metabolism; D051899:Smad2 Protein / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism*
chemical_list: D018125:Receptors, Transforming Growth Factor beta; C497782:SMAD2 protein, human; D051898:Smad1 Protein; D051899:Smad2 Protein; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; C479138:PLOD2 protein, human; D008240:Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human