title: Homocysteine induces serum amyloid A3 in osteoblasts via unlocking RGD-motifs in collagen.
abstract: Hyperhomocysteinemia is a risk factor for osteoporotic fractures. Homocysteine (Hcys) inhibits collagen cross-linking and consequently decreases bone extracellular matrix (ECM) quality. Serum amyloid A (A-SAA), an acute-phase protein family, plays an important role in chronic and inflammatory diseases and up-regulates MMP13, which plays an important role in bone development and remodeling. Here, we investigate the effect of Hcys on expression of SAA3, a member of the A-SAA gene family, in osteoblasts characterizing underlying mechanisms and possible consequences on bone metabolism. MC3T3-E1 osteoblast-like cells were cultured up to 21 d with Hcys (low millimolar range) or reseeded onto ECM resulting from untreated or Hcys-treated MC3T3-E1 cells. Fourier-transformed infrared spectroscopy and a discriminative antibody were used to characterize the resulting ECM. Gene expression and signaling pathways were analyzed by gene chip, quantitative RT-PCR, and immunoblotting. Transcriptional regulation of Saa3 was studied by promoter transfection assays, chromatin immunoprecipitation, and immunofluorescence microscopy. Hcys treatment resulted in reduced collagen cross-linking, uncovering of RGD-motifs, and activation of the PTK2-PXN-CTNNB1 pathway followed by RELA activation. These signaling events led to increased SAA3 expression followed by the production of MMP13 and several chemokines, including Ccl5, Ccl2, Cxcl10, and Il6. Our data suggest Saa3 as link between hyperhomocysteinemia and development of osteoporosis.
authors.affiliation: Ludwig Boltzmann Institute of Osteology at Hanusch Hospital of Wiener Gebietskrankenkasse and Allgemeine Unfallversicherungsanstalt Trauma Center Meidling, 1st Medical Department, Hanusch Hospital, Vienna, Austria.
authors.forename: Roman
authors.initials: R
authors.lastname: Thaler
authors.forename: Jochen
authors.initials: J
authors.lastname: Zwerina
authors.forename: Monika
authors.initials: M
authors.lastname: Rumpler
authors.forename: Silvia
authors.initials: S
authors.lastname: Spitzer
authors.forename: Sonja
authors.initials: S
authors.lastname: Gamsjaeger
authors.forename: Eleftherios P
authors.initials: EP
authors.lastname: Paschalis
authors.forename: Klaus
authors.initials: K
authors.lastname: Klaushofer
authors.forename: Franz
authors.initials: F
authors.lastname: Varga
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D018946:Chemokine CCL5 / Q000235:genetics; D024042:Collagen Type I / Q000737:chemistry / Q000378:metabolism; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism; D051417:Focal Adhesion Kinase 1 / Q000096:biosynthesis; D015870:Gene Expression / Q000187:drug effects; D006710:Homocysteine / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D020138:Hyperhomocysteinemia / Q000150:complications / Q000378:metabolism; D053509:Matrix Metalloproteinase 13 / Q000235:genetics; D051379:Mice; D008954:Models, Biological; D009842:Oligopeptides / Q000737:chemistry; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism*; D010024:Osteoporosis / Q000209:etiology / Q000378:metabolism; D051419:Paxillin / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D000685:Serum Amyloid A Protein / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D051176:beta Catenin / Q000378:metabolism; D019014:fas Receptor / Q000235:genetics
chemical_list: C495267:CTNNB1 protein, mouse; C503460:Ccl5 protein, mouse; D018946:Chemokine CCL5; D024042:Collagen Type I; C491899:Fas protein, mouse; D009842:Oligopeptides; D051419:Paxillin; C496062:Pxn protein, mouse; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C521404:SAA1 protein, human; C076498:Saa3 protein, mouse; D000685:Serum Amyloid A Protein; D051176:beta Catenin; D019014:fas Receptor; D006710:Homocysteine; C047981:arginyl-glycyl-aspartic acid; D051417:Focal Adhesion Kinase 1; C496051:Ptk2 protein, mouse; D053509:Matrix Metalloproteinase 13; C507081:Mmp13 protein, mouse