title: Collagen cross-linking influences osteoblastic differentiation.
abstract: Osteoblasts synthesize collagen matrix, which itself regulates the differentiation of precursor cells into mature osteoblasts. They express lysyl oxidase (LOX), which is involved in the collagen cross-linking process. Lathyrogens, like ss-aminopropionitrile (ssAPN), inhibit the formation of a stable matrix. The aim of the present study was to investigate the influence of cross-linking on osteoblastic differentiation. MC3T3-E1 cells were seeded and treated with or without 400 muM ssAPN for 1 week. Thereafter, living cells were removed and, on this extracellular matrix, new MC3T3-E1 cells were seeded and cultured for 1 week without ssAPN. RNA was isolated, and expression of specific marker genes was determined by quantitative reverse transcription-polymerase chain reaction. Changes in specific cross-links after ssAPN treatment were measured with Fourier-transform infrared spectroscopy. The collagen matrix that formed showed a significant reduction of two major cross-links of bone collagen, deH-DHLNL and pyr, compared to control cultures. Gene expression studies showed an increase of collagen alpha1 (I) (COL1A1) to 150%. Expression of LOX and osteocalcin (OCN) mRNA was significantly downregulated to about 75%. When fresh MC3T3-E1 cells were seeded on this altered matrix without ssAPN, COL1A1 mRNA expression was upregulated (140%), OCN was downregulated (60%), and LOX mRNA expression remained unaffected. These results indicate that ssAPN treatment not only disrupts collagen cross-link formation but also affects osteoblastic activity and expression. In conclusion, the disrupted matrix produced in the presence of lathyrogen influences, even in its absence, the expression of osteoblastic genes.
authors.affiliation: Ludwig Boltzmann Institute of Osteology at the Hanusch Hospital of WGKK and AUVA Trauma Center Meidling, 4th Medical Department, Hanusch Hospital, Heinrich Collinstrasse 30, Vienna, Austria.
authors.forename: C
authors.initials: C
authors.lastname: Turecek
authors.forename: N
authors.initials: N
authors.lastname: Fratzl-Zelman
authors.forename: M
authors.initials: M
authors.lastname: Rumpler
authors.forename: B
authors.initials: B
authors.lastname: Buchinger
authors.forename: S
authors.initials: S
authors.lastname: Spitzer
authors.forename: R
authors.initials: R
authors.lastname: Zoehrer
authors.forename: E
authors.initials: E
authors.lastname: Durchschlag
authors.forename: K
authors.initials: K
authors.lastname: Klaushofer
authors.forename: E P
authors.initials: EP
authors.lastname: Paschalis
authors.forename: F
authors.initials: F
authors.lastname: Varga
mesh_terms: D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D000629:Aminopropionitrile / Q000494:pharmacology; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D003094:Collagen / Q000096:biosynthesis* / Q000737:chemistry; D024042:Collagen Type I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000090602:Collagen Type I, alpha 1 Chain; D003432:Cross-Linking Reagents; D004151:Dipeptides / Q000737:chemistry / Q000378:metabolism; D005109:Extracellular Matrix / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D051379:Mice; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D008249:Protein-Lysine 6-Oxidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000596:Amino Acids; D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D003432:Cross-Linking Reagents; D004151:Dipeptides; D012333:RNA, Messenger; D015675:Osteocalcin; C007428:5,5'-dihydroxylysylnorleucine; D000629:Aminopropionitrile; C015484:pyridinoline; D003094:Collagen; D008249:Protein-Lysine 6-Oxidase