title: Building Elastin. Incorporation of recombinant human tropoelastin into extracellular matrices using nonelastogenic rat-1 fibroblasts as a source for lysyl oxidase.
abstract: The purpose of this study was to assess the feasibility of crosslinking exogenously produced tropoelastin, the precursor of insoluble elastin, into existing elastin. Tritiated recombinant human tropoelastin (rhTE) was added to neonatal rat aorta smooth-muscle cell (NNRSMC) cultures. As much as 12% of the added rhTE was incorporated into the NNRSMC-derived insoluble elastin with the formation of the elastin crosslinks desmosine (DES) and isodesmosine (IDES) in a time-dependent fashion. The ratio of radioactivity found in DES and IDES crosslinks to that found in lysyl residues increased from 0.18 immediately after incubation with rhTE to 0.76 after 14 d. Crosslinking of rhTE into elastin and the accompanying formation of tritiated water was inhibited by beta-aminoproprionitrile, a potent inhibitor of lysyl oxidase, an enzyme critical for the post-translational processing of elastin and collagen. Acellular NNRSMC matrices were produced and replated with Rat-1 fibroblasts, cells that were found to express lysyl oxidase but not tropoelastin. At 14 d after incubation with rhTE, the ratio of DES and IDES radioactivity to that of lysine in the insoluble elastin was 0.38. We show for the first time that cells expressing lysyl oxidase, but not elastin, as well as elastogenic cells can incorporate rhTE into insoluble elastin with the formation of elastin crosslinks. This novel approach might be used to augment elastin repair in certain pathologic states.
authors: Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118, USA. stone@biochem.bumc.bu.edu
authors: P J
authors: PJ
authors: Stone
authors: S M
authors: SM
authors: Morris
authors: S
authors: S
authors: Griffin
authors: S
authors: S
authors: Mithieux
authors: A S
authors: AS
authors: Weiss
mesh_terms: D000818:Animals; D003432:Cross-Linking Reagents; D004549:Elastin / Q000235:genetics / Q000378:metabolism*; D005091:Exons; D005109:Extracellular Matrix / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D008249:Protein-Lysine 6-Oxidase / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D012995:Solubility; D014333:Tropoelastin / Q000235:genetics / Q000378:metabolism*
chemical_list: D003432:Cross-Linking Reagents; D011994:Recombinant Proteins; D014333:Tropoelastin; D004549:Elastin; D008249:Protein-Lysine 6-Oxidase