Studies were designed to investigate the expression and regulation of interstitial collagenase (MMP-1) during endochondral ossification in fetal bovine growth plate chondrocytes in vitro. Fetal bovine growth plate chondrocytes were separated into subpopulations representing different maturational stages by using a Percoll discontinuous gradient. Gene expression and synthesis of MMP-1 in these chondrocyte subpopulations were followed in serum-free cultures as they matured from the prehypertrophic to hypertrophic phenotype. However, MMP-1 mRNA expression and protein were undetectable. This expression was also undetectable in interleukin 1alpha (IL-1alpha) stimulated growth plate chondrocytes. MMP-1 mRNA and protein were however observed in IL-1alpha stimulated bovine fetal skin fibroblasts. Although MMP-1 was absent in growth plate chondrocytes, a collagenolytic activity was detectable in culture medium by using 14C labeled type II collagen. It was only seen when the chondrocytes were hypertrophic. By using the polymerase chain reaction (PCR), employing conserved sequences found in mammalian collagenase-3 (MMP-13), a novel bovine collagenase, was cloned. The cDNA and predicted amino acid sequence shared a 92% and 90% identity to the human MMP-13 cDNA and amino acid sequence respectively. MMP-13 mRNA and protein were found only when the chondrocytes became hypertrophic. The role of MMP-13 during the maturation of these fetal growth plate chondrocytes was further investigated by using a preferential carboxylate inhibitor of collagenase-3. This inhibitor reduced the generation of denatured type II collagen in chondrocyte cell layers and proteoglycan release into conditioned medium in a dose dependent manner. Also, types II and X collagen synthesis and matrix calcification were inhibited. These studies indicate that MMP-13 may play a very important role during the maturation of fetal growth plate chondrocyte in vitro and during endochondral ossification and that cellul
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.35420 |
| Date | January 1997 |
| Creators | Wu, Ching Hua William. |
| Contributors | Poole, A. R. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Division of Surgical Research.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001605476, proquestno: NQ44630, Theses scanned by UMI/ProQuest. |
Page generated in 0.3952 seconds