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Matrix GLA protein is a developmental regulator of chondrocyte mineralization and, when constitutively expressed, blocks endochondral and intramembranous ossification in the limb.

Yagami K, Suh JY, Enomoto-Iwamoto M, Koyama E, Abrams WR, Shapiro IM, Pacifici M, Iwamoto M - J. Cell Biol. (1999)

Bottom Line: Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment.Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization.The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Surgery, Showa University, Dental School, Ohta-Ku, Tokyo 145, Japan.

ABSTRACT
Matrix GLA protein (MGP), a gamma-carboxyglutamic acid (GLA)-rich, vitamin K-dependent and apatite-binding protein, is a regulator of hypertrophic cartilage mineralization during development. However, MGP is produced by both hypertrophic and immature chondrocytes, suggesting that MGP's role in mineralization is cell stage-dependent, and that MGP may have other roles in immature cells. It is also unclear whether MGP regulates the quantity of mineral or mineral nature and quality as well. To address these issues, we determined the effects of manipulations of MGP synthesis and expression in (a) immature and hypertrophic chondrocyte cultures and (b) the chick limb bud in vivo. The two chondrocyte cultures displayed comparable levels of MGP gene expression. Yet, treatment with warfarin, a gamma-carboxylase inhibitor and vitamin K antagonist, triggered mineralization in hypertrophic but not immature cultures. Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment. Scanning electron microscopy, x-ray microanalysis, and Fourier-transform infrared spectroscopy revealed that mineral forming in control and warfarin-treated hypertrophic cell cultures was similar and represented stoichiometric apatite. Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization. Surprisingly, MGP overexpression in the developing limb not only inhibited cartilage mineralization, but also delayed chondrocyte maturation and blocked endochondral ossification and formation of a diaphyseal intramembranous bone collar. The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.

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Northern blot analysis of MGP gene expression. Confluent immature (A) and hypertrophic (B) cultures in medium A or medium B were treated for 4 d with warfarin, vitamin K, or both. Blots (10 μg RNA/lane) were stained with methylene blue to reveal the ribosomal bands and to verify equal loading and transfer efficiency. Blots were hybridized with a 32P-labeled MGP probe and exposed to x-ray films. The two main 3.0- and 1.3-kb MGP transcripts are indicated.
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Figure 3: Northern blot analysis of MGP gene expression. Confluent immature (A) and hypertrophic (B) cultures in medium A or medium B were treated for 4 d with warfarin, vitamin K, or both. Blots (10 μg RNA/lane) were stained with methylene blue to reveal the ribosomal bands and to verify equal loading and transfer efficiency. Blots were hybridized with a 32P-labeled MGP probe and exposed to x-ray films. The two main 3.0- and 1.3-kb MGP transcripts are indicated.

Mentions: To determine the relationship between mineralization responses and endogenous MGP gene expression, cultures of immature and hypertrophic chondrocytes in medium A or medium B were treated as above and were processed for Northern blot analyses. We found that both immature and hypertrophic cultures contained obvious amounts of the 1.3- and 3.0-kb MGP RNAs (Fig. 3A and Fig. B, respectively) (Wiedemann et al. 1998). Cultures in medium A contained two to threefold more MGP transcripts (Fig. 3A and Fig. B, lane 1) than cultures in medium B (Fig. 3A and Fig. B, lane 5). However, these basal levels of expression were not significantly affected by treatment with warfarin, vitamin K, or warfarin plus vitamin K (Fig. 3, A–B, lanes 1–8).


Matrix GLA protein is a developmental regulator of chondrocyte mineralization and, when constitutively expressed, blocks endochondral and intramembranous ossification in the limb.

Yagami K, Suh JY, Enomoto-Iwamoto M, Koyama E, Abrams WR, Shapiro IM, Pacifici M, Iwamoto M - J. Cell Biol. (1999)

Northern blot analysis of MGP gene expression. Confluent immature (A) and hypertrophic (B) cultures in medium A or medium B were treated for 4 d with warfarin, vitamin K, or both. Blots (10 μg RNA/lane) were stained with methylene blue to reveal the ribosomal bands and to verify equal loading and transfer efficiency. Blots were hybridized with a 32P-labeled MGP probe and exposed to x-ray films. The two main 3.0- and 1.3-kb MGP transcripts are indicated.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169349&req=5

Figure 3: Northern blot analysis of MGP gene expression. Confluent immature (A) and hypertrophic (B) cultures in medium A or medium B were treated for 4 d with warfarin, vitamin K, or both. Blots (10 μg RNA/lane) were stained with methylene blue to reveal the ribosomal bands and to verify equal loading and transfer efficiency. Blots were hybridized with a 32P-labeled MGP probe and exposed to x-ray films. The two main 3.0- and 1.3-kb MGP transcripts are indicated.
Mentions: To determine the relationship between mineralization responses and endogenous MGP gene expression, cultures of immature and hypertrophic chondrocytes in medium A or medium B were treated as above and were processed for Northern blot analyses. We found that both immature and hypertrophic cultures contained obvious amounts of the 1.3- and 3.0-kb MGP RNAs (Fig. 3A and Fig. B, respectively) (Wiedemann et al. 1998). Cultures in medium A contained two to threefold more MGP transcripts (Fig. 3A and Fig. B, lane 1) than cultures in medium B (Fig. 3A and Fig. B, lane 5). However, these basal levels of expression were not significantly affected by treatment with warfarin, vitamin K, or warfarin plus vitamin K (Fig. 3, A–B, lanes 1–8).

Bottom Line: Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment.Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization.The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Oral Surgery, Showa University, Dental School, Ohta-Ku, Tokyo 145, Japan.

ABSTRACT
Matrix GLA protein (MGP), a gamma-carboxyglutamic acid (GLA)-rich, vitamin K-dependent and apatite-binding protein, is a regulator of hypertrophic cartilage mineralization during development. However, MGP is produced by both hypertrophic and immature chondrocytes, suggesting that MGP's role in mineralization is cell stage-dependent, and that MGP may have other roles in immature cells. It is also unclear whether MGP regulates the quantity of mineral or mineral nature and quality as well. To address these issues, we determined the effects of manipulations of MGP synthesis and expression in (a) immature and hypertrophic chondrocyte cultures and (b) the chick limb bud in vivo. The two chondrocyte cultures displayed comparable levels of MGP gene expression. Yet, treatment with warfarin, a gamma-carboxylase inhibitor and vitamin K antagonist, triggered mineralization in hypertrophic but not immature cultures. Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment. Scanning electron microscopy, x-ray microanalysis, and Fourier-transform infrared spectroscopy revealed that mineral forming in control and warfarin-treated hypertrophic cell cultures was similar and represented stoichiometric apatite. Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization. Surprisingly, MGP overexpression in the developing limb not only inhibited cartilage mineralization, but also delayed chondrocyte maturation and blocked endochondral ossification and formation of a diaphyseal intramembranous bone collar. The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.

Show MeSH
Related in: MedlinePlus