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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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In situ hybridization analysis of gene expression patterns in control (A–C) and MGP-overexpressing (D–F) tibia. Only about half of the control tibia is shown in A-C, whereas the entire RCAS-MGP tibia is shown in D–F. Note in C the particularly strong MGP expression in articular layer (arrow), along the lateral side of the proliferative and prehypertrophic zones (double arrowheads), and in posthypertrophic zone (double arrow); barely detectable signal is seen throughout the hypertrophic zone (arrowhead). II, type II collagen; X, type X collagen. Art., articular layer; Prol., proliferative zone; Prehyp., prehypertrophic zone; and Posthyp., posthypertrophic zone. Bar, 250 μm.
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Figure 10: In situ hybridization analysis of gene expression patterns in control (A–C) and MGP-overexpressing (D–F) tibia. Only about half of the control tibia is shown in A-C, whereas the entire RCAS-MGP tibia is shown in D–F. Note in C the particularly strong MGP expression in articular layer (arrow), along the lateral side of the proliferative and prehypertrophic zones (double arrowheads), and in posthypertrophic zone (double arrow); barely detectable signal is seen throughout the hypertrophic zone (arrowhead). II, type II collagen; X, type X collagen. Art., articular layer; Prol., proliferative zone; Prehyp., prehypertrophic zone; and Posthyp., posthypertrophic zone. Bar, 250 μm.

Mentions: To confirm and extend these observations, we carried out in situ hybridization on similar longitudinal sections of day 12 control (Fig. 10, A–C) and RCAS-MGP (Fig. 10, D–F) tibia, using probes encoding cartilage characteristic type II and X collagens and MGP. It should be noted that, because of their different lengths, about one half of the control tibia is shown in Fig. 10A–C, whereas the entire RCAS-MGP tibia is shown in Fig. 10D–F. In control tibia (Fig. 10 A), transcripts encoding type II collagen were abundant in the epiphyseal articular layer and underlying growth plate containing proliferating, prehypertrophic, and hypertrophic chondrocytes; the transcripts were reduced in late posthypertrophic chondrocytes (Fig. 10 A), as reported previously (Leboy et al. 1988; Iyama et al. 1991). Transcripts encoding type X collagen, a product typical of hypertrophic chondrocytes, were abundant in hypertrophic and posthypertrophic chondrocytes (Fig. 10 B). MGP transcripts displayed more complex patterns reminiscent of those seen in mouse embryos (Luo et al. 1997; Komori et al. 1997); they were prominent in the articular layer (Fig. 10 C, arrow), in proliferating/prehypertrophic chondrocytes along the lateral side of tibia (Fig. 10 C, double arrowhead), and in posthypertrophic chondrocytes (Fig. 10 C, double arrow). Notably, MGP transcripts were essentially undetectable in the entire hypertrophic zone, either along its lateral side (Fig. 10 C, arrowhead) or its central region.


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)

In situ hybridization analysis of gene expression patterns in control (A–C) and MGP-overexpressing (D–F) tibia. Only about half of the control tibia is shown in A-C, whereas the entire RCAS-MGP tibia is shown in D–F. Note in C the particularly strong MGP expression in articular layer (arrow), along the lateral side of the proliferative and prehypertrophic zones (double arrowheads), and in posthypertrophic zone (double arrow); barely detectable signal is seen throughout the hypertrophic zone (arrowhead). II, type II collagen; X, type X collagen. Art., articular layer; Prol., proliferative zone; Prehyp., prehypertrophic zone; and Posthyp., posthypertrophic zone. Bar, 250 μm.
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Figure 10: In situ hybridization analysis of gene expression patterns in control (A–C) and MGP-overexpressing (D–F) tibia. Only about half of the control tibia is shown in A-C, whereas the entire RCAS-MGP tibia is shown in D–F. Note in C the particularly strong MGP expression in articular layer (arrow), along the lateral side of the proliferative and prehypertrophic zones (double arrowheads), and in posthypertrophic zone (double arrow); barely detectable signal is seen throughout the hypertrophic zone (arrowhead). II, type II collagen; X, type X collagen. Art., articular layer; Prol., proliferative zone; Prehyp., prehypertrophic zone; and Posthyp., posthypertrophic zone. Bar, 250 μm.
Mentions: To confirm and extend these observations, we carried out in situ hybridization on similar longitudinal sections of day 12 control (Fig. 10, A–C) and RCAS-MGP (Fig. 10, D–F) tibia, using probes encoding cartilage characteristic type II and X collagens and MGP. It should be noted that, because of their different lengths, about one half of the control tibia is shown in Fig. 10A–C, whereas the entire RCAS-MGP tibia is shown in Fig. 10D–F. In control tibia (Fig. 10 A), transcripts encoding type II collagen were abundant in the epiphyseal articular layer and underlying growth plate containing proliferating, prehypertrophic, and hypertrophic chondrocytes; the transcripts were reduced in late posthypertrophic chondrocytes (Fig. 10 A), as reported previously (Leboy et al. 1988; Iyama et al. 1991). Transcripts encoding type X collagen, a product typical of hypertrophic chondrocytes, were abundant in hypertrophic and posthypertrophic chondrocytes (Fig. 10 B). MGP transcripts displayed more complex patterns reminiscent of those seen in mouse embryos (Luo et al. 1997; Komori et al. 1997); they were prominent in the articular layer (Fig. 10 C, arrow), in proliferating/prehypertrophic chondrocytes along the lateral side of tibia (Fig. 10 C, double arrowhead), and in posthypertrophic chondrocytes (Fig. 10 C, double arrow). Notably, MGP transcripts were essentially undetectable in the entire hypertrophic zone, either along its lateral side (Fig. 10 C, arrowhead) or its central region.

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