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The Wnt/beta-catenin pathway interacts differentially with PTHrP signaling to control chondrocyte hypertrophy and final maturation.

Guo X, Mak KK, Taketo MM, Yang Y - PLoS ONE (2009)

Bottom Line: Here we show by genetic approaches that chondrocyte hypertrophy and final maturation are two distinct developmental processes that are differentially regulated by Wnt/beta-catenin and PTHrP signaling.Wnt/beta-catenin signaling regulates initiation of chondrocyte hypertrophy by inhibiting PTHrP signaling activity, but it does not regulate PTHrP expression.In addition, Wnt/beta-catenin signaling regulates chondrocyte hypertrophy in a non-cell autonomous manner and Gdf5/Bmp signaling may be one of the downstream pathways.

View Article: PubMed Central - PubMed

Affiliation: Developmental Genetics Section, National Human Genome Research Institute, Bethesda, MD, USA.

ABSTRACT
Sequential proliferation, hypertrophy and maturation of chondrocytes are required for proper endochondral bone development and tightly regulated by cell signaling. The canonical Wnt signaling pathway acts through beta-catenin to promote chondrocyte hypertrophy whereas PTHrP signaling inhibits it by holding chondrocytes in proliferating states. Here we show by genetic approaches that chondrocyte hypertrophy and final maturation are two distinct developmental processes that are differentially regulated by Wnt/beta-catenin and PTHrP signaling. Wnt/beta-catenin signaling regulates initiation of chondrocyte hypertrophy by inhibiting PTHrP signaling activity, but it does not regulate PTHrP expression. In addition, Wnt/beta-catenin signaling regulates chondrocyte hypertrophy in a non-cell autonomous manner and Gdf5/Bmp signaling may be one of the downstream pathways. Furthermore, Wnt/beta-catenin signaling also controls final maturation of hypertrophic chondrocytes, but such regulation is PTHrP signaling-independent.

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Hypertrophic chondrocyte maturation was regulated by β-catenin independently of PTHrP.Consecutive sections of the developing humerus at 14.5 dpc and 16.5 dpc were examined by Von Kossa staining and in situ hybridization with the indicated riboprobes. Mature hypertrophic chondrocytes were stained black by the Von Kossa method and expressed Mmp13 and Opn. (A) Von Kossa staining and Mmp13 and Opn expression were much reduced in both Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (B) Ihh and Pthr1 were expressed in the less mature hypertrophic chondrocytes (double-headed arrow). This domain was expanded significantly only in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (C) At 16.5 dpc, chondrocyte final maturation in the Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant was similar to that of the wild type control and PTHrP−/− mutant, respectively. (D) Expansion of Ihh and Pthr1 expression domains (white line) were still observed in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant at 16.5 dpc. (E) Expression of Mmp9 and Vegf in the developing humerus of the indicated genotypes at 14.5 dpc.
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pone-0006067-g003: Hypertrophic chondrocyte maturation was regulated by β-catenin independently of PTHrP.Consecutive sections of the developing humerus at 14.5 dpc and 16.5 dpc were examined by Von Kossa staining and in situ hybridization with the indicated riboprobes. Mature hypertrophic chondrocytes were stained black by the Von Kossa method and expressed Mmp13 and Opn. (A) Von Kossa staining and Mmp13 and Opn expression were much reduced in both Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (B) Ihh and Pthr1 were expressed in the less mature hypertrophic chondrocytes (double-headed arrow). This domain was expanded significantly only in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (C) At 16.5 dpc, chondrocyte final maturation in the Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant was similar to that of the wild type control and PTHrP−/− mutant, respectively. (D) Expansion of Ihh and Pthr1 expression domains (white line) were still observed in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant at 16.5 dpc. (E) Expression of Mmp9 and Vegf in the developing humerus of the indicated genotypes at 14.5 dpc.

Mentions: In the normal process of long bone development, chondrocyte hypertrophy is followed by hypertrophic chondrocyte maturation to allow the invasion of osteoblasts and blood vessels. Our observation that mineralization, but not hypertrophy, was similarly delayed in the PTHrP−/−; Catnbc/c; Col2a1-Cre and Catnbc/c; Col2a1-Cre embryos led us to think that formation of the early and late populations of hypertrophic chondrocytes are differentially regulated. To test this, we further examined chondrocyte maturation on histological sections. At 14.5 dpc, although Col10a1 expression domains were comparable between PTHrP−/− and PTHrP−/−; Catnbc/c; Col2a1-Cre embryos (Fig. 1D), in stark contrast to PTHrP−/− embryo, where maturation of hypertrophic chondrocytes was also accelerated, hypertrophic chondrocyte maturation detected by von Kossa staining was significantly delayed in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos (Fig. 3A). We then performed in situ hybridizations with probes specific to distinct stages of chondrocyte hypertrophy and maturation. At 14.5 dpc, expression domains of Mmp13 and Opn were much smaller in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos compared to those in the PTHrP−/− embryos (Fig. 3A). By contrast, expression domains of Ihh and Pthr1, which mark prehypertrophic chondrocytes and early hypertrophic chondocytes, were expanded in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos compared to that in the PTHrP−/− embryos (Fig. 3B). Thus, in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryo, loss of PTHrP signaling accelerated chondrocyte hypertrophy, but these hypertrophic chondrocytes were not able to undergo further maturation due to the blockage of Wnt/β-catenin signaling. As a result, the Ihh and Pthr1 expressing early hypertrophic chondrocyte population was significantly expanded. This increased Ihh expression domain in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryo likely caused increased osteoblast differentiation compared to that in the Catnbc/c; Col2a1-Cre embryo (Fig. 3A and S3).


The Wnt/beta-catenin pathway interacts differentially with PTHrP signaling to control chondrocyte hypertrophy and final maturation.

Guo X, Mak KK, Taketo MM, Yang Y - PLoS ONE (2009)

Hypertrophic chondrocyte maturation was regulated by β-catenin independently of PTHrP.Consecutive sections of the developing humerus at 14.5 dpc and 16.5 dpc were examined by Von Kossa staining and in situ hybridization with the indicated riboprobes. Mature hypertrophic chondrocytes were stained black by the Von Kossa method and expressed Mmp13 and Opn. (A) Von Kossa staining and Mmp13 and Opn expression were much reduced in both Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (B) Ihh and Pthr1 were expressed in the less mature hypertrophic chondrocytes (double-headed arrow). This domain was expanded significantly only in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (C) At 16.5 dpc, chondrocyte final maturation in the Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant was similar to that of the wild type control and PTHrP−/− mutant, respectively. (D) Expansion of Ihh and Pthr1 expression domains (white line) were still observed in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant at 16.5 dpc. (E) Expression of Mmp9 and Vegf in the developing humerus of the indicated genotypes at 14.5 dpc.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2698152&req=5

pone-0006067-g003: Hypertrophic chondrocyte maturation was regulated by β-catenin independently of PTHrP.Consecutive sections of the developing humerus at 14.5 dpc and 16.5 dpc were examined by Von Kossa staining and in situ hybridization with the indicated riboprobes. Mature hypertrophic chondrocytes were stained black by the Von Kossa method and expressed Mmp13 and Opn. (A) Von Kossa staining and Mmp13 and Opn expression were much reduced in both Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (B) Ihh and Pthr1 were expressed in the less mature hypertrophic chondrocytes (double-headed arrow). This domain was expanded significantly only in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant embryos at 14.5 dpc. (C) At 16.5 dpc, chondrocyte final maturation in the Catnbc/c; Col2a1-Cre and PTHrP−/−; Catnbc/c; Col2a1-Cre mutant was similar to that of the wild type control and PTHrP−/− mutant, respectively. (D) Expansion of Ihh and Pthr1 expression domains (white line) were still observed in the PTHrP−/−; Catnbc/c; Col2a1-Cre mutant at 16.5 dpc. (E) Expression of Mmp9 and Vegf in the developing humerus of the indicated genotypes at 14.5 dpc.
Mentions: In the normal process of long bone development, chondrocyte hypertrophy is followed by hypertrophic chondrocyte maturation to allow the invasion of osteoblasts and blood vessels. Our observation that mineralization, but not hypertrophy, was similarly delayed in the PTHrP−/−; Catnbc/c; Col2a1-Cre and Catnbc/c; Col2a1-Cre embryos led us to think that formation of the early and late populations of hypertrophic chondrocytes are differentially regulated. To test this, we further examined chondrocyte maturation on histological sections. At 14.5 dpc, although Col10a1 expression domains were comparable between PTHrP−/− and PTHrP−/−; Catnbc/c; Col2a1-Cre embryos (Fig. 1D), in stark contrast to PTHrP−/− embryo, where maturation of hypertrophic chondrocytes was also accelerated, hypertrophic chondrocyte maturation detected by von Kossa staining was significantly delayed in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos (Fig. 3A). We then performed in situ hybridizations with probes specific to distinct stages of chondrocyte hypertrophy and maturation. At 14.5 dpc, expression domains of Mmp13 and Opn were much smaller in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos compared to those in the PTHrP−/− embryos (Fig. 3A). By contrast, expression domains of Ihh and Pthr1, which mark prehypertrophic chondrocytes and early hypertrophic chondocytes, were expanded in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryos compared to that in the PTHrP−/− embryos (Fig. 3B). Thus, in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryo, loss of PTHrP signaling accelerated chondrocyte hypertrophy, but these hypertrophic chondrocytes were not able to undergo further maturation due to the blockage of Wnt/β-catenin signaling. As a result, the Ihh and Pthr1 expressing early hypertrophic chondrocyte population was significantly expanded. This increased Ihh expression domain in the PTHrP−/−; Catnbc/c; Col2a1-Cre embryo likely caused increased osteoblast differentiation compared to that in the Catnbc/c; Col2a1-Cre embryo (Fig. 3A and S3).

Bottom Line: Here we show by genetic approaches that chondrocyte hypertrophy and final maturation are two distinct developmental processes that are differentially regulated by Wnt/beta-catenin and PTHrP signaling.Wnt/beta-catenin signaling regulates initiation of chondrocyte hypertrophy by inhibiting PTHrP signaling activity, but it does not regulate PTHrP expression.In addition, Wnt/beta-catenin signaling regulates chondrocyte hypertrophy in a non-cell autonomous manner and Gdf5/Bmp signaling may be one of the downstream pathways.

View Article: PubMed Central - PubMed

Affiliation: Developmental Genetics Section, National Human Genome Research Institute, Bethesda, MD, USA.

ABSTRACT
Sequential proliferation, hypertrophy and maturation of chondrocytes are required for proper endochondral bone development and tightly regulated by cell signaling. The canonical Wnt signaling pathway acts through beta-catenin to promote chondrocyte hypertrophy whereas PTHrP signaling inhibits it by holding chondrocytes in proliferating states. Here we show by genetic approaches that chondrocyte hypertrophy and final maturation are two distinct developmental processes that are differentially regulated by Wnt/beta-catenin and PTHrP signaling. Wnt/beta-catenin signaling regulates initiation of chondrocyte hypertrophy by inhibiting PTHrP signaling activity, but it does not regulate PTHrP expression. In addition, Wnt/beta-catenin signaling regulates chondrocyte hypertrophy in a non-cell autonomous manner and Gdf5/Bmp signaling may be one of the downstream pathways. Furthermore, Wnt/beta-catenin signaling also controls final maturation of hypertrophic chondrocytes, but such regulation is PTHrP signaling-independent.

Show MeSH
Related in: MedlinePlus