Developmental regulation of Wnt/beta-catenin signals is required for growth plate assembly, cartilage integrity, and endochondral ossification.
Studies have suggested that continuous Wnt/beta-catenin signaling in nascent cartilaginous skeletal elements blocks chondrocyte hypertrophy and endochondral ossification, whereas signaling starting at later stages stimulates hypertrophy and ossification, indicating that Wnt/beta-catenin roles are developmentally regulated. To test this conclusion further, we created transgenic mice expressing a fusion mutant protein of beta-catenin and LEF (CA-LEF) in nascent chondrocytes. Transgenic mice had severe skeletal defects, particularly in limbs. Growth plates were totally disorganized, lacked maturing chondrocytes expressing Indian hedgehog and collagen X, and failed to undergo endochondral ossification. Interestingly, the transgenic cartilaginous elements were ill defined, intermingled with surrounding connective and vascular tissues, and even displayed abnormal joints. However, when activated beta-catenin mutant (delta-beta-catenin) was expressed in chondrocytes already engaged in maturation such as those present in chick limbs, chondrocyte maturation and bone formation were greatly enhanced. Differential responses to Wnt/beta-catenin signaling were confirmed in cultured chondrocytes. Activation in immature cells blocked maturation and actually de-stabilized their phenotype, as revealed by reduced expression of chondrocyte markers, abnormal cytoarchitecture, and loss of proteoglycan matrix. Activation in mature cells instead stimulated hypertrophy, matrix mineralization, and expression of terminal markers such as metalloprotease (MMP)-13 and vascular endothelial growth factor. Because proteoglycans are crucial for cartilage function, we tested possible mechanisms for matrix loss. Delta-beta-catenin expression markedly increased expression of MMP-2, MMP-3, MMP-7, MMP-9, MT3-MMP, and ADAMTS5. In conclusion, Wnt/beta-catenin signaling regulates chondrocyte phenotype, maturation, and function in a developmentally regulated manner, and regulated action by this pathway is critical for growth plate organization, cartilage boundary definition, and endochondral ossification.
Tamamura
Yoshihiro
Y
Department of Orthopaedic Surgery, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Otani
Tomohiro
T
Kanatani
Naoko
N
Koyama
Eiki
E
Kitagaki
Jirota
J
Komori
Toshihisa
T
Yamada
Yoshihiko
Y
Costantini
Frank
F
Wakisaka
Satoshi
S
Pacifici
Maurizio
M
Iwamoto
Masahiro
M
Enomoto-Iwamoto
Motomi
M
D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; D000818:Animals; D002356:Cartilage / Q000196:embryology* / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D019902:Chondrocytes / Q000378:metabolism*; D017364:Collagenases / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000096:biosynthesis* / Q000235:genetics*; D005121:Extremities / Q000196:embryology; D018507:Gene Expression Regulation, Developmental; D006132:Growth Plate / Q000196:embryology* / Q000378:metabolism; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D053509:Matrix Metalloproteinase 13; D053513:Matrix Metalloproteinase 16; D020778:Matrix Metalloproteinase 2 / Q000096:biosynthesis; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis; D020783:Matrix Metalloproteinase 7 / Q000096:biosynthesis; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000096:biosynthesis; D000090023:Metallothionein 3; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D010012:Osteogenesis; D010641:Phenotype; D012313:RNA / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013997:Time Factors; D015534:Trans-Activators / Q000096:biosynthesis* / Q000235:genetics*; D051176:beta Catenin
C495267:CTNNB1 protein, mouse; D003598:Cytoskeletal Proteins; D000090023:Metallothionein 3; C507104:Mmp16 protein, mouse; C000715430:Mt3 protein, mouse; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D051176:beta Catenin; D012313:RNA; D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; C498489:Adamts5 protein, mouse; D017364:Collagenases; D053509:Matrix Metalloproteinase 13; D053513:Matrix Metalloproteinase 16; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; C507081:Mmp13 protein, mouse; D019278:Matrix Metalloproteinase 3; D020783:Matrix Metalloproteinase 7; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9