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Connexin43 deficiency causes delayed ossification, craniofacial abnormalities, and osteoblast dysfunction.

Lecanda F, Warlow PM, Sheikh S, Furlan F, Steinberg TH, Civitelli R - J. Cell Biol. (2000)

Bottom Line: We have shown that overexpression of Cx45 in osteoblasts expressing endogenous Cx43 leads to decreased cell-cell communication (Koval, M., S.T.Cell to cell diffusion of calcein was poor among Cx43-deficient osteoblasts, whose differentiated phenotypic profile and mineralization potential were greatly impaired, compared with wild-type cells.Cell to cell signaling, mediated by Cx43 gap junctions, was critical for normal osteogenesis, craniofacial development, and osteoblastic function.

View Article: PubMed Central - PubMed

Affiliation: Divisions of Bone and Mineral and Infectious Diseases, Department of Internal Medicine, Washington University School of Medicine, Barnes-Jewish Hospital, St. Louis, Missouri 63110, USA.

ABSTRACT
Connexin(Cx)43 is the major gap junction protein present in osteoblasts. We have shown that overexpression of Cx45 in osteoblasts expressing endogenous Cx43 leads to decreased cell-cell communication (Koval, M., S.T. Geist, E.M. Westphale, A.E. Kemendy, R. Civitelli, E.C. Beyer, and T.H. Steinberg. 1995. J. Cell Biol. 130:987-995) and transcriptional downregulation of several osteoblastic differentiation markers (Lecanda, F., D.A. Towler, K. Ziambaras, S.-L. Cheng, M. Koval, T.H. Steinberg, and R. Civitelli. 1998. Mol. Biol. Cell 9:2249-2258). Here, using the Cx43- mouse model, we determined whether genetic deficiency of Cx43 affects skeletal development in vivo. Both intramembranous and endochondral ossification of the cranial vault were delayed in the mutant embryos, and cranial bones originating from migratory neural crest cells were also hypoplastic, leaving an open foramen at birth. Cx43-deficient animals also exhibited retarded ossification of the clavicles, ribs, vertebrae, and limbs, demonstrating that skeletal abnormalities are not restricted to a neural crest defect. However, the axial and appendicular skeleton of Cx43- animals were essentially normal at birth. Cell to cell diffusion of calcein was poor among Cx43-deficient osteoblasts, whose differentiated phenotypic profile and mineralization potential were greatly impaired, compared with wild-type cells. Therefore, in addition to the reported neural crest cell defect, lack of Cx43 also causes a generalized osteoblast dysfunction, leading to delayed mineralization and skull abnormalities. Cell to cell signaling, mediated by Cx43 gap junctions, was critical for normal osteogenesis, craniofacial development, and osteoblastic function.

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Mandible development in Cx43−/− mice. Alizarin red/alcian blue staining of the mandibles at (a and b) E18.5 and at (c and d) birth of wild-type (WT) and Cx43−/− littermates. In newborn mutants, the mandible was smaller, with a more rounded alveolar ridge and a flatter arch. This malformation contributes to the slightly smaller and more pointed snout in mutant animals. In some Cx43- newborn mice, cartilaginous primordia of the temporal bone and the condylar region of the mandible were still present at (d) birth Note, (b, arrow) the lack of the incisor tooth at E18.5 in the Cx43−/− embryo and (d) the less prominent tooth at birth. Also, note the less pronounced alveolar ridge (AR) in the mutant animals and the cartilaginous coronoid process (CP).
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Figure 2: Mandible development in Cx43−/− mice. Alizarin red/alcian blue staining of the mandibles at (a and b) E18.5 and at (c and d) birth of wild-type (WT) and Cx43−/− littermates. In newborn mutants, the mandible was smaller, with a more rounded alveolar ridge and a flatter arch. This malformation contributes to the slightly smaller and more pointed snout in mutant animals. In some Cx43- newborn mice, cartilaginous primordia of the temporal bone and the condylar region of the mandible were still present at (d) birth Note, (b, arrow) the lack of the incisor tooth at E18.5 in the Cx43−/− embryo and (d) the less prominent tooth at birth. Also, note the less pronounced alveolar ridge (AR) in the mutant animals and the cartilaginous coronoid process (CP).

Mentions: The maxilla and mandibula are derived from neural crest cells populating the first pharyngeal arch, and they undergo both endochondral (Meckel's cartilage) or intramembranous ossification (vomer, palatine, mandibular, and premaxilla). At E16.5, Meckel's cartilage was the only mandibular element present in the homozygous Cx43−/− mutants, whereas robust ossification within a membranous primordium in the inferior part of the ramus was already present at this stage in normal animals (Fig. 1, a and b). Those differences disappeared at E18.5 when maxillary bones and mandible were almost completely mineralized in both mutant and wild-type mice (Fig. 1g and Fig. h, and Fig. 2, a and b). However, the mandible was reduced in size in Cx43- mice at E18.5 (Fig. 2, a and b) and at birth (Fig. 2c and Fig. d). As a consequence, the lower incisors were not as prominent as in normal littermates at birth (Fig. 2c and Fig. d, arrow), suggesting a delay in tooth development and eruption. Nonetheless, the temporo-mandibular joint was apparently normal in the mutant mice.


Connexin43 deficiency causes delayed ossification, craniofacial abnormalities, and osteoblast dysfunction.

Lecanda F, Warlow PM, Sheikh S, Furlan F, Steinberg TH, Civitelli R - J. Cell Biol. (2000)

Mandible development in Cx43−/− mice. Alizarin red/alcian blue staining of the mandibles at (a and b) E18.5 and at (c and d) birth of wild-type (WT) and Cx43−/− littermates. In newborn mutants, the mandible was smaller, with a more rounded alveolar ridge and a flatter arch. This malformation contributes to the slightly smaller and more pointed snout in mutant animals. In some Cx43- newborn mice, cartilaginous primordia of the temporal bone and the condylar region of the mandible were still present at (d) birth Note, (b, arrow) the lack of the incisor tooth at E18.5 in the Cx43−/− embryo and (d) the less prominent tooth at birth. Also, note the less pronounced alveolar ridge (AR) in the mutant animals and the cartilaginous coronoid process (CP).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2169447&req=5

Figure 2: Mandible development in Cx43−/− mice. Alizarin red/alcian blue staining of the mandibles at (a and b) E18.5 and at (c and d) birth of wild-type (WT) and Cx43−/− littermates. In newborn mutants, the mandible was smaller, with a more rounded alveolar ridge and a flatter arch. This malformation contributes to the slightly smaller and more pointed snout in mutant animals. In some Cx43- newborn mice, cartilaginous primordia of the temporal bone and the condylar region of the mandible were still present at (d) birth Note, (b, arrow) the lack of the incisor tooth at E18.5 in the Cx43−/− embryo and (d) the less prominent tooth at birth. Also, note the less pronounced alveolar ridge (AR) in the mutant animals and the cartilaginous coronoid process (CP).
Mentions: The maxilla and mandibula are derived from neural crest cells populating the first pharyngeal arch, and they undergo both endochondral (Meckel's cartilage) or intramembranous ossification (vomer, palatine, mandibular, and premaxilla). At E16.5, Meckel's cartilage was the only mandibular element present in the homozygous Cx43−/− mutants, whereas robust ossification within a membranous primordium in the inferior part of the ramus was already present at this stage in normal animals (Fig. 1, a and b). Those differences disappeared at E18.5 when maxillary bones and mandible were almost completely mineralized in both mutant and wild-type mice (Fig. 1g and Fig. h, and Fig. 2, a and b). However, the mandible was reduced in size in Cx43- mice at E18.5 (Fig. 2, a and b) and at birth (Fig. 2c and Fig. d). As a consequence, the lower incisors were not as prominent as in normal littermates at birth (Fig. 2c and Fig. d, arrow), suggesting a delay in tooth development and eruption. Nonetheless, the temporo-mandibular joint was apparently normal in the mutant mice.

Bottom Line: We have shown that overexpression of Cx45 in osteoblasts expressing endogenous Cx43 leads to decreased cell-cell communication (Koval, M., S.T.Cell to cell diffusion of calcein was poor among Cx43-deficient osteoblasts, whose differentiated phenotypic profile and mineralization potential were greatly impaired, compared with wild-type cells.Cell to cell signaling, mediated by Cx43 gap junctions, was critical for normal osteogenesis, craniofacial development, and osteoblastic function.

View Article: PubMed Central - PubMed

Affiliation: Divisions of Bone and Mineral and Infectious Diseases, Department of Internal Medicine, Washington University School of Medicine, Barnes-Jewish Hospital, St. Louis, Missouri 63110, USA.

ABSTRACT
Connexin(Cx)43 is the major gap junction protein present in osteoblasts. We have shown that overexpression of Cx45 in osteoblasts expressing endogenous Cx43 leads to decreased cell-cell communication (Koval, M., S.T. Geist, E.M. Westphale, A.E. Kemendy, R. Civitelli, E.C. Beyer, and T.H. Steinberg. 1995. J. Cell Biol. 130:987-995) and transcriptional downregulation of several osteoblastic differentiation markers (Lecanda, F., D.A. Towler, K. Ziambaras, S.-L. Cheng, M. Koval, T.H. Steinberg, and R. Civitelli. 1998. Mol. Biol. Cell 9:2249-2258). Here, using the Cx43- mouse model, we determined whether genetic deficiency of Cx43 affects skeletal development in vivo. Both intramembranous and endochondral ossification of the cranial vault were delayed in the mutant embryos, and cranial bones originating from migratory neural crest cells were also hypoplastic, leaving an open foramen at birth. Cx43-deficient animals also exhibited retarded ossification of the clavicles, ribs, vertebrae, and limbs, demonstrating that skeletal abnormalities are not restricted to a neural crest defect. However, the axial and appendicular skeleton of Cx43- animals were essentially normal at birth. Cell to cell diffusion of calcein was poor among Cx43-deficient osteoblasts, whose differentiated phenotypic profile and mineralization potential were greatly impaired, compared with wild-type cells. Therefore, in addition to the reported neural crest cell defect, lack of Cx43 also causes a generalized osteoblast dysfunction, leading to delayed mineralization and skull abnormalities. Cell to cell signaling, mediated by Cx43 gap junctions, was critical for normal osteogenesis, craniofacial development, and osteoblastic function.

Show MeSH
Related in: MedlinePlus