Msx1 and Dlx5 function synergistically to regulate frontal bone development.

The Msx and Dlx families of homeobox proteins are important regulators for embryogenesis. Loss of Msx1 in mice results in multiple developmental defects including craniofacial malformations. Although Dlx5 is widely expressed during embryonic development, targeted null mutation of Dlx5 mainly affects the development of craniofacial bones. Msx1 and Dlx5 show overlapping expression patterns during frontal bone development. To investigate the functional significance of Msx1/Dlx5 interaction in regulating frontal bone development, we generated Msx1 and Dlx5 double null mutant mice. In Msx1(-/-) ;Dlx5(-/-) mice, the frontal bones defect was more severe than that of either Msx1(-/-) or Dlx5(-/-) mice. This aggravated frontal bone defect suggests that Msx1 and Dlx5 function synergistically to regulate osteogenesis. This synergistic effect of Msx1 and Dlx5 on the frontal bone represents a tissue specific mode of interaction of the Msx and Dlx genes. Furthermore, Dlx5 requires Msx1 for its expression in the context of frontal bone development. Our study shows that Msx1/Dlx5 interaction is crucial for osteogenic induction during frontal bone development.

Stem cell property of postmigratory cranial neural crest cells and their utility in alveolar bone regeneration and tooth development.

The vertebrate neural crest is a multipotent cell population that gives rise to a variety of different cell types. We have discovered that postmigratory cranial neural crest cells (CNCCs) maintain mesenchymal stem cell characteristics and show potential utility for the regeneration of craniofacial structures. We are able to induce the osteogenic differentiation of postmigratory CNCCs, and this differentiation is regulated by bone morphogenetic protein (BMP) and transforming growth factor-beta signaling pathways. After transplantation into a host animal, postmigratory CNCCs form bone matrix. CNCC-formed bones are distinct from bones regenerated by bone marrow mesenchymal stem cells. In addition, CNCCs support tooth germ survival via BMP signaling in our CNCC-tooth germ cotransplantation system. Thus, we conclude that postmigratory CNCCs preserve stem cell features, contribute to craniofacial bone formation, and play a fundamental role in supporting tooth organ development. These findings reveal a novel function for postmigratory CNCCs in organ development, and demonstrate the utility of these CNCCs in regenerating craniofacial structures.

Postoperative stability after sagittal split ramus osteotomies for a mandibular setback with monocortical plate fixation or bicortical screw fixation.

PURPOSE: This comparative study was performed to analyze mandibular stability after bilateral sagittal split ramus osteotomies for a mandibular setback with a monocortical plate fixation or bicortical screw fixation. PATIENTS AND METHODS: A total of 60 patients with a skeletal Class III malocclusion who underwent sagittal split osteotomies and mandibular setback were included in the present study. Among the patients, 30 were osteosynthesized monocortically with a titanium plate, while the other 30 were osteosynthesized bicortically with positioning screws. This retrospective study used cephalometric radiographs taken preoperatively (T0), and at 1 week (T1), 3 months (T2), and 1 year (T3) postoperatively. The linear and angular changes of the cephalometric landmarks of the chin region were measured at each time period, and the changes of each cephalometric landmark were determined for 4 different time intervals: T0 to T1 (Delta T1), T1 to T2 (Delta T2), T2 to T3 (Delta T3), and T0 to T3 (Delta T4). Postoperative changes in mandibular shape were analyzed using a paired t test to determine the stability of fixation methods. RESULTS: The surgical changes (Delta T1) could be characterized as posterior movements of the mandibular body and posterosuperior movements of the chin landmark. The results of the cephalometric analysis at 3 and 12 months postsurgery (Delta T2 and Delta T3) could be characterized as anterosuperior movements of the mandible. In both treatment groups, surgical changes were relatively well maintained. Likewise, analysis of the horizontal and vertical movements of the chin landmarks indicated a similar tendency in both groups. There were no statistically significant differences between the measurements of postoperative changes at each time period after surgery. CONCLUSIONS: The findings of our study suggest that there were no significant differences in postoperative changes in mandibular shape in both the monocortical and bicortical fixation groups after sagittal split ramus osteotomy. We concluded that monocortical osteosynthesis using a miniplate could be used to obtain stable postoperative changes after mandibular setback.

Smad4 is required to regulate the fate of cranial neural crest cells.

Smad4 is the central mediator for TGF-beta/BMP signals, which are involved in regulating cranial neural crest (CNC) cell formation, migration, proliferation and fate determination. It is unclear whether TGF-beta/BMP signals utilize Smad-dependent or -independent pathways to control the development of CNC cells. To investigate the functional significance of Smad4 in regulating CNC cells, we generated mice with neural crest specific inactivation of the Smad4 gene. Our study shows that Smad4 is not required for the migration of CNC cells, but is required in neural crest cells for the development of the cardiac outflow tract. Smad4 is essential in mediating BMP signaling in the CNC-derived ectomesenchyme during early stages of tooth development because conditional inactivation of Smad4 in neural crest derived cells results in incisor and molar development arrested at the dental lamina stage. Furthermore, Smad-mediated TGF-beta/BMP signaling controls the homeobox gene patterning of oral/aboral and proximal/distal domains within the first branchial arch. At the cellular level, a Smad4-mediated downstream target gene(s) is required for the survival of CNC cells in the proximal domain of the first branchial arch. Smad4 mutant mice show underdevelopment of the first branchial arch and midline fusion defects. Taken together, our data show that TGF-beta/BMP signals rely on Smad-dependent pathways in the ectomesenchyme to mediate epithelial-mesenchymal interactions that control craniofacial organogenesis.

Regulating the role of bone morphogenetic protein 4 in tooth bioengineering.

PURPOSE: Culture of the whole organ and regulation of its development using biologic and engineering principles can be used to produce structures and organs for reconstructing defects. The application of these bioengineering approaches in artificial tooth development may be the alternative way to replace missing dentition. MATERIALS AND METHODS: For the artificial bioengineering of a mouse tooth, tooth buds were dissected and transplanted into the diastema of the developing mandible. The mandiblular primordia containing transplanted tooth buds were culture in vitro and in vivo using a bioengineering method. In addition, to regulate the development of tooth germs, bone morphogenetic protein 4 (BMP4) or its antagonist, Noggin was administered. RESULTS: After the period of in vitro and in vivo culture, the transplanted tooth germ in the diastema showed tooth development with supportive structure formation. In the BMP-treated group, the bioengineered tooth was observed with increased maturation of cusp and enamel matrix. However, in the Noggin-treated tooth germs, the developing molar had a crater-like appearance with the immature development of the cusp and suppressed formation of the enamel matrix. CONCLUSIONS: This study confirmed that tooth germ transplantation in the diastema and culture with administration of BMP4 could lead to the mature development of the dental structures. In addition, these results suggest the possibility of bioengineering the tooth in morphogenesis and differentiation even in the toothless area.

Determination of specific interactions between glucose ligand carrying polymer and glucose transporter type-1 (GLUT-1) using different cell types.

In order to develop a biomimetic polymer for cell recognition, poly [3-O-(4'-vinylbenzyl)-D-glucose] (PVG) and different types of glucose transport (GLUT)-carrying cells, namely, HepG2 cells (GLUT-1), 3T3-L1 fibroblast cells (GLUT-1 and GLUT-4), and MIN6 cells (GLUT-2), were tested for specific interaction. To clarify the nature of interaction between PVG and the different cell types, rhodamine-B isothiocyanate (RITC)-labeled polymers were used to prove and visualize the interactions. In this study, we found that labeled polymer strongly binds to HepG2 cells. The fluorescence intensity of PVG with in the presence of HepG2 cells was found to be stronger than that of 3T3-L1 fibroblast cells (0.11 +/- 0.05) and MIN6 cells, which carry GLUT-2 (0.028 +/- 0.01); a confocal laser microscopic study confirmed this interaction. To confirm the specificity of the interaction mediated by GLUT, the cells were pretreated with phloretin, an inhibitor of GLUT-1, before adding RITC-labeled PVG polymer to the cell culture medium. This treatment suppressed the interaction of PVG with HepG2 cells and partially suppressed its interaction with 3T3-L1 fibroblast cells.