Yoko Takigawa

Sox9 Family Members Negatively Regulate Maturation and Calcification of Chondrocytes through Up-Regulation of Parathyroid Hormone–related Protein

Sox9 is a transcription factor that plays an essential role in chondrogenesis and has been proposed to inhibit the late stages of endochondral ossification. However, the molecular mechanisms underlying the regulation of chondrocyte maturation and calcification by Sox9 remain unknown. In this study, we attempted to clarify roles of Sox9 in the late stages of chondrocyte differentiation. We found that overexpression of Sox9 alone or Sox9 together with Sox5 and Sox6 (Sox5/6/9) inhibited the maturation and calcification of murine primary chondrocytes and up-regulated parathyroid hormone-related protein (PTHrP) expression in primary chondrocytes and the mesenchymal cell line C3H10T1/2. Sox5/6/9 stimulated the early stages of chondrocyte proliferation and development. In contrast, Sox5/6/9 inhibited maturation and calcification of chondrocytes in organ culture. The inhibitory effects of Sox5/6/9 were rescued by treating with anti-PTHrP antibody. Moreover, Sox5/6/9 bound to the promoter region of the PTHrP gene and up-regulated PTHrP gene promoter activity. Interestingly, we also found that the Sox9 family members functionally collaborated with Ihh/Gli2 signaling to regulate PTHrP expression and chondrocyte differentiation. Our results provide novel evidence that Sox9 family members mediate endochondral ossification by up-regulating PTHrP expression in association with Ihh/Gli2 signaling.

The transcription factor Znf219 regulates chondrocyte differentiation by assembling a transcription factory with Sox9

Sox9 is an essential transcription factor for chondrogenesis by regulating the expression of chondrogenic genes. However, its regulatory mechanism is not fully understood. To address this, we attempted to identify the transcriptional partners of Sox9 by screening the cDNA library of the chondrogenic cell line ATDC5 using the collagen 2α1 (Col2α1) gene promoter fused to a luciferase reporter gene. One of the positive clones encoded the Znf219 gene. Whole mount in situ hybridization experiments indicated that Znf219 mRNA was specifically expressed in the developing limb buds where Col2α1 and Sox9 were strongly expressed. Znf219 markedly enhanced the transcriptional activity of Sox9 on the Col2a1 gene promoter. In addition, Znf219 is physically associated with Sox9 and is colocalized with Sox9 in the nucleus. We also found that overexpression of Znf219 profoundly increased Sox9-induced mRNA expression of Col2a1, aggrecan and Col11a2. Consistently, knockdown of Znf219 decreased the Sox9-induced mRNA expression of these genes. Furthermore, a dominant-negative mutant Znf219 inhibited Bmp2-induced chondrocyte differentiation. Our results suggest that Znf219 plays an important role in the regulation of chondrocyte differentiation as a transcriptional partner of Sox9.

Msx2 Stimulates Chondrocyte Maturation by Controlling Ihh Expression

Several studies indicated that a homeobox gene, Msx2, is implicated in regulation of skeletal development by controlling enchondral ossification as well as membranous ossification. However, the molecular basis by which Msx2 conducts chondrogenesis is currently unclear. In this study, we examined the role of Msx2 in chondrocyte differentiation using mouse primary chondrocytes and embryonic metatarsal explants. Treatment with BMP2 up-regulated the expression of Msx2 mRNA along with chondrocyte differentiation in murine primary chondrocytes. Overexpression of wild-type Msx2 stimulated calcification of primary chondrocytes in the presence of BMP2. We also found that constitutively active Msx2 (caMsx2) enhanced BMP2-dependent calcification more efficiently than wild-type Msx2. Consistently, caMsx2 overexpression up-regulated the expression of alkaline phosphatase and collagen type X induced by BMP2. Furthermore, organ culture experiments using mouse embryonic metatarsals indicated that caMsx2 clearly stimulated the maturation of chondrocytes into the prehypertrophic and hypertrophic stages in the presence of BMP2. In contrast, knockdown of Msx2 inhibited maturation of primary chondrocytes. The stimulatory effect of Msx2 on chondrocyte maturation was enhanced by overexpression of Smad1 and Smad4 but inhibited by Smad6, an inhibitory Smad for BMP2 signaling. These data suggest that Msx2 requires BMP2/Smad signaling for its chondrogenic action. In addition, caMsx2 overexpression induced Ihh (Indian hedgehog) expression in mouse primary chondrocytes. Importantly, treatment with cyclopamine, a specific inhibitor for hedgehogs, blocked Msx2-induced chondrogenesis. Collectively, our results indicated that Msx2 promotes the maturation of chondrocytes, at least in part, through up-regulating Ihh expression.

Arid5a cooperates with Sox9 to stimulate chondrocyte-specific transcription

This study shows that Arid5a interacts with Sox9 and subsequently modulates histone 3 acetylation of a chondrogenic gene, Col2a1, and stimulates chondrocyte differentiation.

Regulation of endochondral ossification by transcription factors.

Abstract Endochondral ossification is very unique and complex biological event which is associated with skeletal development and tissue partnering. Genetic studies and gene-targeting approaches identified several transcription factors that play important roles in endochondral ossification. These transcription factors sequentially and harmoniously regulate each step of endochondral ossification, and consequently maintain the spatio-temporal control of the program. Importantly, these transcription factors form large protein complex to control chromatin remodeling, histone modification, transcription and splicing steps during endochondral ossification. It is also important to understand how these transcription factors regulate expression of their target genes. Biochemical and molecular cloning techniques largely contributed to identification of the components of the transcriptional complex and the target genes. Most recently, importance of endoplasmic reticulum (ER) stress in endochondral ossification has been reported. A transcription factor, BBF2H7, functions as an ER stress sensor in chondrocytes through regulation of appropriate secretion of chondrogenic matrices. We would like to discuss how the transcription factors regulate endochondral ossification.

Breakage of internal maxillary distractor: considerable complication of maxillary distraction osteogenesis

Maxillary distraction osteogenesis using intraoral distractors is now one of the standard treatments of maxillary retrusion. This report shows 2 cases of breakage of this internal maxillary distractor in patients with cleft lip and palate; one was observed during the distraction period and the other was during the retention period. The first case required a rotational movement of the distraction segment, and this movement caused the laterally dislocation of the posterior part of the distractor, where the distractor suffered some mechanical forces by mouth opening. In the latter case, breakage of distractor was observed on the radiographs taken 3 months after distraction and this complication may have been caused by mechanical force by occlusion and mastication. Both breakages were found at the joint of the anchorage plate and the extension rod, which has some flexibility for adjusting the plate to the bone surface. Therefore, surgeons should pay special attention for this mechanical weak area in this distractor not only during the advancement period, but also during the retention period and should avoid unnecessary frequent bending for adopting the bone surface, which directly weakens the joint.

Maxillary advancement using distraction osteogenesis with intraoral device.

This article describes the surgical orthodontic treatment of maxillary hypoplasia in a patient with cleft lip and palate using maxillary distraction osteogenesis with internal maxillary distractors. Maxillary advancement was performed to correct the retrusive maxillary facial profile and Class III malocclusion. Rotational movement of the distraction segment was made to correct the upper dental midline. Although maxillary advancement was insufficient because of unexpected breakage of the intraoral distractor after completion of the distraction, skeletal traction with a face mask compensated for the shortage. Successful esthetic improvement and posttreatment occlusal stability were achieved with no discernible relapse after 2 years of retention.

The outcome of a two-phase treatment in a patient with Angle Class II, Division 1 malocclusion and an excessive overjet

Abstract This article describes a two-phase orthodontic approach to treating a young adolescent girl that had Angle Class II, Division 1 malocclusion associated with an excessive incisor overjet caused by prolonged digit sucking. The Phase I treatment utilized a tongue crib and a straight-pull headgear appliance during puberty and achieved significant improvements in the facial profile. The Phase II treatment involved edgewise appliances, a transpalatal arch and a straight-pull headgear appliance without extraction of permanent teeth. Dental and skeletal corrections were made with the two-phase treatment and stable occlusion and good facial proportion were obtained.

Improvement in three-dimensional facial configuration and jaw motion following surgical orthodontic treatment of a case with jaw deviation

Abstract The establishment of normal stomatognathic function and an acceptable facial appearance is considered an important treatment goal in orthodontics. The patient, who demonstrated posterior crossbites and jaw deviation, underwent treatment with a combined orthodontic-surgical approach utilizing maxillary transverse distraction osteogenesis, Le Fort I osteotomy, and bilateral SSRO. The smoothness of the masticatory jaw-closing movement and three-dimensional facial configuration were compared pre- and post-treatment. Correction of the posterior crossbites enabled the patient to perform smoother jaw-closing movements during chewing. Additionally, asymmetry in the morphology of the chin, cheek, and lips was reduced. These favorable changes in masticatory jaw movement kinematics and three-dimensional evaluation of the face justify the application of surgical-orthodontic treatment to improve jaw function and facial appearance.

Nonextraction treatment of open-bite by sequential uses of tongue crib, temporary anchorage devices and myofunctional therapy: A case report of an adolescent

Abstract This report of a case describes the nonsurgical and nonextraction treatment of a female adolescent with severe skeletal open-bite using the tongue crib appliance, the quad-helix appliance, temporary anchorage devices (TADs) and myofunctional therapy. The tongue crib appliance and the quad-helix appliance were used to eliminate the digit sucking habit and to modify the tongue position. With TADs, the overerupted molars were intruded. After the post-active treatment, daily gum-chewing exercise for 15 min and lip closure training were performed to produce effective retention. Successful esthetic improvement from convex type profile to straight type profile was achieved, and post-treatment occlusal stability was maintained after 3-year retention with the strengthened masticatory muscle tone and modified tongue position.