Craniofacial Bone Development Research Articles

Preliminary investigation into the impact of BPA on osteoblast activity and bone development: In vitro and in vivo models

Bisphenol A (BPA), an ingredient in consumer products, has been suggested that it can interfere with bone development and maintenance, whereas the molecule mechanism remains unclear. The objective of this study is to investigate the effect of BPA on early bone differentiation and metabolism, and its potential molecule mechanism by employing hFOB1.19 cell as an in vitro model, as well as larval zebrafish as an in vivo model. The in vitro experiments indicated that BPA decreased cell viability, inhibited osteogenic activity (such as ALP, RUNX2), increased ROS production, upregulated transcriptional or protein levels of apoptosis-related molecules (such as Caspase 3, Caspase 9), while suppressed transcriptional or protein levels of pyroptosis-specific markers (TNF-α, TNF-β, IL-1β, ASC, Caspase 1, and GSDMD). Moreover, the evidences from in vivo model demonstrated that exposure to BPA distinctly disrupted pharyngeal cartilage, craniofacial bone development, and retarded bone mineralization. The transcriptional level of bone development-related genes (bmp2, dlx2a, runx2, and sp7), apoptosis-related genes (bcl2), and pyroptosis-related genes (cas1, nlrp3) were significantly altered after treating with BPA in zebrafish larvae. In summary, our study, combining in vitro and in vivo models, confirmed that BPA has detrimental effects on osteoblast activity and bone development. These effects may be due to the promotion of apoptosis, the initiation of oxidative stress, and the inhibition of pyroptosis.

Loss of Sc5d results in micrognathia due to a failure in osteoblast differentiation

IntroductionMutations in genes related to cholesterol metabolism, or maternal diet and health status, affect craniofacial bone formation. However, the precise role of intracellular cholesterol metabolism in craniofacial bone development remains unclear. ObjectiveThe aim of this study is to determine how cholesterol metabolism aberrations affect craniofacial bone development. MethodsMice with a deficiency in Sc5d, which encodes an enzyme involved in cholesterol synthesis, were analyzed with histology, micro computed tomography (microCT), and cellular and molecular biological methods. ResultsSc5d null mice exhibited mandible hypoplasia resulting from defects in osteoblast differentiation. The activation of the hedgehog and WNT/β-catenin signaling pathways, which induce expression of osteogenic genes Col1a1 and Spp1, was compromised in the mandible of Sc5d null mice due to a failure in the formation of the primary cilium, a cell surface structure that senses extracellular cues. Treatments with an inducer of hedgehog or WNT/β-catenin signaling or with simvastatin, a drug that restores abnormal cholesterol production, partially rescued the defects in osteoblast differentiation seen in Sc5d mutant cells. ConclusionOur results indicate that loss of Sc5d results in mandibular hypoplasia through defective primary cilia-mediated hedgehog and WNT/β-catenin signaling pathways.

Impact of maternal topiramate ingestion on ossification of skull and appendicular bones in rat fetuses

The skull and appendicular bones are derived from different embryological sources during their development. The impact of prenatal exposure of topiramate on ossification of these bones is not adequately studied. The goal of this study was to assess the ossification patterns of the craniofacial bones and bones of the forelimbs and hindlimbs in 20-day-old rat fetuses after maternal exposure to topiramate at doses equivalent to human therapeutic doses. Three groups of Sprague-Dawley pregnant rats were used: control, topiramate 50mg/kg/day (T50) and topiramate 100mg/kg/day (T100). Topiramate was given by oral gavage from day 6 to day19 of gestation. Ossification was evaluated in the bones of 20 days fetuses after staining with Alizarin red. Results showed a significant reduction in complete ossified centers of the metacarpal, metatarsal and craniofacial bones in topiramate-exposed fetuses at both doses when compared to the control group. Also, a significant decrease in the length of ossified part of the long bones of the forelimbs and hindlimbs in topiramate-exposed fetuses at both doses was noted when compared to the control group. Crown-rump length and fetal weight were significantly decreased in topiramate treated groups compared to the control group. In all examined groups, there was a positive correlation between the crown-rump length and the lengths of humerus and femur. No abnormalities in the ossified bones and no significant changes in their ossification pattern were observed between the treated groups. In conclusion, prenatal administration of topiramate in doses equivalent to human therapeutic doses delayed ossification and development of craniofacial and appendicular bones in rat fetuses and their effects are not dose dependent at doses investigated. The implications of these findings in women who require topiramate therapy in pregnancy merit further evaluation.

The Role of Gli1+ Mesenchymal Stem Cells in Osteogenesis of Craniofacial Bone.

Glioma-associated oncogene homolog 1 (Gli1) is a transcriptional activator of hedgehog (Hh) signaling that regulates target gene expression and several cellular biological processes. Cell lineage tracing techniques have highlighted Gli1 as an ideal marker for mesenchymal stem cells (MSCs) in vivo. Gli1+ MSCs are critical for the osteogenesis of the craniofacial bone; however, the regulatory mechanism by which Gli1+ MSCs mediate the bone development and tissue regeneration of craniofacial bone has not been systematically outlined. This review comprehensively elucidates the specific roles of Gli1+ MSCs in craniofacial bone osteogenesis. In addition to governing craniofacial bone development, Gli1+ MSCs are associated with the tissue repair of craniofacial bone under pathological conditions. Gli1+ MSCs promote intramembranous and endochondral ossification of the craniofacial bones, and assist the osteogenesis of the craniofacial bone by improving angiopoiesis. This review summarizes the novel role of Gli1+ MSCs in bone development and tissue repair in craniofacial bones, which offers new insights into bone regeneration therapy.

Enhanced BMP signaling leads to enlarged nasal cartilage formation in mice

Bone morphogenetic proteins (BMPs) are required for craniofacial bone development. However, it remains elusive how BMP signaling regulates craniofacial cartilage development. To address this question, we utilized a genetic system to enhance BMP signaling via one of BMP type I receptors ALK2 in a chondrocyte-specific manner (hereafter Ca-Alk2:Col2-Cre) in mice. Ca-Alk2:Col2-Cre mice died shortly after birth due to severe craniofacial abnormalities including cleft palate, defective tongue, and shorter mandible formation. Histological analysis revealed that these phenotypes were attributed to the extensive chondrogenesis. Compared with controls, enhanced SOX9 and RUNX2 production were observed in nasal cartilage of Ca-Alk2:Col2-Cre mice. To reveal the mechanisms responsible for enlarged nasal cartilage, we examined Smad-dependent and Smad-independent BMP signaling pathways. While the Smad-independent BMP signaling pathway including p38, ERK, and JNK remained silent, the Smad1/5/9 was highly phosphorylated in Ca-Alk2:Col2-Cre mice. Interestingly, Ca-Alk2:Col2-Cre mice showed enhanced S6 kinase phosphorylation, a readout of mammalian target of rapamycin complex 1 (mTORC1). These findings may suggest that enhanced Smad-dependent BMP signaling positively regulates the mTOR pathway and stimulates chondrocytes toward hypertrophic differentiation, thereby leading to enlarged nasal cartilage formation in mice.

Cranial Neural Crest Cells Contribution to Craniofacial Bone Development and Regeneration.

This review aims to summarize (i) the latest evidence on cranial neural crest cells (CNCC) contribution to craniofacial development and ossification; (ii) the recent discoveries on the mechanisms responsible for their plasticity; and (iii) the newest procedures to ameliorate maxillofacial tissue repair. CNCC display a remarkable differentiation potential that exceeds the capacity of their germ layer of origin. The mechanisms by which they expand their plasticity was recently described. Their ability to participate to craniofacial bone development and regeneration open new perspectives for treatments of traumatic craniofacial injuries or congenital syndromes. These conditions can be life-threatening, require invasive maxillofacial surgery and can leave deep sequels on our health or quality of life. With accumulating evidence showing how CNCC-derived stem cells potential can ameliorate craniofacial reconstruction and tissue repair, we believe a deeper understanding of the mechanisms regulating CNCC plasticity is essential to ameliorate endogenous regeneration and improve tissue repair therapies.

FGF signaling in cranial suture development and related diseases.

Suture mesenchymal stem cells (SMSCs) are a heterogeneous stem cell population with the ability to self-renew and differentiate into multiple cell lineages. The cranial suture provides a niche for SMSCs to maintain suture patency, allowing for cranial bone repair and regeneration. In addition, the cranial suture functions as an intramembranous bone growth site during craniofacial bone development. Defects in suture development have been implicated in various congenital diseases, such as sutural agenesis and craniosynostosis. However, it remains largely unknown how intricate signaling pathways orchestrate suture and SMSC function in craniofacial bone development, homeostasis, repair and diseases. Studies in patients with syndromic craniosynostosis identified fibroblast growth factor (FGF) signaling as an important signaling pathway that regulates cranial vault development. A series of in vitro and in vivo studies have since revealed the critical roles of FGF signaling in SMSCs, cranial suture and cranial skeleton development, and the pathogenesis of related diseases. Here, we summarize the characteristics of cranial sutures and SMSCs, and the important functions of the FGF signaling pathway in SMSC and cranial suture development as well as diseases caused by suture dysfunction. We also discuss emerging current and future studies of signaling regulation in SMSCs.

The LRP5 high-bone-mass mutation causes alveolar bone accrual with minor craniofacial alteration.

Mutations in low-density lipoprotein receptor-related protein 5 (LRP5) cause various bone diseases. Several mouse models were generated to study the role of LRP5 in bone development. But most of the studies were confined to the appendicular skeleton. The role of LRP5 in the axial skeleton, especially in the craniofacial skeleton, is largely unknown. The aim of this study was to investigate the craniofacial phenotype with the LRP5G171V mutation. To understand how LRP5 affects craniofacial bone properties, we analyzed LRP5 high-bone-mass mutant mice carrying the G171V missense mutation (LRP5HBM ). Quantitative microcomputed tomographic imaging and histomorphometric analyses were used to study craniofacial phenotypes and bone density. Histology, immunohistochemistry, and in vivo fluorochrome labeling were used to study molecular mechanisms. LRP5HBM mice showed overall minor changes in the craniofacial bone development but with increased bone mass in the interradicular alveolar bone, edentulous ridge, palatine bone, and premaxillary suture. Elevated osteocyte density was observed in LRP5HBM mice, along with increased Runx2 expression and unmineralized bone surrounding osteocytes. Meanwhile, LRP5HBM mice exhibited increased osteoprogenitors, but no significant changes were observed in osteoclasts. This led to a high-bone-mass phenotype, and an increased osteocyte density in the alveolar bone and edentulous ridge. LRP5HBM mice display increased bone mass in the alveolar bone with minor changes in the craniofacial morphology. Collectively, these data elucidated the important role of LRP5 in axial bone development and homeostasis and provided clues into the therapeutical potential of LRP5 signaling in treating alveolar bone loss.

Is mandible derived mesenchymal stromal cells superior in proliferation and regeneration to long bone-derived mesenchymal stromal cells?

Mesenchymal stromal cells (MSCs) are cells with the characteristic ability of self-renewal along with the ability to exhibit multilineage differentiation. Bone marrow (BM) is the first tissue in which MSCs were identified and BM-MSCs are most commonly used among various MSCs in clinical settings. MSCs can stimulate and promote osseous regeneration. Due to the difference in the development of long bones and craniofacial bones, the mandibular-derived MSCs (M-MSCs) have distinct differentiation characteristics as compared to that of long bones. Both mandibular and long bone-derived MSCs are positive for MSC-associated markers such as CD-73, -105, and -106, stage-specific embryonic antigen 4 and Octamer-4, and negative for hematopoietic markers such as CD-14, -34, and -45. As the M-MSCs are derived from neural crest cells, they have embryogenic cells which promote bone repair and high osteogenic potential. In vitro and in vivo animal-based studies demonstrate a higher rate of proliferation and high osteogenic potential for M-MSCs as compared to long-bones MSCs, but in vivo studies in human subjects are lacking. The BM-MSCs have their advantages and limitations. M-MSCs may be utilized as an alternative source of MSCs which can be utilized for tissue engineering and promoting the regeneration of bone. M-MSCs may have potential advantages in the repair of craniofacial or orofacial defects. Considering the utility of M-MSCs in the field of orthopaedics, we have discussed various unresolved questions, which need to be explored for their better utility in clinical practice.

Determining Bone-forming Ability and Frequency of Skeletal Stem Cells by Kidney Capsule Transplantation and Limiting Dilution Assay.

Adult stem cells not only maintain tissue homeostasis but are also critical for tissue regeneration during injury. Skeletal stem cells are multipotent stem cells that can even generate bones and cartilage upon transplantation to an ectopic site. This tissue generation process requires essential stem cell characteristics including self-renewal, engraftment, proliferation, and differentiation in the microenvironment. Our research team has successfully characterized and isolated skeletal stem cells (SSCs) from the cranial suture called suture stem cells (SuSCs), which are responsible for craniofacial bone development, homeostasis, and injury-induced repair. To assess their stemness features, we have demonstrated the use of kidney capsule transplantation for an in vivo clonal expansion study. The results show bone formation at a single-cell level, thus permitting a faithful assessment of stem cell numbers at the ectopic site. The sensitivity in assessing stem cell presence permits using kidney capsule transplantation to determine stem cell frequency by limiting dilution assay. Here, we described detailed protocols for kidney capsule transplantation and limiting dilution assay. These methods are extremely valuable both for the evaluation of skeletogenic ability and the determination of stem cell frequency.

Research progress on the role of transforming growth factor-β in tooth and craniofacial bone development and diseases

Research progress on the role of transforming growth factor-β in tooth and craniofacial bone development and diseases

Influences of Airway Obstruction Caused by Adenoid Hypertrophy on Growth and Development of Craniomaxillofacial Structure and Respiratory Function in Children.

Adenoid hypertrophy (AH) is a common disease in otorhinolaryngology. Children with chronic snoring and hypoxia are susceptible to long-term nasal obstruction, while long-term open-mouth breathing may cause craniofacial bone development disorders and dull facial expressions, the so-called adenoid face. The purpose of this work is to analyze the influence of AH-induced airway obstruction (AO) on the growth and development of craniomaxillofacial structure and respiratory function (RF) in children. The clinical data of 56 AH children (observation group) and 42 healthy children with physical examination (control group) who visited the Hebei Eye Hospital during the same period were retrospectively analyzed. All children received acoustic rhinometry and X-ray cephalometric measurements. The upper airway structure, sleep disorder score, and A/N value of nasopharyngeal lateral X-ray images were compared between cases and controls. For AH children, sleep tests were also performed to assess their RF. X-ray cephalometric measurements of facial morphology showed obvious vertical growth, mandibular retrognathia, and enlarged mandibular angle in AH children. AH mainly affects the size of the nasopharyngeal and oropharyngeal airway. AH children presented with higher nasal airway resistance (5.11 ± 1.95 cmH2O/L min) and lower nasopharyngeal volume (NPV) (16.86 ± 3.93 cm3) than controls. Of the AH children, 45 had abnormal RF, including 4 with obstructive sleep apnea syndrome. The A/N value of nasopharyngeal lateral X-ray images was significantly higher in AH children than in controls. Besides, worse sleep quality was found in AH children. The above differences were all of statistical significance. The above indicates that AH can affect the size of the nasopharyngeal and oropharyngeal airway, change children's respiratory mode and RF, increase nasal resistance, and decrease NPV, resulting in upper respiratory tract stenosis, as well as craniomaxillofacial and oral malformations, which affects children's normal growth and development.

Single nucleotide polymorphisms MYO1H 1001 C>T SNP (rs3825393) is a strong risk factor for mandibular prognathism

Mandibular prognathism (MP) is a common craniofacial disorder of Class III malocclusion that causes esthetic and functional problems. Class III malocclusion diversity is influenced by both environmental and genetic factors. Single nucleotide polymorphisms (SNPs) in genes involved in craniofacial morphogenesis, bone and cartilage development, and metabolism, could play a role as predisposing factors. The present study aimed to establish a potential association between MATN1 -1878 A>G (rs1149048), MYO1H 1001 C>T (rs3825393), and BMP-4 538 A>G (rs17563) SNPs and MP in Serbian population. The study included 110 participants: 55 patients with Class III malocclusion diagnosed with MP and 55 with Class I malocclusion. The 3 SNPs were analyzed using the polymerase chain reaction-restriction fragment length polymorphism method. The genotype frequency of MYO1H showed a highly significant difference between patients and controls. Heterozygous carriers of the T allele had an almost 3-fold increase in odds for the development of MP (odds ratio, 2.79; 95% confidence interval, 1.26-6.19; P= 0.010). No association could be established between MATN1 and BMP-4 polymorphisms and MP. Our results support the concept of gene polymorphisms as risk modulators in mandibular prognathism development, although only the association between MYO1H and MP was found in Serbian patients with Class III malocclusion.

Genetic Interaction of Thm2 and Thm1 Shapes Postnatal Craniofacial Bone.

Ciliopathies are genetic syndromes that link skeletal dysplasias to the dysfunction of primary cilia. Primary cilia are sensory organelles synthesized by intraflagellar transport (IFT)—A and B complexes, which traffic protein cargo along a microtubular core. We have reported that the deletion of the IFT-A gene, Thm2, together with a null allele of its paralog, Thm1, causes a small skeleton with a small mandible or micrognathia in juvenile mice. Using micro-computed tomography, here we quantify the craniofacial defects of Thm2−/−; Thm1aln/+ triple allele mutant mice. At postnatal day 14, triple allele mutant mice exhibited micrognathia, midface hypoplasia, and a decreased facial angle due to shortened upper jaw length, premaxilla, and nasal bones, reflecting altered development of facial anterior-posterior elements. Mutant mice also showed increased palatal width, while other aspects of the facial transverse, as well as vertical dimensions, remained intact. As such, other ciliopathy-related craniofacial defects, such as cleft lip and/or palate, hypo-/hypertelorism, broad nasal bridge, craniosynostosis, and facial asymmetry, were not observed. Calvarial-derived osteoblasts of triple allele mutant mice showed reduced bone formation in vitro that was ameliorated by Hedgehog agonist, SAG. Together, these data indicate that Thm2 and Thm1 genetically interact to regulate bone formation and sculpting of the postnatal face. The triple allele mutant mice present a novel model to study craniofacial bone development.

Expression of Cre recombinase in chondrocytes causes abnormal craniofacial and skeletal development.

The cranial base synchondroses are growth centers that drive cranial and upper facial growth. The intersphenoid synchondrosis (ISS) and the spheno-occipital synchondrosis (SOS) are two major synchondroses located in the middle of the cranial base and are maintained at early developmental stages to sustain cranial base elongation. In this study, we report unexpected premature ossification of ISS and SOS when Cre recombinase is activated in a chondrocyte-specific manner. We used a Cre transgenic line expressing Aggrecan enhancer-driven, Tetracycline-inducible Cre (ATC), of which expression is controlled by a Col2a1 promoter. Neonatal doxycycline injection or doxycycline diet fed to breeders was used to activate Cre recombinase. The premature ossification of ISS and/or SOS led to a reduction in cranial base length and subsequently a dome-shaped skull. Furthermore, the mice carrying either heterozygous or homozygous conditional deletion of Tsc1 or Fip200 using ATC mice developed similar craniofacial abnormalities, indicating that Cre activity itself but not conditional deletion of Tsc1 or Fip200 gene, is the major contributor of this phenotype. In contrast, the Col2a1-Cre mice carrying Cre expression in both perichondrium and chondrocytes and the mice carrying the conditional deletion of Tsc1 or Fip200 using Col2a1-Cre did not manifest the same skull abnormalities. In addition to the defective craniofacial bone development, our data also showed that the Cre activation in chondrocytes significantly compromised bone acquisition in femur. Our data calls for the consideration of the potential in vivo adverse effects caused by Cre expression in chondrocytes and reinforcement of the importance of including Cre-containing controls to facilitate accurate phenotype interpretation in transgenic research.

Dissecting the Role of the Six1‐Associated Genes in Craniofacial Development and Disease

Genetic variants of SIX1, a transcription factor implicated in many cellular processes during craniofacial development, and of its co‐factor EYA1 underlie 50% of Branchio‐oto‐renal syndrome (BOR) cases. BOR is an autosomal dominant disorder characterized by various craniofacial defects, including malformed middle ear ossicles leading to conductive hearing loss. Interestingly, some BOR variants in SIX1 also underlie craniosynostosis (CRS) which indicates that SIX1 has a crucial role in craniofacial bone development. One important aspect of SIX1 function is that it interacts with co‐factors that can induce or repress its transcriptional activity. In this work, we expand our knowledge of the Six1 gene regulatory network with respect to craniofacial development. We first used a Six1 knockout mouse line to assess gene expression profiles of dissected E10.5 mandibular arches which give rise to the middle ear ossicles and lower jaw, via RNA‐sequencing. Our transcriptomic analysis led to the identification of 122 genes, including components of the FGF and WNT signaling pathways, known contributors to craniofacial bone development. In parallel, we used in vitro studies to characterize three novel co‐factors of Six1: Sobp, Mcrs1 and Pa2g4 that were recently identified in Xenopus. Our findings indicate that Sobp and Mcrs1 are bona fide Six1 co‐factors in mouse. We are now studying the genes identified in the RNA‐seq and the novel Six1 co‐factors using in vitro culture of mouse pre‐osteoblastic cells (MC3T3‐E1) to identify their role in bone formation. Altogether, our results identified novel Six1‐associated genes that are potential candidate genes for the unresolved cases of BOR and CRS.

Discerning a Potential Link between MEMO1’s Role in Vascular, Bone, and Craniofacial Development

Embryonic development of craniofacial structures requires the tight integration of multiple tissues and cell‐types. For example, the intersection of neural crest derived mesenchyme with mesoderm derived endothelium (i.e., vasculature) is necessary for craniofacial bone development and mineralization. Despite craniofacial anomalies being the 2nd leading congenital anomaly worldwide, our understanding of how these tissues interact is limited.Through a forward genetic screen, we previously identified a novel role for the protein encoded by the gene mediator of ERBB2‐driven cell motility 1 (Memo1) in craniofacial development. Global loss of Memo1 resulted in a range of defects, including hypo‐mineralization of the craniofacial skeleton, fully penetrant cleft secondary palate, coupled with widespread vascular abnormalities. Interestingly, while neural crest cell specific deletion of Memo1 recapitulated some aspects of global loss of Memo1, several defects were less apparent—suggesting MEMO1 functions in tissues outside the neural crest during craniofacial development. Further, molecular profiling of craniofacial elements in global Memo1mutants identified significantly altered Vegfa expression, coupled with disrupted vascular development, relative to controls. Based on these observations, along with Memo1’s semi‐ubiquitous expression during development, we hypothesized that MEMO1 functions within the endothelial derived vasculature and this function is necessary for proper craniofacial development.To test this hypothesis, we have generated a Memo1 endothelial specific mouse knock‐out model. Similar to Memo1 global knock‐outs, and in contrast to Memo1 neural crest knock‐outs and controls, Memo1 endothelial knock‐outs develop vascular hemorrhaging and edema with variable penetrance. Vascular defects are accompanied by gross alterations to craniofacial shape. We are currently utilizing µCT, cell sorting coupled with gene expression profiling, and ex vivo ‘sprouting’ assays to quantify the impact endothelial loss of Memo1 has on craniofacial bone development and mineralization, MEMO1’s molecular role in craniofacial endothelial cells, and MEMO1’s functional role in endothelial cells, respectively.Overall, in addition to MEMO1’s neural crest cell specific role, our findings extend MEMO1’s function into an additional cell‐type, endothelial cells, during craniofacial development. These data illuminate the molecular pathology for how disrupted neural crest or endothelial cells may lead to human craniofacial disorders.

Meclozine Attenuates the MARK Pathway in Mammalian Chondrocytes and Ameliorates FGF2-Induced Bone Hyperossification in Larval Zebrafish.

Meclozine has been developed as an inhibitor of fibroblast growth factor receptor 3 (FGFR3) to treat achondroplasia (ACH). Extracellular signal regulated kinase (ERK) phosphorylation was attenuated by meclozine in FGF2-treated chondrocyte cell line, but the site of its action has not been elucidated. Although orally administered meclozine promoted longitudinal bone growth in a mouse model of ACH, its effect on craniofacial bone development during the early stage remains unknown. Herein, RNA-sequencing analysis was performed using murine chondrocytes from FGF2-treated cultured tibiae, which was significantly elongated by meclozine treatment. Gene set enrichment analysis demonstrated that FGF2 significantly increased the enrichment score of mitogen-activated protein kinase (MAPK) family signaling cascades in chondrocytes; however, meclozine reduced this enrichment. Next, we administered meclozine to FGF2-treated larval zebrafish from 8 h post-fertilization (hpf). We observed that FGF2 significantly increased the number of ossified vertebrae in larval zebrafish at 7 days post-fertilization (dpf), while meclozine delayed vertebral ossification in FGF2-induced zebrafish. Meclozine also reversed the FGF2-induced upregulation of ossified craniofacial bone area, including ceratohyal, hyomandibular, and quadrate. The current study provided additional evidence regarding the inhibitory effect of meclozine on the FGF2-induced upregulation of MAPK signaling in chondrocytes and FGF2-induced development of craniofacial and vertebral bones.

Functional Matrix Hypothesis: A Review of Literature

Craniofacial growth is a complex process. Most of the treatment approaches have been based on the fundamental biological mechanisms involved in the growth and development of craniofacial bones and teeth. The exact mechanism which controls cranio facial growth has been a matter of debate and research for years together. The concept that “forms follow function” was first proposed by Vander Klaaw. Functional matrix hypothesis (F.M.H.) is actually an extension of this concept given by Moss. F.M.H. includes a functional matrix and skeletal units. Capsule is an envelope that contains functional cranial components sandwiched between the covering layers. It acts indirectly and passively and not by resorption or deposition. Functional matrix cranial analysis can be done in the maxilla and mandible. There are some limitations in F.M.H., so F.M.H. revisited is explained later which include 4 theories in it.Genomic and epi-genetic processes are examples of totally different types of causation- Genomic- formal cause and Epigenetic- efficient cause. Individually both are necessary causes, but neither are sufficient causes alone. They provide both the necessary and sufficient causes for the control of morphogenesis.

Special AT-rich sequence-binding protein 2 (Satb2) synergizes with Bmp9 and is essential for osteo/odontogenic differentiation of mouse incisor mesenchymal stem cells.

ObjectivesMouse incisor mesenchymal stem cells (MSCs) have self‐renewal ability and osteo/odontogenic differentiation potential. However, the mechanism controlling the continuous self‐renewal and osteo/odontogenic differentiation of mouse incisor MSCs remains unclear. Special AT‐rich sequence‐binding protein 2 (SATB2) positively regulates craniofacial patterning, bone development and regeneration, whereas SATB2 deletion or mutation leads to craniomaxillofacial dysplasia and delayed tooth and root development, similar to bone morphogenetic protein (BMP) loss‐of‐function phenotypes. However, the detailed mechanism underlying the SATB2 role in odontogenic MSCs is poorly understood. The aim of this study was to investigate whether SATB2 can regulate self‐renewal and osteo/odontogenic differentiation of odontogenic MSCs.Materials and methods Satb2 expression was detected in the rapidly renewing mouse incisor mesenchyme by immunofluorescence staining, quantitative RT‐PCR and Western blot analysis. Ad‐Satb2 and Ad‐siSatb2 were constructed to evaluate the effect of Satb2 on odontogenic MSCs self‐renewal and osteo/odontogenic differentiation properties and the potential role of Satb2 with the osteogenic factor bone morphogenetic protein 9 (Bmp 9) in vitro and in vivo.Results Satb2 was found to be expressed in mesenchymal cells and pre‐odontoblasts/odontoblasts. We further discovered that Satb2 effectively enhances mouse incisor MSCs self‐renewal. Satb2 acted synergistically with the potent osteogenic factor Bmp9 in inducing osteo/odontogenic differentiation of mouse incisor MSCs in vitro and in vivo.Conclusions Satb2 promotes self‐renewal and osteo/odontogenic differentiation of mouse incisor MSCs. Thus, Satb2 can cooperate with Bmp9 as a new efficacious bio‐factor for osteogenic regeneration and tooth engineering.