A novel mutation in the WNT6 gene of congenital tooth agenesis / 口腔疾病防治

Objective @#This article explores the relationship between congenital tooth agenesis and related gene mutations, providing a reference for early diagnosis of the disease.@*Methods @# Clinical and radiographic examinations of a rare case of congenital tooth agenesis were conducted to evaluate the abnormal morphology and quantity of the teeth, as well as the overall health of the patient. Bidirectional sequencing of the PAX9 and MSX1 genes and whole-exome sequencing were conducted to identify potential genetic abnormalities. Sanger sequencing of the newly discovered mutation site was performed on the proband's son. Subsequently, the impacts of the mutations were evaluated through computational tools and a cell-based gene transfection assay. @*Results @#This is a rare case of tooth agenesis characterized by a congenitally missing first molar, a second molar with one single root and a supernumerary second premolar in the right mandibular dentition. The c.717 C>C/T in PAX9 is synonymous. The c.119C>G in MSX1 is a missense mutation predicted to be “benign” by Polyphen. Through whole-exome sequencing, we found a novel mutation, c.637-7 C>A in intron 3 of the WNT6 gene, which is predicted by MAXENT to influence the splicing of mRNA. Both the proband and his son carry this mutation. A cell-based gene transfection assay demonstrated that it did not alter the mRNA splicing of WNT6. @* Conclusion @#The interaction between single nucleotide polymorphisms may contribute to congenital tooth agenesis.

Detección polimórfica del rs104893850 de MSX1 y rs28933373 de PAX9 en personas con agenesia dental no sindrómica / Polymorphic detection of rs104893850 of MSX1 and rs28933373 of PAX9 in people with non syndromic dental agnesia

Introducción: la agenesia dental no sindrómica (ADNS) genera efec- tos negativos en la salud oral y psicosocial de los seres humanos. El determinante genético desempeña un papel importante en su desarrollo. Objetivo: determinar la frecuencia de los polimorfismos rs104893850 de MSX1 y rs28933373 de PAX9 en pacientes de seis a 18 años con ADNS. Material y métodos: estudio transversal prolectivo en el cual se revisaron individuos de seis a 18 años sin defectos congénitos y originarios del estado de Durango. Después de haber obtenido su con- sentimiento para formar parte del estudio, se estableció el diagnóstico de ADNS a través de una inspección clínica odontológica y un examen radiográfico. Se tomó una muestra de sangre capilar para la genotipi- ficación de los polimorfismos a través de la técnica de qPCR-HRM. Resultados: de un total de 124 individuos, 77 (62%) mujeres y 47 (38%) hombres; sólo 39 presentaron ADNS. En el análisis polimórfico de rs104893850 de MSX1 y rs28933373 de PAX9 se obtuvo 94.9% y 84.6% respectivamente de homocigotos mutados. Conclusiones: se obtuvo una alta frecuencia de hipodoncia, el diente que mostró más agenesia fue el órgano dentario 18. Las mutaciones polimórficas están presentes en una alta proporción de agenesia dental (AU)

Introduction: non-syndromic dental agenesis (NSDA) generates negative oral health and psychosocial effects in humans. The genetic determinant plays an important role in its development. Objective: to determine the frequency of MSX1 rs104893850 and PAX9 rs28933373 polymorphisms in patients aged 6 to 18 years with NSDA. Material and methods: prolective cross-sectional study, in which individuals aged 6 to 18 years without congenital defects and from the city of Durango were reviewed. After obtaining their consent to be part of the study, the diagnosis of NSDA was established through a clinical dental inspection, a radiographic examination and a capillary blood sample was taken for the genotyping of the polymorphisms through the qPCR-HRM technique. Results: out of a total of 124 individuals, 77 (62%) females and 47 (38%) males; only 39 presented ADNS. In the polymorphic analysis of rs104893850 of MSX1 and rs28933373 of PAX9 we obtained 94.9% and 84.6% respectively of mutated homozygotes. Conclusions: a high frequency of hypodontia was obtained, and the tooth that presented the most agenesis was dental organ 18. Polymorphic mutations are present in a high proportion for dental agenesis (AU)

Preliminary study of PAX9 single nucleotide polymorphism (rs8004560) in patients with Class II skeletal base malocclusion contributed by mandibular retrognatism

@#Polymorphism in PAX9 (rs8004560), a gene responsible for craniofacial and tooth development, is often associated with Class II/Div2 malocclusion. This study aimed to detect the presence of PAX9 SNP (rs8004560) and to determine its genotype and allele distribution in Class II skeletal base malocclusion, contributed by retrognathic mandible, in the local Malaysian population. The association of PAX9 SNP (rs8004560) with Class II skeletal base malocclusion was also determined. A case control study was performed on 30 samples; 15 from Class II skeletal base malocclusion, and 15 from Class I skeletal base subject as control. Cephalometric measurements were performed prior to saliva samples collection. Genomic DNA was extracted from unstimulated saliva of all subjects, and the DNA was amplified using specific primers for marker rs8004560, followed by genotyping by sequencing. SHEsis online software was used to analyse Hardy-Weinberg Equilibrium (HWE) for cases and controls. Allelic and genotypic frequencies were compared between cases and controls. Significant difference in allele frequency was observed within the group whereby G allele was over-represented in the analysed population (p<0.05). However, when compared between cases and control; there was no significant association between PAX9 SNP (rs8004560) with Class II skeletal base malocclusion (p=0.56, OR=0.71; 95% CI=0.225-2.246). The distribution of genotype frequency in both groups were consistent with HWE (p>0.05). Although no genetic association between PAX9 SNP (rs8004560) with Class II skeletal base malocclusion was observed, significant difference in allele frequency observed might provide some indication in the involvement of PAX9 polymorphism in Class II skeletal base malocclusion contributed by retrognathic mandible. Further research utilising larger sample size will be required in order to determine the role of PAX9 gene in the aetiology of Class II skeletal base malocclusion observed in the local Malaysian population.

IRF6, RYK, and PAX9 expression in facial tissue of children with cleft palate / Expresión de IRF6, RYK y PAX9 en el tejido facial de niños con paladar fisurado

Cleft lip and palate (CLP) is a congenital anomaly characterized by the inappropriate fusion of the upper lip, alveolus, and secondary palate. This study investigated whether expression of interferon regulatory fac tor 6 (IRF6), receptor-like tyrosine kinase (RYK), and paired-box 9 (PAX9), which are essential for the normal development and morphogenesis of craniofacial structures, is dysregulated in children with CLP. Oral mucosa tissue samples were obtained from patients with complete bilateral (CB) CLP (n= 19) during corrective plastic surgery and unaffected control subjects (n= 7). IRF6, RYK, and PAX9 expression was assessed by immunohistochemistry, and data were analyzed with the Mann-Whitney test. In patients, IRF6 immunoreactivity in the connective tissue was moderate to high, but the overall number of IRF6-positive oral epithelial cells was lower than that in controls (z= -3.41; P= 0.01). RYK expression was observed only sporadically in the oral epithelium of 4 patients, in contrast to the control group (z= -3.75; P< 0.001). PAX9-positive epithelial cells were present in low to moderate numbers in patients with CBCLP, while an abundance of these cells was observed in the basal layer of the oral epithelium in controls (z= -3.60; P<0.001). IRF6 is the main connective tissue regulatory factor in CBCLP, and its low level of expression in the oral epithelium suggests a reduced potential for epitheliocyte differentiation, while low PAX9 and RYK expression may explain the decreased cell migration and cleft remodeling in CBCLP.

La fisura labial y palatina (FLP) son anomalías congénitas caracterizadas por la fusión inadecuada del labio superior, alvéolo y paladar secundario. En este estudio se investigó si en niños con FLP hay una desregulación de la expresión del factor regulador de interferón 6 (IRF6), del receptor de la tirosina quinasa (RYK), y del factor de transcripción PAX9, que son esenciales para el desarrollo normal y la morfogénesis de las estructuras craneofaciales. Se obtuvieron muestras de la mucosa oral de pacientes con FLP completa bilateral (CB) (n= 19), tomadas durante la realización de cirugía plástica correctiva, y de sujetos de control no afectados (n= 7). Se evaluó la expresión de IRF6, RYK y PAX9 por inmunohistoquímica, y los datos se analizaron con la prueba de Mann-Whitney. En los pacientes, la inmunoreactividad de IRF6 en el tejido conectivo fue de moderada a alta, pero el número total de células epiteliales orales positivas para IRF6 fue menor que en los controles (z= -3,41; P= 0,01). La expresión de RYK se observó sólo esporádicamente en el epitelio oral de 4 pacientes, en contraste con el grupo control (z= -3,75; P<0.001). Células epiteliales positivas para PAX9 estaban presentes en números bajos a moderados en pacientes con FLP completa bilateral, mientras que se observó una abundante cantidad de estas células en la capa basal del epitelio oral en los controles (z= -3,60; P<0,001). IRF6 es el principal factor regulador del tejido conectivo con FLP completa bilateral, y su bajo nivel de expresión en el epitelio oral sugiere un potencial reducido para la diferenciación del epitelio, mientras que la expresión baja de PAX9 y RYK pueden explicar la disminución de la migración celular y la remodelación de la fisura con FLP completa bilateral.

Clinical and genetic evaluation of a family with isolated oligodontia / 实用医学杂志

Objectives To investigate the mutational characteristics of MSX1 and PAX9 genes in a family affected by non-syndromic oligodonti so as to study the pathogenesis of oligodontia from a molecular prospective. Methods A family with oligodontia, but of different descent and unrelated healthy controls were enrolled in our study. Genomic DNA was isolated from the blood samples. Mutation analyses were performed by amplifying MSX1 and PAX9 exons and sequencing the products. Results DNA sequencing revealed a novel missense mutation c.348C>T in a highly conserved homeobox sequence of MSX1 and a known polymorphisms c. 469+35- c.469+45del in exon 1 and in intron in the two patients and in two unrelated healthy controls. But we did not detect any mutation in PAX9. Conclusion Our finding suggests the samesense mutation (c.348C>T) and the polymorphisms (c.469+35- c.469+45del) may be responsible for oligodontia phenotype in this Chinese family.

PAX9 polymorphism and susceptibility to sporadic non-syndromic severe anodontia: a case-control study in southwest China

Our research aimed to look into the clinical traits and genetic mutations in sporadic non-syndromic anodontia and to gain insight into the role of mutations of PAX9, MSX1, AXIN2 and EDA in anodontia phenotypes, especially for the PAX9. Material and Methods The female proband and her family members from the ethnic Han families underwent complete oral examinations and received a retrospective review. Venous blood samples were obtained to screen variants in the PAX9, MSX1, AXIN2, and EDA genes. A case-control study was performed on 50 subjects with sporadic tooth agenesis (cases) and 100 healthy controls, which genotyped a PAX9 gene polymorphism (rs4904210). Results Intra-oral and panoramic radiographs revealed that the female proband had anodontia denoted by the complete absence of teeth in both the primary and secondary dentitions, while all her family members maintained normal dentitions. Detected in the female proband were variants of the PAX9 and AXIN2 including A240P (rs4904210) of the PAX9, c.148C>T (rs2240308), c.1365A>G (rs9915936) and c.1386C>T (rs1133683) of the AXIN2. The same variants were present in her unaffected younger brother. The PAX9 variations were in a different state in her parents. Mutations in the MSX1 and EDA genes were not identified. No significant diferences were found in the allele and genotype frequencies of the PAX9 polymorphism between the controls and the subjects with sporadic tooth agenesis. Conclusions These results suggest that the association of A240P with sporadic tooth agenesis still remains obscure, especially for different populations. The genotype/phenotype correlation in congenital anodontia should be verified. .

Bases moleculares de la agenesia dental no sindrómica: revisión de la información bibliográfica / Molecular bases of non-syndromic dental agenesis: a review of the literature

Este trabajo pretende actualizar los conocimientos acerca de las bases moleculares de la agenesia dental no sindrómica. Más de doscientos genes codifican múltiples proteínas con funciones necesarias para el desarrollo dental. Los factores de transcripción MSXC-1 y PAX-9 son fundamentales para activar la expresión proteica sinérgica de la cascada de señalización de las proteínas morfogénicas óseas, responsables de la progresión secuencial de la odontogénesis. En busca de las posibles causas de agenesias dentarias no sindrómicas, se han detectado dieciocho mutaciones de tipo missense de pares de dominio del gen humano PAX-9, mutaciones de haplo-insuficiencia funcional de los genes PAX-9 y MSX-1 y múltiples polimorfismos de localizaciones diversas. En todos los casos fue notable la disminución del nivel de expresión de las proteínas mutantes (a las que los genes antes mencionados codifican como transcriptores), la cual afectó la capacidad de unión al ADN de éstas. El impacto deletéreo de estas mutaciones para generar agenesias dentarias selectivas continúa siendo objeto de estudio.

Hipodoncia y hueso navicular accesorio: una interesante asociación sindrómica / Hypodontia and accessory navicular bone: an interesting syndromic association

Los dientes se desarrollan a partir de múltiples interacciones recíprocas entre células del epitelio oral y el ectomesénquima. Una serie de genes participan en el desarrollo del diente, así como de otros órganos y miembros superiores e inferiores. PAX9, miembro de la familia de factores de transcripción, es uno de los principales responsables de este desarrollo, y juega un rol fundamental en la hipodoncia dental y malformaciones en los huesos del miembro inferior. El objetivo de este reporte es presentar un caso de asociación entre hipodoncia y el hueso navicular accesorio. Se trata de una paciente de 18 años de edad, que acude a la consulta dental por dolor agudo en la pieza dentaria 3.6 y dolor de 9 meses de evolución en la cara interna del pie izquierdo. Al examen radiográfico, se observó ausencia de las piezas dentarias 1.7, 2.7, 2.8, 3.8 y 4.8, además en el pie izquierdo se vio la presencia de un hueso navicular accesorio. Es interesante proponer esta rara asociación, de clara herencia autosómica dominante, dado que la agenesia dental y la presencia del hueso navicular accesorio, poseen una prevalencia semejante, lo que hace que pueda existir una nueva asociación sindrómica probablemente relacionada con la ausencia de PAX9(AU)

Teeth are developed from many interactions between oral epithelium and mesenchymal cells. A number of genes are involved in tooth development, as well as in other organs, and upper and lower limbs. PAX9, a member of the transcriptional factor family, is one of the main drivers of this development, playing a key role in dental hypodontia and malformations in the lower limb bones. The aim of this report was to present the association between hypodontia and the accessory navicular bone based on a case report. This is a 18 years old female patient, who attended a dental clinic because she had acute pain in the 3.6 tooth and also pain on the left foot's inner area. The radiographic examination showed loss of the teeth 1.7, 2.7, 2.8, 3.8 and 4.8; and the presence of an accessory navicular bone in the left foot. It is interesting to suggest that this rare association, with clear dominant autosomal inheritance, might exist; since the dental agenesis and the presence of accessory navicular bone have similar prevalence, which could point to a new syndromic association probably related to the lack of PAX9(AU)

Aspectos clínicos e moleculares da agenesia dentária congênita / Molecular and clinical aspects of congenital dental agenesis

A agenesia dentária consiste em uma anomalia comum de desenvolvimento, que resulta na alteração do número de dentes preentes na cavidade bucal e afeta aproximadamente 20% da população. Sua etiologia está associada a fatores ambientais, como infecções, traumas, quimioterapia, radioterapia e causas genéticas. Atualmente a etiologia mais aceita para explicar a ocorrência das anomalias dentárias é a alteração na expressão de genes específicos. Com base no conhecimento dos genes e fatores de transcrição envolvidos na odontogênese, presume-se que diferentes formas fenotípicas de agenesia dentária são causadas por mutações em diferentes genes. Os genes envolvidos na agenesia dentária em humanos incluem os fatores de transcrição (MSX1 e PAX9) que desempenham um papel crítico durante o desenvolvimento craniofacial e o gene que codifica uma proteína envolvida na via de sinalização canônica Wnt (AXIN2). Dessa maneira, a proposta do presente estudo é discorrer sobre os principais genes que têm sido relatados como reguladores da formação dental e a ocorrência de mutações nestes genes que poderiam resultar em agenesias dentárias

Dental agenesis is a common developmental anomaly which affects approximately 20% of the population and results in a reduction of number of teeth present in the oral cavity. The etiology is associated with environmental factors, such as infections, trauma, chemotherapy, radiotherapy, and genetic causes. Currently the widely accepted theory to explain the occurrence of dental agenesis is the change in the expression of specific genes. Different phenotypic patterns of dental agenesis are caused by mutations in genes and transcription factors involved in odontogenesis. In humans those genes include transcription factors (MSX1 and PAX9) that play a critical role during development and the gene coding for a protein involved in the canonical Wnt signaling (AXIN2). Therefore, the purpose of this study is to discuss about dental agenesis and the key genes that have been reported as regulators of dental formation and how the occurrence of mutations in these genes could result in dental agenesis

Transcriptional analysis of the human PAX9 promoter

OBJECTIVES: PAX9 belongs to the Pax family of transcriptional factor genes. This gene is expressed in embryonic tissues such as somites, pharyngeal pouch endoderm, distal limb buds and neural crest-derived mesenchyme. Polymorphisms in the upstream promoter region of the human PAX9 have been associated with human non-syndromic tooth agenesis. In the present study, we verified the in vitro mRNA expression of this gene and the luciferase activity of two constructs containing promoter sequences of the PAX9 gene. MATERIAL AND METHODS: Embryonic tissues were obtained from digits, face, and midbrain/hindbrain regions. Fragments containing PAX9 promoter sequences were cloned into reporter plasmids and were transfected into the different cell cultures. mRNA were extracted from primary cell cultures. RESULTS: The semi-quantitative RT-PCR results showed that in vitro E13.5 limb bud and CNS cells express PAX9, but cells derived from the facial region do not. Moreover, the luciferase assay showed that protein activity of the constructed vector was weaker than pgl3 -basic alone. CONCLUSIONS: The present results suggest that the promoter sequences analyzed are not sufficient to drive PAX9 gene transcription.

Localization Patterns of Cytokeratin 1, 14 and PAX 9 in Mouse Embryonic Tongue Development / 체질인류학회지

Epithelial differentiation and morphogenesis in skin and oral mucosa were elucidated using various experimental tools. However, tongue epithelial differentiation has not been examined properly yet. In this study, we identified the relationship between morphological changes and localizations of differentiation markers, such as cytokeratins and PAX 9 in mice embryonic tongue development. Protective barrier formation and localization pattern of cytokeratins in tongue epithelium were examined with toluidine blue staining and immunohistochemistry respectively. Localization patterns of PAX 9 and Cytokeratin 14 were coincided during tongue epithelium development. In addition, compared with Ki67 localizations, marker for cell proliferation, localization patterns of PAX 9 and Cytokeratin 14 would suggest that these factors would involve in tongue barrier formation through cell proliferation. Based on these results, tongue epithelial differentiation would begin at E14 with the specific localizations of PAX 9 and Cytokeratin 14 prior to protective barrier formation then Cytokeratin 1, keratinization marker, would involve in protective barrier and filiform papillae formations.

Association between polymorphism in the promoter region (G/C-915) of PAX9 gene and third molar agenesis

PURPOSE: Hypodontia is the congenital absence of one or more (up to six) permanent and/or deciduous teeth, being one of the most common alterations of the human dentition. Genetic polymorphisms are variations of DNA sequences occurring in a population. This study investigated whether G-915C single nucleotide polymorphism (SNPs) in the PAX9 gene promoter is associated with hypodontia in humans. MATERIAL AND METHODS: The polymorphism in region G/C-915 of PAX9 gene (NCBI ref SNP ID: rs 2073247) of 240 patients was analyzed, being 110 controls and 130 individuals with third molar agenesis. After DNA extraction, the region of interest was amplified by PCR technique using two different primers. The significance of the differences in observed frequencies of polymorphisms in both groups was assessed by odds-ratio and chi-squared test with 95 percent confidence interval. RESULTS: Genotype CC was more frequent in patients with agenesis (11.5 percent) compared to the control (1.8 percent), while GG was more prevalent in the control group (39.1 percent) compared to the individuals with agenesis (26.2 percent). CONCLUSION: These data showed that the allele C could be associated with the third molar agenesis.