Polymorphisms associated with oral clefts as potential markers for oral pre and malignant disorders.

OBJECTIVE: To investigate whether genes in the Wnt pathway, which have been previously associated with both oral clefts and oral squamous cell carcinoma, are also associated with oral potentially malignant disorders (leukoplakia, erythroplakia and lichen planus). MATERIALS AND METHODS: Case-control study: Dataset consisted of clinical information linked to DNA samples from affected subjects diagnosed with oral potential malignant disorders and oral cancer and their matched controls. Individual samples, clinical history, and potential risk factors were obtained through the Dental Registry and DNA Repository project of the School of Dental Medicine, University of Pittsburgh. The rs1533767 (WNT11), rs9879992 (GSK3B), and rs3923087 (AXIN2) were tested. After genomic DNA had been extracted, genotyping was performed blindly to clinical diagnosis status. Representation of genotypes and alleles in affected subjects in comparison to the unaffected individuals was determined using PLINK. Additional analysis was performed to investigate associations between environmental (socioeconomic/lifestyle) risk factors and the oral pathologies studied using STATA. RESULTS: Two of the SNPs tested (rs9879992 in GSK3B and rs3923087 in AXIN2) were statistically, significantly associated with the pathologies studied (p = 0.039 and 0.038, respectively). CONCLUSION: Single-nucleotide polymorphisms in genes in the Wnt pathway were associated with oral potentially malignant disorders.

Coast-or Inland Residence and Differences in the Occurrence of Cleft Lip and Cleft Palate.

OBJETIVE: To analyze if differences in lifestyle and environment between coastal and inland areas are associated with differences in frequency of orofacial cleft types. DESIGN: Populational cross-sectional study. SETTING: All live borns with orofacial cleft registered at Brazilian Live Birth Information System between 1999 and 2020. PARTICIPANTS: 33,699 live borns with orofacial cleft. INTERVENTION: Data from borns with orofacial cleft were collected at Brazilian Live Birth Information System. MAIN OUTCOME MEASURE: Differences in frequencies between the cleft types and covariates were determined using chi-square. Bivariate analysis was done to obtain the prevalence ratio of types of clefts by geographic origin. Multiple logistic regression analysis was used to determine adjusted odds ratios, controlling for covariates, establishing a significance level of p value <0.05. RESULTS: The frequency of cleft types was statistically significant different according to geographic origin (inland x coast). For syndromic clefts, the prevalence ratio for cleft lip with/without palate was 3.6 times higher inland (p value = 0.000). Regarding non-syndromics, the prevalence ratio for cleft lip with/without palate was two times higher inland (p value = 0.000). Logistic regression suggested cleft lip with/without palate was 6.33 more likely to occur in inland regions (p value = 0.000). CONCLUSION: Geographic origin was associated with the type of cleft in Brazil, with a higher prevalence of cleft lip with/without palate in inland areas, compared to cleft palate, which was higher in the coast.

Molar-incisor hypomineralization in a cohort of individuals born with cleft lip and palate.

OBJECTIVE: This study aimed to investigate the frequency of molar-incisor hypomineralization (MIH) in individuals born with cleft lip and or cleft palate. SETTINGS AND SAMPLE: Three hundred eighty-six individuals born with cleft lip and/or palate before orthodontic treatment. METHODS: All the individuals were submitted to a clinical examination and intraoral standardized photos. The registration of MIH was taken by two orthodontists and analysed in association with the cleft type and laterality. The Kruskal-Wallis test and the regression test were used to compare the frequency of molars and incisors affected according to cleft type and laterality, sex and age. RESULTS: We found a frequency of 67.87% of MIH in the studied sample. The frequency varied from 25% (in individuals born with cleft palate) to 77% in individuals born with bilateral cleft lip and palate). The number of affected molars was statistically different depending on cleft type and laterality (P < .001- Kruskal-Wallis test). Differences were found between individuals born with unilateral cleft lip and palate and unilateral cleft lip and alveolus (P = .03), and with isolated cleft palate (P = .03), and between individuals born with bilateral cleft lip and palate and born with unilateral cleft lip and alveolus (P = .01), and cleft palate (P = .01). Sex (P = .21) and age (P = .36) had no influence on the frequency of MIH. A positive correlation was found between the number of molars affected and incisors affected (P < .001). CONCLUSION: Individuals born with cleft lip and palate have a higher frequency of MIH, and the complexity of cleft type was associated with the number of affected molars.

CLEFT RR- New app to Calculate Recurrence Risk in Cleft lip and Palate.

OBJECTIVE: To describe the Cleft Recurrence Risk (Cleft RR) App, designed to be used on genetic counseling for cleft lip and/ or palate. DESIGN: A validation study, single cohort. SETTING: Tertiary care children's Hospital. PATIENTS, PARTICIPANTS: The manual obtained the results of 100 cases undergoing genetic counseling at the cleft lip and palate treatment center. INTERVENTIONS: The application for genetic counseling for cleft lip and/ or palate is designed to calculate quickly the recurrence risk considering the ancestry, cleft type, sex, and family history and thus encourage the implementation of genetic counseling in cleft lip and palate centers around the world. MAIN OUTCOME MEASURE(S): The data were submitted to the Bland-Altman statistics. RESULTS: After defining parameters the application development follows the steps: development, prototyping, and documentation. The validation of the calculated data was performed by comparing the results of 100 cases undergoing genetic counseling at the cleft lip and palate treatment center obtained by the manual method with the results obtained by the mobile app method; the data were submitted to the Bland-Altman statistics and a high concordance was found. CONCLUSIONS: The mobile app for use by healthcare professionals proved to be simple to use, easy to apply, and provided accurate results. Cleft Recurrence Risk is an application for smartphones developed for genetic counseling in cleft lip and palate, supplementary use by health professionals, and should not replace professional performance.

Chapter 4: Development Defects of Enamel and Dentine and Coronal Caries.

The development of the human dentition is prone to disruption due to its delicate and complex nature - including variations in tooth number and anatomical form and in the characteristics of enamel, dentine, and cementum. This chapter will focus on developmental defects of dental enamel (DDE) and dentine (DDD), which can be associated with considerable treatment burden on an individual, often related to the change in dental hard tissue characteristics in those at increased caries risk. DDE are prevalent and can be related to genetic conditions such as amelogenesis imperfecta and environmental challenges such as direct physical trauma to the developing tooth or systemic insults during the different phases of amelogenesis. Phenotypical variability can be great, making diagnosis difficult in many cases. There are two major enamel defects - the quantitative defect of hypoplasia and the qualitative defect of hypomineralization. DDDs are less prevalent than DDEs, with two major DDD types: dentinogenesis imperfecta and dentine dysplasia. The main features of the DDDs are enamel fracture exposing the dentine and subsequent wear, with enlarged pulp spaces in some variants. The appearance may be affected, with bulbous teeth and grey-blue to brown opalescent colouring. With respect to dental caries, developmental defects of the teeth, in themselves, do not cause caries risk; however, they can change the manifestation of the disease due to creating niches for biofilm accumulation and thereby increasing cleaning difficulty and changing the physical and chemical characteristics of dental hard tissues and how they react to cariogenic challenges.

Association between molar hypomineralization, genes involved in enamel development, and medication in early childhood: A preliminary study.

BACKGROUND: Molar hypomineralization (MH) is defined as a multifactorial condition, and thus, its presence may be defined by interactions between environmental and genetic factors. AIM: To evaluate the association between MH, genes involved in enamel development, and the use of medication during pregnancy in early childhood. DESIGN: One hundred and eighteen children, 54 with and 64 without MH, were studied. The data collected included demographics, socioeconomic data, and the medical history of mothers and children. Genomic DNA was collected from saliva. Genetic polymorphisms in ameloblastin (AMBN; rs4694075), enamelin (ENAM; rs3796704, rs7664896), and kallikrein (KLK4; rs2235091) were evaluated. These genes were analyzed by real-time polymerase chain reaction using TaqMan chemistry. The software PLINK was used to compare allele and genotype distributions of the groups and to assess the interaction between environmental variables and genotypes (p < .05). RESULTS: The variant allele KLK4 rs2235091 was associated with MH in some children (odds ratio [OR]: 3.75; 95% confidence interval [CI] = 1.65-7.81; p = .001). Taking medications in the first 4 years of life was also associated with MH (OR: 2.94; 95% CI = 1.02-6.04; p = .041) and specifically in association with polymorphisms in ENAM, AMBN, and KLK4 (p < .05). The use of medications during pregnancy was not associated with MH (OR: 1.37; 95% CI = 0.593-3.18; p = .458). CONCLUSION: The results of this study suggest that taking medication in the postnatal period appears to contribute to the etiology of MH in some evaluated children. There may be a possible genetic influence of polymorphisms in the KLK4 gene with this condition.

COMT rs4818 less common allele is associated with psychological and psychiatric worse indicators in a cohort of individuals born with cleft lip and palate.

OBJECTIVE: Individuals born with cleft lip and palate may face difficulties in speech function, nutrition, facial aesthetics, and long-term care. These difficulties may increase the risk of psychological and psychiatric diseases. This work aimed to test if the variant allele of COMT was carried more frequently among individuals that have psychological and psychiatric outcomes within a cohort of patients born with cleft lip and palate. METHOD: DNA extraction from saliva of two hundred and fifteen individuals born with cleft lip with and/or palate and genotyping was performed, and the frequency of COMT rs4818 alleles was determined. The domain 'Psychological Function' of Cleft-Q™ was used to generate scores for analysis. The scores were computed, and differences in genotype or allele frequencies between individuals with psychological function scores 60 or above and 59 or below were compared. The history of psychiatric illness (family history of psychiatric disease or self-reported psychiatric illness) was registered. RESULTS: Genotype and allele frequencies were compared between individuals with and without a family history of psychiatric illness. Individuals with lower Psychological Function (Cleft-Q™) scores were more likely to be GG (P = .04) or carriers of allele G (P < .001). The reported psychiatric illness and positive family history of psychiatric illness were compared to COMT rs4818 allele and genotype frequencies of individuals without these indicators, and individuals with psychiatric illness and positive family history of psychiatric illness were more likely to carry allele G (P = .03 and P = .008, respectively). CONCLUSION: The study confirms previously suggested role of COMT rs4818 in psychiatric and psychological outcomes in a distinct cohort of patients born with cleft lip and palate.

No association between dopamine D2 receptor (DRD2) alleles and crack/cocaine dependence in Brazilians

INTRODUCTION: The causal mechanisms behind crack/cocaine use are still unknown, but genetic influences are suggested. OBJECTIVE: To investigate the relationship between the genetic polymorphism TaqI (rs1800497) in the dopamine D2 receptor (DRD2) gene and susceptibility to crack/cocaine dependence in a group of addicts to crack/cocaine and a non-addicted group. METHODS: The case group (n=515) was composed of crack/cocaine-dependent men and the control group (n=106) comprised men who were considered not dependent on crack/cocaine. The oral hygiene habits, decayed, missing, and filled teeth index, gingival index, and plaque index were evaluated. The reference single nucleotide polymorphism (rs1800497 C/T) of the DRD2 gene was genotyped by a real-time polymerase chain reaction technique. Student's t-tests for independent samples or the non-parametric Mann-Whitney test were used to compare groups regarding quantitative variables. RESULTS: The case group showed a mean time of 9.91±7.03 years of crack use, and 61.06±92.96 stones/week. The socio-demographic profile of the sample was White, single men, with basic education, blue-collar worker, smoker, and reporting alcohol use. There was a high frequency of gingival inflammation, plaque accumulation, and caries experience. For all genetic models tested, there was no significant difference in the genotypic frequency in rs1800497 of the DRD2 gene, between case and control groups (p>0.05). CONCLUSION: The genetic variant in the DRD2 did not increase the vulnerability to develop crack/cocaine dependence. The complex genetic nature of crack/cocaine dependence and a large variation of DRD2 allele frequencies, depending on the population group sampled, could be one explanation for the no association.

A randomized clinical trial of hyaluronic acid gel pulpotomy in primary molars with 1 year follow-up.

OBJECTIVE: This randomized clinical study aimed to evaluate the success of hyaluronic acid (HA) as a pulpotomy medicament of human primary molars and to compare it with formocresol (FC) and ferric sulphate (FS) pulpotomy treatments up to 12 months. MATERIALS AND METHODS: The study was conducted with 130 primary molars of 44 children. The ethical approval and registration to clinical trials (No: NCT04115358) were completed. After the removal of all the coronal pulp tissue, a 0.5% HA gel, or a FC, or a 20% FS solution were applied randomly to the radicular pulp tissues of the primary molars. Then, the pulp chambers were filled with a zinc oxide eugenol cement and restored either with a composite filling material or with a stainless-steel crown. The treatment success rates of the 3 groups were followed and compared clinically and radiographically at 1st-, 3rd-, 6th- and 12th-months. RESULTS: Primary molars treated with FC, FS and HA dressings were clinically successful 77.5%, 86.8% and 87.5% respectively after 12th-month follow-up (p > .05). Radiographic successes of FC, FS and HA groups were lower than clinical successes (57.6%, 68.8%, 57.9% respectively at the 12th-month) but the difference between the groups was not statistically significant (p > .05). Equivalence analysis assuming not more than 10% difference between the materials suggested that HA was not inferior to FC or FS. CONCLUSIONS: Within the limitations of this study, our randomized clinical trial shows that HA is a promising pulpotomy medicament in primary molars. However, further studies are justified to further improve the HA material success.

Use of collaborative learning as atool to evaluate syndromic craniofacial anomalies / Uso da aprendizagem colaborativa como ferramenta para avaliar anomalias craniofaciais sindrômicas

Almost 30% of oral cleftsare associated with other structural abnormalities.However, little is known on orofacial characteristics related tothese cases since they are not systematically reported. To close this gap, we developed a collaborative learning approach supported by an interprofessional team aiming to systematicallydescribe oral findings and impactthe training of future professionals that hopefully will incorporate these descriptionsintotheir clinical practice. The methodological proposal consisted of small group sessions focusing on a particular syndrome or group of syndromes followed by examiningpatients with those conditions. Twenty cases were examined and studied over one semester andaset of conditions to be identified in the orofacial regionwasdefined. Here, we present a guideline that we suggest that dentists and dental institutions use. We also present the advantages of using collaborative learning as a tool in the training of the clinician (AU).

Quase 30% das fissuras orais estão associadas a outras anormalidades estruturais. No entanto, pouco se sabe sobre as características orofaciais relacionadas a esses casos, uma vez que não são relatados de forma sistemática. Para fechar essa lacuna, desenvolvemos uma abordagem de aprendizagem colaborativa apoiada por uma equipe interprofissional com o objetivo de descrever sistematicamente os achados orais e impactar o treinamento de futuros profissionais que, esperançosamente, irão incorporar essas descrições em sua prática clínica. A proposta metodológica consistia em sessões de pequenos grupos enfocando uma determinada síndrome ou grupo de síndromes seguidas de exame de pacientes com essas condições. Vinte casos foram examinados e estudados ao longo de um semestre e foi definido um conjunto de condições a serem identificadas na região orofacial. Aqui, apresentamos uma diretriz que sugerimos que os dentistas e instituições odontológicas utilizem. Também apresentamos as vantagens de usar a aprendizagem colaborativa como uma ferramenta no treinamento do clínico (AU).

PAX7 gene polymorphism in muscular temporomandibular disorders as potentially related to muscle stem cells.

BACKGROUND: Temporomandibular disorders (TMD) are a group of painful and debilitating disorders, involving the masticatory muscles and/or the temporomandibular joint (TMJ). Chronic TMD pain can be associated with genetic changes in the key muscle development genes. OBJECTIVE: To evaluate the association between polymorphisms in the PAX7 (paired box 7) gene and masticatory myalgia in patients with temporomandibular disorders (TMD). MATERIALS AND METHODS: This is a case-control study. Patients with TMD were divided into two groups: (a) presence of muscular TMD (n = 122) and (b) absence of muscular TMD (n = 49). Genomic DNA was obtained from saliva samples from all participants to allow for genotyping single nucleotide polymorphisms in PAX7 (rs766325 and rs6659735). Over-representation of alleles was tested using chi-square or Fisher's exact tests. Values of p < 0.05 were considered to be statistically significant. RESULTS: Individuals without muscular TMD were less likely to have the PAX7 rs6659735 GG genotype (p = 0.03). No associations were found for PAX7 rs766325. CONCLUSIONS: Alterations in PAX7 may influence muscular pathophysiology and individuals with TMD and the rs6659735 homozygous genotype (GG) are seemingly associated with muscular involvement of the disorder. No associations were found in the region rs766325.

Missing and unerupted teeth in osteogenesis imperfecta.

INTRODUCTION: Osteogenesis imperfecta (OI) is a genetic disorder characterized by bone fragility and craniofacial and dental abnormalities such as congenitally missing teeth and teeth that failed to erupt which are believed to be doubled in OI patients than normal populations and were associated with low oral health quality of life. However, the etiology of these abnormalities remains unclear. To understand the factors influencing missing and unerupted teeth, we investigated their prevalence in a cohort of OI patients as a function of the clinical phenotype (OI type), the genetic variant type, the tooth type and the onset of bisphosphonate treatment. METHOD: A total of 144 OI patients were recruited from The Shriners Hospital, Montreal, Canada, between 2016 and 2017. Patients were evaluated using intraoral photographs and panoramic radiographs. Missing teeth were evaluated in all patients, and unerupted teeth were assessed only in patients ≥15 years old (n = 82). RESULTS: On average, each OI patient had 2.4 missing teeth and 0.8 unerupted teeth, and the most common missing and unerupted teeth were the premolars and the upper second molars, respectively. These phenomena were more prominent in OI type III and IV than in OI type I, and were not sex or age-related. Missing teeth were significantly more common in patients with C-propeptide variants than all other variants (p-value <0.05). Unerupted teeth were significantly more common in patients with α1 and α2 glycine variants or substitutions than in those with haploinsufficiency variants. Early-onset of bisphosphonate treatment would significantly increase the risk of unerupted teeth in patients with OI types III and IV (OR = 1.68, 95% CI (1.15-1.53)). CONCLUSION: The prevalence of missing and unerupted teeth at the tooth type level in OI patients varies according to the nature of the collagen variants and the OI type. These findings highlight the role of collagen in tooth development and eruption.

FGF10 and FGF13 genetic variation and tooth-size discrepancies.

OBJECTIVES: To explore whether variations in odontogenesis-related genes are associated with tooth-size discrepancies. MATERIALS AND METHODS: Measurements of the width of permanent teeth were obtained from dental casts of 62 orthodontic patients (age 15.65 ± 6.82 years; 29 males and 33 females). Participants were classified according to the anterior and overall Bolton ratios as without tooth-size discrepancy or with maxillary or mandibular tooth-size excess. Genomic DNA extracted from buccal cells was used, and 13 single nucleotide polymorphisms (SNPs) across nine genes were genotyped by polymerase chain reaction using TaqMan chemistry. χ2 or Fisher exact tests were applied to determine the overrepresentation of genotypes/alleles depending on the type of tooth-size discrepancy (α = .05; corrected P value: P < 5.556 × 10-3). Odds ratios (ORs) and their correspondent 95% confidence intervals (CIs) were also calculated to investigate the risk of this phenotype for the SNPs having significant association. RESULTS: Individuals carrying the FGF10 rs900379 T allele were more likely to have larger mandibular teeth (OR = 3.74; 95% CI: 1.65-8.47; P = .002). This effect appeared to be stronger when two copies of the risk allele (TT) were found (recessive model, OR = 6.16; 95% CI: 1.71-22.16; P = .006). On the other hand, FGF13 rs5931572 rare homozygotes (AA, or male A hemizygotes) had increased risk of displaying tooth-size discrepancies when compared with the common homozygotes (GG, or male G hemizygotes; OR = 10.32; 95% CI: 2.20-48.26; P = .003). CONCLUSIONS: The results suggest that FGF10 and FGF13 may contribute to the presence of tooth-size discrepancies.

Multiple Structural Microform Defects Suggest Role of Modifier Genes.

ABSTRACT: One of the biggest challenges in clinical genetics is establishing associations between specific germline mutations and the resulting spectrum of phenotypes. The careful characterization of clinical presentations continues to be a tool for establishing these genotype phenotype correlations. The authors intend, by presenting a case study, proposing that the concomitant occurrence of a combinations of mild structural anomalies in the same individual may be due to changes in genes that can be linked by related pathways. A new born with cleft lip and palate was referred at the Cleft Lip and Palate Center. The anamnese was performed and collected data of familiar history, parental consanguinity, and information about pregnancy period. The careful characterization of clinical presentations and the genetic pathways was studied. It is possible that there is no single mutation that can be clearly identified as the etiology of the combination of the defects displayed in the present case.

COMT rs4818, pain sensitivity and duration, and alveolar bone grafting of oral clefts.

PURPOSE: Verifying whether the mutation in COMT rs4818 could be involved in pain modulation. METHODS: Thirty-two individuals born with cleft lip and palate that underwent bone graft from the iliac crest bone were assessed at 12, 24, 48, 72 h, and 7 days regarding their pain experience using a visual analogic scale. DNA from each participant was collected from saliva samples, and genotyping of rs4818 was performed using TaqMan chemistry. Overrepresentation of rs4818 alleles was tested using chi-square or Fisher's exact tests with an alpha of 0.05. RESULTS: Of the 32 individuals, eighteen reported long pain duration, nine reported high pain intensity, and fourteen low pain intensity up to 48 h. No differences were found in the distribution of individuals depending on the reported pain by sex (p = 0.12), age (p = 0.42), or cleft type (p = 0.5). The distribution of COMT r4818 alleles was different depending on the intensity and duration of pain. Carriers of the C wild-type allele were four times more likely to show high pain intensity and duration (odds ratio = 4.29, 95% confidence interval 1.13-16.18), meaning that the G variant allele is protective. CONCLUSION: COMT rs4818 is associated with postoperative pain after alveolar bone grafting.

Beyond Amelogenesis Imperfecta Mutations: Hypomorphic Forms.

Mutations in several genes can lead to amelogenesis imperfecta. These same genes and other members of their pathways quite possibly may also contribute to individual susceptibility to dental caries and/or erosive tooth wear. This chapter provides an analysis of the function of the genes which, when mutated, cause amelogenesis imperfecta and discusses how mechanisms involving hypomorphic alleles in one or more genes, methylation changes, and imprinting disorders could be underlying individual susceptibility to dental caries and/or erosive tooth wear.

Individual Susceptibility to Dental Caries: The Vipeholm Study.

The evidence for a genetic component to dental caries is in comparison much less explored in the literature than the other classically described components of the disease: microbiota and diet. Diet can be said to have been conclusively linked to the etiology of dental caries by the results of the Vipeholm study, which did not deal with any microbial aspect of the disease. What is much less emphasized is that these same results provided one of the most robust lines of evidence that exist in support of a genetic component to dental caries. The results of the study showed great variability, with some study participants showing a dramatic increase in the establishment of new carious lesions under conditions that did not affect the majority of the other subjects. Similarly, highly cariogenic conditions due to a diet rich in sugars did not cause an increase in new carious lesions in other participants. The best explanation for these variable results is the existence of individual biological factors (genetic variation) modulating the disease. This chapter also comments on the ethical controversy surrounding the study and the need for finally intervening on the effects of sugars in oral health.

Genetics of Dental Caries: Controlled Animal Models.

Research in animal models, particularly rodents, has been used as a tool for gaining insight into the genetics contribution to dental caries. This chapter dissects some of these data, particularly the early studies motivated by observations in humans, complementing them with more recent ones designed specifically to map genes for dental caries. Finally, it offers a critical view of the rationale and lack of ethical principles of the use of nonhuman species in research.

Heritability of Dental Caries: Twin Studies.

The study of twins is a powerful tool to infer the presence and amount of contribution of genetic variation to a particular trait or disease. The ability to compare identical or monozygotic twins with dizygotic twins permits the direct comparison of pairs of individuals that share 100% of their genomic DNA with pairs that share only 50%, with the assumption that these pairs are under the same environment. In the case of dental caries, the environment is same parents, under the same roof, with the same diet, oral hygiene habits, culture, and lifestyle. These data have consistently suggested that a relevant proportion of the variation of dental caries in populations is due to genetics.

Individual Susceptibility to Erosive Tooth Wear: Wine Tasters.

Erosive tooth wear is the classic gene-environmental model. It requires the exposure to acids, typically from the diet or from the gastric content of the host, and shows variation among individuals and populations, which suggests it to be determined by more than one gene, each with small individual effects. This genetic component is not easily studied since a precise assessment of acidic exposure is complicated. In humans, this is commonly done by self-reported data. In this chapter, evidence is discussed that supports the assumption that a genetic component exists, inspecting data from wine tasters, and ex vivo experiments combining different dental enamel specimens and saliva compositions. These data can be found "hidden" in reports that do not directly deal with individual susceptibility to erosive tooth wear.