Clinicopathological features of myofibromas and myofibromatosis affecting the oral and maxillofacial region: A systematic review.

BACKGROUND: Myofibromas are rare benign neoplasms composed of myoid cells and myofibroblasts. This study aimed to systematically review case reports and a series of myofibromas (MF) and myofibromatosis (MFT) occurring in the oral and maxillofacial regions in order to describe their main clinicopathological features. METHODS: This systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Electronic searches were conducted in 2023 in four databases: MEDLINE/PubMed, Web of Science, Scopus, and EMBASE. A manual search and a search in the grey literature were also conducted. The lesions were classified as MF or MFT according to their original report. RESULTS: A total of 169 cases were included in this systematic review. Men were slightly more affected, with a painless nodule. When occurring in soft tissue, MF usually developed in the gingiva (mean age:29.23 ± 21.93 years) and when it was intra-osseous, it occurred more frequently in the posterior mandible (mean age:14.33 ± 15.62 years). MFT occurred mainly in the mandible and was predominantly described as well-circumscribed masses of spindle cells organized in fascicles with a prominent vascular activity in a hemangiopericytoma-like pattern. The lesions were mainly positive for smooth muscle actin and vimentin immunomarkers. Surgical excision was the treatment of choice in the majority of cases and recurrence was observed in only three cases. CONCLUSION: MF and MFT affect more men, with an indolent clinical course. Intra-osseous tumors and MFT seem to occur more frequently in younger individuals. These lesions seem to have a good prognosis and low recurrence.

Targeted therapies in ameloblastomas and amelobastic carcinoma-A systematic review.

Targeted therapy has the potential to be used in the neoadjuvant setting for odontogenic tumors, reducing the morbidities associated with major surgery. In this regard, the aim of this study was to summarize the current evidence on the different forms of targeted therapy, effectiveness, and drawbacks of this course of treatment. Four databases were searched electronically without regard to publication date or language. Grey literature searches and manual searches were also undertaken. Publications with sufficient clinical data on targeted therapy for odontogenic tumors were required to meet the criteria for eligibility. The analysis of the data was descriptive. A total of 15 papers comprising 17 cases (15 ameloblastomas and 2 ameloblastic carcinomas) were included. Numerous mutations were found, with BRAF V600E being most common. Dabrafenib was the most utilized drug in targeted therapy. Except for one case, the treatment reduced the size of the lesion (16/17 cases), showing promise. Most of the adverse events recorded were mild, such as skin issues, voice changes, abnormal hair texture, dry eyes, and systemic symptoms (e.g., fatigue, joint pain, and nausea). It is possible to reach the conclusion that targeted therapy for ameloblastoma and ameloblastic carcinoma may be a useful treatment strategy, based on the findings of the included studies.

Classificatory updates in verrucous and cuniculatum carcinomas: Insights from the 5th edition of WHO-IARC head and neck tumor classification.

The International Agency for Research on Cancer (IARC) and World Health Organization (WHO) collaboratively produce the 'WHO Blue Books' essential tools standardizing the diagnostic process for human cancers. Regular updates in this classification accommodate emerging molecular discoveries, advances in immunohistochemical techniques, and evolving clinical insights. The 5th edition of the WHO/IARC classification of head and neck tumors refines the 'Oral Cavity and Mobile Tongue' chapter, including sections for non-neoplastic lesions, epithelial tumors, and tumors of uncertain histogenesis. Notably, the epithelial tumors section is rearranged by tumor behavior, starting with benign squamous papillomas and progressing through potentially malignant oral disorders to oral squamous cell carcinoma (OSCC). The section on OSCC reflects recent information on epidemiology, pathogenesis, and histological prognostic factors. Noteworthy is the specific categorization of verrucous carcinoma (VC) and carcinoma cuniculatum (CC), both associated with the oral cavity and distinct in clinical and histologic characteristics. This classification adjustment emphasizes the oral cavity as their predominant site in the head and neck. Designating specific sections for VC and CC aims to provide comprehensive insights into these unique subtypes, elucidating their clinical features, distinct histological characteristics, prevalence, significance, and clinical relevance. By categorizing these subtypes into specific sections, the 5th edition of the WHO classification aims to provide a more nuanced and detailed account, enhancing our understanding of these specific variants within the broader spectrum of head and neck tumors.

Barriers to early diagnosis and management of oral cancer in Latin America and the Caribbean.

OBJECTIVE: This study aimed to explore perceived barriers to early diagnosis and management of oral cancer, as well as potential pathways for improvement in Latin America and the Caribbean (LAC). METHODS: This cross-sectional study used a self-administered online questionnaire created via the Research Electronic Data Capture platform. The survey was distributed to health professionals trained in Oral Medicine, Oral Pathology, Oral and Maxillofacial Surgery, and Dentists with clinical and academic expertise in oral potentially malignant disorder (OPMD) and oral cancer. Data obtained were systematically organized and analyzed descriptively using Microsoft Excel. RESULTS: Twenty-three professionals from 21 LAC countries participated. Major barriers included the limited implementation of OPMD and oral cancer control plans (17.4%), low compulsory reporting for OPMD (8.7%) and oral cancer (34.8%), unclear referral pathways for OPMD (34.8%) and oral cancer (43.5%), and a shortage of trained professionals (8.7%). Participants endorsed the utility of online education (100%) and telemedicine (91.3%). CONCLUSION: The survey highlights major perceived barriers to early diagnosis and management of OPMD and oral cancer in LAC, as well as potential avenues for improvement.

Comprehensive insights into the understanding of hypoxia in ameloblastoma.

Hypoxia is characterized by a disparity between supply and demand of oxygen. The association between hypoxia and head and neck tumors is a topic of significant interest. Tumors frequently encounter areas with inadequate oxygen supply, resulting in a hypoxic microenvironment. Ameloblastoma is one of the most common benign odontogenic tumors of the maxillofacial region. It is a slow-growing but locally invasive tumor with a high recurrence rate. The literature has demonstrated the correlation between hypoxia and ameloblastoma, revealing a discernible link between the heightened expression of hypoxic markers in low oxygen conditions. This association is intricately tied to the tumoral potential for invasion, progression, and malignant transformation. Hypoxia profoundly influences the molecular and cellular landscape within ameloblastic lesions. The present review sheds light on the mechanisms, implications, and emerging perspectives in understanding this intriguing association to clarify the dynamic relationship between hypoxia and ameloblastoma.

Contributions of viral oncogenes of HPV-18 and hypoxia to oxidative stress and genetic damage in human keratinocytes.

Infection with high-risk human papillomaviruses like HPV-16 and HPV-18 is highly associated with the development of cervical and other cancers. Malignant transformation requires viral oncoproteins E5, E6 and E7, which promote cell proliferation and increase DNA damage. Oxidative stress and hypoxia are also key factors in cervical malignant transformation. Increased levels of reactive species of oxygen (ROS) and nitrogen (RNS) are found in the hypoxic tumor microenvironment, promoting genetic instability and invasiveness. In this work, we studied the combined effect of E5, E6 and E7 and hypoxia in increasing oxidative stress and promoting DNA damage and nuclear architecture alterations. HaCaT cells containing HPV-18 viral oncogenes (HaCaT E5/E6/E7-18) showed higher ROS levels in normoxia and higher levels of RNS in hypoxia compared to HaCaT parental cells, as well as higher genetic damage in hypoxia as measured by γH2AX and comet assays. In hypoxia, HaCaT E5/E6/E7-18 increased its nuclear dry mass and both cell types displayed marked heterogeneity in nuclear dry mass distribution and increased nuclear foci. Our results show contributions of both viral oncogenes and hypoxia to oxidative stress, DNA damage and altered nuclear architecture, exemplifying how an altered microenvironment combines with oncogenic transformation to promote tumor progression.

Oral squamous cell carcinomas and oral potentially malignant disorders: A Latin American study.

OBJECTIVE: To determine the frequency of oral squamous cell carcinoma (OSCC) associated or not with oral potentially malignant disorders (OPMD), and the epidemiological profile and traditional risk factors in Latin America. METHODS: A retrospective observational study was conducted in 17 Latin American centres. There were included cases of OSCC, analysing age, gender, OSCC and their association with previous OPMD. Clinicopathological variables were retrieved. The condition of sequential-OSCC versus OSCC-de novo (OSCC-dn) was analysed concerning the aforementioned variables. Quantitative variables were analysed using Student's t-test, and qualitative variables with chi-square. RESULTS: In total, 2705 OSCC were included with a mean age of 62.8 years old. 55.8% were men. 53.75% of the patients were smokers and 38% were common drinkers. The lateral tongue border was the most affected site (24.65%). There were regional variations in OPMD, being leukoplakia the most frequent. Of the overall 2705 OSCC cases, 81.4% corresponded to OSCC-dn, while s-OSCC were 18.6%. Regarding lip vermillion SCC, 35.7% corresponded to de novo lip SCC and 64.3% were associated with previous OPMD. CONCLUSIONS: In Latin America, OSCC-dn seems to be more frequent with regional variations of some clinical and histopathological features. Further prospective studies are needed to analyse this phenomenon.

Correlation of Bony Invasion With Nodal Metastasis, Pattern of Invasion and Survival in Oral Squamous Cell Carcinoma: A Retrospective Analysis of 122 Primary Cases From Oral Cancer Centre of South India.

Background Oral squamous cell carcinoma (OSCC) is considered to be the most common epithelial malignant neoplasm of the oral cavity. Despite advancements in diagnosis and therapeutics the clinical outcome of the disease has not improved much which may be attributed to tumor biology and heterogeneity. Bone invasion by cancer cells is currently staged as a moderately advanced disease. However, many low-grade carcinomas such as verrucous carcinoma and carcinoma cuniculatum show body invasion but less nodal metastases and better overall survival. The present study was orchestrated to analyze if bone invasion in OSCC has any impact on regional nodal metastases and survival. Materials and methods A total of 122 cases of OSCC who underwent excision and neck dissection were retrieved and included. These cases were then divided into two study groups. Group I comprised 56 OSCC cases with bone involvement and 66 cases with no bony involvement. The bone invasion was correlated with nodal metastases, survival and pattern of invasion. Statistical analysis was done using SPSS software (IBM Corp., Armonk, NY, USA). Results There was no statistically significant correlation between bone invasion with either nodal metastases or pattern of invasion, however, the worst pattern of invasion (WPOI)-4,5 showed a statistically higher incidence of nodal involvement in OSCC. No statistical difference was noted in overall survival between the two groups. Conclusion The worst pattern of invasion and not bone involvement, depicts nodal metastases in OSCC and thus, deserves consideration while staging and treatment planning.

Artificial Intelligence for Image Analysis in Oral Squamous Cell Carcinoma: A Review.

Head and neck tumor differential diagnosis and prognosis have always been a challenge for oral pathologists due to their similarities and complexity. Artificial intelligence novel applications can function as an auxiliary tool for the objective interpretation of histomorphological digital slides. In this review, we present digital histopathological image analysis applications in oral squamous cell carcinoma. A literature search was performed in PubMed MEDLINE with the following keywords: "artificial intelligence" OR "deep learning" OR "machine learning" AND "oral squamous cell carcinoma". Artificial intelligence has proven to be a helpful tool in histopathological image analysis of tumors and other lesions, even though it is necessary to continue researching in this area, mainly for clinical validation.

Mechanical profile of human keratinocytes expressing HPV-18 oncogenes.

During tumorigenesis, the mechanical properties of cancer cells change markedly, with decreased stiffness often accompanying a more invasive phenotype. Less is known about the changes in mechanical parameters at intermediate stages in the process of malignant transformation. We have recently developed a pre-tumoral cell model by stably transducing the immortalized but non-tumorigenic human keratinocyte cell line HaCaT with the E5, E6 and E7 oncogenes from HPV-18, one of the leading causes of cervical cancer and other types of cancer worldwide. We have used atomic force microscopy (AFM) to measure cell stiffness and to obtain mechanical maps of parental HaCaT and HaCaT E5/E6/E7-18 cell lines. We observed a significant decrease in Young's modulus in HaCaT E5/E6/E7-18 cells measured by nanoindentation in the central region, as well as decreased cell rigidity in regions of cell-cell contact measured by Peakforce Quantitative Nanomechanical Mapping (PF-QNM). As a morphological correlate, HaCaT E5/E6/E7-18 cells displayed a significantly rounder cell shape than parental HaCaT cells. Our results therefore show that decreased stiffness with concomitant perturbations in cell shape are early mechanical and morphological changes during the process of malignant transformation.

Connexin 43 expression in tooth germ and benign odontogenic tumors.

OBJECTIVE: The aim of this study was to investigate and compare the immunohistochemical expression of connexin 43 (Cx43) in tooth germs (TGs), ameloblastic fibromas (AFs), ameloblastic fibro-odontomas (AFOs), and conventional ameloblastomas (AMs). STUDY DESIGN: Nine TGs, 12 AFs, 12 AFOs, and 27 AMs were evaluated for Cx43 expression by immunohistochemistry. RESULTS: Most of the TGs expressed Cx43 in the mesenchyme (77.6%) and in the late stages of odontogenesis. Cx43 was more highly expressed (P < .05) in the mesenchymal layer of all groups than in the epithelial layer except for the AFOs. When comparing the expression of Cx43 in the different layers of the analyzed groups, statistically significant differences were observed between AFO vs AM (*P = .0158) in the epithelial layer and between AF vs AFO (P** = .0046) in the mesenchymal layer. CONCLUSIONS: The results obtained in this study showed that Cx43 is a protein with important expression in the mesenchymal layer of the embryonic and odontogenic tissues studied. It could be speculated that Cx43 participates in mineralization events based on the relationship of the expression of this protein between the epithelial and mesenchymal layers of odontogenic tissues.

A Systematic Review and Meta-Analysis of the Relationship Between the Severity of Dental Fluorosis and Fluoride Biomarkers in Endemic Areas.

The intake of high concentrations of fluoride, mainly through drinking water, diet and fluoridated dentifrices, produces fluorosis, which in its early stages is manifested as dental fluorosis (DF). To recognize exposure to fluoride in endemic areas and to evaluate the risk of developing health impairment, the WHO has established several biomarkers that are used to determine systemic fluorine (F-) exposure. Thus, the aim of this study was to conduct a systematic review and meta-analysis of the relationship between the severity of DF and fluoride biomarkers in endemic areas. The protocol of this study was previously registered as CRD42021244974. A digital search was carried out in PubMed/Medline, SpringerLink, Scopus, Cochrane and Google Scholar by employing the keywords "urine", "nails", "hair", "plasma", "saliva" and "dental fluorosis" for the original studies with content associated with F- for the biomarkers and DF. The mean difference was established as the effect measure for the meta-analysis. Seven studies fulfilled the eligibility criteria, among which five assessed urine and two employed nails as fluoride biomarkers. A positive significant difference was found between the biomarkers and the severity of DF (0.27, p < 0.001) and individually for each biomarker (urine: 0.14, p = 0.001; nails: 0.88, p < 0.05). The F- concentration in urine and nails is correlated with the severity of DF, with the most evident differences between healthy individuals and those with mild severity. Both biomarkers are adequate to assess this relationship in endemic areas of fluoride and DF.

Antagonistic, synergistic, and additive antibacterial interaction between ciprofloxacin and amoxicillin against Staphylococcus aureus.

The aim of this in vitro study was to evaluate the interaction between ciprofloxacin and amoxicillin against beta-lactamase-producing Staphylococcus aureus. Concentration-dependent curves for each individual drug were carried out to obtain the mean inhibitory concentration in the agar well diffusion assay. Then, different ratios of the ciprofloxacin-amoxicillin combination (0.5:0.5, 0.8:0.2, 0.2:0.8, 0.9:0.1, 0.1:0.9, 0.95:0.05, and 0.05:0.95) were assessed. Data were analyzed using the isobolographic analysis and interaction index. The isobolographic evaluation shows that the 0.9:0.1 and 0.95:0.05 ratios of the ciprofloxacin-amoxicillin combination produced a synergistic antimicrobial interaction, the 0.8:0.2, 0.2:0.8, 0.1:0.9, and 0.05:0.95 proportions showed an additive antibacterial effect, and the 0.5:0.5 proportion induced antagonistic antimicrobial effects. The interaction index showed similar outcomes to the isobolographic analysis. In conclusion, the data of this study mainly show antimicrobial additive results of the ciprofloxacin-amoxicillin combination against beta-lactamase-producing S. aureus.

Spatial distribution of CD34 protein in primordial odontogenic tumour, ameloblastic fibroma and the tooth germ.

BACKGROUND: Primordial odontogenic tumour is a benign mixed neoplasm of recent description, which has histological similarities with other odontogenic tumours such as the ameloblastic fibroma. In this article, we investigate the architecture of the sub-epithelial layer of mesenchymal cells expressing the marker CD34 in primordial odontogenic tumour. OBJECTIVE: Analyse the spatial patterns of CD34 expression in primordial odontogenic tumour and compare them with those in ameloblastic fibroma and the normal tooth germ by means of objective imaging approaches, to better characterise these lesions. METHODS: Two cases of primordial odontogenic tumour, four cases of ameloblastic fibroma and two cases of tooth germ in cap and bell stages were used for morphological, structural and immunohistochemical analyses. RESULTS: CD34 expression was found in vascular endothelium of primordial odontogenic tumour, ameloblastic fibroma and tooth germ. In addition, a characteristic sub-epithelial expression was observed only in primordial odontogenic tumour, corresponding to 84%-86% of the sample boundaries. Moreover, the zone expressing CD34 corresponded with a higher cellularity, which was absent in ameloblastic fibroma and tooth germ. CONCLUSION: Image analysis of the primordial odontogenic tumour architecture revealed characteristics absent in other odontogenic tumours and tooth germs. This study provides additional information to support the idea that this neoplasm is a distinct entity from early stage AF or developing odontoma.

Periodontal Disease in Obese Patients; Interleukin-6 and C-Reactive Protein Study: A Systematic Review.

Periodontal disease (PD) and obesity are characterized by a dysregulated inflammatory state. Both conditions trigger inflammatory and immune responses with an increase in proinflammatory cytokines such as Interleukin 6 (IL-6) and the release of inflammatory mediators such as C-reactive protein (CRP). Individuals with a high body mass index (BMI) present a chronic inflammatory state. The aim of the present study was to perform a systematic review of inflammatory markers (IL-6 and CRP) in obese patients with PD and their possible relationship by analyzing the levels of these markers. A digital literature search was performed in three databases-PubMed, SciElo and Medigraphic-through an advanced search for original articles, employing IL-6 and CRP in obese patients with PD, within a publication period from 2010 to 2021. PRISMA guidelines, the JADAD scale and a qualitative analysis of scientific evidence were performed using the Cochrane collaboration method and the RoB 2 assessment tool. Ten articles were included in this analysis with the variables recorded and associated with subjects with obesity and PD. Of the ten articles included, three analyzed IL-6 and CRP, four analyzed IL-6 and three analyzed CRP. In conclusion, and based on the available evidence, the aforementioned markers of inflammation demonstrate that there is a relationship between PD and obesity.

Hypoxia, acidification and oxidative stress in cells cultured at large distances from an oxygen source.

Hypoxia is a condition frequently encountered by cells in tissues, whether as a normal feature of their microenvironment or subsequent to deregulated growth. Hypoxia can lead to acidification and increased oxidative stress, with profound consequences for cell physiology and tumorigenesis. Therefore, the interplay between hypoxia and oxidative stress is an important aspect for understanding the effects of hypoxic microenvironments on cells. We have used a previously developed variant of the method of coverslip-induced hypoxia to study the process of acidification in a hypoxic microenvironment and to simultaneously visualize intracellular levels of hypoxia and oxidative stress. We observed high accumulation of CO2 in hypoxic conditions, which we show is the main contributor to acidification in our model. Also, increased levels of oxidative stress were observed in moderately hypoxic cells close to the oxygen source, where the mitochondrial membrane potential was preserved. Conversely, cells at large distances from the oxygen source showed higher levels of hypoxia, milder oxidative stress and reduced mitochondrial membrane potential. Our results contribute to characterize the interplay between reduced oxygen levels, acidification and oxidative stress in a simple in vitro setting, which can be used to model cell responses to an altered environment, such as the early tumor microenvironment.

Estudio de los efectos de un microambiente hipóxico en queratinocitos humanos in vitro y su correlato con alteraciones del microambiente en la patología de liquen plano oral

La hipoxia es un factor fundamental en el proceso de génesis tumoral, así como en patologías precursoras de cáncer, como es el Liquen Plano Oral (LPO). Objetivo: Determinar si es posible establecer una correlación entre las alteraciones que sufren queratinocitos normales en un microambiente hipóxico in vitro y alteraciones que aparecen en los queratinocitos en el epitelio de la mucosa oral en el contexto de la patología LPO. Métodos: Se estudiaron los cambios morfológicos mediante microscopía de contraste de fases, y la detección de marcadores asociados a hipoxia de queratinocitos humanos (HaCaT), como modelo celular oral, en un microambiente hipóxico generado por la variante del método "Hipoxia inducida por cubreobjetos". Resultados: Mediante microscopía confocal se observó la presencia de los marcadores de hipoxia GLUT-1 y aductos de pimonidazol (Hipoxyprobe) en los cultivos celulares de HaCaT expuestos a un microambiente hipóxico. Además, se observó la presencia del marcador GLUT-1 mediante inmunohistoquímica en tejido epitelial humano derivado de biopsias de la patología LPO. Conclusiones: Se estableció una correlación entre las alteraciones detectadas en queratinocitos humanos inducidos a un microambiente hipóxico in vitro y las alteraciones detectadas in vivo en tejido epitelial de la mucosa oral.

A hipóxia é um fator fundamental no processo de gênese tumoral, bem como em patologias precursoras do câncer, como o Líquen Plano Oral (LPO). Objetivo: Determinar se é possível estabelecer uma correlação entre as alterações que os queratinócitos normais sofrem em um microambiente hipóxico in vitro e as alterações que aparecem nos queratinócitos no epitélio da mucosa oral no contexto da patologia do LPO. Métodos: As alterações morfológicas foram estudadas por microscopia de contraste de fase e a detecção de marcadores associados à hipóxia de queratinócitos humanos (HaCaT), como modelo de célula oral, em um microambiente hipóxico gerado pela variante do método "Hipóxia induzida por lamínulas". Resultados: Por microscopia confocal, observou-se a presença dos marcadores de hipóxia GLUT-1 e Hipoxyprobe em culturas de células HaCaT expostas a um microambiente hipóxico. Além disso, a presença do marcador GLUT-1 foi observada por imuno-histoquímica em tecido epitelial humano derivado de biópsias de patologia de LPO. Conclusões: Foi estabelecida uma correlação entre as alterações detectadas em queratinócitos humanos induzidas em um microambiente hipóxico in vitro e as alterações detectadas in vivo no tecido epitelial da mucosa oral.

Hypoxia is a fundamental factor in the process of tumor genesis, as well as in precursor pathologies of cancer, such as Oral Lichen Planus (OLP). Objective: To determine if it is possible to establish a correlation between the alterations that normal keratinocytes suffer in a hypoxic microenvironment in vitro and alterations that appear in the keratinocytes in the epithelium of the oral mucosa in the context of OLP pathology. Methods: Morphological changes were studied by phase contrast microscopy, and the detection of markers associated with hypoxia of human keratinocytes (HaCaT), as an oral cell model, in a hypoxic microenvironment generated by the variant of the method "Hypoxia induced by coverslips". Results: Using confocal microscopy, the presence of hypoxia markers GLUT-1 and Hipoxyprobe was observed in HaCaT cell cultures exposed to a hypoxic microenvironment. In addition, the presence of the GLUT-1 marker was observed by immunohistochemistry in human epithelial tissue derived from biopsies of OLP pathology. Conclusions: A correlation was established between the alterations detected in human keratinocytes induced in a hypoxic microenvironment in vitro and the alterations detected in vivo in epithelial tissue of the oral mucosa.

Single Nucleotide Polymorphisms and Dental Fluorosis: A Systematic Review.

Genetic factors contribute to susceptibility and resistance to fluoride exposure. The aim of this systematic review was to identify alleles/genotypes of single nucleotide polymorphisms (SNPs) associated with dental fluorosis (DF) and to identify them as protective or risk factors. PubMed, ScienceDirect, Cochrane Library, Scopus and Web of Science were searched for articles; the last search was performed in August 2022. Human studies that analyzed the relationship between SNPs and DF published in English were included; systematic reviews and meta-analyses were excluded. Methodological quality was graded using the Joanna Briggs Institute checklist and risk of bias was assessed using the Cochrane Collaboration's tool. Eighteen articles were included, 44% of which showed high methodological quality and data from 5,625 participants aged 6 to 75 years were analyzed. The SNPs COL1A2, ESR2, DLX1, DLX2, AMBN, TUFT1, TFIP11, miRNA17, and SOD2 were considered risk factors, and ESR1, MMP20, and ENAM were considered protective factors. In conclusion, there are alleles and genotypes of different single nucleotide polymorphisms involved in increasing or decreasing the risk of developing dental fluorosis.