Prognostic value of the expression and localization of cell proliferation and apoptosis markers in unicystic ameloblastomas.

The aim of this study was to verify whether the expression of cell proliferation and apoptosis markers in different types of unicystic ameloblastoma (UA) is associated with the location of neoplastic cells. Immunohistochemical study with a sample of 32 cases of UA, 11 cases of conventional ameloblastoma (CAM) and ten dental follicles (DF) cases was performed. Cell proliferation was assessed using Ki-67 status, and apoptosis by caspase-3 expression. Mural UA (MUA) showed a higher immunostaining of Ki-67 (p < 0.05) and a lower immunostaining of Caspase-3 (p < 0.05) compared with luminal and intraluminal subtypes of UA and CAM. The neoplastic cells of the MUA's cystic capsule showed a higher expression of Ki-67 protein (p < 0.0001) and a lower expression of Caspase-3 (p < 0.0001) compared with the lumen. DF showed lower Ki-67 and Caspase-3 immunostaining (p < 0.05) than neoplasms. The higher immunoexpression of Ki-67 and the lower immunoexpression of Caspase-3 in MUA, in the parenchyma cells within the cystic capsule, suggest an association between the biological behaviour and location of neoplastic cells in a tumour.

Platelet-rich fibrin stimulates the proliferation and expression of proteins related to survival, adhesion, and angiogenesis in gingival fibroblasts cultured on a titanium nano-hydroxyapatite-treated surface.

OBJECTIVES: This in vitro study aimed to evaluate the cell viability and expression of proteins related to angiogenesis, adhesion, and cell survival (vascular endothelial growth factor, paxillin, vinculin, fibronectin, and protein kinase B) in gingival fibroblasts that were cultured on titanium discs treated with or without nanohydroxyapatite and exposed to platelet-rich fibrin (PRF)-conditioned medium. METHODS: To obtain the conditioned medium, the PRF membranes were prepared and incubated for 48 h in a culture medium without fetal bovine serum. Analyses were performed at 24 and 48 h for the cells cultured on machined-titanium discs or surfaces treated with nanohydroxyapatite in a control medium or PRF-conditioned medium, resulting in four experimental groups (CT-TI, CT-NANO, PRF-TI, and PRF-NANO). RESULTS: A decrease in the viability of the gingival fibroblasts was not observed in any of the experimental groups. The PRF-NANO group showed significantly higher immunoexpression of paxillin and AKT at 24 and 48 h (p < 0.01). The same result was observed for vinculin expression at 24 h (p < 0.001). The expression of fibronectin at 48 h and VEGF at 24 and 48 h was significantly higher when the cells were exposed to the PRF-conditioned medium, regardless of the disc surface (p < 0.05). CONCLUSION: Gingival fibroblasts cultured on a nanohydroxyapatite-treated surface and in a PRF-conditioned medium showed a greater expression of proteins modulating adhesion, angiogenesis, and cell survival. Our results may contribute to the understanding of the mechanisms related to peri-implant soft tissue sealing.

Gene expression in cell lines from human ameloblastoma immortalized using hTERT and HPV16-E6/E7.

OBJECTIVE: The aim of this study was to evaluate and compare alterations in gene expression using two distinct immortalization methods (hTERT and HPV16-E6/E7) in ameloblastoma cell lines. MATERIALS AND METHODS: A primary cell culture derived from human ameloblastoma (AME-1) was established and immortalized by two different methods using a transfection processes to hTERT and HPV-E6/E7. The RNA-seq was used to verify which immortalization method had less influence on gene expression. It was performed in four steps: extraction and collection of mRNA, PCR amplification, comparison with the human reference genome, and analysis of differential expression. The genes with differentiated expression were identified and mapped. RESULTS: RNA-seq revealed genetic alterations in ameloblastoma cell lines after the immortalization process, including increased expression of tumor genes like MYC, E2F1, BRAF, HRAS, and HTERT, and a decrease in tumor suppressor genes like P53, P21, and Rb. CONCLUSIONS: It is possible to affirm that cell immortalization is not an inert method regarding gene regulation mechanisms and the hTERT method (AME-TERT) presented fewer changes in gene expression levels.

Non-Lethal Concentration of MeHg Causes Marked Responses in the DNA Repair, Integrity, and Replication Pathways in the Exposed Human Salivary Gland Cell Line.

In Brazilian northern Amazon, communities are potentially exposed and vulnerable to methylmercury (MeHg) toxicity through the vast ingestion of fish. In vivo and in vitro studies demonstrated that the salivary glands as a susceptible organ to this potent environmental pollutant, reporting alterations on physiological, biochemical, and proteomic parameters. However, the alterations caused by MeHg on the gene expression of the exposed human salivary gland cells are still unknown. Therefore, the goal was to perform the transcriptome profile of the human salivary gland cell line after exposure to MeHg, using the microarray technique and posterior bioinformatics analysis. The cell exposure was performed using 2.5 µM MeHg. A previously published study demonstrated that this concentration belongs to a range of concentrations that caused biochemical and metabolic alterations in this linage. As a result, the MeHg exposure did not cause lethality in the human salivary gland cells line but was able to alter the expression of 155 genes. Downregulated genes (15) are entirety relating to the cell metabolism impairment, and according to KEGG analysis, they belong to the glycosphingolipid (GSL) biosynthesis pathway. On the other hand, most of the 140 upregulated genes were related to cell-cycle progression, DNA repair, and replication pathway, or cellular defenses through the GSH basal metabolism. These genomic changes revealed the effort to the cell to maintain physiological and genomic stability to avoid cell death, being in accordance with the nonlethality in the toxicity test. Last, the results support in-depth studies on nonlethal MeHg concentrations for biomarkers identification that interpret transcriptomics data in toxicological tests serving as an early alert of physiological changes in vitro biological models.

Gene Expression Profile in Immortalized Human Periodontal Ligament Fibroblasts Through hTERT Ectopic Expression: Transcriptome and Bioinformatic Analysis.

Human periodontal ligament fibroblast (hPLF) cells play an important role in maintaining oral cavity homeostasis with special function in tissue regeneration and maintenance of dental alveoli. Although their primary cell cultures are considered a good experimental model with no genetic changes, the finite life span may limit some experimental designs. The immortalization process increases cell life span but may cause genetic changes and chromosomal instability, resulting in direct effects on physiological cell responses. In this way, we aimed to investigate the global gene expression of hPLFs after the immortalization process by the ectopic expression of the catalytic subunit of the enzyme telomerase reverse transcriptase (hTERT) through transcriptome analysis. The embryonic origin of the primary culture of hPLF cells and immortalized hPLF-hTERT was also tested by vimentin staining, hTERT synthesis evaluated by indirect immunocytochemistry, analysis of cell proliferation, and morphology. The results indicated that hPLFs and hPLF-hTERT were positive for vimentin. On the 20th cell passage, hPLFs were in senescence, while hPLF-hTERT maintained their proliferation and morphology characteristics. At the same passage, hPLF-hTERT presented a significant increase in hTERT synthesis, but transcriptome did not reveal overexpression of the hTERT gene. Fifty-eight genes had their expression altered (11 upregulated and 47 downregulated) with the absence of changes in the key genes related to these cell types and in the main cancer-associated genes. In addition, the increase in hTERT protein expression without the overexpression of its gene indicates posttranscriptional level regulation. Successful immortalization of hPLFs through the ectopic expression of hTERT encourages further studies to design experimental protocols to investigate clinical questions from a translational perspective.

Metabolic and oxidative impairments in human salivary gland cells line exposed to MeHg.

BACKGROUND/AIM: The ingestion of contaminated seafood by MeHg is considered the main route of human exposure, turning the salivary gland one important target organ. The salivary glands play critical roles in maintaining oral health homeostasis, producing saliva that maintains the oral microbiota, initiation of the digestion of macromolecules, and being essential in maintaining the integrity of the adjacent soft tissues and teeth. Thus, this study aimed to investigate the effects of MeHg exposure on human salivary gland cells line. METHODS: Cells were exposed to 1-6 µM of MeHg for 24 h, and analysis of toxicity was performed. Based on these results, the LC50 was calculated and two concentrations were chosen (0.25 and 2.5 µM MeHg) to evaluate intracellular mercury (Hg) accumulation (THg), metabolic viability and oxidative stress parameters (GSH:GSSG ratio, lipid peroxidation, protein oxidation and DNA damage). RESULTS: The results demonstrated accumulation of THg as we increased the MeHg concentrations in the exposure and, the higher the dose, the lower is the cell metabolic response. In addition, the 2.5 µM MeHg concentration also triggered oxidative stress in human salivary gland cells by depleting the antioxidant competence of GSH:GSSG ratio and increasing lipid peroxidation and proteins carbonyl levels, but no damages to DNA integrity. CONCLUSION: In conclusion, although these two elected doses did not show lethal effects, the highest dose triggered oxidative stress and new questionings about long-term exposure models are raised to investigate furthers cellular damages to human salivary gland cells caused by MeHg exposure to extrapolate in a translational perspective.

HIF-1α Is Associated with Resistance to Hypoxia-Induced Apoptosis in Ameloblastoma.

BACKGROUND: Ameloblastoma (AMB) is a benign odontogenic tumour, with an aggressive local behaviour and a high rate of recurrence. Previous studies have demonstrated that hypoxia-induced factor alpha 1 (HIF-1α) and activated caspase-3 contribute to tumour invasiveness and cytogenesis in ameloblastoma. Hypoxia increases HIF-1α levels, which triggers a number of signalling pathways. This paper aimed to present data in the study of hypoxia-activated signalling pathways that modulate proapoptotic and antiapoptotic events in AMB. METHODS: Twenty cases of AMB and ten cases of dental follicle (DF) were used to analyse the immunoexpression of HIF-1α, p53, BNIP3, Bcl-2, IAP-2, GLUT1, and Bax. To contribute to the study, an analysis of expression and genetic interaction was performed using the cell line AME-1. RESULTS: AMB and DF expressed the studied proteins. These proteins showed significantly greater immunoexpression in AMB compared with the DF (p < 0.05). HIF-1α showed an important association with GLUT1, and a positive correlation was observed among p53, Bcl-2, and IAP-2. Transcriptomic analysis showed the significant expression of the studied proteins, and the network generated showed a direct association of HIF-1αF with GLUT1 (SLC2A1), TP53, and LDHA. Interestingly, GLUT1 also exhibited direct interaction with TP53 and LDHA. CONCLUSION: In AMB tumorigenesis, hypoxia is possibly related to antiapoptotic events, which suggests an important role for HIF-1α, GLUT1, Bcl-2, IAP-2, and possibly p53.

Metallothionein Expression and its Influence on the In Vitro Biological Behavior of Mucoepidermoid Carcinoma.

Mucoepidermoid carcinoma (MEC) is the most common tumor in the salivary glands, often presenting with recurrence and metastasis due to its high invasive capacity. Metallothionein (MT), a zinc storage protein that supplies this element for protease activity, is probably related to mucoepidermoid carcinoma behavior. This prompted us to characterize a cell line derived from mucoepidermoid carcinoma and to correlate metallothionein expression with transforming growth factor-α (TGF-α), tumor necrosis factor-α (TNF-α) and matrix metalloproteinases (MMPs). Transcriptomic analysis and cytogenetic assays were performed to detect the expression of genes of interest and cellular chromosomal alterations, respectively. MEC cells with a depleted metallothionein 2A (MT2A) gene were subjected to Western blot to correlate metallothionein expression with growth factors and MMPs. Additionally, cells with depleted MT were subjected to migration and invasion assays. The transcriptomic study revealed reads mapped to cytokeratins 19 and AE1/AE3, α-smooth muscle actin, vimentin, and fibronectin. Cytogenetic evaluation demonstrated structural and numerical alterations, including the translocation t(11;19)(q21;p13), characteristic of MEC. Metallothionein depletion was correlated with the decreased expression of TGF-α and MMP-9, while TNF-α protein levels were augmented. Migration and invasion activity were diminished after metallothionein silencing. Our findings suggest an important role of MT in MEC invasion, through the regulation of proteins involved in this process.

Piceatannol Increases Antioxidant Defense and Reduces Cell Death in Human Periodontal Ligament Fibroblast under Oxidative Stress.

Piceatannol is a resveratrol metabolite that is considered a potent antioxidant and cytoprotector because of its high capacity to chelate/sequester reactive oxygen species. In pathogenesis of periodontal diseases, the imbalance of reactive oxygen species is closely related to the disorder in the cells and may cause changes in cellular metabolism and mitochondrial activity, which is implicated in oxidative stress status or even in cell death. In this way, this study aimed to evaluate piceatannol as cytoprotector in culture of human periodontal ligament fibroblasts through in vitro analyses of cell viability and oxidative stress parameters after oxidative stress induced as an injury simulator. Fibroblasts were seeded and divided into the following study groups: control, vehicle, control piceatannol, H2O2 exposure, and H2O2 exposure combined with the maintenance in piceatannol ranging from 0.1 to 20 µM. The parameters analyzed following exposure were cell viability by trypan blue exclusion test, general metabolism status by the 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) method, mitochondrial activity through the ATP production, total antioxidant capacity, and reduced gluthatione. Piceatannol was shown to be cytoprotective due the maintenance of cell viability between 1 and 10 µM even in the presence of H2O2. In a concentration of 0.1 µM piceatannol decreased significantly cell viability but increased cellular metabolism and antioxidant capacity of the fibroblasts. On the other hand, the fibroblasts treated with piceatannol at 1 µM presented low metabolism and antioxidant capacity. However, piceatannol did not protect cells from mitochondrial damage as measured by ATP production. In summary, piceatannol is a potent antioxidant in low concentrations with cytoprotective capacity, but it does not prevent all damage caused by hydrogen peroxide.

HIF-1α, NOTCH1, ADAM12, and HB-EGF are overexpressed in mucoepidermoid carcinoma.

OBJECTIVE: Intratumoral hypoxia (IH) occurs during cellular proliferation of malignant tumors. This phenomenon is characterized by a decrease in oxygen levels in the neoplastic microenvironment. Throughout this condition, the proteins HIF-1α, NOTCH1, ADAM12, and HB-EGF can be activated, triggering signaling pathways associated with tumor invasiveness through invadopodia formation. This study aimed to evaluate the immunostaining of HIF-1α, NOTCH1, ADAM12, and HBEGF in 19 cases of mucoepidermoid carcinoma (MEC) and 10 samples of salivary glands (control group). STUDY DESIGN: The immunoperoxidase technique was employed to detect the proteins of interest. The Student t test was used to compare immunoexpression between MEC samples and the control group. RESULTS: Protein immunostaining was statistically significantly higher in MEC samples than in the control group (P < .01), and the proteins were especially overexpressed in epidermoid cells of MEC. CONCLUSIONS: We suggest that there is an association between the NOTCH1 signaling pathway activated by IH and the biologic behavior of MEC.

Odontologia e saúde mental: experiência do PET Saúde no Centro de Atenção Psicossocial Álcool e Drogas / Odontology and mental health: experience of PET Saúde in the Center for Psychosocial Care Alcohol and Drugs

A fim de discutir a importância de experiências multiprofissionais e o papel de inovação do PET - Saúde na formação acadêmica, este artigo relata a experiência do PET - Odontologia da UFPA junto à equipe multiprofissional de um CAPS AD localizado no município de Belém/PA, além de avaliar as condições de saúde bucal dos dependentes químicos. A experiência mostrou a integração entre os diferentes profissionais e motivou o desenvolvimento de habilidades dos discentes para o planejamento de ações integradas e criativas de promoção de saúde, através de metodologias ativas, apontando novos rumos de atenção à saúde, centrado na lógica da realidade da comunidade e não somente no ambiente clínico. As atividades realizadas pela equipe de Odontologia foram integradas às oficinas e grupos desenvolvidos na unidade, como oficinas culinárias e de confecção e o cuidado de hortas, que propiciaram momentos de educação em saúde bucal. Foram realizadas dinâmicas e rodas de conversas, sendo os usuários agentes ativos no processo de construção compartilhada de conhecimento. Ocorreu o "dia D no CAPS AD", no qual os usuários participaram de atividade de escovação supervisionada e foram examinados por equipe médica e odontológica, com coleta de informações clínicas. Considera-se que a interação entre ensino, serviço e comunidade potencializada pelo PET - Saúde conduz a formação em saúde a um novo perfil, em que os profissionais se tornam mais capazes de responder às necessidades dos usuários e atuar de maneira compartilhada nos diferentes cenários (AU).

In order to discuss the importance of multiprofessional experiences and the innovation role of the PET - Saúde in academic training, this article reports on the experience PET - Odontology of UFPA with the multi-professional team of a CAPS AD located in the city of Belém / PA, besides evaluating the oral health conditions of chemical dependents. The experience showed the integration among the different professionals and motivated the development ofthe students' abilities to the planning of integrated and creative actions of health promotion, through active methodologies, pointing out new directions of health care, centered on the logic of the community reality and not only in the clinical environment. The activities carried out by the Dentistry team were integrated with the workshops and groups developed at the unit, such as cooking and cooking workshops and garden care, which provided moments of education in oral health. Was made dynamics and wheels of conversations, and the users are active agents in the process of shared knowledge construction. "D-day at CAPS AD" occurred, in which users participated in supervised brushing activity and were examined by medical and dental staff, collecting clinical information. It is considered that the interaction between teaching, service and community enhanced by PET ­ Saúde leads health education to a new profile in which professionals become better able to respond to users' needs and act in a shared way in different scenarios (AU).

A novel cell line derived from pleomorphic adenoma expresses MMP2, MMP9, TIMP1, TIMP2, and shows numeric chromosomal anomalies.

Pleomorphic adenoma is the most common salivary gland neoplasm, and it can be locally invasive, despite its slow growth. This study aimed to establish a novel cell line (AP-1) derived from a human pleomorphic adenoma sample to better understand local invasiveness of this tumor. AP-1 cell line was characterized by cell growth analysis, expression of epithelial and myoepithelial markers by immunofluorescence, electron microscopy, 3D cell culture assays, cytogenetic features and transcriptomic study. Expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) was also analyzed by immunofluorescence and zymography. Furthermore, epithelial and myoepithelial markers, MMPs and TIMPs were studied in the tumor that originated the cell line. AP-1 cells showed neoplastic epithelial and myoepithelial markers, such as cytokeratins, vimentin, S100 protein and smooth-muscle actin. These molecules were also found in vivo, in the tumor that originated the cell line. MMPs and TIMPs were observed in vivo and in AP-1 cells. Growth curve showed that AP-1 exhibited a doubling time of 3.342 days. AP-1 cells grown inside Matrigel recapitulated tumor architecture. Different numerical and structural chromosomal anomalies were visualized in cytogenetic analysis. Transcriptomic analysis addressed expression of 7 target genes (VIM, TIMP2, MMP2, MMP9, TIMP1, ACTA2 e PLAG1). Results were compared to transcriptomic profile of non-neoplastic salivary gland cells (HSG). Only MMP9 was not expressed in both libraries, and VIM was expressed solely in AP-1 library. The major difference regarding gene expression level between AP-1 and HSG samples occurred for MMP2. This gene was 184 times more expressed in AP-1 cells. Our findings suggest that AP-1 cell line could be a useful model for further studies on pleomorphic adenoma biology.

Recurrent Infection by varicella-zoster virus with facial scarring / Infecção recorrente pelo vírus varicela-zoster com sequelas cicatriciais na face

Primary contact with the varicella-zoster virus occurs through varicella (chickenpox) and culminates with this virus entering the sensory nerves and remaining latent in the dorsal root ganglion. Transmission occurs by dissemination of infectious particles of the varicella-zoster virus by the aerosol released from nasopharyngeal secretions or skin lesions, or by direct contact with lesions. Herpes zoster occurs after clinically evident reactivation of the virus, affecting the whole distribution of the infected sensory nerve. When compared with primary infection, herpes zoster has a more severe character, requiring the use of pharmaceutical drugs. The cause of reactivation is unknown and may be associated with predisposing factors, such as age, stress or impaired immune system. This study reports a case of a patient who presented clinical manifestations compatible with varicella zoster infection exacerbated by the use of homemade remedies, resulting in a secondary infection and facial scarring.

O contato primário com o vírus varicela-zoster ocorre na varicela (catapora), culminando com a transposição desse vírus para os nervos sensitivos, onde estabelece sua latência no gânglio espinhal dorsal. A transmissão ocorre por disseminação das partículas infecciosas do vírus varicela-zoster através de aerossóis liberados a partir de secreções do nasofaringe ou lesões cutâneas ou, ainda, pelo contato direto com lesões. O herpes-zoster clinicamente evidente ocorre após a reativação do vírus, com o envolvimento da distribuição do nervo sensitivo afetado. Quando comparado com a infecção primária, o herpes-zoster desenvolve um caráter de maior severidade, sendo sempre necessária a administração de uma terapêutica medicamentosa eficaz. A causa dessa reativação é desconhecida, podendo estar relacionada a fatores predisponentes como a faixa etária, estresse ou imunodeficiências. Neste trabalho relata-se um caso clínico em que a paciente apresentou manifestações clínicas condizentes com um quadro característico de infecção por varicela-zoster, complicado por uso de medicação caseira, resultando em infecção secundária e cicatrizes faciais.

Odontodisplasia regional: relato de caso em maxila com transpasse de linha média / Regional odontodysplasia: case report in maxilla crossing the midline

Odontodisplasia regional é uma afecção rara, não-hereditária, a qual afeta o desenvolvimento do esmalte e dentina na dentição decídua e/ou permanente. A etiologia ainda é desconhecida. Em geral, esta condição é diagnosticada durante exames radiográficos de rotina e muitos casos são diagnosticados erroneamente como dentes malformados ou odontomas. Esta condição afeta duas vezes mais o sexo feminino do que o masculino e envolve com mais frequência à maxila do que a mandíbula. Esta afecção é também mencionada como "dentes fantasmas", uma vez que os dentes afetados, radiograficamente, não apresentam distinção dos limites entre o esmalte e a dentina; e a câmara pulpar se apresenta alargada. Clinicamente os dentes são vistos com uma coloração castanha ou amarelada. O presente trabalho apresenta um caso clínico de odontodisplasia regional, não usual, que cruza a linha média da maxila, em uma criança do sexo feminino de quatro anos de idade. Todos os dentes decíduos e permanentes superiores do lado esquerdo eram displásicos, assim como também os incisivos centrais, decíduo e permanente do lado direito.

Regional odontodysplasia is a rare, nonhereditary condition that affects the development of the enamel and dentin of the deciduous and/or permanent teeth. The etiology is still unknown. In general, this condition is diagnosed during routine radiographic assessments and many cases are mistaken for malformed teeth or odontomas. This condition affects twice as many females as males and is more common in the maxilla than the jaw. This condition is also called "ghost teeth" because the enamel-dentine border of the affected teeth cannot be distinguished in the radiograph and the pulp chambers are wide. Clinically, the teeth are brownish or yellowish. The present study reports an unusual case of maxillary regional odontodysplasia crossing the midline in a 4-year-old female. All deciduous and permanent teeth in the left side of the maxilla and the central incisors in the right side of the maxilla were dysplastic.

Classificaçäo da DCM em pacientes do serviço de disfunçäo craniomandibular do Curso de Odontologia da Universidade Federal do Pará / Classification of CMD in patients seen at the craniomandibular dysfunction service, Federal University of Pará

Uma amostra de 120 pacientes, colhida no Serviço de Disfunçäo Craniomandibular do Curso de Odontologia da Universidade Federal do Pará, foi submetida ao questionário proposto por Martti Helkimo (1974). Foi considerado o sexo, a idade, cada resposta ao questionário e o nível de DCM obtido pela somatória das notas de cada pergunta. As respostas ao questionário foram caracteristicamente dicotômicas, na grande maioria dos casos ou houve aceitaçäo total ou rejeiçäo total das perguntas. E, com base nesse resultado, infere-se sobre o caráter influente do questionário. Os dados referentes a cada resposta foram submetidos à análises multivariadas, sendo as dez perguntas reduzidas a quatro fatores e aos 120 pacientes a 11 grupos. As perguntas que apresentaram caráter discriminante puderam ser definidas como referentes a (i) dor na cavidade bucal, (ii) sintomas indiretos e comportamentais, (iii) articulaçäo dos dentes e (iv) bruxismo. Apenas os grupos (i) e (iii) apresentaram correlaçäo com o nível de DCM, sendo que o primeiro a apresentou de maneira mais contudente