Alteración en la integridad de la membrana basal en agrandamiento gingival inducido por tratamiento ortodóncico

Introducción: El agrandamiento gingival inducido por tratamiento de ortodoncia es un aumento progresivo, localizado o generalizado del tejido gingival. Objetivo: Determinar aspectos morfológicos en la membrana basal del tejido gingival de pacientes con agrandamiento gingival inducido por tratamiento de ortodoncia. Métodos: Estudio descriptivo de corte transversal, donde se analizaron tejidos gingivales de pacientes con agrandamiento gingival inducido por tratamiento de ortodoncia (grupo test: n=5) e individuos sanos (grupo control: n=5) mediante análisis histológicos e inmunohistoquímico con anticuerpo policlonal anti-citoqueratina 14. Las interrupciones de la membrana basal grado 1 y grado 2 fueron identificadas. Fue utilizado el programa estadístico R versión 4.0.2 para Windows. Se declaró significancia si p <0,05. Resultados: Se constató la presencia de rupturas de la membrana basal en todos los pacientes del grupo test. Estos individuos presentaron una mayor cantidad de cambios morfológicos en el tejido gingival. Exponiendo así, valores estadísticamente significativos de rupturas de la membrana basal (Grado I) y rupturas rodeadas de células epiteliales y/o fibroblastos gingivales (Grado II) en comparación con el grupo control (p <0,001). Conclusión: El tejido epitelial de pacientes con agrandamiento gingival inducido por tratamiento de ortodoncia presenta una evidente pérdida en la integridad de la membrana basal. Estas discontinuidades sugieren un aumento considerable de la plasticidad del epitelio en pacientes con agrandamiento gingival inducido por tratamiento de ortodoncia.

Introduction: Orthodontic treatment-induced gingival enlargement is a progressive, localized or generalized increase in gingival tissue. Objective: To determine morphologic aspects in the basal membrane of the gingival tissue in patients with orthodontic treatment-induced gingival enlargement. Methods: A descriptive and cross-sectional study was carried out, in which gingival tissues of patients with orthodontic treatment-induced gingival enlargement (test group: n=5) and healthy individuals (control group: n=5) were analyzed by histological and immunohistochemical analysis with the polyclonal antibody anticytokeratin 14. Grade 1 and grade 2 disrupted basal membrane were identified. The statistical program R (version 4.0.2) for Windows was used. Significance was declared if p was greater than 0.05. Results: The presence of disrupted basal membranes was observed in all the patients from the test group. These individuals presented a greater number of morphological changes in the gingival tissue. Compared to the control group (p < 0.001), statistically significant values were observed for cases of disrupted basal membrane (grade I) and disruptions surrounded by epithelial cells or gingival fibroblasts (grade II). Conclusion: The epithelial tissue of patients with orthodontic treatment-induced gingival enlargement shows an evident loss of the basal membrane integrity. These discontinuities are suggestive of a considerable increase in epithelial plasticity in patients with orthodontic treatment-induced gingival enlargement.

Inmunolocalización de citoqueratina 14, 19 y Ki-67 en el epitelio de pacientes con agrandamiento gingival

R E S U M E N En pacientes con ortodoncia aparecen eventos patológicos no deseados como agrandamiento gingival inducido por tratamiento de ortodoncia (AGTO) o hipertrofia gingival. El objetivo del estudio es identificar la distribución inmunohistoquímica de citoqueratina CK-14, CK-19 y Ki-67 en epitelio gingival de pacientes con AGTO. Se seleccionaron I3 pacientes divididos en: grupo control (n=6), conformado por individuos periodontalmente sanos no portadores de aparatología ortodóntica y grupo test (n=7), integrado por pacientes con AGTO. Los marcadores CK-14, CK-19 y Ki-67 fueron identificados mediante inmunohistoquímica con anticuerpos monoclonales y observados en un microscopio óptico Leica DM 500. En los pacientes del grupo test el tejido epitelial se mostró hipertrófico con pérdida en la continuidad de la membrana basal. La CK-14 y CK-19 fue positiva en el epitelio de todos los sujetos evaluados, con una expresión positiva de alta intensidad en células de la lámina basal del grupo test. El promedio de células positivas para Ki-67 en el grupo test fue de 56%. En conclusión, la CK-14, CK-19 y Ki-67 son marcadores con elevada inmunoreactividad en tejido gingival de pacientes con AGTO portadores de ortodoncia.

During orthodontic treatment, unwanted pathological events such as gingival overgrowth induced by orthodontic treatment or gingival hypertrophy may appear The objective of this study is to identify immunohistochemical distribution of cytokeratin CK-14, CK-19 and Ki-67 in the gingival epithelium of patients with gingival overgrowth induced by orthodontic treatment. Thirteen patients were selected divided into: control group (n = 6), conformed of periodontally healthy individuals without orthodontic appliances and the test group (n = 7), conformed of patients with gingival overgrowth induced by orthodontic treatment. The biomarkers CK-14, CK-19 and Ki-67 were identified by immunohistochemistry with monoclonal antibodies and observed in a Leica DM 500 optical microscope. Hypertrophic epithelial tissue with loss of continuity of the basement membrane was found in the test group patients. CK-14 and CK-19 were positive in the epithelial tissue of all the subjects evaluated, with a high intensity positive expression in the cells of the basal lamina of the test group. The average number of cells positive for Ki-67 in test group was 56%. In conclusion, CK-14, CK-19 and Ki-67 are biomarkers with high immunoreactivity in the gingival tissue of patients with gingival overgrowth induced by orthodontic treatment.

Durante o tratamento ortodôntico, eventos patológicos indesejados como o crescimento gengival induzido pelo tratamento ortodôntico (CGTO) ou hipertrofia gengival podem aparecer: O objetivo deste estudo é identificar a distribuição imuno-histoquímica das citoqueratinas CK -14, CK-19 e Ki-67 no epitélio gengival de pacientes com CGTO. Foram selecionados 13 pacientes divididos em: grupo controle (n=6), conformado por indivíduos periodontalmente saudáveis sem aparelhos ortodônticos e o grupo teste (n=7), conformado por pacientes com CGTO. Os biomarcadores CK-14, CK-19 e Ki-67 foram identificados por imuno-histoquímica com anticorpos monoclonais e observados em microscópio óptico Leica DM 500. Tecido epitelial hipertrófico com perda de continuidade da membrana basal foi encontrado nos pacientes do grupo teste. CK-14 e CK-19 foram positivos no tecido epitelial de todos os sujeitos avaliados, com expressão positiva de alta intensidade nas células da lâmina basal do grupo teste. O número médio de células positivas para Ki-67 no grupo teste foi de 56%. Em conclusão, CK-14, CK-19 e Ki-67 são biomarcadores com alta imunorreatividade no tecido gengival de pacientes com CGTO.

Agrandamiento gingival asociado al tratamiento de ortodoncia: análisis histológico e inmunohistoquímico de dos casos clínicos

El agrandamiento gingival asociado al tratamiento de ortodoncia (AGTO) es el crecimiento no controlado de la encía. Aquí reportamos dos casos clínicos de pacientes masculinos sistèmicamente sanos con AGTO generalizado, con asociación a la biopelícula dental y sin esta. En ambos pacientes se identificó un tejido epitelial hiperplásico con abundantes células positivas para Ki-67 y tejido conectivo rico en fibras de colágeno distribuidas aleatoriamente. Futuros estudios serán útiles para dilucidar las diferencias fisiopatológicas del AGTO con relación con el biofilm dental y sin esta.

Orthodontic treatment-induce gingival overgrowth (OTGO) is uncontrolled growth of the gingiva. Here, we report two clinical cases of systemically healthy male patients with generalized GH undergoing orthodontic treatment, with and without association with dental biofilm. In both patients, hyperplastic epithelial tissue was identified with abundant Ki-67 positive cells and connective tissue rich in randomly distributed collagen fibers. Future studies will be useful to elucidate the pathophysiological differences of OTGO with and without relation to dental biofilm.

Enamel and dentin in Enamel renal syndrome: A confocal Raman microscopy view.

Enamel Renal Syndrome (ERS) is a rare genetic disorder caused by biallelic mutations in Family with sequence similarity 20A (FAM20A) gene encoding the secretory pathway pseudokinase FAM20A. ERS is characterized by hypoplastic amelogenesis imperfecta (AI), impaired tooth eruption, intra-pulpal calcifications, gingival fibromatosis and nephrocalcinosis of various severity. Previous studies showed that the hypoplastic enamel was also hypomineralized but its chemical composition has not been extensively studied. Furthermore it is currently unclear whether dentinal defects are associated with AI in ERS patients. The objective of the study was to provide a structural and chemical analysis of enamel, dentin and dentin enamel junction (DEJ) in ERS patients carrying four, previously reported, distinct mutations in FAM20A. Chemical cartography obtained with Raman microscopy showed that compared to control samples, ERS enamel composition was severely altered and a cementum-like structure was observed in some cases. Chemical composition of peripulpal dentin was also affected and usual gradient of phosphate intensity, shown in DEJ profile, was absent in ERS samples. DEJ and dentinal anomalies were further confirmed by scanning electron microscopy analysis. In conclusion, our study shows that enamel formation is severely compromised in ERS patients and provides evidence that dentinal defects are an additional feature of the ERS dental phenotype.

Colágeno tipo I en el agrandamiento gingival inducido por tratamiento ortodóntico: un estudio inmunohistoquímico piloto / Type I Collagen in Gingival Overgrowth Induced by Orthodontic Treatment: A Pilot Immunohistochemical Study / Colágeno do tipo I no aumento gengival induzido por tratamento ortodôntico: um estudo imuno-histoquímico piloto

el tratamiento ortodóntico es responsable del agrandamiento gingival (ag), una condición clínica caracterizada por el crecimiento patológico, difuso o localizado del tejido gingival. La acumulación excesiva de la matriz extracelular (mec), incluyendo el colágeno tipo I, parece contribuir a las manifestaciones patológicas del ag. El objetivo del artículo es identificar y describir la distribución del colágeno tipo I en el tejido gingival de pacientes con ag por ortodoncia fija. Materiales y métodos: estudio de tipo descriptivo que analizó los tejidos gingivales de sujetos diagnosticados con ag portadores de ortodoncia (test, n = 5) e individuos periodontalmente sanos (control, n = 5). Las muestras se obtuvieron mediante gingivectomía. Todas las biopsias fueron fijadas, incluidas en parafina, cortadas y analizadas por medio de la coloración rojo picrosirius/verde rápido, con el propósito de distinguir las fibras de colágeno. Mediante una reacción inmunohistoquímica, el colágeno tipo I fue identificado con anticuerpo monoclonal. Resultados: en los pacientes con ag por tratamiento ortodóntico, se identificó un tejido epitelial hiperplásico con aumento evidente de las prolongaciones epiteliales y un tejido conectivo con abundantes haces de fibras de colágenos, principalmente en la lámina basal y la zona subyacente. Las fibras de colágeno tipo I en los tejidos de pacientes con ag por ortodoncia fueron gruesas de aspecto desorganizado, con una tinción inmunohistoquímica intensa, en comparación con las fibras del grupo control. Conclusiones: el aumento de fibras de colágenos, en especial de colágeno de tipo I, es un hallazgo histológico que caracteriza a los pacientes con ag por ortodoncia fija.

Orthodontic treatment is responsible for gingival overgrowth (go), a clinical condition charac-terized by pathological, diffuse, or localized growth of gingival tissue. Excessive accumulation of the extra-cellular matrix, including type I collagen, contributes to the pathological manifestations of go. The objective of this study is to identify and describe the distribution of type I collagen in the gingival tissue of patients with go because of fixed orthodontics. Materials and Methods: A descriptive study that analyzed the gingival tissues of subjects diagnosed with go with orthodontic (test, n = 5) and periodontally healthy individuals (control, n = 5). The samples were obtained by gingivectomy. All the biopsies were fixed, embedded in paraf-fin, and cut and analyzed using picrosirius red/fast green staining, in order to distinguish the collagen fiber. By means of an immunohistochemical reaction, type I collagen was identified with a monoclonal antibody. Results: A hyperplastic epithelial tissue was identified with an evident increase in epithelial processes and connective tissue with abundant bundles of collagen fiber, mainly in the basal lamina and the underlying area in patients with go because of orthodontic treatment. Type I collagen fiber in the tissues of patients with orthodontic go were thick and disorganized in appearance with intense immunohistochemical stain-ing, compared to the fibers of the control group. Conclusions:The increase in collagen fibers, particularly type I collagen, is a histological finding that characterizes patients with go because of fixed orthodontics.

• tratamento ortodôntico é responsável pelo aumento gengival (ag), uma condição clínica caracterizada pelo crescimento patológico difuso ou localizado do tecido gengival. O acúmulo excessivo de matriz extracelular (mec), incluindo colágeno tipo I, parece contribuir para as manifestações patoló-gicas do ag. O objetivo deste trabalho é identificar e descrever a distribuição do colágeno do tipo I no tecido gengival de pacientes com AG devido à ortodontia fixa. Materiais e métodos: estudo descritivo que analisou os tecidos gengivais de indivíduos diagnosticados com ag em uso de ortodontia (teste, n = 5) e indivíduos periodontalmente saudáveis (controle, n = 5). As amostras foram obtidas por gengivectomia. Todas as biópsias foram fixadas, embebidas em parafina, cortadas e analisadas com coloração picrosirius vermelho/verde rápido, a fim de distinguir as fibras colágenas. Usando uma reação imuno-histoquímica, o colágeno tipo I foi identificado com anticorpo monoclonal. Resultados: em pacientes com ag devido ao tratamento ortodôntico, foi identificado tecido epitelial hiperplásico com evidente aumento das exten-sões epiteliais e tecido conjuntivo com abundantes feixes de fibras colágenas, principalmente na lâmina basal e região subjacente. As fibras de colágeno tipo I em tecidos de pacientes com ag ortodôntico eram espessas com aspecto desorganizado e intensa coloração imuno-histoquímica, em comparação com as fibras do grupo controle. Conclusões: o aumento das fibras colágenas, principalmente do colágeno do tipo I, é um achado histológico que caracteriza os pacientes com ag devido à ortodontia fixa.

Fibrina rica en plaquetas aplicada en conjunto con biomodificación alveolar en la regeneración ósea guiada. Reporte de caso / Platelet rich fibrin applied in conjunction with alveolar biomodification in guided bone regeneration. Case report

Resumen Debido a los diferentes problemas existentes a la hora de realizar procedimientos implantológicos por la calidad del remanente óseo y el estado de los tejidos periodontales en la zona a rehabilitar, existen opciones de regeneración ósea inducida que muestran tasas de éxito considerablemente favorables al ser empleadas en casos donde es necesario una regeneración ósea extensa capaz de soportar un implante dental. El objetivo del presente estudio es describir el éxito en la regeneración ósea por medio de alternativas combinadas de fibrina rica en plaquetas y modificación biológica con tetraciclina. Reporte del caso clínico: paciente femenina de 67 años que consulta por inflamación, supuración y dolor en la zona antero-superior. Es diagnosticada como absceso periapical crónico, se hace cirugía de extracción de órganos dentales, posterior a eso se hace descontaminación y adaptación de los alveolos con tetraciclina para realizar la regeneración ósea guiada con fibrina rica en plaquetas, hueso tipo Lumina Bone Porous y Lumina Coat, luego se espera el proceso de evolución para colocación de implantes dentales. El éxito obtenido con el uso de fibrina rica en plaquetas y la biomodificacion ósea como coadyuvante en la desinfección de la zona a tratar, logró mostrar resultados altamente favorables en el proceso de regeneración ósea guiada.

Abstract As the different problems that exist when carrying out implant procedures due to the quality of the bone remnant and the state of the existing periodontal tissues in the area to be rehabilitated, there are induced bone regeneration options that show considerably favorable success rates as they are used in cases where extensive bone regeneration is necessary to support a dental implant. The objective of the present study is to describe the success in bone regeneration using combined alternatives of platelet-rich fibrin and biological modification with tetracycline. Report of the clinical case: 67-year-old female patient consulted for inflammation and suppuration and pain in the upper antero-superior area, she was diagnosed as chronic periapical abscess, dental organ extraction surgery was performed, after that decontamination and adaptation of the alveoli was performed with tetracycline to perform guided bone regeneration with platelet-rich fibrin, Coat membrane and Lumina bone criteria, then wait for the evolution process to place dental implants. The success obtained with the use of FRP and bone biomodification as an adjuvant in the disinfection of the area to be treated achieved to show highly favorable results in the guided bone regeneration process.

Comparación de métodos histoquímicos para el análisis histológico de muestras de encía humana / Comparison of histochemical methods for the histological analysis of human gingival samples

Introducción: Las técnicas de coloración histológica son útiles en el análisis ultraestructural de muestras tisulares, incluyendo el tejido gingival. Objetivo: Comparar la utilidad de tres métodos histoquímicos (hematoxilina-eosina, Masson Goldner y rojo sirio) en la identificación de elementos celulares y otros constituyentes del tejido gingival. Métodos: Estudio experimental in vitro que comprendió el análisis de tejidos gingivales de donantes sanos sin signos de inflamación gingival y con indicación de cirugía periodontal. Las muestras de encía se obtuvieron mediante gingivectomía, se procesaron e incluyeron en parafina, posteriormente se realizaron cortes con un micrótomo y se depositaron en portaobjetos de adhesión con polisina. Las muestras se agruparon y fueron teñidas con hematoxilina-eosina, Masson Goldner y rojo sirio, finalmente fueron visualizadas en un microscopio óptico Leica DM 750®. La lectura de los hallazgos fue realizada por patólogos orales. Resultados: La coloración hematoxilina-eosina evidencia elementos celulares y extracelulares del tejido epitelial y conectivo. Núcleos de color azul violeta, citoplasmas rosados, fibras de colágeno de matiz rosa claro, arteriolas y vénulas con túnica adventicia, media e íntima diferenciadas. La coloración Masson Goldner diferencia núcleos de coloración púrpura y citoplasma fucsia, presenta especificidad en identificar fibras de colágeno con tonalidad verde, distribuidas densa, homogénea y paralelamente en el tejido conectivo gingival. La tinción rojo sirio, permitió identificar las fibras de colágeno de color rosa brillante, mientras que el tejido epitelial y los vasos sanguíneos fueron de color amarillo. Conclusión: Cada coloración histológica evaluada en el presente trabajo tiene cierta afinidad y sensibilidad por estructuras celulares y componentes de la matriz extracelular específica. Su empleo es útil en el estudio de tejidos gingivales y podrían contribuir en el análisis de biopsias gingivales(AU)

Introduction: Histological staining techniques are useful in the ultrastructural analysis of tissue samples, including gingival tissue. Objective: Compare the usefulness of three histochemical methods (hematoxylin-eosin, Masson-Goldner and sirius red) for identification of cellular elements and other constituents of gingival tissue. Methods: An in vitro experimental study was conducted which included the analysis of gingival tissue from healthy donors without gingival inflammation signs and indication of periodontal surgery. The gum samples were obtained by gingivectomy, processed with paraffin, cut with a microtome and placed on Polysine adhesion slides. The samples were grouped, stained with hematoxylin-eosin, Masson Goldner and sirius red, and visualized under a Leica DM 750® microscope. Reading of the findings was performed by oral pathologists. Results: Hematoxylin-eosin staining found cellular and extracellular elements of the epithelial and connective tissue: violet-blue nuclei, pink cytoplasms, light rose collagen fibers, and arterioles and venules with differentiated tunica adventitia, media and intima. Masson-Goldner staining differentiated purple nuclei and fuchsia cytoplasm. It displayed specificity identifying green collagen fibers with dense, homogeneous and parallel distribution in the gingival connective tissue. Sirius red staining allowed identification of bright rose collagen fibers, whereas epithelial tissue and blood vessels were yellow. Conclusion: Each of the histological staining methods evaluated in the study shows a certain affinity with and sensitivity to cellular structures and components of the specific extracellular matrix. All three are useful for the study of gingival tissue and could contribute to the analysis of gingival biopsies(AU)

Pathogenesis of Enamel-Renal Syndrome Associated Gingival Fibromatosis: A Proteomic Approach.

The enamel renal syndrome (ERS) is a rare disorder featured by amelogenesis imperfecta, gingival fibromatosis and nephrocalcinosis. ERS is caused by bi-allelic mutations in the secretory pathway pseudokinase FAM20A. How mutations in FAM20A may modify the gingival connective tissue homeostasis and cause fibromatosis is currently unknown. We here analyzed conditioned media of gingival fibroblasts (GFs) obtained from four unrelated ERS patients carrying distinct mutations and control subjects. Secretomic analysis identified 109 dysregulated proteins whose abundance had increased (69 proteins) or decreased (40 proteins) at least 1.5-fold compared to control GFs. Proteins over-represented were mainly involved in extracellular matrix organization, collagen fibril assembly, and biomineralization whereas those under-represented were extracellular matrix-associated proteins. More specifically, transforming growth factor-beta 2, a member of the TGFß family involved in both mineralization and fibrosis was strongly increased in samples from GFs of ERS patients and so were various known targets of the TGFß signaling pathway including Collagens, Matrix metallopeptidase 2 and Fibronectin. For the over-expressed proteins quantitative RT-PCR analysis showed increased transcript levels, suggesting increased synthesis and this was further confirmed at the tissue level. Additional immunohistochemical and western blot analyses showed activation and nuclear localization of the classical TGFß effector phospho-Smad3 in both ERS gingival tissue and ERS GFs. Exposure of the mutant cells to TGFB1 further upregulated the expression of TGFß targets suggesting that this pathway could be a central player in the pathogenesis of the ERS gingival fibromatosis. In conclusion our data strongly suggest that TGFß -induced modifications of the extracellular matrix contribute to the pathogenesis of ERS. To our knowledge this is the first proteomic-based analysis of FAM20A-associated modifications.

Complicaciones asociadas a la anestesia bucal: diagnóstico y manejo / Complications associated with oral anesthesia: diagnosis and management

La seguridad en el uso de los anestésicos locales y de los diferentes abordajes anestésicos en odontología, hacen que la práctica de la anestesia dental sea considerada segura. Los autores reportan las diferentes complicaciones inherentes al uso de la anestesia dental y las consideradas sistémicas, describiendo en cada una de ellas sus características, signos, síntomas y estrategias para solventarlas. (AU)

The safety in the use of local anesthetics and the different anesthetic approaches in dentistry make the practice of dental anesthesia considered safe. The authors report the different complications inherent to the use of dental anesthesia and those considered systemic, describing in each of them their characteristics, signs, symptoms and strategies to solve them. (AU)

Alteración del colágeno en la Hipertrofia Gingival en pacientes con ortodoncia: caracterización histológica e inmunohistoquímica / Collagen alteration in gingival hypertrophy in orthodontic patients: histological and immunohistochemical characterization

Resumen Introducción: La hipertrofia gingival (HG) es el aumento del volumen de la encía asociado a ciertas enfermedades sistémicas, hereditarias (idiopático), ingesta de algunos medicamentos o a factores locales como el tratamiento ortodóntico, capaz de provocar cambios histológicos en el tejido conectivo gingival. Objetivo: Describir las características histológicas e identificar el colágeno tipo I y tipo III en tejidos gingivales de sujetos con hipertrofia gingival portadores de ortodoncia. Materiales y método: Se diseñó un estudio de casos y controles que incluyó el análisis de biopsias de tejido gingival de 12 pacientes sometidos a cirugías periodontales. La muestra se dividió en dos grupos: individuos sanos (control; n= 6) y pacientes con HG portadores de ortodoncia (pacientes; n= 6). Las muestras fueron procesadas e incluidas en parafina. Las tinciones Masson-Goldner y rojo sirius/verde rápido fueron empleadas. El colágeno tipo I y tipo III fueron identificados mediante inmunohistoquímica con anticuerpos monoclonales. Resultado: En los pacientes con HG portadores de ortodoncia se observó un epitelio hiperplásico y tejido conectivo denso con abundantes fibras de colágeno distribuidos aleatoriamente. La inmunodetención de colágeno tipo I indicó la presencia de abundantes fibras desorganizadas y el colágeno tipo III fue inmunolocalizado subyacente a la membrana basal, vasos sanguíneos y toda la extensión del tejido conectivo de los pacientes con HG con tratamiento ortodóntico. Conclusión: La acumulación de fibras de colágeno, particularmente del colágeno tipo I y tipo III, son hallazgos histológicos que caracterizan la HG en pacientes portadores de ortodoncia. Futuros estudios son necesarios para dilucidar el fenotipo de los fibroblastos gingivales y la probable pérdida homeostática entre la producción y degradación de colágeno en esta patología.

Abstract Introduction: Gingival hypertrophy (GH) is the increase in the volume of the gingiva associated with certain systemic, hereditary (idiopathic) diseases, the intake of some medications or local factors such as orthodontic treatment, capable of causing histological changes in the gingival connective tissue. Objective: To describe the histological characteristics and identify type I and type III collagen in gingival tissues of subjects with gingival hypertrophy wearing orthodontics. Method: A case-control study was designed that included the analysis of gingival tissue biopsies from 12 patients submitted to periodontal surgeries. The sample was divided into two groups: healthy individuals (Control; n= 6) and patients with GH wearing orthodontics (Patients; n= 6). The samples were processed and embedded in paraffin. Masson-goldner and sirius red/fast green stains were used. Type I and type III collagen were identified by immunohistochemistry with monoclonal antibodies. Result: A hyperplastic epithelium and dense connective tissue with abundant randomly distributed collagen fibers were observed in patients with orthodontic GH. Immunodetention of type I collagen indicated the presence of abundant disorganized fibers and type III collagen was inmunolocalized underlying the basement membrane, blood vessels and the entire extension of the connective tissue of patients with GH orthodontic. Conclusion: The accumulation of collagen fibers, particularly type I and type III collagen, are histological findings that characterize GH in orthodontic wearers. Future studies are necessary to elucidate the phenotype of gingival fibroblasts and the probable homeostatic loss between collagen production and degradation in this pathology.

Immunolocalización de s100a4 y α-sma en tejidos gingivales de pacientes con hipertrofia gingival por tratamiento ortodóntico: estudio preliminar / Immunolocalization of s100a4 and α-sma in gingival tissues of patients with gingival hypertrophy due to orthodontic treatment: preliminary study

Objective: to determine the presence and distribution of markers of the epithelialmesenchymal transition (EMT) (S-100A4 and alpha-smooth muscle actin-α-SMA) in gingival tissues of patients affected by Gingival hypertrophy (GH) due to orthodontics.GH is an exaggerated increase in gingival tissue whose pathogenesis is unknown. However, it has been reported that the epithelial-mesenchymal transition as a process involved in other types of GH. Materials and methods: descriptive study that included the analysis of gingival tissues of healthy individuals (n = 6) and patients with GH by orthodontic treatment (n = 6). Before gingival surgery, the patients underwent a periodontal hygiene phase. The gingival tissue samples obtained were processed and embedded in paraffin. The cuts were made with a microtome and deposited on polysine adhesion slides. Histological hematoxylin-eosin staining was performed.The identification and location of S-100A4 and α-SMA markers was determined by immunohistochemistry with monoclonal antibodies. The reading of the findings was carried out by oral pathologists. Results: in healthy individuals, an S100A4 label was observed in Langerhans cells, while α-SMA was identified in the vascular endothelium of all samples analysed. However, in patients with GH due to orthodontics, they registered an intense staining of S100A4 in gingival fibroblasts, Langerhans cells, vascular endothelium, and areas adjacent to the rupture of blood vessel. α-SMA expression in GO was detected in the vascular endothelium and gingival fibroblasts. Conclusion: the differential immunostaining of EMT markers in gingival tissues of patients with orthodontic GH suggests an eventual role of EMT in the pathogenesis of this pathology..Au

Objective: to determine the presence and distribution of markers of the epithelialmesenchymal transition (EMT) (S-100A4 and alpha-smooth muscle actin-α-SMA) in gingival tissues of patients affected by Gingival hypertrophy (GH) due to orthodontics. GH is an exaggerated increase in gingival tissue whose pathogenesis is unknown. However, it has been reported that the epithelial-mesenchymal transition as a process involved in other types of GH. Materials and methods: descriptive study that included the analysis of gingival tissues of healthy individuals (n = 6) and patients with GH by orthodontic treatment (n = 6). Before gingival surgery, the patients underwent a periodontal hygiene phase. The gingival tissue samples obtained were processed and embedded in paraffin. The cuts were made with a microtome and deposited on polysine adhesion slides. Histological hematoxylin-eosin staining was performed. The identification and location of S-100A4 and α-SMA markers was determined by immunohistochemistry with monoclonal antibodies. The reading of the findings was carried out by oral pathologists. Results: in healthy individuals, an S100A4 label was observed in Langerhans cells, while α-SMA was identified in the vascular endothelium of all samples analysed. However, in patients with GH due to orthodontics, they registered an intense staining of S100A4 in gingival fibroblasts, Langerhans cells, vascular endothelium, and areas adjacent to the rupture of blood vessel. α-SMA expression in GO was detected in the vascular endothelium and gingival fibroblasts. Conclusion: the differential immunostaining of EMT markers in gingival tissues of patients with orthodontic GH suggests an eventual role of EMT in the pathogenesis of this pathology..Au

Diferencias fenotípicas de fibroblastos gingivales en sujetos con hiperplasia gingival idiopática frente a sujetos periodontalmente sanos: estudio piloto / Phenotypic differences of gingival fibroblasts in subjects with idiopathic gingival hyperplasia versus periodontally healthy subjects: a pilot study

Objetivo: identificar, describir y diferenciar las características fenotípicas de los fibroblastos gingivales (FGs) en pacientes con hiperplasia gingival idiopática (HGI) e individuos periodontalmente sanos. Métodos: los FGs fueron aislados a partir de tejido gingival de individuos periodontalmente sanos (n=2) y pacientes con HGI (n=2). Los FGs se cultivaron en el medio DMEM (Dulbecco's Modified of Eagle Medium) a 37°C con 5% de CO2. La identificación y localización de la actina, vimentina y mitocondrias en FGs fue realizada y evaluada microscópicamente mediante inmunofluorescencia con anticuerpos monoclonales. La capacidad de migración de los FGs en los pacientes con HGI e individuos sanos también fue estudiada. Resultados: todos los FGs fueron mononucleares, fusiformes y con prolongaciones citoplasmáticas visibles. La faloidina permitió identificar una densa red de actina en los FGs de pacientes con HGI, contrariamente a los FGs de individuos periodontalmente sanos. La vimentina y mitocondrias fueron identificadas en los FGs de individuos sanos y pacientes con HGI sin ninguna alteración en su expresión y localización. La migración de la monocapa de los FGs indicó una actividad de migración celular importante en los FGs de los pacientes con HGI, en relación a los FGs de los individuos periodontalmente sanos. Conclusión: los FGs de pacientes con HGI conservan características fenotípicas celulares similares a los FGs de individuos periodontalmente sanos. Sin embargo, los FGs de pacientes con HGI simulan tener una mayor capacidad migratoria que amerita ser explorada en futuros trabajos de investigación.

Objective: To identify and to describe the phenotypic characteristics of gingival fibroblasts from patients with idiopathic gingival hyperplasia (IGH) and periodontally healthy individuals. Methods: Gingival fibroblasts (GFs) were isolated from gingival tissue from periodontally healthy individuals (n=2) and patients with IGH (n=2). The GFs were grown in DMEM (Dulbecco's Modified of Eagle Medium) at 37°C with 5% CO2. The identification and location of actin, vimentin and mitochondria in GFs were performed and evaluated microscopically by immunofluorescence with monoclonal antibodies. The migration capacity of GFs from IGH and healthy individuals was also studied. Results: All the GFs were mononuclear, fusiform and with visible cytoplasmic extensions. The phalloidin allowed to identify a dense actin network in the GFs of patients with IGH, contrary to the GFs of periodontally healthy individuals. Vimentin and mitochondria were identified in the GFs of healthy individuals and patients with IGH without any alteration in their expression and location. Monolayer migration of GFs indicates significant cell migration activity in the GFs of patients with IGH in relation to the GFs of periodontally healthy individuals. Conclusion: GFs from patients with IGH retain cellular phenotypic characteristic similar to GFs from periodontally healthy individuals. However, the GFs of patients with IGH simulate having a greater migratory capacity that deserves to be explored in future research works.

Biodentine: ¿sustituto de la dentina? / Biodentine: a dentine substitute?

RESUMEN La biodentine es un biomaterial a base de silicato de calcio que ha sido concebido como un sustituto de la dentina. Es frecuentemente utilizada en diversas situaciones clínicas, como el recubrimiento pulpar (directo e indirecto), la pulpotomía, perforaciones endodónticas, apexificación, entre otras. El objetivo de esta revisión es proporcionar un análisis detallado de las propiedades fisicoquímicas, biológicas y las principales aplicaciones clínicas de la biodentine. Se realizó una búsqueda electrónica de literatura en las bases de datos Medline (Pubmed), EBS-CO-Host y Science direct (Elsevier) hasta julio de 2020. El 92 % de los artículos referenciados en esta revisión fueron seleccionados a través de esta búsqueda. Los resultados indican que la biodentine es un cemento de uso odontológico con excelentes propiedades que se caracteriza por una manipulación relativamente fácil, bajo costo, alta resistencia a la compresión y flexión, alta biocompatibilidad y una excelente bioactividad. En conclusión, la biodentine es un material recomendado en el tratamiento de diversas situaciones clínicas de la práctica odontológica dadas sus características fisicoquímicas y biológicas. Sin embargo, se hace necesario ensayos clínicos controlados aleatorizados que permitan determinar y confirmar que la biodentine es realmente un material odontológico capaz de reemplazar la dentina.

ABSTRACT Biodentine is a calcium silicate-based biomaterial that has been conceived as a substitute for dentin. It is frequently used in various clinical situations such as pulp lining (direct and indirect), pulpotomy, endodontic perforations, apexification, among others. The objective of this review is to provide a detailed analysis of the physical-chemical, biological properties and the main clinical applications of biodentine. An electronic literature search was carried out in the databases Medline (Pubmed), EBSCO-Host and Science direct (Elsevier) until July 2020. 92 % of the articles referenced in this review were selected through this search. The results indicate that biodentine is a cement for dental use with excellent properties characterized by relatively easy handling, low cost, high compressive and flexural strength, high biocompatibility and excellent bioactivity. In conclusion, biodentine is a recommended material in the treatment of various clinical situations in dental practice given its physicochemical and biological characteristics. However, randomized controlled clinical trials are necessary to determine and confirm that biodentine is indeed a dental material capable of replacing dentin.

Aspectos morfológicos in vivo e in vitro de fibroblastos gingivales en pacientes con agrandamiento gingival / Morphological aspects in vivo and in vitro of gingival fibroblasts in patients with gingival overgrowth / Aspectos morfológicos in vivo e in vitro de fibroblastos gengivais em pacientes com aumento gengival

RESUMEN El agrandamiento gingival (AG) es el incremento excesivo del volumen del tejido gingival. Comúnmente el AG se asocia con enfermedades sistémicas, ingesta prolongada de medicamentos y factores locales como el tratamiento por ortodoncia. El propósito de este trabajo fue identificar y describir los aspectos morfológicos in vivo e in vitro de los tejidos y fibroblastos gingivales en pacientes con AG por ortodoncia e individuos sanos. La identificación de fibras de colágeno y elásticas de la encía de cuatro pacientes con AG por ortodoncia y cinco individuos sanos fueron realizados mediante las tinciones Masson-Goldner y Orceina, respectivamente. A partir de cultivos celulares, la morfología de fibroblastos gingivales se estudió por inmunofluorescencia. Para este fin, fueron empleados los anticuerpos anti-actina y anti-vimentina. En los pacientes con AG por ortodoncia fue observado tejido epitelial con acantosis e hiperqueratinizado, acompañado de red compacta de fibras de colágeno. Las fibras elásticas de pacientes con AG tuvieron una disposición difusa y aleatoria en el tejido conjuntivo. Los fibroblastos gingivales in vitro de pacientes con AG por ortodoncia e individuos sanos fueron morfológicamente similares.

ABSTRACT Gingival overgrowth (GO) is the excessive increase in the volume of the gingival tissue. GO is commonly associated with systemic diseases, prolonged intake of medications, and local factors such as orthodontic treatment. The purpose of this work was to identify and describe the in vivo and in vitro morphological aspects of gingival tissues and fibroblasts in patients with GO due to orthodontics and healthy individuals. The identification of collagen and elastic fibers of the gingiva of four patients with GO by orthodontics and five healthy individuals were performed using Masson-Golgner and Orcein stains, respectively. From cell cultures, the morphology of gingival fibroblasts was studied by immunofluorescence. For this purpose, anti-actin and anti-vimentin antibodies were used. In patients with GO due to orthodontia, epithelial tissue with acanthosis and hyperkeratinization was observed, accompanied by a compact network of collagen fibers. The elastic fibers of GO patients had a diffuse and random arrangement in the connective tissue. In vitro, gingival fibroblast from orthodontic GO patients and healthy individuals were morphologically similar

RESUMO O aumento gengival (AG) é o aumento excessivo do volume do tecido gengival. A AG é comumente associada a doenças sistémicas, ingestão prolongada de medicamentos e fatores locais, como tratamento ortodôntico. O objetivo deste trabalho foi identificar e descrever os aspectos morfológicos in vivo e in vitro dos tecidos gengivais e fibroblastos em pacientes com AG ortodôntica e indivíduos saudáveis. A identificação de fibras colágenas e elásticas das gengivas de quatro pacientes com AG por ortodontia e, cinco indivíduos saudáveis, foram realizados com as manchas de Masson-Goldner e Orceina, respectivamente. A partir de culturas celulares, a morfologia dos fibroblastos gengivais foi estudada por imunofluorescência. Para este fim, foram utilizados anticorpos anti-actina e anti-vimentina. Nos pacientes com AG devido à ortodontia, observou-se tecido epitelial com acantose e hiperqueratinização, acompanhado por uma rede compacta de fibras colágenas. As fibras elásticas dos pacientes com AG tiveram uma disposição difusa e aleatória no tecido conjuntivo. Os fibroblastos gengivais in vitro de pacientes com AG ortodôntica e indivíduos saudáveis foram morfologicamente semelhantes.

Supervivencia de células fibroblásticas humanas en ausencia de suplementación / Survival of Human Fibroblastic Cells in the Absence of Supplementation

Resumen Introducción. Los fibroblastos gingivales (FGs) son células del tejido conjuntivo gingival que han tomado en los últimos años una relevancia promisoria por su probable utilización en la terapia celular, dadas sus capacidades de multipotencialidad y de autorrenovación. Objetivo. Conocer y describir el impacto de la ausencia en la suplementación de Suero Fetal Bovino (SFB) en la supervivencia de fibroblastos gingivales en cultivos. Materiales y métodos. Fibroblastos gingivales fueron aislados de tejido gingival de pacientes sanos y cultivados en medios de cultivos DMEM (Dulbecco's Modified of Eagle Medium) en ausencia y suplementados con 0.2% de SFB a 37°C en una atmósfera húmeda con 5% de CO2. Se llevó a cabo una evaluación morfológica, de supervivencia y proliferación de los FGs, así como la identificación mediante la técnica de inmunofluorescencia de marcadores del citoesqueleto celular como la actina y mitocondrias. Resultados. Los FGs cultivados en ausencia y con suplementación de 0.2% de SFB evidenciaron una forma fusiforme, con núcleos ovalados y numerosas prolongaciones citoplasmáticas durante el tiempo de cultivo. Un leve aumento en la proliferación de FGs fue observado en aquellas células en contacto con el medio DMEM+0.2% de SFB comparadas con el medio donde estuvo ausente la suplementación. El inmunomarcaje de la actina y las mitocondrias dejó en evidencia que la ausencia y suplementación a 0.2% de SFB no afectó su localización en los FGs evaluados. Conclusión. Los fibroblastos gingivales sobreviven y proliferan en ausencia de SFB, conservando sus características morfológicas celulares.

Abstract Introduction. Gingival fibroblasts (GF) are cells of gingival connective tissue that have taken promising relevance in recent years due to their probable use in cell therapy, given their multipotencial and self-renewal capabilities. Objective. To know and to describe the impact of the absence of Fetal Bovine Serum (FBS supplementation on the survival of gingival fibroblasts in cultures. Materials and methods. Gingival fibroblasts were isolated from gingival tissue of healthy patients and cultured in DMEM (Dulbecco's Modified of Eagle Medium) culture media in absence and supplemented with 0.2% FBS at 37 ° C in a humid atmosphere with 5% CO2. A morphological evaluation, survival and proliferation of GF were carried out, as well as the identification by the immunofluorescence technique of cellular cytoskeleton markers such as actin and mitochondria. Results. The GF grown in the absence and with supplementation of 0.2% FBS showed a fusiform shape, with oval nuclei and numerous cytoplasmic extensions during the culture time. A slight increase in the proliferation of GF was observed in those cells in contact with the DMEM medium +0.2% FBS compared to the medium where the supplementation was absent. Immunostaining of actin and mitochondria showed that the absence and supplementation to 0.2% of FBS did not affect its location in the evaluated. Conclusion. Gingival fibroblasts survive and proliferate in the absence of FBS, preserving their cellular morphological characteristics.

Expression of type III collagen in hypertrophic gingival tissue of patients with orthodontic treatment: a pilot study / Expresión del colágeno tipo III en tejido con hipertrofia gingival de pacientes con tratamiento ortodóntico: un estudio piloto

ABSTRACT Introduction: gingival hypertrophy (GH) is the uncontrolled increase in gingival volume induced by different etiological factors, including orthodontic treatment. This pathology is characterized by changes in epithelial and connective tissue, including modifications in the extracellular matrix. The present study determined the presence and distribution of type III collagen in tissues of patients with GH wearing fixed orthodontic appliances. Methods: 12 samples of gingival tissue were obtained from patients undergoing periodontal surgery. They were divided into two groups, the first with healthy patients (control; n = 6) and the second with patients diagnosed with GH and orthodontic treatment (patients; n = 6). Each obtained sample was subjected to the hematoxylin-eosin stain, Masson-Goldner staining, and type III collagen immunohistochemistry. Results: the hematoxylin-eosin and Masson-Goldner histological stains showed hypertrophia of epithelial tissue and connective tissue with a marked collagen fiber increase in the gingival tissue of orthodontic wearers with GH compared to individuals in the control group. The gingival tissue of patients with GH caused by orthodontic treatment showed a distribution and location of type III collagen near the basal lamina, around the blood vessels, but unlike the control group, its location was noticeable throughout the connective tissue. Conclusion: the gingival tissues of orthodontic wearers with GH experience an increase in the number and density of collagen fibers. Type III collagen seems to lose its usual location in the gingival tissues of orthodontic wearers with GH.

RESUMEN Introducción: la hipertrofia gingival (HG) es el aumento descontrolado del volumen de la encía debido a diversos factores etiológicos, entre ellos el tratamiento ortodóntico. Esta patología se caracteriza por cambios del tejido epitelial y conectivo, incluyendo modificaciones en la matriz extracelular. El presente estudio determinó la presencia y distribución de colágeno tipo III en tejidos de pacientes con HG portadores de ortodoncia fija. Métodos: se obtuvieron 12 muestras de tejido gingival de pacientes sometidos a cirugías periodontales. Se dividieron en dos grupos, el primero, integrado por pacientes sanos (control; n=6), y el segundo por pacientes diagnosticados con HG con ortodoncia (pacientes; n=6). Cada muestra obtenida fue sometida a la coloración hematoxilina-eosina, Masson-Goldner e inmunohistoquímica del colágeno tipo III. Resultados: las tinciones histológicas hematoxilina-eosina y Masson-Goldner permitieron constatar hiperplasia del tejido epitelial y un tejido conectivo denso con notable aumento de las fibras de colágeno en el tejido gingival de los pacientes con HG portadores de ortodoncia en comparación con los individuos del grupo control. El tejido gingival de pacientes con HG por ortodoncia evidenció una distribución y localización del colágeno tipo III cerca de la lámina basal, alrededor de los vasos sanguíneos, pero a diferencia del grupo control, su localización fue notoria en toda la extensión del tejido conectivo. Conclusión: los tejidos gingivales de pacientes con HG portadores de ortodoncia experimentan aumento en número y densidad de las fibras de colágeno. El colágeno tipo III parece perder su localización habitual en los tejidos gingivales de pacientes con HG portadores de ortodoncia.

Cultivo de células epiteliales dentales: impacto del suero fetal bovino / Culture of dental epithelial cells: impact of fetal bovine serum

RESUMEN Objetivo. Describir la influencia del Suero Fetal Bovino (SFB) en la supervivencia, crecimiento y expresión de organelas celulares en las células epiteliales dentales de rata. Materiales y métodos. Cultivos de células epiteliales dentales de rata fueron llevados a cabo a 37°C en una atmosfera húmeda, en ausencia y a una concentración de 10% de SFB. Una evaluación morfológica fue realizada durante la proliferación y confluencia de las células en cultivo. Dobles marcajes por inmunofluorencia fueron efectuados haciendo uso de anticuerpos anti-actina, anti-TOMM20 y anti-LAMP1. Resultados. Se evidenciaron células epiteliales dentales circulares u ovoides con núcleos voluminosos durante la proliferación y confluencias de manera similar en las células cultivas en presencia y ausencia de SFB. La carencia de SFB impactó negativamente la proliferación de las células epiteliales. No fueron observadas alteraciones en la localización de los inmunomarcajes anti-actina, anti-TOMM20 y anti-LAMP1 en las dos condiciones de cultivos experimentales. Conclusiones. La supresión del SFB en el cultivo de células epiteliales dentales de rata disminuyó la supervivencia, proliferación y sugiere no tener un impacto sobre las organelas evaluadas.

ABSTRACT Objective. Describe the influence of Fetal bovine serum (FBS) on the survival, growth and expression of cellular organelles in rat dental epithelial cells. Material and methods. Cell cultures of rat dental epithelial cells were carried out at 37°C in a humid atmosphere, in the absence and at a concentration of 10% FBS. Morphological evaluation was performed during the proliferation and confluence of cell in culture. Double immunofluorescence labels were made using anti-Actin, anti-TOMM20A, and anti-LAMP1 antibodies. Results. Circular or ovoid dental epithelial cells with bulky nuclei were evidenced during proliferation and confluences in a similar manner in culturing cells in the presence and absence of FBS. The lack of FBS negatively impacts the proliferation of epithelial cells. No alterations were observed in the localization of the anti-actin, anti-TOMM20 and anti-LAMP1 immunomarkers in both conditions of experimental cultures. Conclusion. FBS suppression in rat dental epithelial cells decreased survival, proliferation and suggests not having an impact on the organelles evaluated.

Expresión inmunohistoquímica de la proteína de membrana asociada a lisosomas en ameloblastos de ratones

Introducción: La formación del esmalte dental comprenden cambios morfofuncionales de los ameloblastos. Estas modificaciones involucran la participación activa de organelas celulares, entre ellas los lisosomas. Objetivo: Identificar la distribución de la proteína de membrana asociada a lisosomas en los ameloblastos. Materiales y métodos: Incisivos en crecimiento continuo de ratones con edades de 7, 14 días y células epiteliales dentales de rata fueron utilizadas para identificar la proteína de membrana asociada a lisosomas tipo 1 (Lamp1) mediante la técnica de inmunohistoquímica. Resultados: Un marcaje citoplasmático de Lamp1 fue identificado en las células ameloblásticas durante todas las etapas del proceso de diferenciación de los ameloblastos, independientemente de sus edades. Lamp1 también fue detectado en las células epiteliales dentales. Conclusiones: Se evidencia la expresión de Lamp1 en los ameloblastos en incisivos de crecimiento continuo y células epiteliales dentales de rata. Próximos estudios deberían abordar el rol funcional de Lamp1 en la formación del esmalte.

Introduction: Tooth enamel formation includes morphological changes in ameloblasts. These modifications involve the active participation of cellular organelles, including lysosomes. Objective: To identify the lysosomal membrane protein distribution in ameloblasts. Materials and method: The lysosomal membrane protein type 1 (Lamp 1) was identified in continuous growth incisor from mice (7, 14 days old) and epithelial rat's cells using the immunohistochemistry technic. Results: A cytoplasmic Lamp 1 marker was identified in ameloblast cells during all the stages of the process, no matter the age. Lamp 1 was detected, too, in the epithelial dental cells. Conclusion: It is evident the expression of Lamp 1 in ameloblasts of continuous growth incisor and epithelial dental rat's cells. Next studies must be done about the functional role of Lamp 1 in tooth enamel formation.

Lack of FAM20A, Ectopic Gingival Mineralization and Chondro/Osteogenic Modifications in Enamel Renal Syndrome.

Enamel renal syndrome (ERS) is a rare recessive disorder caused by loss-of-function mutations in FAM20A (family with sequence similarity 20 member A, OMIM #611062). Enamel renal syndrome is characterized by amelogenesis imperfecta, delayed or failed tooth eruption, intrapulpal calcifications, gingival overgrowth and nephrocalcinosis. Although gingival overgrowth has consistently been associated with heterotopic calcifications the pathogenesis, structure and interactions of the mineral deposits with the surrounding connective tissue are largely unknown. We here report a novel FAM20A mutation in exon 1 (c.358C > T) introducing a premature stop codon (p.Gln120*) and resulting in a complete loss of FAM20A. In addition to the typical oral findings and nephrocalcinosis, ectopic calcified nodules were also seen in the cervical and thoracic vertebrae regions. Histopathologic analysis of the gingiva showed an enlarged papillary layer associated with aberrant angiogenesis and a lamina propria displaying significant changes in its extracellular matrix composition, including disruption of the collagen I fiber network. Ectopic calcifications were found throughout the connective gingival tissue. Immunomorphological and ultrastructural analyses indicated that the calcification process was associated with epithelial degeneration and transformation of the gingival fibroblasts to chondro/osteoblastic-like cells. Mutant gingival fibroblasts cultures were prone to calcify and abnormally expressed osteoblastic markers such as RUNX2 or PERIOSTIN. Our findings expand the previously reported phenotypes and highlight some aspects of ERS pathogenesis.