Diferencia en la expresión del ligando de receptor activador para el factor nuclear K B, en tejido tumoral frente a la infección por diferentes patógenos periodontales / Difference in receptor activator of nuclear factor K B expression in tumoral tissue by different periodontal pathogens

La enfermedad periodontal es una de las principales causas de pérdida dentaria. Clínicamente, esta patología, mediada por la desregulación del sistema inmune producto de una disbiosis ocurrida en el surco gingival, inicia con la inflamación de la encía y evoluciona con el daño irreversible de los tejidos que rodean el diente. El hueso alveolar es uno de los tejidos afectados esta patología, esto debido a la activación de osteoclastos por la sobreexpresión de la proteína RANKL en el huésped. El propósito de este trabajo es determinar el nivel de sobreexpresión de RANKL, en un modelo de células tumorales U2OS, frente a la infección con Porphyromonas gingivalis y Prevotella intermedia. Para identificar el nivel de RANKL, se definieron cuatro grupos: Un grupo control, no tratado; Grupo PG, tratado con P. gingivalis; Grupo PI, tratado con P. Intermedia; y un grupo PG+PI, tratado con ambas bacterias. El nivel relativo de la proteína RANKL fue determinado en el sobrenadante y en los extractos celulares de manera independiente, mediante la técnica Western blot. En sobrenadantes, el grupo PG mostró mayores niveles de RANKL comparados con PI (p < 0,05). En extractos celulares los niveles fueron mayores en el grupo PG+PI (p < 0,05). El grupo PI mostró los niveles más bajos de RANKL. La infección polimicrobiana resulta en una mayor expresión de RANKL en células tumorales U2OS, mientras que frente a la infección P. gingivalis, se observó mayor cantidad de RANKL soluble.

SUMMARY: Periodontal disease is one of the main causes of tooth loss. Clinically, this pathology, mediated by the deregulation of the immune system due to a dysbiosis occurred in the gingival sulcus, begins with the inflammation of the gum and evolves with the irreversible damage of the tissues that surround the tooth. Alveolar bone is one of the most affected tissues by this disease, due to the activation of osteoclasts by the upregulation of RANKL in the host. The aim of this study is to determine the increase of RANKL, in a U2OS tumor cells model, inoculated with Porphyromonas gingivalis and Prevotella intermedia. To identify the level of RANKL, four groups were defined: A control group, not treated; PG group, treated with P.gingivalis; PI group, treated with P. intermedia; and a PG+PI group, treated with both bacteria. The relative level of RANKL was determined in the supernatant and cell extracts independently, using the Western blot technique. In supernatants, the PG group showed higher RANKL levels compared to PI (p < 0.05). In cell extracts the levels were higher in the PG+PI group (p < 0.05.). The PI group showed the lowest levels of RANKL.Polymicrobial infection results in a greater expression of of soluble RANKL was observed.

Study of resveratrol against bone loss by using in-silico and in-vitro methods / Estudo do resveratrol contra a perda óssea usando métodos in silico e in vitro

Abstract By applying the in-silico method, resveratrol was docked on those proteins which are responsible for bone loss. The Molecular docking data between the resveratrol and Receptor activator of nuclear factor-kappa-Β ligand [RANKL] receptors proved that resveratrol binds tightly to the receptors, showed the highest binding affinities of −6.9, −7.6, −7.1, −6.9, −6.7, and −7.1 kcal/mol. According to in-vitro data, Resveratrol reduced the osteoclasts after treating Marrow-Derived Macrophages [BMM] with Macrophage colony-stimulating factor [MCSF] 20ng / ml and RANKL 50ng / ml, with different concentrations of resveratrol (2.5, 10 μg / ml) For 7 days, the cells were treated with MCSF (20 ng / ml) and RANKL (40 ng / ml) together with concentrated trimethyl ether and resveratrol (2.5, 10 μg / ml) within 12 hours. Which, not affect cell survival. After fixing osteoclast cells with formaldehyde fixative on glass coverslip followed by incubation with 0.1% Triton X-100 in PBS for 5 min and after that stain with rhodamine phalloidin staining for actin and Hoechst for nuclei. Fluorescence microscopy was performed to see the distribution of filaments actin [F.actin]. Finally, resveratrol reduced the actin ring formation. Resveratrol is the best bioactive compound for drug preparation against bone loss.

Resumo Com a aplicação do método in-silico, o resveratrol foi ancorado nas proteínas responsáveis ​​pela perda óssea. Os dados de docking molecular entre o resveratrol e o ligante do receptor ativador do fator nuclear kappa-Β [Receptor Activator of Nuclear Factor kappa-B Ligant (RANKL)] provaram que o resveratrol se liga fortemente aos receptores, mostraram as afinidades de ligação mais altas de −6,9, −7,6, −7,1, −6,9, - 6,7 e -7,1 kcal / mol. De acordo com dados in-vitro, o resveratrol reduziu os osteoclastos após o tratamento de macrófagos derivados da medula óssea [Bone Marrow-derived Macrophage (BMM)] com fator estimulador de colônias de macrófagos [Macrophage Colony-Stimulating Factor (MCSF)] 20ng / ml e RANKL 50ng / ml, com diferentes concentrações de resveratrol (2,5, 10 μg / ml). Durante sete dias, as células foram tratadas com MCSF (20 ng / ml) e RANKL (40 ng / ml) juntamente com éter trimetílico concentrado e resveratrol (2,5, 10 μg / ml) em 12 horas, processo que não afeta a sobrevivência celular. Após a fixação de células de osteoclastos com fixador de formaldeído em lamela de vidro seguido de incubação com 0,1% Triton X-100 em PBS por 5 min, foi realizado posteriormente o procedimento para corar com rodamina faloidina a actina e Hoechst os núcleos. A microscopia de fluorescência foi realizada para ver a distribuição dos filamentos de actina [F.actina]. Finalmente, o resveratrol reduziu a formação do anel de actina. O resveratrol é o melhor composto bioativo para o preparo de medicamentos contra a perda óssea.

RANKL+ senescent cells under mechanical stress: a therapeutic target for orthodontic root resorption using senolytics / 国际口腔科学杂志·英文版

In dentistry, orthodontic root resorption is a long-lasting issue with no effective treatment strategy, and its mechanisms, especially those related to senescent cells, remain largely unknown. Here, we used an orthodontic intrusion tooth movement model with an L-loop in rats to demonstrate that mechanical stress-induced senescent cells aggravate apical root resorption, which was prevented by administering senolytics (a dasatinib and quercetin cocktail). Our results indicated that cementoblasts and periodontal ligament cells underwent cellular senescence (p21+ or p16+) and strongly expressed receptor activator of nuclear factor-kappa B (RANKL) from day three, subsequently inducing tartrate-resistant acid phosphatase (TRAP)-positive odontoclasts and provoking apical root resorption. More p21+ senescent cells expressed RANKL than p16+ senescent cells. We observed only minor changes in the number of RANKL+ non-senescent cells, whereas RANKL+ senescent cells markedly increased from day seven. Intriguingly, we also found cathepsin K+p21+p16+ cells in the root resorption fossa, suggesting senescent odontoclasts. Oral administration of dasatinib and quercetin markedly reduced these senescent cells and TRAP+ cells, eventually alleviating root resorption. Altogether, these results unveil those aberrant stimuli in orthodontic intrusive tooth movement induced RANKL+ early senescent cells, which have a pivotal role in odontoclastogenesis and subsequent root resorption. These findings offer a new therapeutic target to prevent root resorption during orthodontic tooth movement.

Regulatory function and mechanism of autophagy on osteoclast / 中国骨伤

Osteoclast (OC) is multinucleated, bone-resorbing cells originated from monocyte/macrophage lineage of cells, excessive production and abnormal activation of which could lead to many bone metabolic diseases, such as osteoporosis, osteoarthritis, etc. Autophagy, as a highly conserved catabolic process in eukaryotic cells, which plays an important role in maintaining cell homeostasis, stress damage repair, proliferation and differentiation. Recent studies have found that autophagy was also involved in the regulation of osteoclast generation and bone resorption. On the one hand, autophagy could be induced and activated by various factors in osteocalsts, such as nutrient deficiency, hypoxia, receptor activator of nuclear factor(NF)-κB ligand(RANKL), inflammatory factors, wear particles, microgravity environment, etc, different inducible factors, such as RANKL, inflammatory factors, wear particles, could interact with each other and work together. On the other hand, activated autophagy is involved in regulating various stages of osteoclast differentiation and maturation, autophagy could promote proliferation of osteoclasts, inhibiting apoptosis, and promoting differentiation, migration and bone resorption of osteoclast. The classical autophagy signaling pathway mediated by mammalian target of rapamycin complex 1(mTORC1) is currently a focus of research, and it could be regulated by upstream signalings such as phosphatidylinositol 3 kinase(PI-3K)/protein kinase B (PKB), AMP-activated protein kinase(AMPK). However, the paper found that mTORC1-mediated autophagy may play a bidirectional role in regulating differentiation and function of osteoclasts, and its underlying mechanism needs to be further ciarified. Integrin αvβ3 and Rab protein families are important targets for autophagy to play a role in osteoclast migration and bone resorption, respectively. In view of important role of osteoclast in the occurrence of various bone diseases, it is of great significance to elucidate the role of autophagy on osteoclast and its mechanism for the treatment of various bone diseases. The autophagy pathway could be used as a new therapeutic target for the treatment of clinical bone diseases such as osteoporosis.

Resveratrol Modulates Bone Mineral Density and Bone Mineral Content in A Rat Model of Male Hypogonadism / 中国结合医学杂志

OBJECTIVE@#To determine whether resveratrol (Res) can correct osteoporosis induced in a rat model of male hypogonadism.@*METHODS@#Thirty-two rats were randomly divided into 4 groups, 8 in each group; 1) a control sham group: underwent a similar surgical procedure for induction of orchiectomy (ORCD) without ligation of any arteries or veins or removal of the testis and epididymis; 2) a control + Res-treated group (Con+Res): underwent sham surgery similar to the control, but was then treated with Res, as described below; 3) an ORCD-induced group: bilateral ORCD surgery as described above, and 4) a ORCD+Res-treated group: bilateral ORCD surgery followed by Res treatment. Res treatment began 4 weeks after ORCD and continued for 12 weeks. After 12 weeks, bone mineral density (BMD) and bone mineral content (BMC) were measured in the tibia and femur of each rat's right hind leg. Blood levels of bone turnover indicators such as deoxypyridinoline (Dpd), N-telopeptide of type I collagen (NTX I), alkaline phosphatase (ALP), and osteocalcin (OC), as well as receptor activator of nuclear factor kappa B (RANK) and osteoprotegerin (OPG) were assessed.@*RESULTS@#ORCD significantly decreased BMD (P<0.01) and significantly increased bone resorption, manifested by increased RANK. In addition, it inhibited serum levels of OPG and OC. Res treatment after ORCD effectively increased serum levels of bone formation markers such as OPG and OC, compared with testisectomized rats (P<0.05).@*CONCLUSION@#Res could ameliorate bone loss induced by male hypogonadism, possible via restoration of the normal balance between RANK and OPG.

Revisión sistemática de los efectos de microvibración de baja magnitud alta frecuencia en osteocitos cultivados / Systematic review of the effects of low intensity high frequency microvibration on osteocytes cultivated

Introducción: El hueso, reservorio de minerales y moléculas orgánicas, es un tejido dinámico que detecta y se adapta a las cargas mecánicas de los órganos y tejidos del cuerpo, el cual mantiene la estructura ósea del esqueleto durante el crecimiento y a través de la vida del ser humano. Las células óseas son sensibles a las cargas mecánicas y microvibra- ciones que recibe el esqueleto. Objetivo: El propósito de este estudio fue realizar una revisión sistemática acerca de los efectos que ejerce la microvibración de alta frecuencia-baja intensidad, en osteocitos cultivados in vitro sobre la síntesis de factores solubles, con el propósito de entender si la microvibración tiene influencia en la aceleración del movimiento dentario. Material y métodos: Se realizó una búsqueda de artículos de revisión de osteocitos y otras células óseas in vitro, a través de la estrategia PICO (Paciente, Intervención, Comparación, Resultado [Outcome]), con el empleo de palabras clave como: «os- teocitos¼, «microvibración¼, «remodelación¼, «osteoclastogénesis¼, «citocinas¼ y «osteoblastos¼. Se estructuró por medio de PRISMA (informe de revisiones sistemáticas y meta-análisis). La captación de datos finales se hizo por medio del método de puntuación de calidad Jadad y Cochrane (modelo de correlación) como herramientas para evaluar el riesgo de sesgo de cada uno de los artículos. Se incluyeron 11 artículos con alta calidad metodológica. Resultados: La mayoría de los experimentos in vitro demostraron que la microvibración tuvo un aumento estadísticamente significativo en la proliferación y dife- renciación de las células madre mesenquimales (MSC), en osteoblastos (MC3T3-E1), en la expresión de proteínas para inducir osteogénesis y en los osteocitos (MLO-Y4). Asimismo, sobrerregularon la expresión de osteoprotegerina (OPG), prostaglandina (PGE2) y óxido nitroso (NO) al alterar y regular los factores solubles como las citocinas, factores de crecimiento y quimiocinas, de las demás células, además de mostrar una disminución en la actividad de los osteoclastos (RAW246.7) en la resorción ósea. Conclusión: La microvibración induce remodelación ósea. Los osteocitos son sensibles a los estímulos mecánicos y producen factores solubles para inducir la remodelación ósea, razón por la cual se emplea la microvibración como una terapia innovadora y prometedora, no invasiva y no farmacológica en la estimulación de la formación ósea de la superficie del hueso (AU)

Osteonecrosis de los maxilares asociada a drogas antirresortivas en pacientes osteoporóticos: presentación de un caso clínico / Antiresorptive drugs-related jaw osteonecrosis in osteoporotic patients: report of a clinical case

La osteoporosis se caracteriza por una masa ósea baja con deterioro de la microarquitectura del tejido que conduce a la fragilidad, lo que aumenta el riesgo de fracturas. Después de la menopausia, la deficiencia de estrógenos aumenta la exposición del tejido al ligan- do RANK, lo que resulta en un aumento de la reabsorción y pérdida ósea, que pueden provocar osteoporosis. (1) Los bifosfonatos y el denosumab son utilizados para el tratamiento de la osteoporosis debido a su capacidad anticatabólica, que reducen la remodelación previniendo la pérdida de masa ósea, disminuyendo la probabilidad de fracturas y aumentando la densidad mineral del tejido. (2) La osteonecrosis de los maxilares asociadas a drogas antirresortivas es una situación que se presenta en pacientes que consumen de manera crónica antirresortivos para el tratamiento de enfermedades como: osteoporosis, osteogénesis imperfecta, enfermedad de Paget, displasia fi- brosa, hipercalcemia maligna asociada a tratamiento oncológico (AU)

Osteoporosis is characterized by low bone mass with deterioration of the tissue microarchitec- ture leading to fragility, which increases the risk of fractures. After menopause, estrogen deficiency increases tissue exposure to the RANK ligand, resulting in increased bone loss and resorption, which can lead to osteoporosis. (1) Bisphosphonates and denosumab are used for the treatment in low concentration, due to their anticatabolic capacity, which reduce remodeling, preventing loss of bone mass and fractures besides, antiresorptives drugs increase the mineral density of the tissue. (2) Osteonecrosis of the jaw associated with antiresorptives drugs occurs in patients whose chro- nically consume these drugs for the treatment of diseases such as: osteoporosis, imperfect osteogenesis, Paget's disease, fibrous dysplasia, malignant hypercalcemia associated with oncological treatment (AU)

Aberrant NF-κB activation in odontoblasts orchestrates inflammatory matrix degradation and mineral resorption / 国际口腔科学杂志·英文版

Inflammation-associated proteinase functions are key determinants of inflammatory stromal tissues deconstruction. As a specialized inflammatory pathological process, dental internal resorption (IR) includes both soft and hard tissues deconstruction within the dentin-pulp complex, which has been one of the main reasons for inflammatory tooth loss. Mechanisms of inflammatory matrix degradation and tissue resorption in IR are largely unclear. In this study, we used a combination of Cre-loxP reporter, flow cytometry, cell transplantation, and enzyme activities assay to mechanistically investigate the role of regenerative cells, odontoblasts (ODs), in inflammatory mineral resorption and matrices degradation. We report that inflamed ODs have strong capabilities of matrix degradation and tissue resorption. Traditionally, ODs are regarded as hard-tissue regenerative cells; however, our data unexpectedly present ODs as a crucial population that participates in IR-associated tissue deconstruction. Specifically, we uncovered that nuclear factor-kappa b (NF-κB) signaling orchestrated Tumor necrosis factor α (TNF-α)-induced matrix metalloproteinases (Mmps) and Cathepsin K (Ctsk) functions in ODs to enhance matrix degradation and tissue resorption. Furthermore, TNF-α increases Rankl/Opg ratio in ODs via NF-κB signaling by impairing Opg expression but increasing Rankl level, which utterly makes ODs cell line 17IIA11 (A11) become Trap+ and Ctsk+ multinucleated cells to perform resorptive actions. Blocking of NF-κB signaling significantly rescues matrix degradation and resorptive functions of inflamed ODs via repressing vital inflammatory proteinases Mmps and Ctsk. Utterly, via utilizing NF-κB specific small molecule inhibitors we satisfactorily attenuated inflammatory ODs-associated human dental IR in vivo. Our data reveal the underlying mechanisms of inflammatory matrix degradation and resorption via proteinase activities in IR-related pathological conditions.

Caracterização de polimorfismos nos genes OPG, RANK e RANKL em uma população brasileira / Characterization of OPG, RANK and RANKL gene polymorphisms in a Brazilian population

Introdução: o sistema RANKL (receptor-ativador do fator nuclear-ligante κB)/RANK (receptor ativador do NF-kB)/OPG (osteoprotegrina) Introdução: o sistema OPG (osteoprotegrina)/RANK (receptor ativador do NF-kB)/RANKL (receptor-ativador do fator nuclear-ligante κB) regula os processos fisiológicos e patológicos da remodelação óssea. Polimorfismos genéticos nos genes OPG, RANK e RANKL têm sido associados a doenças, em diferentes populações. Objetivo: Descrever a frequência e o potencial regulatório dos polimorfismos do sistema OPG, RANK e RANKL em uma população brasileira; avaliar o seu potencial como marcadores genéticos informativos de ancestralidade; comparar com patologias associadas em outras populações. Metodologia: neste estudo, 506 indivíduos adultos, participantes de uma coorte acometidos de asma e periodontite, tiveram o DNA genômico extraído e genotipado, utilizando-se a plataforma Illumina. As plataformas NCBI, RegulomeDB, Haploview 4.2 e rSNPBase foram consultadas e utilizadas para análises. Resultados e Discussão: os polimorfismos mais frequentes na população estudada foram o rs3102724 no gene OPG, com frequência de menor alelo (MAF) de 46%; o rs4941129 em RANK, MAF 50%; e o rs9525641 em RANKL, MAF 46%. Os rs3134063 (1f) em OPG, rs17069898 (1f) em RANK e rs2200287 (1d) em RANKL apresentaram maior impacto funcional. Em OPG e RANK, nove polimorfismos se caracterizaram como marcadores genéticos informativos de ancestralidade, com predomínio nas populações YRI (africanos) e CEU (europeus). Os nove polimorfismos, com função intrônica, apresentaram MAF entre 2 a 46% na população-alvo e foram associados a patologias do metabolismo ósseo em outras populações. Conclusão: polimorfismos dos genes estudados se mostraram frequentes na população estudada e tiveram seus alelos mais frequentes associados a doenças em populações ancestrais. Sugere-se que sejam realizados mais estudos.

Introduction: The OPG (osteoprotegerin)/ RANK (NF-kB activating receptor)/ RANKL (nuclear-binding factor κB receptor-activating system regulates the physiological and pathological processes of bone remodeling. Genetic polymorphisms (SNPs) in OPG, RANK and RANKL genes have been associated with diseases in different populations. Objective: Describe the regulatory frequency and potential of SNPs in OPG, RANK and RANKL in a Brazilian population; assess their potential as informative genetic markers of ancestry; compare with pathologies associated with these polymorphisms in other populations. Methods: in this study, 506 adult individuals, participating in a cohort involving asthma and periodontitis, had genomic DNA extracted and genotyped using the Illumina platform. The NCBI, RegulomeDB, Haploview 4.2 and rSNPBase platforms were consulted and used for analysis. Results and discussion: the most frequent polymorphisms in the studied population were the rs3102724 in the OPG gene, with the lowest allele frequency (MAF) of 46%; rs4941129 in RANK, MAF 50% and rs9525641 in RANKL, MAF 46%. The rs3134063 (1f) in OPG, rs17069898 (1f) in RANK and rs2200287 (1d) in RANKL, had greater functional impact. In OPG and RANK, 9 SNPs were characterized as informative genetic markers of ancestry, predominantly in YRI (African) and CEU (European) populations. These 9 SNPs, with intronic function, presented MAF between 2 and 46% in our population, and were associated with pathologies in bone metabolism in other populations. Conclusion: SNPs of the studied genes were found to be frequent in the studied population and had their most frequent alleles associated with diseases in ancestral populations. It is suggested that further studies be carried out

Salivary molecules of bone remodeling and tissue repair after head and neck radiotherapy

Abstract Head and neck radiotherapy causes quantitative and qualitative changes in saliva. The objective of this case-control study was to evaluate the salivary biomarkers associated with bone remodeling and tissue repair in patients submitted to radiotherapy for head and neck cancer treatment, compared with non-irradiated individuals. Total unstimulated saliva was collected for ELISA assay analysis of receptor activator for nuclear factor κ B (RANK) and its ligand (RANK-L), osteoprotegerin, matrix metalloproteinase-9/ tissue inhibitor of metalloproteinase-2, vascular endothelial growth factor, and epidermal growth factor. Statistics were performed, and revealed that salivary RANK (p = 0.0304), RANK-L (p = 0.0005), matrix metalloproteinase-9/ tissue inhibitor of metalloproteinase-2 (p = 0.0067), vascular endothelial growth factor (p = 0.0060), and epidermal growth factor (p < 0.0001) were reduced in patients, compared with the control group. Osteoprotegerin did not differ between the groups (p = 0.3765). Salivary biomarkers did not differ according to radiotherapy completion time (p > 0.05). In conclusion, the lower output of the salivary molecules - essential for bone remodeling and tissue repair - may disrupt tissue homeostasis and play a role in the pathogenesis of the radiotherapy-induced deleterious effects in the oral cavity.

Advances in molecular mechanisms of bone invasion by oral cancer / 华西口腔医学杂志

Bone invasion by oral cancer is a common clinical problem, which affects the choice of treatment and predicts a poor prognosis. Unfortunately, the molecular mechanism of this phenomenon has not been fully elucidated. Current studies have revealed that oral cancer cells modulate the formation and function of osteoclasts through the expression of a series of signal molecules. Many signal pathways are involved in this process, of which receptor activator of nuclear factor-κB ligand/receptor activator of nuclear factor-κB/osteoprotegerin signaling pathway attracted much attention. In this review, we introduce recent progress in molecular mechanisms of bone invasion by oral cancer.

TRAF6/ERK/p38 pathway is involved in interleukin-17-mediated autophagy to promote osteoclast precursor cell differentiation / 浙江大学学报·医学版

To investigate the effects of interleukin (IL)-17-mediated autophagy on the TNF receptor associated factor (TRAF6)/extracellular signal-regulated kinase (ERK)/p38 pathway and osteoclast differentiation. Mouse bone marrow-derived macrophages (BMM) were cultured with a medium containing 30 ng/mL macrophage colony stimulating factor and 50 ng/mL receptor activator of nuclear factor-kappa B ligard (RANKL), and IL-17 (0.01, 0.1, 1.0, 10 ng/mL) was added for intervention (IL-17 group). Tartrate-resistant acid phosphatase (TRAP) staining was used to observe TRAP positive multinucleated cells; phalloidin fluorescent staining was used to detect actin ring circumference; toluidine blue staining was used to analyze bone resorption lacuna formation. To further examine the mechanism of the effect of IL-17-mediated autophagy on the differentiation of osteoclasts, the control group used RANKL medium to culture mouse macrophage RAW264.7 cells, while the IL-17 group was treated with IL-17 (0.01, 0.1, 1.0, /mL). Western blot was used to detect the expression of autophagy-related proteins Beclin-1, microtubule-associated protein 1 light chain 3 (LC3) and osteoclast-related proteins c-fos and nuclear factor of activated T cell 1 (NFATc1) after treatment with different concentrations of IL-17. The expression of LC3, NFATc1, TRAF6/ERK/p38 signaling pathway related proteins were detected in IL-17 and autophagy inhibitor 3-MA group. The number of TRAP positive multinucleated cells, the circumference of the actin ring and the area of bone resorption lacuna in IL-17 group treated with IL-17 (0.01, 0.1, were significantly higher than those in the control group. In IL-17 treated RAW264.7 cells, the expression of c-fos, NFATc1, Beclin-1, LC3, TRAF6, p-ERK, and p-p38 was all significantly up-regulated (all 0.05). After treatment with the autophagy inhibitor 3-MA, the expression levels of LC3, NFATc1, TRAF6, p-ERK, and p-p38 all decreased significantly (all 0.05). IL-17 can promote the expression of autophagy proteins and enhance the differentiation ability of osteoclast precursor cells, and the TRAF6/ERK/p38 signaling pathway may be involved in this process.

Actualidad en ortodoncia: las micro-osteoperforaciones en la aceleración del tratamiento ortodóncico / An update in orthodontics: micro-osteoperforations for accelerating orthodontic treatment

La búsqueda por encontrar métodos para acortar la duración de los tratamientos de ortodoncia tiene un pasado reciente, un presente y un futuro. Las fuerzas ortodóncicas que se ejercen sobre la membrana periodontal producen movimientos dentarios por modificaciones histológicas y biomoleculares. El conocimiento de los procesos biológicos da lugar a implementar cambios para favorecer la aceleración de los procesos resortivos y neoformativos. El objetivo de esta publicación es hacer una breve síntesis de lo acontecido con este tema y exponer el procedimiento de las micro-osteoperforaciones (MOPs) como una opción complementaria al tratamiento de ortodoncia convencional. Aún no existe suficiente apoyo de ensayos clínicos en humanos para aseverar su éxito. Más aún, distintos autores publican conclusiones contradictorias. Es de esperar que, en breve, nuevas investigaciones contribuyan a respaldarlo o desestimarlo (AU)

The quest to find methods to shorten the duration of orthodontic treatments has a recent past, a present, and a future. Orthodontic forces exerted on the periodontal membrane produce tooth movements by histological and biomolecular modifications. Knowledge of biological processes results in changes to promote the acceleration of spring and neoformative processes. The objective of this publication is to make a brief synthesis of what happened with this topic and expose the micro-osteoperforations (MOPs) procedure as a complementary option to conventional orthodontic treatment. There is not yet enough support from human clinical trials to assert its success. Moreover, different authors publish conflicting conclusions. It is to be expected that, shortly, further investigations will help to support or dismiss it (AU)

Lymphocyte B and Th17 chemotactic cytokine levels in peri-implant crevicular fluid of patients with healthy, peri-mucositis, and peri-implantitis implants / Niveles de citoquinas quimiotácticas de linfocitos B y Th17 en el fluido crevicular periimplantario de pacientes con implantes sanos, perimucositis y periimplantitis

Peri-implantitis is one of the leading causes of implant failure and loss, and its early diagnosis is not currently feasible due to the low sensitivity of currents methods. In the current exploratory cross-sectional study, we explored the diagnostic potential of lymphocyte B and Th17-chemotactic cytokine levels in peri-implant crevicular fluid (PICF) in 54 patients with healthy, peri-mucositis, or peri-implantitis implants. Peri-implant crevicular fluid was collected, and the levels of the molecules under study were quantified by Luminex assay. The concentrations of CCL-20 MIP-3 alpha, BAFF/BLYS, RANKL and OPG concentration in PICF were analyzed in the context of patient and clinical variables (smoking status, history of periodontitis, periodontal diagnosis, implant survival, suppuration, bleeding on probing, periodontal probing depth, clinical attachment level, mean of implant probing depth, and plaque index). Patients with peri-implantitis, appear to have an overregulation of the RANKL/BAFF-BLyS axis. This phenomenon needs to be investigated in depth in further studies with a larger sample size.

La periimplantitis es una de las principales causas de falla y pérdida del implante, y su diagnóstico temprano no es factible debido a la baja sensibilidad de los métodos actuales. En este estudio transversal exploratorio, se estudió el potencial diagnóstico de los niveles de citocinas quimiotácticas de linfocitos B y Th17 en el líquido crevicular periimplantario (LCPI) en 54 pacientes con implantes sanos, peri-mucositis o periimplantitis. Se recogió líquido crevicular periimplantario y se cuantificaron los niveles de las moléculas estudiadas mediante Luminex assay. Las concentraciones de CCL-20 MIP-3 alfa, BAFF/BLYS, RANKL y la concentración de OPG en LCPI se analizaron en el contexto de las variables clínicas y del paciente (tabaquismo, antecedentes de periodontitis, diagnóstico periodontal, supervivencia del implante, supuración, sangrado al sondaje, profundidad de sondeo periodontal, nivel de inserción clínica, media de la profundidad de sondeo del implante e índice de placa). Los pacientes con periimplantitis parecen tener una sobrerregulación del eje RANKL/BAFF-BLyS. Este fenómeno debe investigarse en profundidad en futuros estudios con un tamaño de muestra mayor.

Association between genetic polymorphisms in rank, rankl and opg and peri-implant diseases in patients from the Amazon Region

Abstract The present study evaluated polymorphisms in RANK, RANKL and OPG-encoding genes to assess whether they are associated with mucositis and peri-implantitis in a population from the Brazilian Amazon region. One hundred and fourteen patients with dental implants were included in the study. After clinical and radiographic examination, the sample was categorized into 4 groups, according to the peri-implant status: Healthy (n=71), Mucositis (n=30), Peri-implantitis (n=13) and Diseased (Mucositis + Peri-implantitis, n=43). Genomic DNA was extracted from buccal cells from saliva, and the genetic polymorphism in osteoprotegerin (OPG), Kappa nuclear factor activator receptor (RANKL) and nuclear kappa factor activator receptor (RANK) were genotyped by the real time PCR. Univariate and multivariate statistical analyses were performed to compare clinical variables among groups and to evaluate genotypes and alleles distributions and the established alpha was 5%. Age, peri-implant biotype, diabetes and presence of peri-implant biofilm were associated with mucositis (p<0.05) and peri-implantitis (p<0.05). Smoking, alcoholism, and periodontal biofilms were also associated with the presence of peri-implantitis (p<0.05). Univariate and multivariate analysis did not demonstrate an association of peri-implantitis or mucositis with any genetic polymorphism in RANK (rs3826620), RANKL (rs9594738) and OPG (rs2073618) (p>0.05). The studied genetic polymorphism in RANK, RANKL and OPG were not associated with mucositis and peri-implantitis in a Brazilian population from the Amazon region.

Resumo O presente estudo avaliou a associação da predisposição clínica e dos fatores genéticos com a presença de doenças peri-implantares. Cento e quatorze pacientes com implantes dentais instalados na Clínica de Especialização do Amazonas, Brazil, foram incluidos no estudo. Após exame clínico e radiográfico, a amostra foi categorizada em 4 grupos, de acordo com o Status peri-implantar: saúde (n=71), mucosite (n=30), peri-implantite (n=13) e doentes (mucosite + peri-implantite). DNA genômico foi extraído de células orais da saliva, e o polimorfismo genético em osteoprotegerina (OPG), ligante do receptor ativador do fator Kappa nuclear (RANKL) e receptor ativador do fator Kappa nuclear (RANK) foram genotipados por PCR em tempo real. O estudo se propôs a avaliar se os polimorfismos em RANK, RANKL e OPG estão envolvidos na patogênese da mucosite e da peri-implantite, e avaliar também a presença de fatores de risco moduladores da resposta em uma população brasileira. Idade, biotipo peri-implantar, diabetes e presença de biofilme peri-implantar foram associados a mucosite (p<0.05) e peri-implantite (p<0.05). Tabagismo, alcoolismo e biofilme periodontal também foram associados com a presença de peri-implantite (p<0.05). Análise univariada e multivariada não demonstraram associação de peri-implantite ou mucosite com os polimorfismos genéticos em RANK (rs3826620), RANKL (rs9594738) e OPG (rs2073618) (p>0.05). Os polimorfismos genéticos estudados não foram associados com mucosite e peri-implantite em uma população brasileira da região Amazônica.

Receptor Activator of Nuclear Factor-Kappa Ligand and Osteoprotegerin Expressions on Hyperglycemic Wistar Rats (Rattus Novergicus) During Orthodontic Tooth Movement

Abstract Objective: To investigate the differences of receptor activator of nuclear factor-κB ligand (RANKL) and Osteoprotegerin (OPG) expressions between normoglycemic and hyperglycemic Wistar rats (Rattus Novergicus) during Orthodontic Tooth Movement (OTM). Material and Methods: This study was true experimental with post-test group only. Thirty-two healthy male Wistar rats, weighted around 200-250 grams, 12-20 weeks old, were used as OTM animal study. They were divided into 2 groups (n=16), normoglycemic rats (normal blood glucose 80-120 mg/dl) and hyperglycemic rats (>250 mg/dl) induced by Streptozotocin with a dose of 30 mg in PBS injection intraperitoneally. A NiTi closed coil spring was mounted between maxillary first molar and incisors with the light force 10gf/mm2 in both groups to induce OTM. The studied animals were then terminated on days 1, 3, 6, and 9, respectively, and premaxilla was extracted. RANKL and OPG expression were examined utilizing immunohistochemistry (IHC) analysis. One-way ANOVA and Tukey HSD (p<0.05) were utilized to analyze the differences in the expression of RANKL and OPG between groups. Results: The hyperglycemic group on day 1, 9 rats showed a significant increase in the expression of RANKL, whereas OPG expression decreased significantly on days 1, 3, and 9. Conclusion: There was a significant increase of RANKL expression and a decrease of OPG expression in hyperglycemic rats as documented immunohistochemically.

Effect of enhanced masticatory force on OPG, RANKL and MGF in alveolar bone of ovariectomized rats

Abstract Menopause induces oral bone loss, leading to various oral diseases. Mastication importantly affects bone metabolism in the jawbone. Objective: To analyze the effect of enhanced masticatory force on osteoprotegerin (OPG), receptor activator of nuclear factor kappa B ligand (RANKL), and mechano-growth factor (MGF) in alveolar bone of ovariectomized rats and to study the mechanics mechanism of the alveolar bone of ovariectomized rats response to enhanced masticatory force. Methodology: Thirty Sprague Dawley rats were randomly divided into three groups: sham-operation group (fat around the removed ovary + normal hard diet), model group (ovariectomy + normal hard diet), and experimental group (ovariectomy + high hard diet). It was a 2-month experiment. Enzyme-linked immunosorbent assay (ELISA) detected serum estradiol (E2), osteocalcin (BGP) and alkaline phosphatase (ALP) in rats. Bone histomorphometric indices in the third molar region of maxilla were detected by micro-CT; protein expressions of OPG, RANKL, and MGF in the third molar region of maxilla was detected by Western blot; and gene expression of OPG, RANKL, and MGF in the third molar region of maxilla was detected by Quantitative Real-Time PCR. Results: Comparing with model group, serum E2 in experimental group increased but not significantly, serum BGP and serum ALP in experimental group decreased but not significantly, OPG in experimental group in alveolar bone increased significantly, RANKL in experimental group in alveolar bone decreased significantly, RANKL/OPG ratio in experimental group decreased significantly, MGF in experimental group in alveolar bone increased significantly, bone volume to total volume fraction increased significantly in experimental group, trabecular thickness increased significantly in experimental group, and trabecular separation decreased significantly in experimental group. Conclusion: Enhanced masticatory force affected the expression of OPG, RANKL, and MGF in alveolar bone of ovariectomized rats, improved the quality of jaw bone of ovariectomized rats, and delayed oral bone loss by ovariectomy.

Impact of history of periodontitis on gene expression of bone-related factors in young patients

Abstract Although dental implants and bone regenerative procedures are important approaches for the reestablishment of esthetics and function in young patients with a history of generalized aggressive periodontitis (GAP), no predictable outcomes have been reported, and the host osteo-immunoinflammatory response may play a relevant role in this context. In view of the lack of molecular investigations into the bone tissue condition of young patients with periodontitis, the aim of this study was to evaluate the gene expression of bone-related factors in this population. Bone biopsies were obtained from the posterior mandible in 16 individuals previously diagnosed with GAP and on periodontal support therapy and from 17 periodontally healthy (PH) patients. The gene expression of tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β, receptor activator of the NF-κB ligand (RANKL), osteoprotegerin (OPG), osteocalcin (OC), bone sialoprotein (BSP), and type I collagen (COL-I), important biomarkers of bone turnover, was evaluated by qRT-PCR. Lower TGF-β and OPG mRNA levels were observed in GAP patients compared to PH individuals (p ≤ 0.05). There were no between-group differences in levels of TNF-α, BSP, RANKL, OC, or COL-I mRNA (p>0.05). In young adults, a history of periodontal disease can negatively modulate the gene expression of important bone-related factors in alveolar bone tissue. These molecular outcomes may contribute to the future development of therapeutic approaches to benefit bone healing in young patients with history of periodontitis via modulation of osteo-immuno-inflammatory biomarkers.

Avaliação dos níveis séricos e salivares de RANKL, OPG, IL-1?, IL-6 e TNF-? e sua relação com o índice da cortical mandibular em pacientes com cirrose hepática / Evaluation of serum and salivary levels of RANKL, OPG, IL-1?, IL-6 and TNF-? and their association with mandibular cortical index in patients with liver cirrhosis

Pacientes com cirrose hepática podem desenvolver osteoporose como complicação relacionada ao déficit funcional hepático. O objetivo desse estudo é o de avaliar a presença de proteínas relacionadas ao metabolismo ósseo (RANKL, OPG, IL-1?, IL-6 e TNF-?) na saliva e no sangue de pacientes cirróticos e compará-los com o índice da cortical mandibular (ICM), relacionado à densidade mineral óssea, em radiografias panorâmicas. Trinta e oito pacientes cirróticos foram submetidos à anamnese, exame físico e tiveram amostras de sangue e de saliva não estimulada coletadas. Desses, 22 apresentavam radiografias panorâmicas que foram avaliadas segundo o ICM. As proteínas foram avaliadas através da tecnologia LuminexTM xMAP. Foram considerados estatisticamente significativos valores de p <=0.05. A maioria dos pacientes era do sexo masculino (68,4%), com idade média de 50,61 anos, média de MELD de 18 pontos no momento da consulta e média de 21,34 meses na fila de espera para o transplante. A análise das radiografias panorâmicas permitiu constatar que a maior parte da amostra apresentava alguma modificação na cortical mandibular sugestiva de alterações osteoporóticas (72,7%). O perfil de expressão das proteínas foi bastante variável no sangue e na saliva. Correlação estatisticamente significativa foi observada entre a expressão salivar de RANKL e OPG em pacientes com ICM alterado. Não foi observada relação estatisticamente significante entre o perfil das proteínas e a etiologia da cirrose, tempo na fila de transplante hepático, idade e sexo dos pacientes. As proteínas RANKL, OPG, IL-1?, IL-6 e TNF-? se comportam de maneira diferente na saliva e no sangue. Valores elevados de RANKL e OPG em saliva estão relacionados com ICM sugestivo de osteoporose em pacientes cirróticos, e propõe-se que esse aumento ocorra devido ao microambiente inflamatório local.

LncRNA Nron regulates osteoclastogenesis during orthodontic bone resorption / 国际口腔科学杂志·英文版

Activation of osteoclasts during orthodontic tooth treatment is a prerequisite for alveolar bone resorption and tooth movement. However, the key regulatory molecules involved in osteoclastogenesis during this process remain unclear. Long noncoding RNAs (lncRNAs) are a newly identified class of functional RNAs that regulate cellular processes, such as gene expression and translation regulation. Recently, lncRNAs have been reported to be involved in osteogenesis and bone formation. However, as the most abundant noncoding RNAs in vivo, the potential regulatory role of lncRNAs in osteoclast formation and bone resorption urgently needs to be clarified. We recently found that the lncRNA Nron (long noncoding RNA repressor of the nuclear factor of activated T cells) is highly expressed in osteoclast precursors. Nron is downregulated during osteoclastogenesis and bone ageing. To further determine whether Nron regulates osteoclast activity during orthodontic treatment, osteoclastic Nron transgenic (Nron cTG) and osteoclastic knockout (Nron CKO) mouse models were generated. When Nron was overexpressed, the orthodontic tooth movement rate was reduced. In addition, the number of osteoclasts decreased, and the activity of osteoclasts was inhibited. Mechanistically, Nron controlled the maturation of osteoclasts by regulating NFATc1 nuclear translocation. In contrast, by deleting Nron specifically in osteoclasts, tooth movement speed increased in Nron CKO mice. These results indicate that lncRNAs could be potential targets to regulate osteoclastogenesis and orthodontic tooth movement speed in the clinic in the future.