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Biol. Res ; 57: 4-4, 2024. ilus, graf
Article in English | LILACS | ID: biblio-1550059


BACKGROUND: Fibrous scars frequently form at the sites of bone nonunion when attempts to repair bone fractures have failed. However, the detailed mechanism by which fibroblasts, which are the main components of fibrous scars, impede osteogenesis remains largely unknown. RESULTS: In this study, we found that fibroblasts compete with osteogenesis in both human bone nonunion tissues and BMP2-induced ectopic osteogenesis in a mouse model. Fibroblasts could inhibit the osteoblastic differentiation of mesenchymal stem cells (MSCs) via direct and indirect cell competition. During this process, fibroblasts modulated the nuclear-cytoplasmic shuttling of YAP in MSCs. Knocking down YAP could inhibit osteoblast differentiation of MSCs, while overexpression of nuclear-localized YAP-5SA could reverse the inhibition of osteoblast differentiation of MSCs caused by fibroblasts. Furthermore, fibroblasts secreted DKK1, which further inhibited the formation of calcium nodules during the late stage of osteogenesis but did not affect the early stage of osteogenesis. Thus, fibroblasts could inhibit osteogenesis by regulating YAP localization in MSCs and secreting DKK1. CONCLUSIONS: Our research revealed that fibroblasts could modulate the nuclear-cytoplasmic shuttling of YAP in MSCs, thereby inhibiting their osteoblast differentiation. Fibroblasts could also secrete DKK1, which inhibited calcium nodule formation at the late stage of osteogenesis.

Humans , Animals , Mice , Osteogenesis/physiology , Mesenchymal Stem Cells , Osteoblasts , Cell Differentiation , Calcium , Cicatrix , Intercellular Signaling Peptides and Proteins , Fibroblasts
Actual. osteol ; 19(1): 18-29, ago. 2023. tab
Article in English | LILACS, UNISALUD, BINACIS | ID: biblio-1511400


MicroRNAs (miRNAs) are small non-coding RNA molecules that play critical roles in post-transcriptional gene regulation. They function by binding to target messenger RNA (mRNA) molecules, leading to their degradation or inhibiting their translation into proteins. In the context of skeletal diseases, such as osteoporosis, osteoarthritis, and bone metastasis, there is growing evidence osteoblastic miRNAs, are involved in the regulation of bone formation and maintenance.Osteoblasts are bone-forming cells responsible for synthesizing and depositing the extracellular matrix, which ultimately mineralizes to form bone tissue. Osteoblastic miRNAs modulate various aspects of osteoblast function, including proliferation, differentiation, mineralization, and apoptosis. Dysregulation of these miRNAs can disrupt the balance between bone formation and resorption, leading to skeletal diseases.The therapeutic implications of targeting osteoblastic miRNAs in skeletal diseases are significant. Modulating the expression levels of specific miRNAs holds promise for developing novel therapeutic strategies to enhance bone formation, prevent bone loss, and promote bone regeneration. Potential therapeutic approaches include the use of synthetic miRNA mimics to restore miRNA expression in diseases associated with miRNA downregulation or the use of anti-miRNA oligonucleotides to inhibit miRNA function in diseases associated with miRNA upregulation.miRNA-based therapies are still in the early stages of development, and further research is needed to fully understand the complexity of miRNA networks. Additionally, the delivery of miRNAs to specific target tissues and cells remains a challenge that needs to be addressed for effective clinical translation. Nonetheless, targeting osteoblastic miRNAs represents a promising avenue for future therapeutic interventions in skeletal diseases. (AU)

Los micro-ARNs (miARNss) son pequeños ARN no codificantes que desempeñan un papel fundamental en la regulación génica postranscripcional. Ejercen su función al unir-se a moléculas de ARN mensajero (ARNm), promoviendo su degradación e inhibiendo su traducción en proteínas. En el contexto de las enfermedades esqueléticas, como la osteoporosis, la osteoartritis y la metástasis ósea existe evidencia de que los miARNs osteoblásticos están involucrados en la regulación de la formación y del mantenimiento óseo. Los osteoblastos son células formadoras de hueso responsables de sintetizar y depositar la matriz extracelular, que finalmente se mineraliza para formar el hueso. Los miARNs derivados de osteoblastos modulan varios aspectos de la función de estas células, incluida la proliferación, diferenciación, mineralización y la apoptosis. La desregulación de estos miARNs puede alterar el equilibrio entre la formación y la resorción ósea, lo que lleva a enfermedades óseas. Las implicaciones terapéuticas de los miARNs osteoblásticos en enfermedades esqueléticas son significativas. La modulación de los niveles de expresión de miARNs específicos es prometedora para desarrollar nuevas estrate-gias terapéuticas a fin de mejorar la formación, prevenir la pérdida y promover la regeneración ósea. Los enfoques terapéuticos potenciales incluyen el uso de miméticos de miARNs para restaurar la expresión de miARNs o el uso de oligonucleótidos anti-miARNs para inhibir su función. Las terapias basadas en miARNs aún se encuentran en las primeras etapas de desarrollo. La administración de miARNs a las células y los tejidos específicos sigue siendo un desafío para lograr una aplicación clínica eficaz. (AU)

Humans , Osteoblasts/cytology , Osteogenesis/genetics , MicroRNAs/genetics , Osteoclasts/cytology , Bone Diseases/prevention & control , Signal Transduction , Gene Expression Regulation , MicroRNAs/biosynthesis , MicroRNAs/physiology , MicroRNAs/therapeutic use
Journal of Southern Medical University ; (12): 39-45, 2023.
Article in Chinese | WPRIM | ID: wpr-971492


OBJECTIVE@#To investigate the effect of teriparatide on the differentiation of MC3T3-E1 cells in high-glucose microenvironment and explore the possible mechanism.@*METHODS@#MC3T3-E1 cells cultured in normal glucose or high-glucose (25 mmol/L) medium were treated with 10 nmol/L teriparatide with or without co-treatment with H-89 (a PKA inhibitor). CCK-8 assay was used to detect the changes in cell proliferation, and cAMP content in the cells was determined with ELISA. Alkaline phosphatase (ALP) activity and mineralized nodules in the cells were detected using ALP kit and Alizarin red staining, respectively. The changes in cell morphology were detected by cytoskeleton staining. Real-time PCR was used to detect the mRNA expressions of PKA, CREB, RUNX2 and Osx in the treated cells.@*RESULTS@#The treatments did not result in significant changes in proliferation of MC3T3-E1 cells (P > 0.05). Compared with the cells in routine culture, the cells treated with teriparatide showed significantly increased cAMP levels (P < 0.05) with enhanced ALP activity and increased area of mineralized nodules (P < 0.05). Teriparatide treatment also resulted in more distinct visualization of the cytoskeleton in the cells and obviously up-regulated the mRNA expressions of PKA, CREB, RUNX2 and Osx (P < 0.05). The opposite changes were observed in cells cultured in high glucose. In cells exposed to high glucose, treatment with teriparatide significantly increased cAMP levels (P < 0.05), ALP activity and the area of mineralized nodules (P < 0.05) and enhanced the clarity of the cytoskeleton and mRNA expressions of PKA, CREB, RUNX2 and Osx; the effects of teriparatide was strongly antagonized by co-treatment with H-89 (P < 0.05).@*CONCLUSION@#Teriparatide can promote osteoblast differentiation of MC3T3-E1 cells in high-glucose microenvironment possibly by activating the cAMP/PKA/CREB signaling pathway.

Animals , Mice , Cell Differentiation , Core Binding Factor Alpha 1 Subunit , Glucose/pharmacology , Osteoblasts/drug effects , RNA, Messenger , Signal Transduction , Teriparatide , Cell Line
China Journal of Orthopaedics and Traumatology ; (12): 393-398, 2023.
Article in Chinese | WPRIM | ID: wpr-981703


Pentaxin 3 (PTX3), as a multifunctional glycoprotein, plays an important role in regulating inflammatory response, promoting tissue repair, inducing ectopic calcification and maintaining bone homeostasis. The effect of PTX3 on bone mineral density (BMD) may be affected by many factors. In PTX3 knockout mice and osteoporosis (OP) patients, the deletion of PTX3 will lead to decrease of BMD. In Korean community "Dong-gu study", it was found that plasma PTX3 was negatively correlated with BMD of femoral neck in male elderly patients. In terms of bone related cells, PTX3 plays an important role in maintaining the phenotype and function of osteoblasts (OB) in OP state;for osteoclast (OC), PTX3 in inflammatory state could stimulate nuclear factor κ receptor activator of nuclear factor-κB ligand (RANKL) production and its combination with TNF-stimulated gene 6(TSG-6) could improve activity of osteoclasts and promote bone resorption;for mesenchymal stem cells (MSCs), PTX3 could promote osteogenic differentiation of MSCs through PI3K/Akt signaling pathway. In recent years, the role of PTX3 as a new bone metabolism regulator in OP and fracture healing has been gradually concerned by scholars. In OP patients, PTX3 regulates bone mass mainly by promoting bone regeneration. In the process of fracture healing, PTX3 promotes fracture healing by coordinating bone regeneration and bone resorption to maintain bone homeostasis. In view of the above biological characteristics, PTX3 is expected to become a new target for the diagnosis and treatment of OP and other age-related bone diseases and fracture healing.

Animals , Male , Mice , Bone Resorption/metabolism , Cell Differentiation , Fracture Healing/genetics , Osteoblasts , Osteoclasts , Osteogenesis , Osteoporosis/genetics , Phosphatidylinositol 3-Kinases/pharmacology
China Journal of Chinese Materia Medica ; (24): 2522-2529, 2023.
Article in Chinese | WPRIM | ID: wpr-981328


This study aimed to investigate the effects of Erxian Decoction(EXD)-containing serum on the proliferation and osteogenic differentiation of MC3T3-E1 cells under oxidative stress through BK channels. The oxidative stress model was induced in MC3T3-E1 cells by H_2O_2, and 3 mmol·L~(-1) tetraethylammonium(TEA) chloride was used to block the BK channels in MC3T3-E1 cells. MC3T3-E1 cells were divided into a control group, a model group, an EXD group, a TEA group, and a TEA+EXD group. After MC3T3-E1 cells were treated with corresponding drugs for 2 days, 700 μmol·L~(-1) H_2O_2 was added for treatment for another 2 hours. CCK-8 assay was used to detect cell proliferation activity. The alkaline phosphatase(ALP) assay kit was used to detect the ALP activity of cells. Western blot and real-time fluorescence-based quantitative PCR(RT-qPCR) were used to detect protein and mRNA expression, respectively. Alizarin red staining was used to detect the mineralization area of osteoblasts. The results showed that compared with the control group, the model group showed significantly blunted cell proliferation activity and ALP activity, reduced expression of BK channel α subunit(BKα), collagen Ⅰ(COL1), bone morphogenetic protein 2(BMP2), osteoprotegerin(OPG), and phosphorylated Akt, decreased mRNA expression levels of Runt-related transcription factor 2(RUNX2), BMP2, and OPG, and declining area of calcium nodules. EXD-containing serum could significantly potentiate the cell proliferation activity and ALP activity, up-regulate the protein expression of BKα, COL1, BMP2, OPG, and phosphorylated Akt, and forkhead box protein O1(FoxO1), promote the mRNA expression of RUNX2, BMP2, and OPG, and enlarge the area of calcium nodules. However, BK channel blockage by TEA reversed the effects of EXD-containing serum in promoting the protein expression of BKα, COL1, BMP2, OPG, and phosphorylated Akt and FoxO1, increasing the mRNA expression of RUNX2, BMP2, and OPG, and enlarging the area of calcium nodules. EXD-containing serum could improve the proliferation activity, osteogenic differentiation, and mineralization ability of MC3T3-E1 cells under oxidative stress, which might be related to the regulation of BK channels and downstream Akt/FoxO1 signaling pathway.

Osteogenesis , Core Binding Factor Alpha 1 Subunit/pharmacology , Large-Conductance Calcium-Activated Potassium Channels/pharmacology , Proto-Oncogene Proteins c-akt/metabolism , Calcium/metabolism , Cell Differentiation , RNA, Messenger/metabolism , Cell Proliferation , Osteoblasts
Acta Physiologica Sinica ; (6): 429-438, 2023.
Article in Chinese | WPRIM | ID: wpr-981018


It has been well documented that exercise can improve bone metabolism, promote bone growth and development, and alleviate bone loss. MicroRNAs (miRNAs) are widely involved in the proliferation and differentiation of bone marrow mesenchymal stem cells, osteoblasts, osteoclasts and other bone tissue cells, and regulation of balance between bone formation and bone resorption by targeting osteogenic factors or bone resorption factors. Thus miRNAs play an important role in the regulation of bone metabolism. Recently, regulation of miRNAs are shown to be one of the ways by which exercise or mechanical stress promotes the positive balance of bone metabolism. Exercise induces changes of miRNAs expression in bone tissue and regulates the expression of related osteogenic factors or bone resorption factors, to further strengthen the osteogenic effect of exercise. This review summarizes relevant studies on the mechanism whereby exercise regulates bone metabolism via miRNAs, providing a theoretical basis for osteoporosis prevention and treatment with exercise.

Humans , MicroRNAs/metabolism , Osteogenesis/genetics , Cell Differentiation , Osteoblasts , Bone Resorption/metabolism
Chinese Medical Journal ; (24): 1468-1477, 2023.
Article in English | WPRIM | ID: wpr-980943


BACKGROUND@#Congenital scoliosis (CS) is a complex spinal malformation of unknown etiology with abnormal bone metabolism. Fibroblast growth factor 23 (FGF23), secreted by osteoblasts and osteocytes, can inhibit bone formation and mineralization. This research aims to investigate the relationship between CS and FGF23.@*METHODS@#We collected peripheral blood from two pairs of identical twins for methylation sequencing of the target region. FGF23 mRNA levels in the peripheral blood of CS patients and age-matched controls were measured. Receiver operator characteristic (ROC) curve analyses were conducted to evaluate the specificity and sensitivity of FGF23. The expression levels of FGF23 and its downstream factors fibroblast growth factor receptor 3 (FGFr3)/tissue non-specific alkaline phosphatase (TNAP)/osteopontin (OPN) in primary osteoblasts from CS patients (CS-Ob) and controls (CT-Ob) were detected. In addition, the osteogenic abilities of FGF23-knockdown or FGF23-overexpressing Ob were examined.@*RESULTS@#DNA methylation of the FGF23 gene in CS patients was decreased compared to that of their identical twins, accompanied by increased mRNA levels. CS patients had increased peripheral blood FGF23 mRNA levels and decreased computed tomography (CT) values compared with controls. The FGF23 mRNA levels were negatively correlated with the CT value of the spine, and ROCs of FGF23 mRNA levels showed high sensitivity and specificity for CS. Additionally, significantly increased levels of FGF23, FGFr3, OPN, impaired osteogenic mineralization and lower TNAP levels were observed in CS-Ob. Moreover, FGF23 overexpression in CT-Ob increased FGFr3 and OPN levels and decreased TNAP levels, while FGF23 knockdown induced downregulation of FGFr3 and OPN but upregulation of TNAP in CS-Ob. Mineralization of CS-Ob was rescued after FGF23 knockdown.@*CONCLUSIONS@#Our results suggested increased peripheral blood FGF23 levels, decreased bone mineral density in CS patients, and a good predictive ability of CS by peripheral blood FGF23 levels. FGF23 may contribute to osteopenia in CS patients through FGFr3/TNAP / OPN pathway.

Humans , Osteopontin/genetics , Alkaline Phosphatase/metabolism , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Scoliosis/genetics , Osteoblasts/metabolism , Calcinosis , RNA, Messenger/metabolism , Bone Diseases, Metabolic/metabolism , Fibroblast Growth Factors/genetics
Natal; s.n; 28/11/2022. 55 p. ilus, tab, graf.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1510707


A engenharia de tecidos ósseos é um ramo importante da medicina regenerativa e envolve o desenvolvimento de arcabouços com composição e arquitetura favoráveis à integração celular, além do estudo de fatores capazes de promover a adesão e proliferação celular, incluindo estímulos químicos e biofísicos. O objetivo do estudo foi avaliar a utilização do laser de baixa intensidade (LBI) como uma ferramenta para promover a bioestimulação in vitro de células osteoblásticas cultivadas em arcabouços nanofibrosos de ácido polilático (PLA). Os arcabouços foram produzidos pela técnica de eletrofiação e caracterizados quanto à molhabilidade, composição pela espectroscopia no infravermelho por transformada de Fourier (FTIR), morfologia da superfície por microscópica eletrônica de varredura (MEV), caracterização termogravimétrica (TGA), calorimetria diferencial exploratória (DSC) e cristalinidade por difração de raios-X (DRX). Os ensaios biológicos foram conduzidos com osteoblastos da linhagem OFCOL II cultivados na superfície dos arcabouços e submetidos ou não (grupo controle) a irradiação com laser diodo InGaAIP na potência de 30 mW, nas doses de 1, 4 e 6 J/cm² e nos comprimentos de onda de 660 nm (grupos V1, V4, V6, respectivo as doses) e 780 nm (grupos I1, I4 e I6, respectivo as doses). Os efeitos do LBI na proliferação dos osteoblastos foram avaliados através do método bioquímico Alamar Blue, nos intervalos de 24, 48 e 72h, enquanto a viabilidade e a morfologia celular foram analisadas no intervalo de 72h, através do ensaio Live/Dead e da microscopia eletrônica de varredura (MEV), respectivamente. Os dados do ensaio bioquímico de Alamar Blue mostraram uma maior proliferação celular nos grupos V6 em todos os intervalos analíticos em comparação ao grupo controle (p<0,05). Outras diferenças entre o grupo controle e irradiados foram encontradas apenas nos intervalos de 48h e 72h para V1, e para o grupo IV6 em 72h. O ensaio Live/Dead revelou um aumento na viabilidade celular nos grupos trados com LBI, sendo significativamente maior no grupo V1 quando comparado ao grupo controle. A análise por MEV mostrou adequada interação dos osteoblastos aos arcabouços, com o corpo celular se espalhando ao longo do eixo da nanofibra e a presença de contatos físicos mais evidentes, através da formação de ligação por meio de filopódios e lamelipódios, nos grupos V1, V6 e I6. Em conjunto, os dados do presente trabalho mostraram que o LBI promove a bioestimulação de osteoblastos cultivados sobre nanofibras de PLA, o que aponta para o seu uso potencial nas técnicas de engenharia tecidual óssea, sobretudo no que se refere ao uso do comprimento de onda de 660 nm, a qual apresentou grupos com mais resultados significativos (AU).

Bone tissue engineering is a relevant branch of regenerative medicine and involves the development of scaffolds with composition and architecture favorable to cell integration, in addition to studying factors capable of promoting cell adhesion and proliferation, including chemical and biophysical stimuli. The study aimed to evaluate the use of low-level laser irradiation (LLLI) to promote in vitro biostimulation of osteoblastic cells cultured on polylactic acid (PLA) nanofibrous scaffolds. The scaffolds were produced by the electrospinning technique and characterized in terms of wettability, composition by Fourier transform infrared spectroscopy (FTIR), surface morphology by scanning electron microscopy (SEM), thermogravimetric characterization (TGA), differential scanning calorimetry (DSC) and crystallinity by Xray diffraction (XRD). The biological assays were conducted with osteoblasts of the OFCOL II lineage cultured on the surface of the scaffolds and submitted or not (control group) to irradiation with InGaAIP diode laser, power of 30 mW, with doses of 1, 4 and 6 J/cm² and wavelengths of 660 nm (groups V1, V4, V6, respectively doses) and 780 nm (groups I1, I4 and I6, respectively doses). The effects of LLLT from the perspective of osteoblasts were evaluated using the biochemical method Alamar Blue assay, at intervals of 24, 48 and 72h, while cell viability and morphology were observed at 72h, using the Live/Dead assay and electron microscopy. scan (SEM), respectively. The Alamar Blue assay data showed more significant cell proliferation in groups in the V6 groups at all analytical intervals compared to the control group (p<0.05). Other differences between the control and irradiated groups were found only at intervals of 48h and 72h for V1, and for group IV6 at 72h. The Live/Dead assay revealed an increase in cell viability in the groups treated with LLLT, being significantly higher in the V1 group when compared to the control group. SEM analysis showed adequate interaction between osteoblasts and scaffolds, with the cell body spreading along the nanofiber axis and the presence of more evident physical contacts, through the formation of bonds through filopodia and lamellipodia, in groups V1, V6 and I6. Together, the data from the present study observed that LLLT promotes the biostimulation of osteoblasts cultured on PLA nanofibers, which pointed to its potential use in bone tissue engineering techniques, especially with regard to the use of the wavelength of 660 nm, which presented groups with more significant results (AU).

Osteoblasts , Bone Regeneration , Low-Level Light Therapy/instrumentation , Tissue Engineering , In Vitro Techniques/methods , Calorimetry, Differential Scanning/instrumentation , Microscopy, Electron, Scanning/instrumentation , Spectroscopy, Fourier Transform Infrared/instrumentation
Braz. dent. sci ; 25(1): 1-9, 2022. ilus
Article in English | LILACS, BBO | ID: biblio-1353788


Objective: The objective of this study was to evaluate in vitro the influence of the anodized surface of Ti35Nb7Zr alloy on the behavior of osteogenic cells, for future application in biomedical implants. Material and Methods: For the development of this research, samples of commercially pure titanium (TiCp) and samples of Ti35Nb7Zr alloy were anodized, both were characterized by scanning electron microscopy (SEM) and were plated afterwards with human osteoblast-like cells (MG63 line) (2 x 104). Cell adhesion, cytotoxicity test, formation of mineralization nodules and a comet assay were also performed in different periods. The bottom of the plate was used as a control, without a sample. Results: SEM analysis showed that the topography of both samples presented surfaces covered by nanotubes. Cellular morphology exhibited spreading in both samples proposing an intimate cell- material liaison. After 3 days, the Ti35Nb7Zr group exhibited greater cell viability than the TiCp group (p<0.01). Regarding calcium content, there was no statistical difference between the anodized groups, but there was a difference between the experimental groups and the control group (p<0.01). In the comet assay, the percentage of DNA in the comet tail did not exhibit any significant difference (p>0.05) among the groups in the evaluated periods. Conclusion: It was concluded that this process of anodization was efficient to form nanotubes, as well as promote a positive influence on the behavior of osteogenic cells without promoting cell damage. (AU)

Objetivo: O objetivo deste estudo foi avaliar in vitro a influência da superfície anodizada da liga Ti35Nb7Zr no comportamento de células osteogênicas, para futura aplicação em implantes biomédicos. Material e Métodos: Para o desenvolvimento desta pesquisa, amostras de titânio comercialmente puro (TiCp) e amostras da liga Ti35Nb7Zr foram anodizadas, ambas foram caracterizadas por microscopia eletrônica de varredura (MEV) e posteriormente plaqueadas com células semelhantes a osteoblastos humanos (linha MG63) (2 x 104). Foram realizados em diferentes períodos a adesão celular, teste de citotoxicidade, formação de nódulos de mineralização e ensaio do cometa. O fundo da placa foi usado como controle, sem amostra. Resultados: A análise em MEV mostrou que a topografia de ambas as amostras apresentava superfícies cobertas por nanotubos. A morfologia celular exibiu espalhamento em ambas as amostras, propondo uma ligação íntima célula-material. Após 3 dias, o grupo Ti35Nb7Zr exibiu maior viabilidade celular do que o grupo TiCp (p<0.01). Em relação ao teor de cálcio, não houve diferença estatística entre os grupos anodizados, mas houve diferença entre os grupos experimentais e o grupo controle (p<0.01). No ensaio do cometa, a porcentagem de DNA na cauda do cometa não apresentou diferença significativa (p> 0.05) entre os grupos nos períodos avaliados. Conclusão:Concluiu-se que esse processo de anodização foi eficiente para formar nanotubos, além de promover uma influência positiva no comportamento das células osteogênicas sem promover dano celular. (AU)

Osteoblasts , Titanium
Journal of Southern Medical University ; (12): 988-996, 2022.
Article in Chinese | WPRIM | ID: wpr-941031


OBJECTIVE@#To explore whether the effect of low-frequency pulsed electromagnetic fields (PEMFs) in promoting osteoblast mineralization and maturation is related to the primary cilia, polycystin2 (PC2) and sAC/PKA/CREB signaling pathway.@*METHODS@#We detected the expression levels of PC2, sAC, PKA, CREB and their phosphorylated proteins in primary rat calvarial osteoblasts exposed to 50 Hz 0.6 mT PEMFs for 0, 5, 15, 30, 60, 90, and 120 min. We blocked PC2 function with amiloride hydrochloride and detected the changes in the activity of sAC/PKA/CREB signal pathway and the mineralization and maturation of the osteoblasts. These examinations were repeated in the osteoblasts after specific knockdown of PC2 via RNA interference and were the co-localization of PC2, sAC, PKA, CREB and their phosphorylated proteins with the primary cilia were using immunofluorescence staining. The expressions of PC2 and the signaling proteins of sAC/PKA/CREB pathway were detected after inhibition of primary ciliation by RNA interference.@*RESULTS@#The expression levels of PC2, sAC, p-PKA and p- CREB were significantly increased in the osteoblasts after exposure to PEMFs for different time lengths (P < 0.01). Blocking PC2 function or PC2 knockdown in the osteoblasts resulted in failure of sAC/PKA/CREB signaling pathway activation and arrest of osteoblast mineralization and maturation. PC2, sAC, p-PKA and p-CREB were localized to the entire primary cilia or its roots, but PKA and CREB were not detected in the primary cilia. After interference of the primary cilia, PEMFs exposure no longer caused increase of PC2 expression and failed to activate the sAC/PKA/CREB signaling pathway or promote osteoblast mineralization and maturation.@*CONCLUSION@#PC2, located on the surface of the primary cilia of osteoblasts, can perceive and transmit the physical signals from PEMFs and promote the mineralization and maturation of osteoblasts by activating the PC2/ sAC/PKA/CREB signaling pathway.

Animals , Rats , Cell Differentiation , Electromagnetic Fields , Osteoblasts , Osteogenesis/genetics , Signal Transduction
Journal of Southern Medical University ; (12): 108-115, 2022.
Article in Chinese | WPRIM | ID: wpr-936291


OBJECTIVE@#To explore the interaction between reactive oxygen species (ROS) and ferroptosis in methylglyoxalinduced injury of mouse embryonic osteoblasts (MC3T3-E1 cells).@*METHODS@#MC3T3-E1 cells were treated with methylglyoxal to establish a cell model of diabetic osteoporosis. CCK-8 assay was used to detect the viability of MC3T3-E1 cells. Rhodamine 123 staining followed by photofluorography was used to examine mitochondrial membrane potential (MMP). The intracellular ROS level was detected by 2', 7'-dichlorodihydrofluorescein diacetate staining with photofluorograph. Alkaline phosphatase (ALP) activity in the cells was detected using an ALP kit, the number of mineralized nodules was determined with alizarin red S staining, and the level of iron ions was detected using a detection kit. The expression level of glutathione peroxidase 4 (GPX4, a marker protein that inhibits ferroptosis) in the osteoblasts was determined using Western blotting.@*RESULTS@#Treatment of MC3T3-E1 cells with 0.6 mmol/L methylglyoxal for 24 h significantly inhibited the expression level of GPX4 (P < 0.001), increased intracellular iron ion concentration, decreased the cell viability, increased the loss of MMP and intracellular ROS level, decreased both ALP activity and the number of mineralized nodules in the cells (P < 0.001). Co-treatment of MC3T3-E1 cells with 2 mmol/L N-acetylcysteine (NAC, a ROS scavenger) and methylglyoxal significantly increased the expression level of GPX4 (P < 0.01); co-treatment with 4 mmo/L FER-1 (a ferroptosis inhibitor) and methylglyoxal obviously decreased the intracellular ROS level (P < 0.001). Co-treatment of the cells either with NAC and methylglyoxal or with FER-1 and methylglyoxal attenuated methylglyoxal-induced injuries in the osteoblasts (P < 0.001).@*CONCLUSION@#The interaction between ROS and ferroptosis pathway plays an important role in methylglyoxal-induced injury of mouse embryonic osteoblasts.

Animals , Mice , Cell Survival , Ferroptosis , Osteoblasts , Pyruvaldehyde/metabolism , Reactive Oxygen Species/metabolism
Journal of Peking University(Health Sciences) ; (6): 31-39, 2022.
Article in Chinese | WPRIM | ID: wpr-936109


OBJECTIVE@#To investigate the effect of porous surface morphology of zirconia on the proliferation and differentiation of osteoblasts.@*METHODS@#According to different manufacturing and pore-forming methods, the zirconia specimens were divided into 4 groups, including milled sintering group (M-Ctrl), milled porous group (M-Porous), 3D printed sintering group (3D-Ctrl) and 3D printed porous group (3D-Porous). The surface micromorphology, surface roughness, contact angle and surface elements of specimens in each group were detected by scanning electron microscope (SEM), 3D laser microscope, contact angle measuring device and energy-dispersion X-ray analysis, respectively. MC3T3-E1 cells were cultured on 4 groups of zirconia discs. The cell morphology of MC3T3-E1 cells on zirconia discs was eva-luated on 1 and 7 days by SEM. The cell proliferation was detected on 1, 3 and 5 days by cell counting kit-8 (CCK-8). After osteogenic induction for 14 days, the relative mRNA expression of alkaline phosphatase (ALP), type Ⅰ collagen (Colla1), Runt-related transcription factor-2 (Runx2) and osteocalcin (OCN) in MC3T3-E1 cells were detected by real-time quantitative polymerase chain reaction.@*RESULTS@#The pore size [(419.72±6.99) μm] and pore depth [(560.38±8.55) μm] of 3D-Porous group were significantly larger than the pore size [(300.55±155.65) μm] and pore depth [(69.97±31.38) μm] of M-Porous group (P < 0.05). The surface of 3D-Porous group appeared with more regular round pores than that of M-Porous group. The contact angles of all the groups were less than 90°. The contact angles of 3D-Ctrl (73.83°±5.34°) and M-Porous group (72.7°±2.72°) were the largest, with no significant difference between them (P>0.05). Cells adhered inside the pores in M-Porous and 3D-Porous groups, and the proliferation activities of them were significantly higher than those of M-Ctrl and 3D-Ctrl groups after 3 and 5 days' culture (P < 0.05). After 14 days' incubation, ALP, Colla1, Runx2 and OCN mRNA expression in 3D-Porous groups were significantly lower than those of M-Ctrl and 3D-Ctrl groups (P < 0.05). Colla1, Runx2 and OCN mRNA expressions in M-Porous group were higher than those of 3D-Porous group (P < 0.05).@*CONCLUSION@#The porous surface morphology of zirconia can promote the proliferation and adhesion but inhibit the differentiation of MC3T3-E1 cells.

Cell Differentiation , Cell Proliferation , Ceramics , Osteoblasts , Osteogenesis , Porosity , Zirconium
Actual. osteol ; 18(1): 40-52, 2022. ilus, tab
Article in Spanish | LILACS, UNISALUD, BINACIS | ID: biblio-1396075


El "microbioma" no solo está constituido por los microbios, sino por todos los componen-tes que viven en el mismo hábitat conforman-do un nicho ecológico. Es decir, está conformado por los microorganismos (bacterias, hongos, protozoos, etc.), todo el espectro de moléculas producidas por ellos tales como sus componentes estructurales (ácidos nucleicos, proteínas, lípidos y glúcidos), meta-bolitos, toxinas, etc., y las moléculas producidas por el huésped. El microbioma intestinal (MI) ha emergido como un factor que tiene un gran efecto sobre la cantidad, calidad y fuerza del hueso. Las investigaciones revelan que la homeostasis ósea está ligada al micro-bioma saludable, mientras que la disbiosis (alteración en la biodiversidad microbiana) puede exacerbar la actividad osteoclástica y promover la osteoporosis. Los mecanismos potenciales involucrados en la interacción del microbioma intestinal y el hueso son la influencia del metabolismo del huésped, el mantenimiento de la integridad intestinal y regulación de la absorción de nutrientes, la regulación del eje intestino-sistema inmune y la modulación del sistema endocrino. Es decir que hay múltiples vías por las cuales el MI influye sobre el hueso, pero estos y otros mecanismos deben profundizarse más aún. También es necesario que se identifiquen y caractericen mejor los microorganismos que están asociados a las enfermedades óseas. El conocimiento de estos aspectos podría ser útil para el desarrollo de herramientas terapéuticas basadas en el MI que puedan mejorar la eficacia de los distintos tratamientos existentes. (AU)

The microbiome is not only constituted by microbes, but by all the components that live in the same habitat forming an ecological niche. It is conformed by the microorganisms ( bacteria, fungi, protozoa, etc), the entire spectrum of molecules produced by them (nucleic acids, proteins, lipid and carbohydrates, metabolites, toxins, etc) and the molecules produced by the host. The intestinal microbiome (IM) has emerged as a factor with great effects on the quantity, quality and strength of bone. The investigations reveal that bone homeostasis is linked to the healthy microbiome, while the dysbiosis (alteration in the microbial biodiversity) can exacerbate the osteoclastic activity and promote osteoporosis. The potential mechanisms involved in the interaction between IM and bone are the influence of the host metabolism, the maintenance of the intestinal integrity and regulation of the nutrient absorption, the regulation of the intestine/ immune system axis and the modulation of the endocrine system. That is, there are multiple ways through which IM influences on bone, but these and other mechanisms need to be further studied. It is also necessary to identify and characterize the microorganisms associated with the bone diseases. Knowledge of these aspects could be useful to develop therapeutical tools based on the IM that could improve the efficacy of the current treatments. (AU)

Humans , Osteoblasts/immunology , Osteoclasts/immunology , Bone and Bones/immunology , Dysbiosis/complications , Gastrointestinal Microbiome/immunology , Osteoblasts/metabolism , Osteoclasts/metabolism , Bone and Bones/metabolism , Intestines/immunology , Intestines/microbiology
Chinese Journal of Biotechnology ; (12): 1159-1172, 2022.
Article in Chinese | WPRIM | ID: wpr-927771


It is known that low-frequency pulsed electromagnetic fields (PEMFs) can promote the differentiation and maturation of rat calvarial osteoblasts (ROBs) cultured in vitro. However, the mechanism that how ROBs perceive the physical signals of PEMFs and initiate osteogenic differentiation remains unknown. In this study, we investigated the relationship between the promotion of osteogenic differentiation of ROBs by 0.6 mT 50 Hz PEMFs and the presence of polycystin2 (PC2) located on the primary cilia on the surface of ROBs. First, immunofluorescence staining was used to study whether PC2 is located in the primary cilia of ROBs, and then the changes of PC2 protein expression in ROBs upon treatment with PEMFs for different time were detected by Western blotting. Subsequently, we detected the expression of PC2 protein by Western blotting and the effect of PEMFs on the activity of alkaline phosphatase (ALP), as well as the expression of Runx-2, Bmp-2, Col-1 and Osx proteins and genes related to bone formation after pretreating ROBs with amiloride HCl (AMI), a PC2 blocker. Moreover, we detected the expression of genes related to bone formation after inhibiting the expression of PC2 in ROBs using RNA interference. The results showed that PC2 was localized on the primary cilia of ROBs, and PEMFs treatment increased the expression of PC2 protein. When PC2 was blocked by AMI, PEMFs could no longer increase PC2 protein expression and ALP activity, and the promotion effect of PEMFs on osteogenic related protein and gene expression was also offset. After inhibiting the expression of PC2 using RNA interference, PEMFs can no longer increase the expression of genes related to bone formation. The results showed that PC2, located on the surface of primary cilia of osteoblasts, plays an indispensable role in perceiving and transmitting the physical signals from PEMFs, and the promotion of osteogenic differentiation of ROBs by PEMFs depends on the existence of PC2. This study may help to elucidate the mechanism underlying the promotion of bone formation and osteoporosis treatment in low-frequency PEMFs.

Animals , Rats , Alkaline Phosphatase/metabolism , Electromagnetic Fields , Osteoblasts/metabolism , Osteogenesis/genetics , TRPP Cation Channels/physiology
Electron. j. biotechnol ; 52: 52-58, July. 2021. tab, ilus
Article in English | LILACS | ID: biblio-1283505


BACKGROUND: Osteoporosis attacks approximately 10% of the population worldwide. Sika Deer (Cervus nippon), one of China's precious traditional medicinal animals, has been widely recorded in ancient Chinese medical books and claimed for centuries to have numerous medical benefits including bone strengthening. This study aimed to find the use of Sika Deer bone in treating osteoporosis according to traditional records and to investigate the protective effect of Sika Deer bone polypeptide extract on glucocorticoidinduced osteoporosis (GIOP) in rats. RESULTS: Sika Deer bone polypeptide extract could increase serum Ca2+ and BGP, decrease serum P3+, ALP, PTH, and CT, but had no effect on serum NO in rats with GIOP. The immunohistochemical iNOS results of the rats' distal femur were negative in each group. Besides the model group, the eNOS color reaction in osteoblasts was strongly positive in the other three groups. CONCLUSIONS: Sika Deer bone polypeptide extract can improve pathological changes in the microstructure and stimulate the expression of eNOS in osteoblasts. The protective effect on bone might be mediated by eNOS-dependent NO generation.

Animals , Male , Rats , Osteoporosis/prevention & control , Peptides/pharmacology , Bone and Bones/metabolism , Deer , Osteoblasts , Dexamethasone , Rats, Wistar , Nitric Oxide Synthase Type III/drug effects
J. oral res. (Impresa) ; 10(2): 1-9, abr. 30, 2021. ilus, tab
Article in English | LILACS | ID: biblio-1381731


Objective: To determine the cytotoxicity and effects of graphene oxide (GO) on cellular proliferation of gingival-fibroblasts, pulp-dental cells and human osteoblasts in culture, and to determine the physical, mechanical and biological properties of poly (methyl methacrylate) (PMMA) enriched with GO. Material and Methods: The GO was characterized with SEM. Cytotoxicity and cell proliferation were determined by the MTT bioassay. The physical mechanical tests (flexural strength and elastic modulus) were carried out with a universal testing machine. Sorption and solubility were determined by weighing before and after drying and immersion in water. Porosity was evaluated by visual inspection. Data were analyzed with Student's t-test and Tukey's posthoc ANOVA. Results: The GO has a heterogeneous morphology and a particle size of 66.67±64.76 µm. GO has a slight to no-cytotoxicity (>50-75% viability) at 1-30 days, and at 24 hours incubation of PMMA with GO significantly stimulates osteoblasts (45±8%, p<0.01). The physical and mechanical properties of PMMA with GO increase considerably without altering sorption, solubility and porosity. Conclusion: GO alone or with PMMA has an acceptable biocompatibility, could contribute to cell proliferation, cell regeneration and improve the physical mechanical properties of PMMA.

Objective: To determine the cytotoxicity and effects of graphene oxide (GO) on cellular proliferation of gingival-fibroblasts, pulpdental cells and human osteoblasts in culture, and to determine the physical, mechanical and biological properties of poly (methyl methacrylate) (PMMA) enriched with GO. Material and Methods: T he G O w as c haracterized with SEM. Cytotoxicity and cell proliferation were determined by the MTT bioassay. The physical-mechanical tests (flexural strength and elastic modulus) were carried out with a universal testing machine. Sorption and solubility were determined by weighing before and after drying and immersion in water. Porosity was evaluated by visual inspection. Data were analyzed with Student's t-test and Tukey's post-hoc ANOVA. Results: The GO has a heterogeneous morphology and a particle size of 66.67±64.76 ?m. GO has a slight to no-cytotoxicity (>50-75% viability) at 1-30 days, and at 24 hours incubation of PMMA with GO significantly stimulates osteoblasts (45±8%, p<0.01). The physical and mechanical properties of PMMA with GO increase considerably without altering sorption, solubility and porosity. Conclusion: GO alone or with PMMA has an acceptable biocompatibility, could contribute to cell proliferation, cell regeneration and improve the physical-mechanical properties of PMMA.

Humans , Biocompatible Materials , Polymethyl Methacrylate/chemistry , Graphite/chemistry , Osteoblasts , Oxides , Regeneration , Biological Assay , Cell Proliferation , Flexural Strength
Araçatuba; s.n; 2021. 91 p. ilus, graf.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1413767


A ozonioterapia vem se demonstrando uma ferramenta promissora na prevenção de infecções e no auxílio da reparação tecidual, conciliando com os desafios no tratamento da osteonecrose dos maxilares induzida por medicamentos (ONM-M), este projeto objetiva analisar os efeitos da ozonioterapia, em 55 ratas senis (18 meses), entre 300-350g, induzidas a osteonecrose via medicamentosa (Zoledronato 100µg/kg), após exodontia do primeiro molar inferior. Os animais foram divididos em 4 grupos equitativos (10 ratas por grupo), o primeiro grupo SAL, recebeu aplicações de soro fisiológico por 7 semanas, grupo SAL + OZ recebeu aplicações de soro fisiológico por 7 semanas e o tratamento com a ozonioterapia (0,7mg/kg) a cada 2 dias por 28 dias, o grupo ZOL recebeu aplicações de zoledronato (100µg/kg) por 7 semanas e por último o grupo ZOL + OZ recebeu também aplicações de zoledronato no mesmo protocolo e foi tratado com a ozonioterapia (0,7mg/kg) a cada 2 dias por 28 dias. Todos as ratas receberam a antibioticoterapia (Cristacilina® 0,1ml/kg por dia) iniciando 3 dias antes do procedimento de extração, se estendendo até 4 dias de pós-operatório, passaram pela extração do molar na terceira semana de experimento e foram submetidas a eutanásia na sétima semana de experimento. Após a eutanásia as mandíbulas foram ressecadas, reduzidas e preparadas para as análises microtomográficas (caracterização óssea do osso senil (MCT0) e após terapia com zoledronato (MCT1ZOL) contra seu par controle (MCT1SAL), parâmetros volumétricos (Bv,Bv.Tv,Tb.Th,Tb.N,Tb.Sp,Po.Tot) dos grupos experimentais), histométricas (porcentagem de osso neoformado e porcentagem de osso não vital) e imunoistoquímicas (expressão de TNFa, IL-1b, VEGF, OCN e TRAP). Os resultados da caracterização óssea não apresentaram diferença quando comparado os grupos experimentais (p> 0,05), possivelmente devido ao pouco tempo decorrido na terapia com zoledronato. Os demais resultados comparando os grupos experimentais mostrou com diferenças estatisticamente significativas (p< 0,05) uma característica de osso vítreo, denso, sem vitalidade, pobre em vascularização, com elevados valores para marcadores de inflamação, traduzindo isso em osteonecrose dos maxilares relacionada com a medicação, destoando principalmente do grupo controle SAL, que apresentou melhora na reparação alveolar e características de osso vital e vascularizado. A ozonioterapia (ZOL+OZ, SAL+OZ) apresentou valores significantes estatisticamente quando comparado ao grupo sem tratamento, traduzindo em melhora na vascularização do tecido ósseo, em melhora reparacional do alvéolo, modulação da inflamação local e o aparecimento/manutenção de células osteoblásticas ativas (p< 0,05). Mostrando-se uma terapia viável no controle/tratamento da osteonecrose dos maxilares relacionado com medicamentos(AU)

Ozone therapy has been shown to be a promising tool in the prevention of infections and in the aid of tissue repair, reconciling with the challenges in the treatment of medication-induced jaw osteonecrosis (ONM-M), this project aims to analyze the effects of ozone therapy in 55 rats senile (18 months), between 300-350g, induced to osteonecrosis via medication (Zoledronate 100µg / kg), after extraction of the lower first molar. The animals were divided into 4 equitable groups (ten rats per group), the first SAL group, received saline applications for 7 weeks, SAL + OZ group received saline applications for 7 weeks and ozone therapy (0, 7mg / kg) every 2 days for 28 days, the ZOL group received applications of zoledronate (100µg / kg) for 7 weeks and lastly the ZOL + OZ group also received applications of zoledronate in the same protocol and was treated with ozone therapy (0.7mg / kg) every 2 days for 28 days. All rats received antibiotic therapy (Cristacilina® 0.1ml / kg per day) starting 3 days before the extraction procedure, extending up to 4 days after the operation, underwent molar extraction in the third week of the experiment and were submitted to euthanasia in the seventh week of experiment. After euthanasia, the mandibles were resected, reduced and prepared for microtomographic analysis (bone characterization of senile bone (MCT0) and after therapy with zoledronate (MCT1ZOL) against its control pair (MCT1SAL), volumetric parameters (Bv, Bv.Tv, Tb .Th, Tb.N, Tb.Sp, Po.Tot) of the experimental groups), histometric (percentage of newly formed bone and percentage of non-vital bone) and immunohistochemistry (expression of TNFa, IL-1b, VEGF, OCN and TRAP) . The results of bone characterization did not show any difference when comparing the experimental groups (P> 0.05), possibly due to the short time elapsed in zoledronate therapy. The other results comparing the experimental groups showed with statistically significant differences (P < 0.05) a characteristic of vitreous bone, dense, without vitality, poor in vascularization, with high values for inflammation markers, translating this into a related jaw osteonecrosis with medication, disagreeing mainly with the SAL control group, which showed improvement in alveolar repair and characteristics of a vital and vascularized bone. Ozone therapy (ZOL + OZ, SAL + OZ) showed statistically significant values when compared to the untreated group, translating into an improvement in bone tissue vascularization, a reparational improvement of the alveolus, modulation of local inflammation and the appearance/maintenance of cells active osteoblasts (P < 0.05). Showing to be a viable therapy in the control/treatment of osteonecrosis of the jaws related to drugs(AU)

Animals , Rats , Osteonecrosis/chemically induced , Abnormalities, Drug-Induced , Bisphosphonate-Associated Osteonecrosis of the Jaw , Zoledronic Acid/adverse effects , Zoledronic Acid/poisoning , Zoledronic Acid/toxicity , Ozone Therapy , Mandible/abnormalities , Maxilla/abnormalities , Osteoblasts , Bone and Bones , Rats, Wistar , Jaw
Araçatuba; s.n; 2021. 79 p. ilus, graf, tab.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1435855


O objetivo deste estudo foi sintetizar um substituto ósseo a base de Hidroxiapatita (HAp), modificá-lo superficialmente com hexametafosfato (HMP) e colágeno tipo I (COL) e analisar o comportamento in vitro e in vivo. A síntese de HAp foi realizada pelo método de coprecipitação controlada a partir de H3PO4, CaCl2 e NH4OH. Após processamento foram realizadas as modificações superficiais em soluções de HMP e COL. As partículas de hidroxiapatita e suas modificações foram caracterizadas através das técnicas de potencial-zeta (ζ), tamanho de partícula, espectroscopia de infravermelho com transformada Fourrier (FTIR) e difração de raios X (DRX), as quais evidenciaram alta semelhança química com a HAp biológica. A morfologia foi avaliada através da técnica de microscopia eletrônica de varredura (MEV), a qual mostrou que as nanopartículas de HAp obtidas possuíam aproximadamente 130 nm, pode ser visualizada uma película recobrindo as superfícies modificadas com HMP e COL. Foi realizada cultura de células MC3T3, com análises de MTT, ALP e nódulos de mineralização. Nas análises in vivo, foram realizados defeitos críticos em calvaria de 150 ratos, divididos em 5 grupos (GC:autógeno; G1:HAp; G2:HMP; G3:COL; G4:BioOss) e submetidos a eutanásia após 7,14,30,60 dias. Os espécimes foram avaliados em cortes calcificados MicroCt e confocal, apresentando fechamento do defeito e formação óssea significante em G1,G3 e G4. Portanto conclui-se que G1 e G3 apresentaram comportamento favorável e viável na neoformação óssea comparado ao G4 substituto ósseo comercialmente disponível, tornando-se uma futura alternativa para regeneração óssea(AU)

The aim of this study was to synthesize a bone substitute based on Hydroxyapatite (HAp), superficially modify it with hexametaphosphate (HMP) and collagen type I (COL) and analyze its behavior in vitro and in vivo. The synthesis of HAp was carried out by the controlled co-precipitation method from H3PO4, CaCl2 and NH4OH. After processing, surface modifications were performed in HMP and COL solutions. HAp particles and their modifications were characterized using the techniques of zeta-potential (ζ), particle size, Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD), which showed high chemical similarity with the Biological HAp. The morphology was evaluated using the technique of scanning electron microscopy (SEM), which showed that the HAp nanoparticles obtained had approximately 130 nm, a film covering the modified surfaces with HMP and COL can be visualized. Culture of MC3T3 cells was performed with analysis of MTT, ALP and mineralization nodules. In the in vivo analysis, critical calvarian defects were performed in 150 rats, divided into 5 groups (GC:autogenous bone; G1:HAp; G2:HMP; G3:COL; G4:BioOss) and euthanized after 7,14,30,60 days. The specimens were evaluated in calcified MicroCt and confocal sections, showing defect closure and significant bone formation in G1,G3 and G4. Therefore, it is concluded that G1 and G3 presented favorable and viable behavior in bone neoformation compared to the commercially available G4 bone substitute, becoming a future alternative for bone regeneration(AU)

Animals , Rats , Osteoblasts , Bone Regeneration , Durapatite , Bone Substitutes , Phosphates , Biocompatible Materials , Rats, Wistar , Collagen Type I , Nanoparticles
Araçatuba; s.n; 2021. 52 p. ilus, graf.
Thesis in Portuguese | BBO, LILACS | ID: biblio-1381565


A perda óssea dentária e a formação de lesões periapicais surgem como uma consequênc ia do desequilíbrio da homeostase óssea. Os osteoblastos, juntamente com os osteoclastos e osteócitos, atuam na formação e na reabsorção óssea. Vários marcadores de formação óssea são produzidos por osteoblastos ativos e refletem diferentes aspectos da dif erenciação osteoblástica e da remodelação óssea. Com isso, muitos autores têm explorado o uso de fitoterápicos, visando obter novos compostos que apresentem propriedades terapêuticas, como os flavonoides, e que estimulem a neoformação óssea e o reparo da r egião periapical. O objetivo deste estudo foi avaliar in vitro a citotoxicidade e efeito indutor de mineralização de flavonoides sobre células osteoblásticas humanas. Para isso, células osteoblásticas da linhagem Saos expostas aos seguintes flavono2 foram ides: quercetina, miricetina e seus derivados taxifolina, isoquercitrina, rutina, ampelopsina e EGCG, além de pinocembrina, crisina e canferol, de forma isolada e combinada. Foi avaliado o efeito citotóxico, a atividade de fosfatase alcalina e indução de n mé todo de Shapiroódulos de mineralização. Os resultados foram analisados p elo Wilk, e as variáveis foram submetidas à análise de ANOVA seguida pelo teste de Tukey para comparar entre os grupos e/ou concentrações ou teste de Dunnett para comparar entre cada grupo e o controle, com nível de significância de 5%. A viabilidade da cultura de osteoblastos não teve uma redução estatisticamente significativa na presença da maioria dos compostos, exceto crisina a 100µM. Taxifolina, isoquercitrina, rutina, ampelopsina e EGCG foram os compostos que estimularam significativamente a atividade da fosfatase alcalina, juntamente com as combinações taxifolina+isoquercitrina, taxifolina+ampelopsina e taxifolina+rutina a 25/25 µM. Quanto a formação de nódulos de mine ralização, ampelopsina, isoquercitrina, rutina, pinocembrina e miricetina isolados e taxifolina+isoquercitrina, taxifolina+ampelopsina e taxifolina+rutina combinados obtiveram os melhores resultados, variando de acordo com as concentrações. Concluise que a taxifolina, isoquercitrina, rutina e ampelopsina e combinações de taxifolina com esses flavonoides são citocompatíveis e apresentam efeito indutor de mineralização em osteoblastos Saos-2(AU)

Dental bone loss and the formation of periapical lesions arise as a consequence of imbalance of bone homeostasis. Osteoblasts, together with osteoclasts and osteocytes, act in bone formation and resorption. Several markers of bone formation are produced by active osteoblasts and reflect different aspects of osteoblastic differentiation and bone remodeling. Thus, many authors have explored the use of phytotherapics in order to obtain new compounds with therapeutic properties, such as flavonoids, and also stimulate bone neoformation and periapical region repair. The objective of this study was to evaluate in vitro the cytotoxicity and inducing effect of flavonoid mineralization on human osteoblastic cells. For this, osteoblastic cells of the Saos-2 lineage were exposed to the following flavonoids: quercetin, myricetin and its derivatives taxifoline, isoquercitrin, rutin, ampelopsin and EGCG, in addition to pinocembrin, chrysin and kaempferol, in an isolated and combined manner. The cytotoxic effect, the activity of alkaline phosphatase and the induction of mineralization nodules were evaluated. The results were analyzed using the Shapiro-Wilk method, and the variables were submitted to ANOVA analysis followed by the Tukey test to compare between groups and/or concentrations or Dunnett's test to compare between each group and the control, with a level of 5% significance. The viability of the osteoblast culture did not have a statistically significant reduction in the presence of most compounds, except 100 µM chrysin. Taxifoline, isoquercitrin, rutin, ampelopsin and EGCG were the compounds that significantly stimulated the activity of alkaline phosphatase, together with the combinations taxifoline+isoquercitrin, taxifoline+ampelopsin and taxifoline+rutin at 25/25 µM. As for the formation of mineralization nodules, ampelopsin, isoquercitrin, rutin, pinocembrin and myricetin alone and taxifoline+isoquercitrin, taxifoline+ampelopsin and taxifoline+rutin combined obtained the best results, varying according to the concentrations. It is concluded that taxifoline, isoquercitrin, rutin and ampelopsin and combinations of taxifolin with these flavonoids are cytocompatible and have a mineralization-inducing effect on Saos-2 osteoblasts(AU)

Osteoblasts , Periapical Periodontitis , Flavonoids , Bone Resorption , Osteoclasts , Osteocytes , Quercetin , Rutin , Flavonoids/toxicity , Flavonoids/therapeutic use , Bone and Bones , Calcification, Physiologic , Bone Remodeling , Flavanones , Homeostasis
J. appl. oral sci ; 29: e20200870, 2021. tab, graf
Article in English | LILACS | ID: biblio-1250190


Abstract Bio-C Pulpecto (Bio-CP) was recently developed as the first bioceramic root filling material for primary teeth. Objective To evaluate the physicochemical properties of radiopacity, setting time, pH, cytocompatibility and potential of Bio-CP to induce mineralisation, compared with (1) Calen thickened with zinc oxide (Calen-ZO), and (2) zinc oxide and eugenol (ZOE). Methodology Physicochemical properties were evaluated according to ISO 6876. Saos-2 (human osteoblast-like cell line) exposed to extracts of the materials were subjected to assays of methyl thiazolyl tetrazolium, neutral red, alkaline phosphatase (ALP) activity and mineralised nodule production. The results were analysed using one-way or two-way ANOVA and Tukey's or Bonferroni's post-tests (α=0.05). Results All the materials showed radiopacity higher than 3 mm Al. Bio-CP had lower pH than Calen-ZO, but higher pH than ZOE. Calen-ZO and Bio-CP did not set. The setting time for ZOE was 110 min. The cytocompatibility order was Calen-ZO > Bio-CP > ZOE (1:2, 1:4 dilutions) and Calen-ZO > Bio-CP = ZOE (1:12, 1:24 dilutions) and Calen-ZO = Bio-CP > ZOE (1:32 dilution). Bio-CP induced greater ALP activity at 7 days, and greater mineralised nodule production, compared to Calen-ZO (p<0.05). Conclusions Bio-CP showed adequate physicochemical properties, cytocompatibility and potential to induce mineralisation.

Humans , Root Canal Filling Materials , Dental Pulp Cavity , Osteoblasts , Root Canal Obturation , Tooth, Deciduous , Zinc Oxide-Eugenol Cement , Biology