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1.
Article in English | WPRIM | ID: wpr-811429

ABSTRACT

OBJECTIVES: This study aimed to evaluate the cell viability and migration of Endosequence Bioceramic Root Canal Sealer (BC Sealer) compared to MTA Fillapex and AH Plus.MATERIALS AND METHODS: BC Sealer, MTA Fillapex, and AH Plus were placed in contact with culture medium to obtain sealers extracts in dilution 1:1, 1:2 and 1:4. 3T3 cells were plated and exposed to the extracts. Cell viability and migration were assessed by 3-(4,5-dimethyl-thiazoyl)-2,5-diphenyl-tetrazolium bromide (MTT) and Scratch assay, respectively. Data were analyzed by Kruskal-Wallis and Dunn's test (p < 0.05).RESULTS: The MTT assay revealed greater cytotoxicity for AH Plus and MTA Fillapex at 1:1 dilution when compared to control (p < 0.05). At 1:2 and 1:4 dilutions, all sealers were similar to control (p > 0.05) and MTA Fillapex was more cytotoxic than BC Sealer (p < 0.05). Scratch assay demonstrated the continuous closure of the wound according to time. At 30 hours, the control group presented closure of the wound (p < 0.05). At 36 hours, only BC Sealer presented the closure when compared to AH Plus and MTA Fillapex (p < 0.05). At 42 hours, AH Plus and MTA Fillapex showed a wound healing (p > 0.05).CONCLUSIONS: All tested sealers demonstrated cell viability highlighting BC Sealer, which showed increased cell migration capacity suggesting that this sealer may achieve better tissue repair when compared to other tested sealers.


Subject(s)
3T3 Cells , Animals , Calcium , Cell Movement , Cell Proliferation , Cell Survival , Dental Pulp Cavity , Endodontics , Fibroblasts , Mice , Pemetrexed , Wound Healing , Wounds and Injuries
2.
Acta Physiologica Sinica ; (6): 175-180, 2020.
Article in Chinese | WPRIM | ID: wpr-827070

ABSTRACT

The present study was aimed to clarify the signaling molecular mechanism by which fibroblast growth factor 21 (FGF21) regulates leptin gene expression in adipocytes. Differentiated 3T3-F442A adipocytes were used as study object. The mRNA expression level of leptin was detected by fluorescence quantitative RT-PCR. The phosphorylation levels of proteins of signal transduction pathways were detected by Western blot. The results showed that FGF21 significantly down-regulated the mRNA expression level of leptin in adipocytes, and FGF21 receptor inhibitor BGJ-398 could completely block this effect. FGF21 up-regulated the phosphorylation levels of ERK1/2 and AMPK in adipocytes. Either ERK1/2 inhibitor SCH772984 or AMPK inhibitor Compound C could partially block the inhibitory effect of FGF21, and the combined application of these two inhibitors completely blocked the effect of FGF21. Neither PI3K inhibitor LY294002 nor Akt inhibitor AZD5363 affected the inhibitory effect of FGF21 on leptin gene expression. These results suggest that FGF21 may inhibit leptin gene expression by activating ERK1/2 and AMPK signaling pathways in adipocytes.


Subject(s)
3T3 Cells , Adenylate Kinase , Adipocytes , Metabolism , Animals , Down-Regulation , Fibroblast Growth Factors , Metabolism , Leptin , Metabolism , MAP Kinase Signaling System , Mice , Phosphorylation , Signal Transduction
3.
Article in Chinese | WPRIM | ID: wpr-774189

ABSTRACT

In this study, we aim to investigat the effect of microgravity on osteoblast differentiation in osteoblast-like cells (MC3T3-E1). In addition, we explored the response mechanism of nuclear factor-kappa B (NF-κB) signaling pathway to "zero- " in MC3T3-E1 cells under the simulated microgravity conditions. MC3T3-E1 were cultured in conventional (CON) and simulated microgravity (SMG), respectively. Then, the expression of the related osteoblastic genes and the specific molecules in NF-κB signaling pathway were measured. The results showed that the mRNA and protein levels of alkaline phosphatase (ALP), osteocalcin (OCN) and type Ⅰ collagen (CoL-Ⅰ) were dramatically decreased under the simulated microgravity. Meanwhile, the NF-κB inhibitor α (IκB-α) protein level was decreased and the expressions of phosphorylation of IκB-α (p-IκB-α), p65 and phosphorylation of p65 (p-p65) were significantly up-regulated in SMG group. In addition, the IL-6 content in SMG group was increased compared to CON. These results indicated that simulated microgravity could activate the NF-κB pathway to regulate MC3T3-E1 cells differentiation.


Subject(s)
3T3 Cells , Animals , Cell Differentiation , Mice , NF-kappa B , Physiology , Osteoblasts , Signal Transduction , Weightlessness Simulation
4.
Article in Chinese | WPRIM | ID: wpr-773491

ABSTRACT

OBJECTIVE@#To investigate the relationship between necroptosis and apoptosis in MCET3-E1 cell death induced by glucocorticoids.@*METHODS@#MC3T3-E1 cells were incubated with 10-6 mol/L dexamethasone followed by treatment with the apoptosis inhibitor z-VAD-fmk (40 μmol/L) or the necroptosis inhibitor necrostatin-1 (40 μmol/L) for 2 h. At 72 h after incubation with dexamethasone, the cells were harvested to determine the cell viability using WST-1 assay and the rate of necrotic cells using annexin V/PI double staining; the percentage of apoptotic cells was determined using Hoechst staining. The mitochondrial membrane potential and the level of ATP in the cells were also evaluated. Transmission electron microscopy was used to observe the microstructural changes of the cells. The expressions of RIP-1 and RIP-3 in the cells were detected by Western blotting.@*RESULTS@#At a concentration of 10-6 mol/L, dexamethasone induced both apoptosis and necroptosis in MC3T3- E1 cells. Annexin V/PI double staining showed that inhibition of cell apoptosis caused an increase in cell necrosis manifested by such changes as mitochondrial swelling and plasma membrane disruption, as shown by electron microscopy; Hoechst staining showed that the percentage of apoptotic cells was significantly reduced. When necroptosis was inhibited by necrostatin-1, MC3T3-E1 cells showed significantly increased apoptosis as shown by both AV/PI and Hoechst staining, and such changes were accompanied by changes in mitochondrial membrane potential and ATP level in the cells.@*CONCLUSIONS@#In the process of dexamethasone-induced cell death, necroptosis and apoptosis can transform reciprocally accompanied by functional changes of the mitochondria.


Subject(s)
3T3 Cells , Adenosine Triphosphate , Animals , Apoptosis , Cell Death , Dexamethasone , Membrane Potential, Mitochondrial , Mice , Microscopy, Electron , Mitochondria , Necrosis
5.
Article in English | WPRIM | ID: wpr-773368

ABSTRACT

OBJECTIVE@#We aimed to explore how fermented barley extracts with Lactobacillus plantarum dy-1 (LFBE) affected the browning in adipocytes and obese rats.@*METHODS@#In vitro, 3T3-L1 cells were induced by LFBE, raw barley extraction (RBE) and polyphenol compounds (PC) from LFBE to evaluate the adipocyte differentiation. In vivo, obese SD rats induced by high fat diet (HFD) were randomly divided into three groups treated with oral gavage: (a) normal control diet with distilled water, (b) HFD with distilled water, (c) HFD with 800 mg LFBE/kg body weight (bw).@*RESULTS@#In vitro, LFBE and the PC in the extraction significantly inhibited adipogenesis and potentiated browning of 3T3-L1 preadipocytes, rather than RBE. In vivo, we observed remarkable decreases in the body weight, serum lipid levels, white adipose tissue (WAT) weights and cell sizes of brown adipose tissues (BAT) in the LFBE group after 10 weeks. LFBE group could gain more mass of interscapular BAT (IBAT) and promote the dehydrogenase activity in the mitochondria. And LFBE may potentiate process of the IBAT thermogenesis and epididymis adipose tissue (EAT) browning via activating the uncoupling protein 1 (UCP1)-dependent mechanism to suppress the obesity.@*CONCLUSION@#These results demonstrated that LFBE decreased obesity partly by increasing the BAT mass and the energy expenditure by activating BAT thermogenesis and WAT browning in a UCP1-dependent mechanism.


Subject(s)
3T3 Cells , Adipocytes , Physiology , Adipose Tissue, Brown , Physiology , Adipose Tissue, White , Physiology , Animal Feed , Animals , Anti-Obesity Agents , Metabolism , Cell Differentiation , Diet , Fermentation , Hordeum , Chemistry , Lactobacillus plantarum , Chemistry , Male , Mice , Obesity , Drug Therapy , Genetics , Plant Extracts , Chemistry , Probiotics , Metabolism , Random Allocation , Rats , Rats, Sprague-Dawley , Uncoupling Protein 1 , Genetics , Metabolism
6.
An. acad. bras. ciênc ; 90(1): 195-204, Mar. 2018. tab, graf
Article in English | LILACS | ID: biblio-886907

ABSTRACT

ABSTRACT Demand for medical implants is rising day by day as the world becomes the place for more diseased and older people. Accordingly, in this research, metallocene polyethylene (mPE), a commonly used polymer was treated with UV rays for improving its biocompatibility. Scanning electron microscopy (SEM) images confirmed the formation of crests and troughs, which depicts the improvement of surface roughness of mPE substrates caused by UV etching. Accordingly, the contact angle measurements revealed that the wettability of mPE-2.5 J/cm2 (68.09º) and mPE-5 J/cm2 (57.93º) samples were found to be increased compared to untreated mPE (86.84º) indicating better hydrophilicity. Further, the UV treated surface exhibited enhanced blood compatibility as determined in APTT (untreated mPE- 55.3 ± 2.5 s, mPE-2.5 J/cm2 - 76.7 ± 4.1 s and mPE-5 J/cm2 - 112.3 ± 2 s) and PT (untreated mPE - 24.7 ± 1.5 s, mPE- 2.5 J/cm2 - 34.3 ± 1.1 s and mPE-5 J/cm2 - 43 ± 2 s) assay. Moreover, the treated mPE-2.5 J/cm2 (4.88%) and mPE-5 J/cm2 (1.79%) showed decreased hemolytic percentage compared to untreated mPE (15.40%) indicating better safety to red blood cells. Interestingly, the changes in physicochemical properties of mPE are directly proportional to the dosage of the UV rays. UV modified mPE surfaces were found to be more compatible as identified through MTT assay, photomicrograph and SEM images of the seeded 3T3 cell population. Hence UV-modified surface of mPE may be successfully exploited for medical implants.


Subject(s)
Animals , Rabbits , Rats , Ultraviolet Rays , Materials Testing , Metallocenes/radiation effects , Surface Properties/radiation effects , Cattle , Microscopy, Electron, Scanning , 3T3 Cells , Hydrophobic and Hydrophilic Interactions , Metallocenes/chemistry , Hemolysis , Histocompatibility
7.
Braz. j. med. biol. res ; 51(12): e7574, 2018. graf
Article in English | LILACS | ID: biblio-974257

ABSTRACT

Bone fracture is a common medical condition, which may occur due to traumatic injury or disease-related conditions. Evidence suggests that microRNAs (miRNAs) can regulate osteoblast differentiation and function. In this study, we explored the effects and mechanism of miR-221 on the growth and migration of osteoblasts using MC3T3-E1 cells. The expression levels of miR-221 in the different groups were measured by qRT-PCR. Then, miR-221 mimic and inhibitor were transfected into MC3T3-E1 cells, and cell viability and migration were measured using the CCK-8 assay and the Transwell migration assay. Additionally, the expression levels of differentiation-related factors (Runx2 and Ocn) and ZFPM2 were measured by qRT-PCR. Western blot was used to measure the expression of cell cycle-related proteins, epithelial-mesenchymal transition (EMT)-related proteins, ZFPM2, and Wnt/Notch, and Smad signaling pathway proteins. miR-221 was significantly up-regulated in the patients with lumbar compression fracture (LCM) and trochanteric fracture (TF). miR-221 promoted ALP, Runx2, and OPN expressions in MC3T3-E1 cells. miR-221 overexpression significantly increased cell proliferation, migration, differentiation, and matrix mineralization, whereas suppression of miR-221 reversed these effects. Additionally, the results displayed that ZFPM2 was a direct target gene of miR-221, and overexpression of ZFPM2 reversed the promoting effects of miR-221 overexpression on osteoblasts. Mechanistic study revealed that overexpression of miR-221 inactivated the Wnt/Notch and Smad signaling pathways by regulating ZFPM2 expression. We drew the conclusions that miR-221 overexpression promoted osteoblast proliferation, migration, and differentiation by regulation of ZFPM2 expression and deactivating the Wnt/Notch and Smad signaling pathways.


Subject(s)
Humans , Animals , Rabbits , Cell Differentiation/physiology , Cell Movement/physiology , MicroRNAs/physiology , Cell Proliferation/physiology , DNA-Binding Proteins/physiology , Fractures, Bone/blood , Osteoblasts/physiology , Reference Values , Transcription Factors/blood , Cell Survival/physiology , Blotting, Western , Analysis of Variance , 3T3 Cells , MicroRNAs/blood , DNA-Binding Proteins/blood
8.
Article in Chinese | WPRIM | ID: wpr-690006

ABSTRACT

<p><b>OBJECTIVE</b>To establish osteoblast-osteoclast cell co-culture system in a Transwell chamber, and detect cell viability of osteoblasts and osteoclasts in system.</p><p><b>METHODS</b>Osteoblast MC3T3-E1 and mouse monocytes RAW264.7 were cultivated in vitro. RANKL-induced mouse RAW264.7 monocytes differentiated into mature osteoclasts, osteoblast-osteoclast cell co-culture system was established in Transwell chamber. Cell activity of osteoblasts and osteoclasts were detected by CCK-8 experimenting, Alizarin Red staining, TRAP staining. The expression of OPG, ALP, RANKL, TGF-b1 gene and RANKL protein in osteoblast MC3T3-E1 were detected by PCR, Western-Blot methods. Also, the expression of RANK, NF-κB in gene and protein level in osteoclast were measured through the same method respectively.</p><p><b>RESULTS</b>The co-culture system of Mouse MC3T3-E1 cells and RAW264.7 cell were established in Transwell chamber. Co-culture system affected cell division activities of osteoblasts and osteoclasts. Differentiation of osteoblasts were increased, while differentiation of osteoclast division were slight decreased under microscope observation. OPG (0.65±0.08) and ALP (0.16±0.01) gene expression of co-culture system were less than single culture OPG(1.00±0.08) and ALP (1.01±0.16); TGF-b1(4.42±0.21) and RANKL(4.12±1.04) of osteoblasts in co-culture system were higher than TGF-b1(1.00±0.10) and RANKL(1.00±0.09) under single culture. However, gene expression of RANK(0.63±0.06) and NF-κB(0.64±0.08) in co-culture system were decreased than RANK(1.00±0.08) and NF-κB(1.00±0.09), in single culture, and had significant differences. Similarly, protein expression of OPG(0.43±0.05) and NF-κB(0.59±0.05) of co-culture system were less than OPG(0.84±0.06) and NF-κB(1.13±0.03) of single culture. While RANKL protein expression (0.54±0.03)of co-culture system was more than single culture RANKL(0.31±0.03), and had statistically differences, which was in agreement of the trend of gene expression change.</p><p><b>CONCLUSIONS</b>Co-culture system of mouse MC3T3-E1 cells and RAW264.7 cell was viable in Transwell chamber, and the activity of osteoblasts is higher than osteoclasts in co-culture system.</p>


Subject(s)
3T3 Cells , Animals , Cell Differentiation , Coculture Techniques , Mice , NF-kappa B , Metabolism , Osteoblasts , Cell Biology , Osteoclasts , Cell Biology , Osteoprotegerin , Metabolism , RANK Ligand , Metabolism , Receptor Activator of Nuclear Factor-kappa B , Metabolism , Transforming Growth Factor beta1 , Metabolism
9.
Braz. dent. j ; 28(6): 744-748, Nov.-Dec. 2017. tab
Article in English | LILACS | ID: biblio-888705

ABSTRACT

Abstract To examine the effect of the alternative coinitiator 4,4'bis dimethylamino benzydrol (BZN) in degree of conversion (DC), mechanical and biological properties of experimental composites. The coinitiator BZN was used in three concentrations (0.2, 0.5 and 1.2%), and the coinitiator DMAEMA was used as control at the same concentrations as above. The molar concentration of camphorquinone (CQ) and coinitiators was kept constant (1:1). The composites were manipulated and submitted to microhardness test (VHN), flexural and compressive strength (in MPa), elastic modulus (GPa), DC (FT-IR) and in vitro cytotoxicity (against 3T3 fibroblastic cells) of the experimental resins. Data were subjected to two-way ANOVA and Tukey post-test (α=0.05). The experimental composite resin with BZN showed higher DC values compared to control DMAEMA groups. For the mechanical properties, microhardness values were higher in BZN groups; flexural strength and elastic modulus were similar between all the groups. Compressive strength for groups BZN0.5 and DMAEMA0.5 were not statistically different, being the lowest values attributed to group BZN0.2. The experimental resins with BZN and DMAEMA were considered nontoxic against 3T3 fibroblasts. The inclusion of the coinitiator BZN in experimental composites was considered nontoxic against 3T3 fibroblast cells, without compromising DC and mechanical properties.


Resumo Analisar o efeito do co-iniciador alternativo 4,4'bisdimetilaminobenzidrol (BZN) no grau de conversão (GC) e nas propriedades mecânicas e biológicas de resinas compostas experimentais. O co-iniciador BZN foi utilizado em três concentrações (0,2, 0,5 e 1,2), e o co-iniciador DMAEMA como controle, nas mesmas concentrações acima. A concentração molar entre canforoquinona (CQ) e os co-iniciadores foi mantida constante (1:1). As resinas compostas foram manipuladas e submetidas aos testes de microdureza (VHN), resistência à compressão e flexural (em MPa), módulo de elasticidade (em GPa), GC (em %, por meio de espectroscopia micro-Raman e FTIR com KBr), citotoxicidade in vitro (frente às células fibroblásticas 3T3) das resinas experimentais. Os resultados foram submetidos ao teste ANOVA 1 fator e pós-teste de Tukey (α=0,05). As resinas compostas experimentais com o BZN apresentaram GC e propriedades mecânicas satisfatórias, além de serem consideradas atóxicas a fibroblastos 3T3. A inclusão do co-iniciador BZN à resina composta foi considerada não tóxica frente a células fibroblásticas 3T3 e sem comprometer o grau de conversão e as propriedades mecânicas da mesma.


Subject(s)
Animals , Mice , Amines/chemistry , Composite Resins , Materials Testing , 3T3 Cells
10.
Article in Chinese | WPRIM | ID: wpr-819079

ABSTRACT

Objective: To investigate the effect of icaritin on maturation and mineralization of mouse osteoblast MC3T3-E1 cells and its mechanism. Methods: The cultured MC3T3-E1 cells were divided into blank control group, CXC chemokine receptor type 4 (CXCR4) inhibitor (AMD3100) group, icaritin group, and icaritin plus AMD3100 group. The expression of CXCR4, stromal cell-derived factor 1 (SDF-1) and osteogenesis-related genes and proteins were detected by real-time RT-PCR and Western blotting after drug treatment for 24 h. The alkaline phosphatase (ALP) activity was determined with ALP kit on d3 and d6; calcium nodules were detected by alizarin red staining after drug treatment for 14 d. Results: Real time RT-PCR showed that compared with the blank control group, relative expressions of CXCR4, SDF-1 and osteogenesis-related genes in icaritin group were significantly increased (PPCXCR4 gene was decreased (PPPPPConclusion: Icaritin may promote maturation and mineralization of mouse osteoblast MC3T3-E1 cells through CXCR4/SDF-1 signaling pathway.


Subject(s)
3T3 Cells , Animals , Calcification, Physiologic , Chemokine CXCL12 , Metabolism , Flavonoids , Pharmacology , Gene Expression Regulation, Developmental , Mice , Osteoblasts , Cell Biology , Receptors, CXCR4 , Metabolism , Signal Transduction
11.
Article in Chinese | WPRIM | ID: wpr-819076

ABSTRACT

Objective: To analysis the biomechanical and biocompatible properties of calcium phosphate cement (CPC) enhanced by chitosan short nanofibers(CSNF) and Arg-Gly-Asp (RGD). Methods: Chitosan nanofibers were prepared by electrospinning, and cut into short fibers by high speed dispersion. CPC with calcium phosphorus ratio of 1.5:1 was prepared by Biocement D method. The composition and structure of CPC, CSNF, RGD modified CSNF (CSNF-RGD), CSNF enhanced CPC (CPC-CSNF), RGD modified CPC-CSNF (CPC-CSNF-RGD) were observed by infrared spectrum, X-ray diffraction (XRD) and scan electron microscopy (SEM). The mechanical properties were measured by universal mechanical testing instrument. The adhesion and proliferation of MC3T3 cells were assessed using immunofluorescence staining and MTT method. Results: The distribution of CSNF in the scaffold was homogeneous, and the porous structure between the nanofibers was observed by SEM. The infrared spectrum showed the characteristic peaks at 1633 nm and 1585 nm, indicating that RGD was successfully grafted on chitosan nanofibers. The XRD pattern showed that the bone cement had a certain curability. The stain-stress test showed that break strengths were (17.74±0.54) MPa for CPC-CSNF and (16.67±0.56) MPa for CPCP-CSNF-RGD, both were higher than that of CPC(all PPConclusion: CSNF-RGD can improve the biomechanical property and biocompatibility of CPC, indicating its potential application in bone tissue repair.


Subject(s)
3T3 Cells , Animals , Biocompatible Materials , Bone Cements , Chemistry , Metabolism , Pharmacology , Calcium Phosphates , Metabolism , Cell Proliferation , Chitosan , Chemistry , Pharmacology , Mice , Nanofibers , Chemistry , Oligopeptides , Chemistry
12.
Braz. dent. j ; 27(6): 652-656, Nov.-Dec. 2016. tab, graf
Article in English | LILACS | ID: biblio-828068

ABSTRACT

Abstract The aim of the present study was to evaluate the cytotoxic effects of five endodontic sealers (AH Plus, Endomethasone N, EndoSequence BC, MTA Fillapex and Pulp Canal Sealer EWT) using a three-dimensional (3D) cell culture model. A conventional bi-dimensional (2D) cell culture model was used as reference technique for comparison. Balb/c 3T3 fibroblasts were cultured in conventional bi-dimensional cell culture and in rat-tail collagen type I three-dimensional cell culture models. Then, both cell cultures were incubated with elutes of freshly mixed endodontic sealers for 24 h. Cell viability was measured by the methyl-thiazol-diphenyltetrazolium assay (MTT). Data were statistically analyzed using ANOVA and the Tukey test at a significance level of p<0.05. All tested sealers exhibited cytotoxic effects; however, cytotoxic effect was culture model- and sealer-dependent. Sealers showed higher cytotoxicity in 2D than in 3D cell culture model (p<0.05). In both conditions, EndoSequence BC showed the lowest cytotoxicity (p<0.05). MTA Fillapex was much more cytotoxic than the other tested endodontic sealers (p<0.05), with the exception of AH Plus in the 2D cell culture model (p>0.05). Endomethasone N and Pulp Canal Sealer EWT showed lower cytotoxic effects than AH Plus in 2D cell culture model (p<0.05); however no statistical differences was observed among these sealers in 3D cell culture model. It may be concluded that cytotoxicity was higher in 2D cell culture compared to 3D cell culture. EndoSequence BC sealer exhibited the highest cytocompatibility and MTA Fillapex the lowest cytocompatibility.


Resumo O objetivo do presente estudo foi avaliar os efeitos citotóxicos de cinco cimentos endodônticos (AH Plus, Endomethasone N, EndoSequence BC, MTA Fillapex e Pulp Canal Sealer EWT) utilizando um modelo de cultura celular tridimensional (3D). Utilizou-se um modelo convencional de cultura de células bidimensionais (2D) como técnica de referência para comparação. Os fibroblastos Balb/c 3T3 foram cultivados em culturas de células bidimensionais convencionais e em modelos de cultura de células tridimensionais de colagéno de cauda de rato do tipo I. Em seguida, ambas as culturas de células foram incubadas com eluções dos cimentos endodônticos recém manipulados, durante 24 h. A viabilidade celular foi medida pelo ensaio de MTT. Os dados foram analisados estatisticamente utilizando ANOVA e o teste de Tukey com nível de significância de p<0,05. Todos os cimentos testados exibiram efeitos citotóxicos. Contudo, o efeito citotóxico foi dependente do modelo de cultura e do cimento testado. Os cimentos apresentaram maior citotoxicidade no modelo 2D do que no modelo 3D (p<0,05). Em ambas as condições, a EndoSequence BC apresentou a menor citotoxicidade (p<0,05). MTA Fillapex foi mais citotóxico do que os outros cimentos endodônticos testados (p<0,05), com exceção do AH Plus no modelo de cultura de células 2D (p>0,05). Endomethasone N e EWT mostraram efeitos citotóxicos mais baixos do que AH Plus no modelo de cultura de células 2D (p<0,05); entretanto, não houve diferenças estatísticas entre esses cimentos no modelo de cultura de células 3D. Pode concluir-se que a citotoxicidade foi maior na cultura de células 2D em comparação com a cultura de células 3D. EndoSequence BC selante exibiu a maior citocompatibilidade e MTA Fillapex a menor citocompatibilidade.


Subject(s)
Animals , Mice , Root Canal Filling Materials , 3T3 Cells , Mice, Inbred BALB C
13.
Rev. peru. med. integr ; 1(2): 5-11, 2016. tab, graf
Article in Spanish | MTYCI, LILACS, MTYCI | ID: biblio-876354

ABSTRACT

Objetivos: Evaluar la actividad citotóxica del extracto etanólico de Alternanthera mexicana sobre las líneas celulares 3T3 y HUTU80. Materiales y métodos: Estudio experimental. Se empleó líneas celulares embrionarias de fibroblastos de ratón 3T3 y células de adenocarcinoma gástrico humano HUTU80. Para evaluar la citotoxicidad del extracto etanólico de Alternanthera mexicana se utilizó el método colorímetro SRB, y para establecer la concentración inhibitoria 50 (CI50) se realizó un análisis de regresión lineal. Se comparó el efecto del extracto etanólico de Alternanthera mexicana frente al 5-fluorouracilo (5FU). Resultados: La línea 3T3 que recibió Alternanthera mexicana creció ligeramente con un CI50 mayor de 500 mg/mL mientras que la que recibió 5-FU mostró una CI50 menor a 0,1 ug/mL. En la línea HUTU80, el CI50 del 5-FU y Alternanthera mexicana fue de 0,32 ug/ mL y 87,1 ug/mL, respectivamente. Alternanthera mexicana mostró un índice de selectividad >5,74 evidenciando mayor citotoxicidad en la línea celular tumoral (HUTU80), mientras que el 5-FU con un índice de selectividad menor que 0,31 nos indica que es más tóxico para las líneas normales. Conclusiones: Los resultados sugieren que el extracto etanólico de Alternanthera mexicana podría ser citotóxico en la línea celular HUTU80 más no en la 3T3.


Subject(s)
Humans , Animals , Mice , 3T3 Cells , Cell Survival/drug effects , Plant Extracts/toxicity , Models, Animal
14.
Article in Chinese | WPRIM | ID: wpr-286898

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of the non-PLC-dependent protein kinase C (PKC) pathway of parathyroid hormone (PTH) on the apoptosis and proliferation of osteoblast MC-3T3E1 cells.</p><p><b>METHODS</b>MC-3T3E1 cells were seeded in 96-well plates at the density of 1.5×10(4) cells/mL and incubated for 3 day. The cells were then exposed to 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-28), 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-34), 100 nmol/L of [Gly(1), Arg(19)]hPTH(1-34)+1 µmol/L Go6983, 1 µmol/L Go6983, or deionized water (control) for 1, 24 or 48 h. After the treatments, cell counting kit-8 (CCK-8) and Caspase-Glo® 3/7 Assay (Caspase-3) were used to examine the proliferation and apoptosis of MC3T3-E1 cells.</p><p><b>RESULTS</b>CCK-8 results showed that hPTH(1-34) increased the number of MC3T3-E1 cells compared with hPTH(1-34)+Go6983 at 1 h and 24 h, but this difference was not statistically different. At 48 h, treatment with hPTH(1-34), as compared with hPTH(1-28), significantly increased the number of MC3T3-E1 cells (P<0.05), and this effect was blocked by the PKC inhibitor Go6983 (P<0.05). hPTH(1-34) did not result in significant inhibition of MC3T3-E1 cell apoptosis at 1 h and 24 h as compared with hPTH(1-34)+Go6983, but significantly inhibited the cell apoptosis as compared with hPTH(1-28) (P<0.05); this inhibitory effect was blocked by Go6983 (P<0.05).</p><p><b>CONCLUSION</b>s A relatively long time (for 48 h) of exposure to PTH can inhibit apoptosis and promote the proliferation of MC3T3-E1cells through a non-PLC-dependent PKC pathway.</p>


Subject(s)
3T3 Cells , Animals , Apoptosis , Cell Proliferation , Indoles , Pharmacology , Maleimides , Pharmacology , Mice , Osteoblasts , Parathyroid Hormone , Pharmacology , Protein Kinase C , Metabolism , Signal Transduction
15.
Chinese Medical Journal ; (24): 2576-2581, 2016.
Article in English | WPRIM | ID: wpr-230918

ABSTRACT

<p><b>BACKGROUND</b>Three-dimensional (3D) printing technology holds great promise for treating diseases or injuries that affect human bones with enhanced performance over traditional techniques. Different patterns of design can lead to various mechanical properties and biocompatibility to various degrees. However, there is still a long way to go before we can fully take advantage of 3D printing technologies.</p><p><b>METHODS</b>This study tailored 3D printed scaffolds with gelatin and platelets to maximize bone regeneration. The scaffolds were designed with special internal porous structures that can allow bone tissue and large molecules to infiltrate better into the scaffolds. They were then treated with gelatin and platelets via thermo-crosslinking and freeze-drying, respectively. Vascular endothelial growth factor (VEGF) and transforming growth factor (TGF)-β1 were measured at different time points after the scaffolds had been made. Cell proliferation and cytotoxicity were determined via cell counting kit-8 (CCK-8) assay.</p><p><b>RESULTS</b>There was a massive boost in the level of VEGF and TGF-β1 released by the scaffolds with gelatin and platelets compared to that of scaffolds with only gelatin. After 21 days of culture, the CCK-8 cell counts of the control group and treated group were significantly higher than that of the blank group (P < 0.05). The cytotoxicity test also indicated the safety of the scaffolds.</p><p><b>CONCLUSIONS</b>Our experiments confirmed that the 3D printed scaffolds we had designed could provide a sustained-release effect for growth factors and improve the proliferation of preosteoblasts with little cytotoxicity in vitro. They may hold promise as bone graft substitute materials in the future.</p>


Subject(s)
3T3 Cells , Animals , Biocompatible Materials , Chemistry , Cell Proliferation , Cell Survival , Gelatin , Chemistry , Mice , Printing, Three-Dimensional , Tissue Engineering , Methods , Tissue Scaffolds , Chemistry , Transforming Growth Factor beta1 , Chemistry , Pharmacology , Vascular Endothelial Growth Factor A , Chemistry , Pharmacology
16.
Article in English | WPRIM | ID: wpr-290150

ABSTRACT

Osteoarthritis is recognised to be an interactive pathological process involving the cartilage, subchondral bone and synovium. The signals from the synovium play an important role in cartilage metabolism, but little is known regarding the influence of the signalling from bone. Additionally, the collagenases and stromelysin-1 are involved in cartilage catabolism through mitogen-activated protein kinase (MAPK) signalling, but the role of the gelatinases has not been elucidated. Here, we studied the influence of osteoclastic signals on chondrocytes by characterising the expression of interleukin-1β (IL-1β)-induced gelatinases through MAPK signalling. We found that osteoclast-conditioned media attenuated the gelatinase activity in chondrocytes. However, IL-1β induced increased levels of gelatinase activity in the conditioned media group relative to the mono-cultured chondrocyte group. More specifically, IL-1β restored high levels of gelatinase activity in c-Jun N-terminal kinase inhibitor-pretreated chondrocytes in the conditioned media group and led to lower levels of gelatinase activity in extracellular signal-regulated kinase or p38 inhibitor-pretreated chondrocytes. Gene expression generally correlated with protein expression. Taken together, these results show for the first time that signals from osteoclasts can influence gelatinase activity in chondrocytes. Furthermore, these data show that IL-1β restores gelatinase activity through MAPK inhibitors; this information can help to increase the understanding of the gelatinase modulation in articular cartilage.


Subject(s)
3T3 Cells , Animals , Cartilage, Articular , Cell Biology , Cell Survival , Physiology , Cells, Cultured , Chondrocytes , Coculture Techniques , Culture Media, Conditioned , Gelatinases , Interleukin-1beta , Pharmacology , JNK Mitogen-Activated Protein Kinases , MAP Kinase Signaling System , Physiology , Matrix Metalloproteinase 2 , Matrix Metalloproteinase 9 , Mice , Mitogen-Activated Protein Kinases , Monocytes , Cell Biology , NF-kappa B , Osteoclasts , Physiology , Protease Inhibitors , Tissue Inhibitor of Metalloproteinase-1 , Tissue Inhibitor of Metalloproteinase-2 , p38 Mitogen-Activated Protein Kinases
17.
Article in English | WPRIM | ID: wpr-109972

ABSTRACT

OBJECTIVE: To synthesize mesoporous silica-core-shell magnetic nanoparticles (MNPs) encapsulated by liposomes (Lipo [MNP@m-SiO2]) in order to enhance their stability, allow them to be used in any buffer solution, and to produce trastuzumab-conjugated (Lipo[MNP@m-SiO2]-Her2Ab) nanoparticles to be utilized in vitro for the targeting of breast cancer. MATERIALS AND METHODS: The physiochemical characteristics of Lipo[MNP@m-SiO2] were assessed in terms of size, morphological features, and in vitro safety. The multimodal imaging properties of the organic dye incorporated into Lipo[MNP@m-SiO2] were assessed with both in vitro fluorescence and MR imaging. The specific targeting ability of trastuzumab (Her2/neu antibody, Herceptin(R))-conjugated Lipo[MNP@m-SiO2] for Her2/neu-positive breast cancer cells was also evaluated with fluorescence and MR imaging. RESULTS: We obtained uniformly-sized and evenly distributed Lipo[MNP@m-SiO2] that demonstrated biological stability, while not disrupting cell viability. Her2/neu-positive breast cancer cell targeting by trastuzumab-conjugated Lipo[MNP@m-SiO2] was observed by in vitro fluorescence and MR imaging. CONCLUSION: Trastuzumab-conjugated Lipo[MNP@m-SiO2] is a potential treatment tool for targeted drug delivery in Her2/neu-positive breast cancer.


Subject(s)
3T3 Cells , Animals , Antibodies, Monoclonal, Humanized/administration & dosage , Antineoplastic Agents/administration & dosage , Breast Neoplasms/chemistry , Cell Line, Tumor , Drug Delivery Systems/methods , Female , Ferric Compounds/chemistry , Humans , Liposomes , Magnetite Nanoparticles/administration & dosage , Mice , Molecular Targeted Therapy/methods , Nanoconjugates/administration & dosage , Nanoparticles/chemistry , Receptor, ErbB-2/immunology , Silicon Dioxide/administration & dosage
18.
Article in Chinese | WPRIM | ID: wpr-341838

ABSTRACT

Previous studies have shown that ginsenoside Rb1 (Rb1), one of active components in ginseng, can activate insulin signaling pathway and promote translocation of glucose transporters (GLUTs) to increase glucose uptake in adipocytes. However, the effect of Rb1 on the expressions of GLUTs remains unknown. In this study, the effects of Rb1 on GLUT1 and GLUT4 were observed in 3T3-L1 adipocytes and epididymal adipose tissue of db/db obese diabetic mice. Male db/db mice were treated with Rb1 by intraperitoneal injection at the dosage of 20 mg x kg(-1) for 14 d. Rb1 reduced HOMA-IR significantly (P < 0.05, n = 5), and FBG and FINS sowed declining trend after treatment with Rb1. Rb1 recovered the expressions of GLUT1 and GLUT4 and phosphorylation of AKT in adipose tissue of db/db mice. In vitro, glucose consumption in 3T3-L1 adipocytes treated with 10 micromol x L(-1) Rb1 for 24 h was elevated (P < 0.05, n=3), and mRNA of GLUT1 and GLUT4 were up-regulated (P < 0.05, n=3) and proteins of GLUT1 and GLUT4 were also increased. AKT was activated in adipocytes treated with Rb1 for 3 h. It can be concluded that ginsenoside Rb1 can up-regulate the expression of GLUTs in adipose tissue, in addition to activate insulin signalling pathway, which may partially account for its insulin sensitizing activity and regulating effect of glucose metabolism.


Subject(s)
3T3 Cells , Adipocytes , Animals , Cell Line , Diabetes Mellitus, Experimental , Metabolism , Ginsenosides , Pharmacology , Glucose , Metabolism , Glucose Transport Proteins, Facilitative , Metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Inbred NOD , Up-Regulation
19.
Article in English | WPRIM | ID: wpr-358145

ABSTRACT

Oestrogen is essential for maintaining bone mass, and it has been demonstrated to induce osteoblast proliferation and bone formation. In this study, complementary DNA (cDNA) microarrays were used to identify and study the expression of novel genes that may be involved in MC3T3-E1 cells' response to 17-β estradiol. MC3T3-E1 cells were inoculated in minimum essential media alpha (α-MEM) cell culture supplemented with 17-β estradiol at different concentrations and for different time periods. MC3T3-E1 cells treated with 10⁻⁸ mol⋅L⁻¹ 17-β estradiol for 5 days exhibited the highest proliferation and alkaline phosphatase (ALP) activity; thus, this group was chosen for microarray analysis. The harvested RNA was used for microarray hybridisation and subsequent real-time reverse transcription polymerase chain reaction (RT-PCR) to validate the expression levels for selected genes. The microarray results were analysed using both functional and pathway analysis. In this study, microarray analysis detected 5403 differentially expressed genes, of which 1996 genes were upregulated and 3407 genes were downregulated, 1553 different functional classifications were identified by gene ontology (GO) analysis and 53 different pathways were involved based on pathway analysis. Among the differentially expressed genes, a portion not previously reported to be associated with the osteoblast response to oestrogen was identified. These findings clearly demonstrate that the expression of genes related to osteoblast proliferation, cell differentiation, collagens and transforming growth factor beta (TGF-β)-related cytokines increases, while the expression of genes related to apoptosis and osteoclast differentiation decreases, following the exposure of MC3T3-E1 cells to α-MEM supplemented with 17-β estradiol. Microarray analysis with functional gene classification is critical for a complete understanding of complementary intracellular processes. This microarray analysis provides large-scale gene expression data that require further confirmatory studies.


Subject(s)
3T3 Cells , Alkaline Phosphatase , Animals , Apoptosis , Genetics , Cell Culture Techniques , Cell Differentiation , Genetics , Cell Proliferation , Cell Survival , Genetics , Collagen , Genetics , Coloring Agents , Cytokines , Genetics , Estradiol , Pharmacology , Estrogens , Pharmacology , Gene Expression Profiling , Mice , Oligonucleotide Array Sequence Analysis , Osteoblasts , Signal Transduction , Genetics , Tetrazolium Salts , Thiazoles , Transforming Growth Factor beta , Genetics
20.
Article in English | WPRIM | ID: wpr-262631

ABSTRACT

<p><b>OBJECTIVE</b>To study, at the cytological level, the basic concept of Chinese medicine that "the Kidney (Shen) controls the bone".</p><p><b>METHODS</b>Kaempferol was isolated form Rhizoma Drynariae (Gu Sui Bu, GSB) and at several concentrations was incubated with opossum kidney (OK) cells, osteoblasts (MC3T3 E1) and human fibroblasts (HF) at cell concentrations of 2×10(4)/mL. Opossum kidney cell-conditioned culture media with kaempferol at 70 nmol/L (70kaeOKM) and without kaempferol (0OKM) were used to stimulate MC3T3 E1 and HF proliferation. The bone morphological protein receptors I and II (BMPR I and II) in OK cells were identified by immune-fluorescence staining and Western blot analysis.</p><p><b>RESULTS</b>Kaempferol was found to increase OK cell growth (P<0.05), but alone did not promote MC3T3 E1 or HF cell proliferation. However, although OKM by itself increased MC3T3 E1 growth by 198% (P<0.01), the 70kaeOKM further increased the growth of these cells by an additional 127% (P<0.01). It indicates that the kidney cell generates a previously unknown osteoblast growth factor (OGF) and kaempferol increases kidney cell secretion of OGF. Neither of these media had any significant effect on HF growth. Kaempferol also was found to increase the level of the BMPR II in OK cells.</p><p><b>CONCLUSIONS</b>This lends strong support to the original idea that the Kidney has a significant influence over bone-formation, as suggested by some long-standing Chinese medical beliefs, kaempferol may also serve to stimulate kidney repair and indirectly stimulate bone formation.</p>


Subject(s)
3T3 Cells , Animals , Cell Line , Culture Media, Conditioned , Intercellular Signaling Peptides and Proteins , Bodily Secretions , Kaempferols , Pharmacology , Kidney Tubules , Physiology , Bodily Secretions , Medicine, Chinese Traditional , Mice , Opossums , Osteoblasts , Chemistry
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