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1.
Braz Oral Res ; 33: e079, 2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31531565

RESUMO

Cell therapy associated with guided bone regeneration (GBR) can be used to treat bone defects under challenging conditions such as osteoporosis. This study aimed to evaluate the effect of mesenchymal stem cells (MSCs) in combination with a poly(vinylidene-trifluoroethylene)/barium titanate (PVDF-TrFE/BT) membrane on bone repair in osteoporotic rats. Osteoporosis was induced in female rats by bilateral removal of the ovaries (OVX) or sham surgery (SHAM), and the osteoporotic condition was characterized after 5 months by microtomographic and morphometric analyses. Calvarial defects were created in osteoporotic rats that immediately received the PVDF-TrFE/BT membrane. After 2 weeks, bone marrow-derived MSCs from healthy rats, characterized by the expression of surface markers using flow cytometry, or phosphate-buffered saline (PBS) (Control) were injected into the defects and bone formation was evaluated 4 weeks post-injection by microtomographic, morphometric, and histological analyses. A reduction in the amount of bone tissue in the femurs of OVX compared with SHAM rats confirmed the osteoporotic condition of the experimental model. More bone formation was observed when the defects were injected with MSCs compared to that with PBS. The modification that we are proposing in this study for the classical GBR approach where cells are locally injected after a membrane implantation may be a promising therapeutic strategy to increase bone formation under osteoporotic condition.


Assuntos
Compostos de Bário/farmacologia , Regeneração Tecidual Guiada/métodos , Células-Tronco Mesenquimais/fisiologia , Osteogênese/efeitos dos fármacos , Osteoporose/terapia , Polivinil/farmacologia , Titânio/farmacologia , Animais , Compostos de Bário/química , Densidade Óssea , Regeneração Óssea/efeitos dos fármacos , Regeneração Óssea/fisiologia , Feminino , Citometria de Fluxo , Imagem Tridimensional , Células-Tronco Mesenquimais/química , Osteogênese/fisiologia , Osteoporose/fisiopatologia , Ovariectomia , Polivinil/química , Distribuição Aleatória , Ratos Wistar , Reprodutibilidade dos Testes , Fatores de Tempo , Titânio/química , Resultado do Tratamento
2.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi ; 33(7): 901-906, 2019 Jul 15.
Artigo em Chinês | MEDLINE | ID: mdl-31298011

RESUMO

Objective: To review the literature reports on research progress of Heme oxygenase 1 (HO-1) modified mesenchymal stem cells (MSCs). Methods: The significance, effects, and related mechanism of HO-1 modification of MSCs were summarized by consulting the related literatures and reports of HO-1 modification of MSCs. Results: HO-1 modification of MSCs has important research value. It can effectively enhance the anti-oxidative stress and anti-apoptotic properties of MSCs in complex internal environment after transplantation into vivo. It can also effectively enhance the immune regulation function of MSCs. It can improve the anti-injury, repair, and immune regulation effect of MSCs in various disease models and research fields. Conclusion: The basic research of HO-1 modified MSCs has made remarkable progress, which is expected to be applied in clinical trials and provide theoretical basis and reference value for stem cell therapy.


Assuntos
Heme Oxigenase-1 , Células-Tronco Mesenquimais , Estresse Oxidativo , Apoptose , Heme Oxigenase-1/fisiologia , Células-Tronco Mesenquimais/fisiologia , Pesquisa
3.
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue ; 31(6): 750-755, 2019 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-31315736

RESUMO

OBJECTIVE: To explore the effects of Hippo pathway on differentiation, proliferation, and migration of bone marrow mesenchymal stem cells (BMSCs) in vitro. METHODS: BMSCs of C57BL/6 mice were identified using fluorescence-activated cellsorting analysis and the capabilities of osteogenic, chondrogenic and adipogenic differentiation were evaluated. The differentiation of BMSCs to type II alveolar epithelial cells (AEC II) was induced by indirect co-culture with mouse lung epithelial cells (MLE-12) and small airway epithelial cell growth medium (SAGM). The Hippo pathway was regulated by 2-deoxy-D-glucose (2-DG) and 9E1, the effects of 2-DG and 9E1 on the expression of BMSCs surface proteins (SPB, SPC and SPD) mRNA and pro-SPC protein were detected by real time quantitative polymerase chain reaction (qRT-PCR) and Western Blot. The effect of Hippo pathway on differentiation of BMSCs to AEC II cells was evaluated. The effect of Hippo pathway on the proliferation of BMSCs was evaluated by methyl thiazolyl tetrazolium (MTT) assay (intervention of 0.1, 0.5, 1.0, 5.0 mmol/L 2-DG). The scratch test and Transwell chamber test were used to analyze the effect of Hippo pathway on migration ability of BMSCs to conditioned medium of acute respiratory distress syndrome (ARDS) lung tissue. RESULTS: 2-DG could activate Hippo pathway in a dose-dependent manner and promote the differentiation to AEC II and proliferation of BMSCs, the maximum effects were observed after 5 mmol/L of 2-DG treatment [SPB mRNA (2-ΔΔCT): 2.42±0.28 vs. 1.89±0.11, SPC mRNA (2-ΔΔCT): 8.06±0.68 vs. 6.59±0.79, SPD mRNA (2-ΔΔCT): 6.45±0.37 vs. 5.27±0.28, pro-SPC/ß-actin: 5.80±1.86 vs. 4.93±1.18, proliferation rate: (145.46±18.18)% vs. (98.91±4.36)%, all P < 0.05], but 9E1 could reverse those effects through inhibition of Hippo pathway [SPB mRNA (2-ΔΔCT): 1.32±0.17 vs. 1.89±0.11, SPC mRNA (2-ΔΔCT): 3.91±0.34 vs. 6.59±0.79, SPD mRNA (2-ΔΔCT): 3.38±0.25 vs. 5.27±0.28, pro-SPC/ß-actin: 2.48±0.17 vs. 4.93±1.18, proliferation rate: (80.00±7.27)% vs. (98.91±4.36)%, all P < 0.05]. The ability of horizontal migration [wound healing: (27.17±3.53)% vs. (52.45±6.52)%, P < 0.05] and homing BMSCs to conditioned medium of ARDS lung tissue [cell count (fold, relative to control): 2.77±0.21 vs. 1.90±0.19, P < 0.05] were increased after activation of Hippo pathway by 2-DG treatment, but those effects were reversed after inhibition of Hippo pathway by 9E1 treatment [wound healing: (79.89±8.42)% vs. (52.45±6.52)%, cell count (fold, relative to control): 1.69±0.13 vs. 1.90±0.19, both P < 0.05]. CONCLUSIONS: Activation of Hippo pathway could enhance differentiation of BMSCs to AEC II, promote proliferation and ability of horizontal migration and homing BMSCs to conditioned medium of ARDS lung tissue in vitro.


Assuntos
Células da Medula Óssea/fisiologia , Células-Tronco Mesenquimais/fisiologia , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Animais , Diferenciação Celular , Movimento Celular , Proliferação de Células , Camundongos , Camundongos Endogâmicos C57BL
4.
Life Sci ; 232: 116625, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31276691

RESUMO

AIMS: The chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) is critical for cartilage regeneration. Tissues constructed from BMSCs through cartilage tissue engineering still exhibit some histological, morphological, and biomechanical differences from normal cartilage tissues. Cyclic tensile strain (CTS) can increase chondrogenic gene expression and reduce hypertrophic gene expression in chondrocytes. miR-365 has been identified as a mechanoresponsive microRNA and is an important regulator of both chondrocyte hypertrophy and differentiation. Therefore, we hypothesized that CTS may promote the chondrogenesis of BMSCs by upregulating the expression of miR-365. METHODS: BMSCs were subjected to CTS to investigate the effects and mechanism on chondrogenesis. An Agilent miRNA microarray was used to profile miRNAs in the CTS-treated BMSCs and 3D-cultured control BMSCs. miR-365 was shown to interact with HDAC4 mRNA through a luciferase reporter assay. An animal cartilage defect model was constructed and different groups of BMSCs were implanted to investigate their in vivo effect. KEY FINDINGS: CTS promoted BMSC chondrogenesis. miR-365 was significantly upregulated in CTS-treated cells and played an important role in CTS-mediated chondrogenesis. Luciferase assays showed that HDAC4 is a direct target of miR-365. An in vivo study showed that CTS treatment and miR-365 overexpression could promote cartilage regeneration from BMSCs. SIGNIFICANCE: CTS can promote the expression of miR-365, a crucial mechanosensitive microRNA involved in the chondrogenesis of BMSCs, which directly inhibits the expression of HDAC4, in turn, enhancing the chondrogenesis of BMSCs.


Assuntos
Condrogênese/genética , Células-Tronco Mesenquimais/fisiologia , MicroRNAs/genética , Animais , Medula Óssea/metabolismo , Células da Medula Óssea/citologia , Cartilagem/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Condrócitos/metabolismo , Condrogênese/fisiologia , MicroRNAs/metabolismo , Ratos , Transdução de Sinais , Resistência à Tração/fisiologia , Engenharia Tecidual
5.
Yakugaku Zasshi ; 139(6): 867-871, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31155527

RESUMO

The mesenchymal stem cell (MSC) is a type of tissue stem cell. In clinical studies, cultured MSCs have shown important therapeutic effects on diseases via both the reduction of neurological defects and the regulation of immune responses. However, in vivo MSC localization, function, and properties are poorly understood; therefore, the molecular understanding of MSC hierarchy is less advanced compared to hematopoietic stem cell hierarchy. Runt-related transcription factor 2 (Runx2) is an essential transcriptional regulator of osteoblast differentiation from MSCs. Runx2 deficiency in Paired-related homeobox 1 (Prrx1)-derived cells (Runx2Prrx1-/- mice) results in defective intramembranous ossification. Double-positive cells for Prrx1-GFP, and stem cell antigen-1 (Sca1) (Prrx1+Sca1+ cells) in the calvaria, express Runx2 at lower levels, and are more homogeneous and primitive compared with Prrx1+Sca1- cells. Our results suggest that osteoblast differentiation in vivo may begin at the Prrx1+Sca1+ MSC stage, with sequential progression to Prrx1+Sca1- cells, followed by Osterix+Prrx1-Sca1- osteoblast precursors, which eventually form mature α1(I)-collagen+ osteoblasts. This research will enable us to better understand the in vivo molecular biology features of MSCs, leading to their therapeutic applications for tissue repair and regeneration.


Assuntos
Linhagem da Célula , Descoberta de Drogas , Células-Tronco Mesenquimais , Camundongos/genética , Medicina Regenerativa , Animais , Diferenciação Celular , Subunidade alfa 1 de Fator de Ligação ao Core/fisiologia , Proteínas de Homeodomínio/fisiologia , Células-Tronco Mesenquimais/fisiologia , Biologia Molecular , Osteoblastos , Osteogênese/genética
6.
Nat Commun ; 10(1): 2876, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253842

RESUMO

Osteoarthritis (OA) is a common, painful disease. Currently OA is incurable, and its etiology largely unknown, partly due to limited understanding of OA as a whole-joint disease. Here we report that two homologous microRNAs, miR-204 and miR-211, maintain joint homeostasis to suppress OA pathogenesis. Specific knockout of miR-204/-211 in mesenchymal progenitor cells (MPCs) results in Runx2 accumulation in multi-type joint cells, causing whole-joint degeneration. Specifically, miR-204/-211 loss-of-function induces matrix-degrading proteases in articular chondrocytes and synoviocytes, stimulating articular cartilage destruction. Moreover, miR-204/-211 ablation enhances NGF expression in a Runx2-dependent manner, and thus hyper-activates Akt signaling and MPC proliferation, underlying multiplex non-cartilaginous OA conditions including synovial hyperplasia, osteophyte outgrowth and subchondral sclerosis. Importantly, miR-204/-211-deficiency-induced OA is largely rescued by Runx2 insufficiency, confirming the miR-204/-211-Runx2 axis. Further, intraarticular administration of miR-204-expressing adeno-associated virus significantly decelerates OA progression. Collectively, miR-204/-211 are essential in maintaining healthy homeostasis of mesenchymal joint cells to counteract OA pathogenesis.


Assuntos
Células-Tronco Mesenquimais/fisiologia , MicroRNAs/metabolismo , Osteoartrite/metabolismo , Animais , Condrócitos/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Camundongos , Camundongos Knockout , MicroRNAs/genética , Osteoartrite/etiologia , Osteoartrite/patologia , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/metabolismo , Sinoviócitos/metabolismo
7.
Chem Commun (Camb) ; 55(60): 8772-8775, 2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31172998

RESUMO

We introduced a new concept to the control of wetting characteristics by modulating the degree of atomic defects of two-dimensional transition metal dichalcogenide nanoassemblies of molybdenum disulfide. This work shed new light on the role of atomic vacancies on wetting characteristic that can be leveraged to develop a new class of superhydrophobic surfaces for various applications without altering their topography.


Assuntos
Adesão Celular , Dissulfetos/química , Células-Tronco Mesenquimais/fisiologia , Molibdênio/química , Nanoestruturas/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Molhabilidade
8.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi ; 33(6): 743-749, 2019 Jun 15.
Artigo em Chinês | MEDLINE | ID: mdl-31198004

RESUMO

Objective: To investigate the effect of daphnetin (DAP) combined with insulin-like growth factor 1 (IGF-1) gene transfection on chondrogenic differentiation of adipose-derived mesenchymal stem cells (ADSCs) in rats. Methods: Rat ADSCs were isolated and amplified by enzymatic digestion. The third generation ADSCs were treated with IGF-1 gene transfection as experimental group and normal ADSCs as control group. The cells of the two groups were treated with different concentrations of DAP (0, 30, 60, 90 µg/mL), respectively. Cell proliferation was detected by cell counting kit 8 (CCK-8) after cultured for 72 hours. After 14 days, real-time fluorescence quantitative PCR and Western blot were used to detect the mRNA and protein expressions of chondrocyte markers (collagen type Ⅱ and Aggrecan) in each group; and toluidine blue staining and collagen type Ⅱ immunohistochemical staining were performed. Results: CCK-8 assay showed that with the increase of DAP concentration, the cell absorbance ( A) value of the control group and the experimental group increased gradually ( P<0.05). At the same DAP concentration, the cell A value of the experimental group was significantly higher than that of the control group ( P<0.05). Real-time fluorescence quantitative PCR and Western blot showed that with the increase of DAP concentration, the relative mRNA and protein expressions of collagen type Ⅱ and Aggrecan in the control group did not change significantly, and there was no significant difference among the different concentration groups ( P>0.05). But the mRNA and protein expressions of collagen type Ⅱ and Aggrecan in the experimental group increased gradually, and the 60 and 90 µg/mL DAP concentration groups were significantly higher than 0 µg/mL DAP concentration group ( P<0.05). At the same DAP concentration, the relative mRNA and protein expressions of collagen type Ⅱ and Aggrecan were significantly higher in the experimental group than in the control group ( P<0.05). Toluidine blue staining showed that with the increase of DAP concentration, there was no significant difference in cell staining between the control group and the experimental group. At the same DAP concentration, the cells in the experimental group were slightly darker than those in the control group. Immunohistochemical staining of collagen type Ⅱ showed that with the increase of DAP concentration, there was no significant difference in the cytoplasmic brown-yellow coloring of the cells in the control group. The cytoplasmic brown-yellow coloring of the cells in the experimental group gradually deepened, with 60 and 90 µg/mL DAP concentration groups significantly deeper than 0 µg/mL DAP concentration group. At the same DAP concentration, the color of the cells in the experimental group was significantly deeper than that in the control group. Conclusion: DAP can promote the proliferation of ADSCs in rats. The differentiation of ADSCs into chondrocytes induced by DAP alone was slightly, but DAP combined with IGF-1 gene transfection has obvious synergistic effect to promote chondrogenic differentiation of ADSCs.


Assuntos
Diferenciação Celular , Fator de Crescimento Insulin-Like I , Células-Tronco Mesenquimais , Umbeliferonas , Tecido Adiposo/citologia , Animais , Células Cultivadas , Fator de Crescimento Insulin-Like I/farmacologia , Células-Tronco Mesenquimais/fisiologia , Ratos , Transfecção , Umbeliferonas/farmacologia
9.
Nat Commun ; 10(1): 2705, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31221969

RESUMO

Folded single chain polymeric nano-objects are the molecular level soft material with ultra-small size. Here, we report an easy and scalable method for preparing single-chain nanogels (SCNGs) with improved efficiency. We further investigate the impact of the dynamic molecular conformational change of SCNGs on cellular interactions from molecular to bulk scale. First, the supramolecular unfoldable SCNGs efficiently deliver siRNAs into stem cells as a molecular drug carrier in a conformation-dependent manner. Furthermore, the conformation changes of SCNGs enable dynamic and precise manipulation of ligand tether structure on 2D biomaterial interfaces to regulate the ligand-receptor ligation and mechanosensing of cells. Lastly, the dynamic SCNGs as the building blocks provide effective energy dissipation to bulk biomaterials such as hydrogels, thereby protecting the encapsulated stem cells from deleterious mechanical shocks in 3D matrix. Such a bottom-up molecular tailoring strategy will inspire further applications of single-chain nano-objects in the biomedical area.


Assuntos
Engenharia Celular/métodos , Portadores de Fármacos/química , Hidrogéis/química , Nanopartículas/química , Polímeros/química , Materiais Biocompatíveis/química , Diferenciação Celular/genética , Linhagem Celular , Humanos , Células-Tronco Mesenquimais/fisiologia , Conformação Molecular , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/metabolismo
10.
Life Sci ; 231: 116556, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31194990

RESUMO

Triiodothyronine (T3) and estrogen (E2) play important roles in the bone remodeling process and signaling of receptor activator of the nuclear factor-kappa ß (RANKL) and osteoprotegerin (OPG) expressed by osteoblasts. However, little is known of the molecular action of these hormones in conditions of hyperthyroidism and associated E2 in human cells. AIMS: This study evaluated the effects of the physiological concentration of E2 (10 nM), alone or in association with physiological (1 nM) and supraphysiological (10 nM) concentrations of T3, on RANKL and OPG gene expression in human osteoblasts. MAIN METHODS: Alkaline phosphatase and osteocalcin assays were performed to verify the presence of mature osteoblasts. After mimicking the experimental hyperthyroidism in osteoblasts untreated or treated with E2, RANKL and OPG gene expression was analyzed by real-time PCR and protein expression by western Blot and ELISA. Alizarin Red staining analyzed the amount of bone matrix after hormonal treatments. KEY FINDINGS: E2 enhanced the gene expression of OPG when associated with 1 nM and 10 nM T3. E2 was able to restore the bone matrix after an initial decrease using 1 nM and 10 nM T3. The protective effect of E2 on the RANKL and OPG signaling pathway was demonstrated. E2 restored the bone matrix induced by experimental hyperthyroidism. SIGNIFICANCE: The data highlight the importance of E2 to maintain OPG expression and osteoblast activity against possible loss of bone mass, especially in conditions where T3 is in excess.


Assuntos
Remodelação Óssea/efeitos dos fármacos , Estrogênios/fisiologia , Osteoblastos/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Remodelação Óssea/fisiologia , Osso e Ossos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Estrogênios/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hipertireoidismo/metabolismo , Células-Tronco Mesenquimais/fisiologia , Osteoclastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Osteoprotegerina/metabolismo , Ligante RANK/metabolismo , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Tri-Iodotironina/metabolismo , Tri-Iodotironina/fisiologia
11.
Life Sci ; 231: 116564, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31202840

RESUMO

Alzheimer's disease (AD) as a dementia and neurodegenerative disease, is mostly prevalent among people more than 65 years. AD is mostly manifested in the form of degraded mental function, such as losing memory and impaired cognitive function. Due to inefficiency of traditional pharmacological therapeutic approaches with no long-term cure, cell therapy can be considered as a capable approach in AD management. Therapies based on mesenchymal stem cells (MSCs) have provided hopeful results in experimental models regarding several disorders. MSCs enhance the levels of functional recoveries in pathologic experimental models of central nervous system (CNS) and are being investigated in clinical trials in neurological disorders. However, there is limited knowledge on the protective capabilities of MSCs in AD management. Almost, several experiments have suggested positive effects of MSCs and helped to better understand of AD-related dementia mechanism. MSCs have the potential to be used in AD treatment through amyloid-ß peptide (AB), Tau protein and cholinergic system. This review aimed to clarify the promising perspective of MSCs in the context of AD.


Assuntos
Doença de Alzheimer/terapia , Transplante de Células-Tronco Mesenquimais/métodos , Animais , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Ensaios Clínicos Controlados Aleatórios como Assunto
12.
Life Sci ; 229: 36-45, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31085242

RESUMO

AIMS: Bone marrow mesenchymal stem cells (BMSCs) show great potential in clinical applications such as in intervertebral disc degeneration. Nevertheless, environmental stress during the BMSC transplant or in the injured tissues is a catastrophic factor that causes cell toxicity and poor survival of BMSCs. Mitophagy plays a vital role in maintaining cellular homeostasis and defending against oxidative stress because this process could control mitochondrial quality and quantity by eliminating dysfunctional or damaged mitochondria that can cause cell death. However, the accurate mechanisms of mitophagy in protecting BMSCs against the harshness of oxidative stress remain largely unknown. MAIN METHODS: BMSCs were treated with H2O2 for various time periods. Mitophagy response was evaluated through the expression levels of LC3-II, p62 and mitophagosomal formation by using Western blot and fluorescence analysis. Cell apoptosis was examined by flow cytometry and TUNEL assay. The interactions of mitophagy and apoptosis and the possible signalling pathways were investigated through the co-treatment of mitophagy inhibitor or mitophagy activator with H2O2. KEY FINDINGS: Oxidative stress rapidly facilitated mitophagy through JNK at an early stage but decreased mitophagy and increased apoptosis at a late stage. Furthermore, mitophagy inhibition significantly enhanced the apoptosis in the cells treated by H2O2. SIGNIFICANCE: Induced mitophagy may play pivotal roles in protecting cells against oxidative stress in BMSCs.


Assuntos
Apoptose , Autofagia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Mitocôndrias/patologia , Estresse Oxidativo , Substâncias Protetoras , Animais , Sobrevivência Celular , Células Cultivadas , Mitocôndrias/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais
13.
Mol Med Rep ; 19(6): 4719-4726, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059010

RESUMO

Leukemia inhibitory factor (LIF) modulates various biological processes. Although previous studies have described the effects of LIF on adipocyte differentiation, the role of LIF receptor (LIFR) on adipocyte differentiation remains unclear. Using reverse transcription­quantitative PCR (RT­qPCR), LIFR expression was demonstrated to increase during adipogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs), indicating that LIFR may be involved in this process. To further evaluate the association between LIFR and adipogenic differentiation, lentivirus­mediated LIFR knockdown was performed in hMSCs. Cells were divided into two groups: Negative control group and LIFR­knockdown group. During the adipogenic differentiation process, intracellular lipid accumulation was assessed with Oil Red O staining at various time points (days 3, 6 and 9). Additionally, the mRNA and protein expression levels of LIF, LIFR and three molecular indicators of adipogenesis, peroxisome proliferator­activated receptor Î³ (PPARγ), CCAAT enhancer binding protein α (C/EBPα) and fatty acid binding protein 4 (FABP4/aP2), were assessed by RT­qPCR and western blotting. The culture supernatant was collected to evaluate the concentration of LIF using ELISA. The present results suggested that LIFR expression progressively increased during adipogenic differentiation of hMSCs. Conversely, LIFR knockdown significantly suppressed this process. Additionally, PPARγ, C/EBPα and aP2 were inhibited following LIFR knockdown. In contrast with LIFR, the expression levels of LIF were significantly decreased after the initiation of adipogenic differentiation. Therefore, the expression levels of LIF and LIFR exhibited opposite trends. Collectively, the present results suggested that LIFR promoted adipogenic differentiation, whereas LIF may negatively regulate this process.


Assuntos
Adipogenia/fisiologia , Diferenciação Celular/fisiologia , Regulação da Expressão Gênica , Subunidade alfa de Receptor de Fator Inibidor de Leucemia/metabolismo , Células-Tronco Mesenquimais/fisiologia , Adipócitos/metabolismo , Adipócitos/patologia , Adipogenia/genética , Células da Medula Óssea , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Diferenciação Celular/genética , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Fator Inibidor de Leucemia/genética , Fator Inibidor de Leucemia/metabolismo , Subunidade alfa de Receptor de Fator Inibidor de Leucemia/genética , Células-Tronco Mesenquimais/patologia , PPAR gama/genética , PPAR gama/metabolismo , RNA Mensageiro/metabolismo
14.
Res Vet Sci ; 124: 393-398, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31077967

RESUMO

Osteoarthritis associated with hip dysplasia is one of the most common orthopedic abnormalities in dogs, with an incidence of up to 40% in some breeds. Tissue therapy of cartilage has received great attention, with use of mesenchymal stromal cells and different types of biomaterials. The present study aimed to evaluate the effect of platelet lysate (PL) on the proliferation and differentiation of canine adipose tissue-derived mesenchymal stromal cells (ASCs), in liquid culture or hydrogels. PL was prepared from blood collected from healthy dogs and submitted to freezing-thawing cycles, and hydrogel was formed with canine thrombin. The effect of PL on the proliferation and differentiation of canine ASCs was evaluated in liquid and hydrogel systems, with microscopy, quantification of dsDNA, histology and quantification of glycosaminoglycans. The addition of 5% or 10% PL to the culture medium induced a greater proliferation rate than the presence of 10% fetal bovine serum. The cultivation of ASCs in PL gel, with normal or chondrogenic medium, resulted in maintenance of proliferation level similar to the conventional 2D cultivation, and induction of chondrogenic differentiation, especially in the presence of the chondrogenesis induction medium.


Assuntos
Tecido Adiposo/fisiologia , Condrogênese/fisiologia , Liases/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Animais , Plaquetas/enzimologia , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cães , Relação Dose-Resposta a Droga , Liases/administração & dosagem , Células-Tronco Mesenquimais/fisiologia
16.
Plast Reconstr Surg ; 143(5): 993e-1002e, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31033820

RESUMO

BACKGROUND: Cell-seeded biomaterial scaffolds have been proposed as a future option for reconstruction of bone tissue. The ability to generate larger, functional volumes of bone has been a challenge that may be addressed through the use of perfusion bioreactors. In this study, the authors investigated use of a tubular perfusion bioreactor system for the growth and differentiation of bone marrow stromal (mesenchymal stem) cells seeded onto fibrin, a highly angiogenic biomaterial. METHODS: Cells were encapsulated within fibrin beads and cultured either within a tubular perfusion bioreactor system or statically for up to 14 days. Scaffolds were analyzed for osteogenic differentiation. A rodent cranial defect model (8-mm diameter) was used to assess the bone regeneration of scaffolds cultured in the bioreactor, statically, or used immediately after formation. Immunohistochemistry was used to visualize CD31 vessel density. Micro-computed tomographic imaging was used to visualize mineral formation within the defect volume. RESULTS: Tubular perfusion bioreactor system-cultured samples showed significantly greater osteodifferentiation, indicated by an increase in VEGF expression and mineral deposition, compared with statically cultured samples. Increased expression of OPN, RUNX2, VEGF, and CD90 was seen over time in both culture methods. After implantation, bioreactor samples exhibited greater bone formation and vessel density compared with all other groups. Analysis of micro-computed tomographic images showed full union formation through the greatest diameter of the defect in all bioreactor samples and the highest levels of mineralized volume after 8 weeks. CONCLUSION: Mesenchymal stem cells encapsulated in fibrin beads and cultured in the tubular perfusion bioreactor system resulted in increased vascularization and mineralized tissue formation in vivo relative to static culture.


Assuntos
Regeneração Óssea , Técnicas de Cultura de Células/métodos , Osteogênese/fisiologia , Crânio/lesões , Tecidos Suporte , Animais , Reatores Biológicos , Células da Medula Óssea/fisiologia , Técnicas de Cultura de Células/instrumentação , Diferenciação Celular/fisiologia , Células Cultivadas , Traumatismos Craniocerebrais/cirurgia , Modelos Animais de Doenças , Humanos , Masculino , Células-Tronco Mesenquimais/fisiologia , Procedimentos Ortopédicos/instrumentação , Procedimentos Ortopédicos/métodos , Perfusão/métodos , Ratos , Ratos Sprague-Dawley , Procedimentos Cirúrgicos Reconstrutivos/instrumentação , Procedimentos Cirúrgicos Reconstrutivos/métodos , Crânio/cirurgia , Resultado do Tratamento
17.
Med Sci Monit ; 25: 2923-2934, 2019 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-31005957

RESUMO

BACKGROUND Rheumatoid arthritis model (CIA) rats were treated by tail vein injection of IL-10-modified bone marrow mesenchymal stem cells (BMSCs) to investigate its feasibility and intrinsic molecular mechanism. MATERIAL AND METHODS The CIA rat model was established by induction type II collagen, and IL-10-modified BMSCs was established by transfecting BMSCs with adenovirus. IL-10-modified BMSCs were used to treat the CIA rats. The therapeutic effect was evaluated by measuring the changes in body weight, ankle swelling, and forced swimming time, as well as observation of synovial hyperplasia and cartilage tissue repair by HE staining. Western blot analysis and ELISA were used to detect gene expression. RESULTS After 4 weeks and 8 weeks of treatment with IL10-BMSCs, the body weight, swelling value, resting time, and forced swimming struggle time of CIA rats were significantly higher than those of BMSCs-treated and -untreated CIA rats (P<0.05). Compared to BMSCs-treated CIA model rats, after treatment with IL10-BMSCs, the repair rate of osteoarticular cartilage was higher and the inhibition of synovial proliferation was better, and serum IL-17, IL-1ß, and TNF-alpha levels were lower. We found that the protein level of SIRT1 in peripheral blood mononuclear cells was lower, the protein level in spleen was higher, and phosphorylation of p65 protein in peripheral blood mononuclear cells was reduced. CONCLUSIONS The efficacy of tail vein injection of IL-10-modified BMSCs in treatment of CIA rats was superior to that of BMSCs alone, which may be related to the more pronounced suppression of IL-10-modified BMSCs in peripheral blood inflammation and spleen immune response.


Assuntos
Artrite Reumatoide/tratamento farmacológico , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/fisiologia , Animais , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/induzido quimicamente , Medula Óssea , Células da Medula Óssea , Cartilagem/efeitos dos fármacos , Colágeno Tipo II/farmacologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Mediadores da Inflamação/metabolismo , Interleucina-10/metabolismo , Interleucina-10/farmacologia , Interleucina-17/sangue , Interleucina-17/metabolismo , Interleucina-1beta/sangue , Interleucina-1beta/metabolismo , Leucócitos Mononucleares/metabolismo , Células-Tronco Mesenquimais/patologia , Ratos , Ratos Wistar , Sirtuína 1/metabolismo , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/metabolismo
18.
In Vitro Cell Dev Biol Anim ; 55(5): 387-394, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30993556

RESUMO

This study aimed at investigating the expression of osteoblast and chondrocyte-related genes in mesenchymal stem cells (MSCs), derived from rabbit adipose tissue, under mechanical vibration. The cells were placed securely on a vibrator's platform and subjected to 300 Hz of sinusoidal vibration, with a maximum amplitude of 10 µm, for 45 min per day, and for 14 consequent days, in the absence of biochemical reagents. The negative control group was placed in the conventional culture medium with no mechanical loading. The expression of osteoblast and chondrocyte-related genes was investigated using real-time polymerase chain reaction (real-time PCR). In addition, F-actin fiber structure and alignment with the help of actin filament fluorescence staining were evaluated, and the level of metabolic activity of MSCs was determined by the methyl thiazolyl tetrazolium assay. The real-time PCR study showed a significant increase of bone gene expression in differentiated cells, compared with MSCs (P < 0.05). On the other hand, the level of chondrocyte gene expression was not remarkable. Applying mechanical vibration enhanced F-actin fiber structure and made them aligned in a specific direction. It was also found that during the differentiation process, the metabolic activity of the cells increased (P < 0.05). The results of this work are in agreement with the well-accepted fact that the MSCs, in the absence of growth factors, are sensitive to low-amplitude, high-frequency vibration. Outcomes of this work can be applied in cell therapy and tissue engineering, when regulation of stem cells is required.


Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Mesenquimais/fisiologia , Estresse Mecânico , Vibração/uso terapêutico , Actinas/genética , Tecido Adiposo/metabolismo , Tecido Adiposo/fisiologia , Animais , Células da Medula Óssea/fisiologia , Diferenciação Celular/genética , Condrócitos/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Células-Tronco Mesenquimais/metabolismo , Osteogênese/genética , Osteogênese/fisiologia , Coelhos , Engenharia Tecidual
19.
J Orthop Surg Res ; 14(1): 102, 2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30975173

RESUMO

BACKGROUND: With the development of indirect three-dimensional (3D) printing technology, it is possible to customise individual scaffolds to be used in bone transplantation and regeneration. In addition, materials previously limited to the 3D printing (3DP) process due to their own characteristics can also be used well in indirect 3DP. In this study, customised ß-TCP/chitosan scaffolds with the shape of rabbit radial head were produced by indirect 3D printing technology. METHODS: Swelling ability, porosity, mechanical characterisation, and degradation rate analysis were performed, and in vitro studies were also implemented to evaluate the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells (MSCs) on the scaffolds. CCK8 cell proliferation assay kit and alkaline phosphatase (ALP) staining solution were used to study cell proliferation and early ALP content at the scaffold surface. Moreover, the osteogenic differentiation of MSCs on scaffolds was also evaluated through the scanning electron microscopy analysis. RESULTS: ß-TCP/chitosan scaffold has good performance and degradation rate, and in vitro cell experiments also confirm that the scaffold has adequate cytocompatibility and bioactivity. CONCLUSION: This study provides a promising new strategy for the design of customised scaffolds for the repair of complex damaged tissues.


Assuntos
Fosfatos de Cálcio/química , Quitosana/química , Impressão Tridimensional , Rádio (Anatomia)/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Fosfatos de Cálcio/administração & dosagem , Diferenciação Celular/fisiologia , Células Cultivadas , Quitosana/administração & dosagem , Células-Tronco Mesenquimais/química , Células-Tronco Mesenquimais/fisiologia , Impressão Tridimensional/tendências , Coelhos , Rádio (Anatomia)/diagnóstico por imagem , Rádio (Anatomia)/fisiologia
20.
Vet Res ; 50(1): 25, 2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30975214

RESUMO

Staphylococcus aureus is the most commonly isolated pathogen from clinical bovine mastitis samples and a difficult pathogen to combat. Mesenchymal stem cells (MSC) are multipotent progenitor cells equipped with a variety of factors that inhibit bacterial growth. The aim of the present study was to evaluate the in vitro antibacterial potential against S. aureus of conditioned medium (CM) from MSC derived from fetal bovine bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC, AT-MSC and fetal fibroblasts (FB) cultures were activated by infection with S. aureus. Bacterial growth was evaluated in presence of CM, concentrated CM (CCM), activated CM (ACM) and concentrated ACM (CACM) from BM-MSC, AT-MSC and FB. Gene expression of ß-defensin 4A (bBD-4A), NK-lysine 1 (NK1), cathelicidin 2 (CATHL2), hepcidin (HEP) and indoleamine 2,3 dioxygenase (IDO) and protein expression of bBD-4A were determined in activated and non-activated cells. The majority of BM-MSC and AT-MSC expressed CD73, Oct4 and Nanog, and were negative for CD34. Growth of S. aureus decreased when it was exposed to CM from BM-MSC, AT-MSC and FB. Moreover, growth of S. aureus in CCM, ACM and CACM was lower compared to controls of CM from BM-MSC and AT-MSC. Activated AT-MSC increased mRNA levels of bBD4A and NK1, and protein levels of bBD4A in CM. Thus, CM from fetal bovine BM-MSC and AT-MSC has the capacity to reduce in average ~30% of S. aureus relative growth under in vitro conditions. The in vitro antibacterial effect of fetal bovine MSC may be mediated by bBD4A and NK1 activity.


Assuntos
Bovinos/fisiologia , Mastite Bovina/fisiopatologia , Células-Tronco Mesenquimais/fisiologia , Infecções Estafilocócicas/veterinária , Staphylococcus aureus/efeitos dos fármacos , Tecido Adiposo/fisiologia , Animais , Medula Óssea/fisiologia , Feto , Técnicas In Vitro , Infecções Estafilocócicas/fisiopatologia
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