scholarly journals A Comparative Study on the Effect of Exosomes Secreted by Mesenchymal Stem Cells Derived from Adipose and Bone Marrow Tissues in the Treatment of Osteoarthritis-Induced Mouse Model

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Hoda Fazaeli ◽  
Naser Kalhor ◽  
Leila Naserpour ◽  
Faezeh Davoodi ◽  
Mohsen Sheykhhasan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Real Time ◽  
Real Time Pcr ◽  
Histological Analysis ◽  
Collagen Type ◽  
Therapeutic Option ◽  
Health Condition ◽  
Histological Staining

Background. Exosomes as extracellular vesicles (EVs) are nanoscale intercellular messengers secreted from cells to deliver biological signals. Today, exosomes have become a new field of research in regenerative medicine and are considered as potential therapies to control inflammation and wound healing and enhance and improve healing in many diseases. Given the global burden of osteoarthritis (OA) as the fastest-growing health condition and one of the major causes of physical disability in the aging population, research to establish EVs as therapeutic products can meet the basic clinical needs in the management of osteoarthritis and provide a therapeutic solution. Objectives. The present study is aimed at evaluating the regenerative potentials of the exosomes secreted from adipose and bone marrow tissue-derived mesenchymal stem cells (AD- and BM-MSCs) in ameliorating the symptoms of OA. Method. In this experimental study, AD- and BM-MSCs were isolated and cultured in the laboratory until passage 3. Finally, these cells’ secreted exosomes were isolated from their conditioned medium. Ciprofloxacin-induced OA mouse models underwent intra-articular injection of exosomes from AD-MSCs and BM-MSCs. Finally, the expression levels of collagen I and II, sox9, and aggrecan genes using real-time PCR, histological analysis, and immunohistochemical (IHC) studies were performed. Results. Real-time PCR data showed that although the expression level of collagen type II was lower in both exosome-treated groups than the normal, but it was significantly increased in comparison with the sham and OA, with higher expression in BM-Exo rather than AD-Exo group. Similarly, the histological staining and IHC results have provided almost identical data, emphasizing on better therapeutic effect of BM-MSCs-exosome than AD-MSCs-exosome. Conclusion. BM-MSCs secreted exosomes in comparison with AD-MSCs could be considered as a better therapeutic option to improve osteoarthritis and exhibit potential as a disease-modifying osteoarthritis cell-free product.

Effect of αCGRP on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Through Smad/Runx2 Signaling Pathway

2020 ◽  
Vol 10 (6) ◽  
pp. 868-873
Author(s):  
Shengxiang Huang ◽  
Haibo Mei ◽  
Rongguo He ◽  
Kun Liu ◽  
Jin Tang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Real Time Pcr ◽  
Caspase 3 ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

The α-calcitonin gene-related peptide (α-CGRP) regulates bone metabolism and has potential applications in enhancing bone remodeling in vivo. However, α-CGRP's role in bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation remain unclear. Rat BMSCs were separated into control group, α-CGRP group and α-CGRP siRNA group, in which BMSCs were transfected with α-CGRP plasmid and α-CGRP siRNA respectively followed by analysis of α-CGRP level by real time PCR and ELISA, cell proliferation by MTT assay, Caspase 3 activity, ALP activity, formation of calcified nodules by alizarin red staining, Smad1 and Smad7 level by Western blot and Runx2 by real time PCR. αCGRP transfection into BMSCs significantly up-regulated CGRP, which could promote cell proliferation, inhibit Caspase 3 activity, promote ALP activity, increase calcified nodules formation and upregulate Smad1, Smad7 and Runx2 compared to control (P < 0.05); transfection of αCGRP siRNA significantly down-regulated CGRP in BMSCs, inhibited cell proliferation, promoted Caspase 3 activity, inhibited ALP activity, inhibited calcified nodules formation and downregulate Smad1, Smad7 and Runx2 (P < 0.05). αCGRP overexpression promotes the Smad/Runx2 signaling, which in turn promotes BMSCs proliferation and osteogenesis. Decreased αCGRP level inhibits Smad/Runx2 signaling, promotes BMSCs apoptosis, inhibits proliferation and osteogenic differentiation.

Effect of SOX9 on Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Through WNTβ/Catenin Pathway

2019 ◽  
Vol 9 (10) ◽  
pp. 1429-1434
Author(s):  
Qing Yang ◽  
Cheng Li ◽  
Manli Yan ◽  
Chunhua Fang
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Protein Expression ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Real Time Pcr ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

Bone marrow mesenchymal stem cells (BMSCs) can be differentiated into different types of cells. SOX9 involves in the development and progression of various diseases. Our study aims to assess SOX9's effect on osteogenic differentiation of BMSCs and its related regulatory mechanisms. Rat BMSCs were isolated and randomly divided into control group, SOX9 group and SOX9 siRNA group, which was transfected with pcDNA-SOX9 plasmid or SOX9 siRNA respectively followed by analysis of SOX9 expression by Real time PCR, cell proliferation by MTT assay, Caspase3 and ALP activity, GSK-3β expression and Wntβ/Catenin Signaling pathway protein expression by Western blot, and expression of osteogenic genes Runx2 and BMP-2 by Real time PCR. Transfection of pcDNA-SOX9 plasmid into BMSCs significantly inhibited cell proliferation, promoted Caspase3 activity, decreased ALP activity and downregulated Runx2 and BMP-2, increased GSK-3β expression and decreased Wntβ/Catenin expression protein expression (P< 0.05). SOX9 siRNA transfection significantly promoted cell proliferation, inhibited Caspase3 activity, increased ALP activity and upregulated Runx2 and BMP-2, downregulated GSK-3β and increased Wntβ/Catenin expression. SOX9 regulates BMSCs proliferation and osteogenic differentiation through Wntβ/Catenin signaling pathway.

scholarly journals Osteogenic Differentiation of Pre-Conditioned Bone Marrow Mesenchymal Stem Cells with Nisin on the Modified Poly-L-Lactic-Acid Nanofibers

2021 ◽  
Author(s):  
Fariba Sadraei ◽  
Marzieh Ghollasi ◽  
Fatemeh Khakpai ◽  
Raheleh Halabian
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Lactic Acid ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Real Time Pcr ◽  
Alizarin Red ◽  
Marrow Mesenchymal Stem Cells

Abstract Background: Human bone marrow-derived mesenchymal stem (MSCs) cells are undifferentiated cells with the self-renewing ability and multi-lineage differentiation beneficial for regenerative medicine. Nano scaffolds are novel materials employed in bone repair and regeneration. Nisin is a prebiotic that can increase stem cells’ life span and proliferation. This study attempted to provide a proper strategy for bone marrow mesenchymal stem cells differentiation into the Osteocytes on a Poly‐L‐lactic‐acid scaffold (PLLA) after pretreating with probiotic Nisin. Methods: MSC osteogenic differentiation was evaluated by measuring Calcium, Alkaline phosphatase, and quantitative tests such as Real-Time PCR, Acridine Orange, Alizarin Red, Von Kossa, and others. Results: The result of the MTT test showed that the optimal dose of Nisin probiotic for the MSCs’ preconditioning was 200 IU/mL on the 1st, 3rd, and 5th days of culture. Real-time PCR data indicated that the expression rate of ALP, Osteonectin, Osteocalcin, and Collagen I have increased in the presence of Nisin, while the RUNX-2 gene expression has decreased. Furthermore, the results of Alizarin Red and Von Kossa tests, as well as Scanning electron microscopy (SEM), revealed that the cell proliferation in the preconditioned samples with Nisin increased significantly. Conclusions: The study concluded that the cell proliferation and differentiation increased in samples pretreated with Nisin on the PLLA Nano scaffolds.

Down Regulation of GATA-2 and the Inversely-Correlated Expression of Adipogenic Specific Gene-PPARγ in Mesenchymal Stem Cells from Patients with Aplastic Anemia.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1414-1414
Author(s):  
Yinyan Xu ◽  
Yoshiyuki Takahashi ◽  
Yue Wang ◽  
Hirokazu Hidaka ◽  
Hiroshi Yagasaki ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Aplastic Anemia ◽  
Cell Line ◽  
Real Time ◽  
Real Time Pcr ◽  
Specific Gene ◽  
Quantitative Real Time Pcr ◽  
Normal Individuals

Abstract (Introduction) Aplastic anemia (AA) is characterized by a reduced number of hematopoietic stem cells resulting in peripheral pancytopenia and fatty marrow replacement. Although immune-mediated suppression of hematopoiesis is considered to play an important role in most patients with AA, the mechanism of fatty marrow replacement remains unclear. Transcription factor GATA-2 is known to play important roles in hematopoiesis. Gene Chip analysis has implicated the decreased expression of GATA-2 in AA CD34 cells, a finding confirmed by quantitative real-time PCR analysis. In a mouse stromal preadipocyte cell line, the suppression of GATA-2 expression accelerated the differentiation of stromal preadipocytes. In order to explore the impact of GATA-2 expression on adipogenesis in AA, the expression of GATA-2, the adipogenic specific gene adipsin, CCAAT/enhancer binding protein alpha (C/EBPα), and peroxisome proliferator-activated receptor γ (PPARγ) was examined in mesenchymal stem cells (MSCs) derived from bone marrow from AA patients and normal individuals. (Patients and methods) Thirty-four AA patients and 15 healthy individuals were included in this study. The median patient age was 10 years, ranging from 2 to 21 years. Twelve patients were studied before immunosuppressive therapy (IST) or bone marrow transplantation. Twenty-two patients were studied after treatment, of whom 11 responded to IST and 11 did not respond. Mononuclear cells were separated from bone marrow aspirates and 1x107 cells used for MSCs culture. MesenCult (StemCell Technologies, USA) was used to culture and expand the MSCs in vitro, after which the cells retain their ability to differentiate along various lineages upon the addition of the appropriate stimulatory supplements. Total RNA was extracted from the third passage of MSCs and synthesis of cDNA was performed using the Thermoscript RT-PCR system. Quantitative real-time PCR was conducted using Taqman primers and probes. The K562 cell line for GATA-2, the EOL1 cell line for adipsin and the MOLM-13 cell line for C/EBPα and PPARγ were used as controls. (Results) GATA-2 expression by MSCs from patients with AA at diagnosis was significantly decreased compared with normal individuals (p=0.031). However, one of the adipogenic specific genes, PPARγ, was significantly increased in MSCs in AA at diagnosis (p=0.017). The expression of adipsin and C/EBPα in MSCs did not differ significantly between AA patients and healthy volunteers. Notably, in the 22 AA patients who received IST, the expression of GATA-2 and PPARγ did not significantly decrease if the patient had responded and hematopoiesis had improved (n=11). However, after the IST the 11 non-responders still had significantly lower GATA-2 and higher PPARγ expression compared to normal individuals. (Conclusion) The mechanism of fatty replacement in the bone marrow in AA may be explained by the down regulation of GATA-2 and the inversely-correlated expression of PPARγ in MSCs. The decreased expression of GATA-2 may be responsible for the pathogenesis and development of the clinical features of this disease.

scholarly journals The Osteogenic Differentiation of Mice Bone Marrow Mesenchymal Stem Cells with Fisetin Phytoestrogen

2017 ◽  
Vol 7 (1) ◽  
pp. 162
Author(s):  
Elahe Vadaye Kheiry ◽  
Kazem Parivar ◽  
Javad Baharara ◽  
Alireza Iranbakhsh
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Real Time Pcr ◽  
Activity Measurement ◽  
Specific Marker ◽  
Alizarin Red ◽  
Marrow Mesenchymal Stem Cells

Objective(s): The aim of this study was to evaluate the effects of fisetin to promote osteogenic differentiation in mice bone marrow mesenchymal stem cells (BMSCs).Materials and Methods: In this study cytotoxicity and viability of fisetin was measured by MTT assay. The differentiation effects of fisetin on BMSCs into osteoblast was assessed with alkaline phosphatase (ALP) activity measurement. Alizarin red staining and Real time PCR for osteoblast specific marker, Osteocalcin (OCN) ,Osteopontin (OPN), Runt-related transcription factor 2 (RUNX2), ERK and MAPK were investigatedResults: The results showed that fisetin does not have toxicity effect on BMSCs and it causes cell proliferation; hence 200, 400 and 800 µg/ml of fisetin was selected for the assessment of differentiation progress. Alizarin red staining (ARS) showed that fisetin promotes osteogenic differentiation on BMSCs at 21st day; dependently also higher alkaline phosphates activity was observed in the treatment groups of 10 days culture, compared to the control groups. The evaluation of Real time -PCR result evaluated showed that OCN OPN, RUNX2,ERK and MAPK genes expressions were increased.Conclusion: The results of this method, showed that differentiation in bone marrow stem cells took place through p38 MAPK, and ERK1 gene activation.

scholarly journals TOB1 Deficiency Enhances the Effect of Bone Marrow-Derived Mesenchymal Stem Cells on Tendon-Bone Healing in a Rat Rotator Cuff Repair Model

2016 ◽  
Vol 38 (1) ◽  
pp. 319-329 ◽  
Author(s):  
Yulei Gao ◽  
Yinquan Zhang ◽  
Yanghu Lu ◽  
Yi Wang ◽  
Xingrui Kou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Rotator Cuff ◽  
Bone Healing ◽  
Rotator Cuff Repair ◽  
Collagen Type ◽  
Collagen Type I ◽  
Type I ◽  
Cuff Repair

Background/Aims: This study investigated the effect of silencing TOB1 (Transducer of ERBB2, 1) expression in bone marrow-derived mesenchymal stem cells (MSCs) on MSC-facilitated tendon-bone healing in a rat supraspinatus repair model. Methods: Rat MSCs were transduced with a recombinant lentivirus encoding short hairpin RNA (shRNA) against TOB1. MSC cell proliferation was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. The effect of MSCs with TOB1 deficiency on tendon-bone healing in a rat rotator cuff repair model was evaluated by biomechanical testing, histological analysis and collagen type I and II gene expression. An upstream regulator (miR-218) of TOB1 was determined in MSCs. Results: We found that knockdown of TOB1 significantly increased the proliferative activity of rat MSCs in vitro. When MSCs with TOB1 deficiency were injected into injured rat supraspinatus tendon-bone junctions, the effect on tendon-bone healing was enhanced compared to treatment with control MSCs with normal TOB1 expression, as evidenced by elevated levels of ultimate load to failure and stiffness, increased amount of fibrocartilage and augmented expression of collagen type I and type II genes. In addition, we found that the TOB1 3′ untranslated region is a direct target of miR-218. Similar to the effect of TOB1 deficiency, overexpression of miR-218 effectively promoted tendon-bone healing in rat. Conclusion: These results suggest that TOB1 may play a negative role in the effect of MSCs on tendon-bone healing, and imply that expression of TOB1 may be regulated by miR-218.

scholarly journals Conditioned Medium from Human Tonsil-Derived Mesenchymal Stem Cells Enhances Bone Marrow Engraftment via Endothelial Cell Restoration by Pleiotrophin

Cells ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 221
Author(s):  
Yu-Hee Kim ◽  
Kyung-Ah Cho ◽  
Hyun-Ji Lee ◽  
Minhwa Park ◽  
Sang-Jin Shin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Conditioned Medium ◽  
Histological Analysis ◽  
Conditioning Regimen ◽  
Mouse Bone Marrow ◽  
Human Tonsil ◽  
Hematopoietic Stem ◽  
Bone Marrow Engraftment

Cotransplantation of mesenchymal stem cells (MSCs) with hematopoietic stem cells (HSCs) has been widely reported to promote HSC engraftment and enhance marrow stromal regeneration. The present study aimed to define whether MSC conditioned medium could recapitulate the effects of MSC cotransplantation. Mouse bone marrow (BM) was partially ablated by the administration of a busulfan and cyclophosphamide (Bu–Cy)-conditioning regimen in BALB/c recipient mice. BM cells (BMCs) isolated from C57BL/6 mice were transplanted via tail vein with or without tonsil-derived MSC conditioned medium (T-MSC CM). Histological analysis of femurs showed increased BM cellularity when T-MSC CM or recombinant human pleiotrophin (rhPTN), a cytokine readily secreted from T-MSCs with a function in hematopoiesis, was injected with BMCs. Microstructural impairment in mesenteric and BM arteriole endothelial cells (ECs) were observed after treatment with Bu–Cy-conditioning regimen; however, T-MSC CM or rhPTN treatment restored the defects. These effects by T-MSC CM were disrupted in the presence of an anti-PTN antibody, indicating that PTN is a key mediator of EC restoration and enhanced BM engraftment. In conclusion, T-MSC CM administration enhances BM engraftment, in part by restoring vasculature via PTN production. These findings highlight the potential therapeutic relevance of T-MSC CM for increasing HSC transplantation efficacy.

scholarly journals Biological Characteristics and Osteogenic Differentiation of Ovine Bone Marrow Derived Mesenchymal Stem Cells Stimulated with FGF-2 and BMP-2

2020 ◽  
Vol 21 (24) ◽  
pp. 9726
Author(s):  
Sandra Gromolak ◽  
Agnieszka Krawczenko ◽  
Agnieszka Antończyk ◽  
Krzysztof Buczak ◽  
Zdzisław Kiełbowicz ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Collagen Type ◽  
Collagen Type I ◽  
Biological Characteristics ◽  
Immunofluorescent Staining ◽  
Type I ◽  
Differentiation Potential

Cell-based therapies using mesenchymal stem cells (MSCs) are a promising tool in bone tissue engineering. Bone regeneration with MSCs involves a series of molecular processes leading to the activation of the osteoinductive cascade supported by bioactive factors, including fibroblast growth factor-2 (FGF-2) and bone morphogenetic protein-2 (BMP-2). In this study, we examined the biological characteristics and osteogenic differentiation potential of sheep bone marrow MSCs (BM-MSCs) treated with 20 ng/mL of FGF-2 and 100 ng/mL BMP-2 in vitro. The biological properties of osteogenic-induced BM-MSCs were investigated by assessing their morphology, proliferation, phenotype, and cytokine secretory profile. The osteogenic differentiation was characterized by Alizarin Red S staining, immunofluorescent staining of osteocalcin and collagen type I, and expression levels of genetic markers of osteogenesis. The results demonstrated that BM-MSCs treated with FGF-2 and BMP-2 maintained their primary MSC properties and improved their osteogenic differentiation capacity, as confirmed by increased expression of osteocalcin and collagen type I and upregulation of osteogenic-related gene markers BMP-2, Runx2, osterix, collagen type I, osteocalcin, and osteopontin. Furthermore, sheep BM-MSCs produced a variety of bioactive factors involved in osteogenesis, and supplementation of the culture medium with FGF-2 and BMP-2 affected the secretome profile of the cells. The results suggest that sheep osteogenic-induced BM-MSCs may be used as a cellular therapy to study bone repair in the preclinical large animal model.

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