scholarly journals Adipose-Derived Stem Cells Conditioned Media Promote In Vitro Osteogenic Differentiation of Hypothyroid Mesenchymal Stem Cells

2020 ◽  
Vol 7 (3) ◽  
Author(s):  
Tayebeh Sanchooli ◽  
Mohsen Norouzian ◽  
Mahtab Teimouri ◽  
Abdolreza Ardeshirylajimi ◽  
Abbas Piryaei
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Calcium Content ◽  
Male Rats ◽  
Osteogenic Potential ◽  
Significant Difference

Background: Thyroid hormones have many effects on the physiological functions of cells, including growth, differentiation, and metabolism. Objectives: Recently, studies have shown that the adipose-derived mesenchymal stem cells conditioned medium (ADMSCs-CM) has many osteogenic factors, such as IGF-1, IL-6, and FGFs. Methods: In the current study, mesenchymal stem cells (MSCs) were isolated from two sources; the adipose tissue of the testicular fat pad and the bone marrow of rat, and then characterized by flow cytometry. ADMSCs-CM was collected from the ADMSC in the healthy adult male rats. Hypothyroidism was induced by the administration of the Methimazole during 60 days and confirmed by the analysis of the serum level of T4 and TSH hormones. Cell proliferation and osteogenic differentiation potential of bone marrow stem cells (BMSCs) derived from hypothyroid rats were investigated in the presence and absence of the CM by MTT assay, alkaline phosphatase (ALP) activity, calcium content assay, and bone-related gene expression. Healthy BMSCs were assigned to the control group. Results: Although Cell proliferation was decreased in the hypothyroid BMSCs, there was no significant difference between the control and the hypothyroid-CM groups. Similarly, osteogenic potential was significantly reduced in the hypothyroid group compared to the control and hypothyroid-CM groups according to the ALP, calcium content assays, and gene expression results. There was no significant difference between the hypothyroid-CM group and control. Conclusions: Our results indicated that hypothyroidism can decrease cell proliferation and osteogenic differentiation of BMSCs. Although ADMSCs-CM improved these parameters, it may be a promising candidate for the bone regeneration of the hypothyroidism cases.

scholarly journals MicroRNA treatment modulates osteogenic differentiation potential of mesenchymal stem cells derived from human chorion and placenta

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kulisara Marupanthorn ◽  
Chairat Tantrawatpan ◽  
Pakpoom Kheolamai ◽  
Duangrat Tantikanlayaporn ◽  
Sirikul Manochantr
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Transcription Factor ◽  
Osteogenic Differentiation ◽  
Differentiation Potential ◽  
Osteogenic Gene Expression ◽  
Osteogenic Gene

AbstractMesenchymal stem cells (MSCs) are important in regenerative medicine because of their potential for multi-differentiation. Bone marrow, chorion and placenta have all been suggested as potential sources for clinical application. However, the osteogenic differentiation potential of MSCs derived from chorion or placenta is not very efficient. Bone morphogenetic protein-2 (BMP-2) plays an important role in bone development. Its effect on osteogenic augmentation has been addressed in several studies. Recent studies have also shown a relationship between miRNAs and osteogenesis. We hypothesized that miRNAs targeted to Runt-related transcription factor 2 (Runx-2), a major transcription factor of osteogenesis, are responsible for regulating the differentiation of MSCs into osteoblasts. This study examines the effect of BMP-2 on the osteogenic differentiation of MSCs isolated from chorion and placenta in comparison to bone marrow-derived MSCs and investigates the role of miRNAs in the osteogenic differentiation of MSCs from these sources. MSCs were isolated from human bone marrow, chorion and placenta. The osteogenic differentiation potential after BMP-2 treatment was examined using ALP staining, ALP activity assay, and osteogenic gene expression. Candidate miRNAs were selected and their expression levels during osteoblastic differentiation were examined using real-time RT-PCR. The role of these miRNAs in osteogenesis was investigated by transfection with specific miRNA inhibitors. The level of osteogenic differentiation was monitored after anti-miRNA treatment. MSCs isolated from chorion and placenta exhibited self-renewal capacity and multi-lineage differentiation potential similar to MSCs isolated from bone marrow. BMP-2 treated MSCs showed higher ALP levels and osteogenic gene expression compared to untreated MSCs. All investigated miRNAs (miR-31, miR-106a and miR148) were consistently downregulated during the process of osteogenic differentiation. After treatment with miRNA inhibitors, ALP activity and osteogenic gene expression increased over the time of osteogenic differentiation. BMP-2 has a positive effect on osteogenic differentiation of chorion- and placenta-derived MSCs. The inhibition of specific miRNAs enhanced the osteogenic differentiation capacity of various MSCs in culture and this strategy might be used to promote bone regeneration. However, further in vivo experiments are required to assess the validity of this approach.

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.

scholarly journals Alpha-5 Integrin Mediates Simvastatin-Induced Osteogenesis of Bone Marrow Mesenchymal Stem Cells

2019 ◽  
Vol 20 (3) ◽  
pp. 506 ◽  
Author(s):  
Pei-Lin Shao ◽  
Shun-Cheng Wu ◽  
Zih-Yin Lin ◽  
Mei-Ling Ho ◽  
Chung-Hwan Chen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Protein Expression ◽  
Osteogenic Differentiation ◽  
Calcium Deposition ◽  
Marker Genes ◽  
Expression Levels ◽  
Gene Expression Levels

Simvastatin (SVS) promotes the osteogenic differentiation of mesenchymal stem cells (MSCs) and has been studied for MSC-based bone regeneration. However, the mechanism underlying SVS-induced osteogenesis is not well understood. We hypothesize that α5 integrin mediates SVS-induced osteogenic differentiation. Bone marrow MSCs (BMSCs) derived from BALB/C mice, referred to as D1 cells, were used. Alizarin red S (calcium deposition) and alkaline phosphatase (ALP) staining were used to evaluate SVS-induced osteogenesis of D1 cells. The mRNA expression levels of α5 integrin and osteogenic marker genes (bone morphogenetic protein-2 (BMP-2), runt-related transcription factor 2 (Runx2), collagen type I, ALP and osteocalcin (OC)) were detected using quantitative real-time PCR. Surface-expressed α5 integrin was detected using flow cytometry analysis. Protein expression levels of α5 integrin and phosphorylated focal adhesion kinase (p-FAK), which is downstream of α5 integrin, were detected using Western blotting. siRNA was used to deplete the expression of α5 integrin in D1 cells. The results showed that SVS dose-dependently enhanced the gene expression levels of osteogenic marker genes as well as subsequent ALP activity and calcium deposition in D1 cells. Upregulated p-FAK was accompanied by an increased protein expression level of α5 integrin after SVS treatment. Surface-expressed α5 integrin was also upregulated after SVS treatment. Depletion of α5 integrin expression significantly suppressed SVS-induced osteogenic gene expression levels, ALP activity, and calcium deposition in D1 cells. These results identify a critical role of α5 integrin in SVS-induced osteogenic differentiation of BMSCs, which may suggest a therapeutic strategy to modulate α5 integrin/FAK signaling to promote MSC-based bone regeneration.

scholarly journals Irradiation Haematopoiesis Recovery Orchestrated by IL-12/IL-12Rβ1/TYK2/STAT3-Initiated Osteogenic Differentiation of Mouse Bone Marrow-Derived Mesenchymal Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Fengjie Li ◽  
Rong Zhang ◽  
Changpeng Hu ◽  
Qian Ran ◽  
Yang Xiang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Osteogenic Potential ◽  
Mouse Bone Marrow ◽  
Hematopoietic Stem ◽  
Stat3 Signaling ◽  
Tyrosine Kinase 2 ◽  
Early Use

PurposeRepairing the irradiation-induced osteogenic differentiation injury of bone marrow mesenchymal stem cells (BM-MSCs) is beneficial to recovering haematopoiesis injury in radiotherapy; however, its mechanism is elusive. Our study aimed to help meet the needs of understanding the effects of radiotherapy on BM-MSC osteogenic potential.Methods and MaterialsBalb/c mice and the BM-MSCs were used to evaluate the irradiation-induced osteogenic differentiation injury in vivo. The cellular and molecular characterization were applied to determine the mechanism for recovery of irradiation-derived haematopoiesis injuries.ResultsWe report a functional role of IL-12 in acute irradiation hematopoietic injury recovery and intend to dissect the possible mechanisms through BM-MSC, other than the direct effect of IL-12 on hematopoietic stem and progenitor cells (HSPCs). Specifically, we show that early use of IL-12 enhanced the osteogenic differentiation of BM-MSCs through IL-12Rβ1/TYK2/STAT3 signaling; furthermore, IL-12 induced osteogenesis facilitated bone formation and irradiation hematopoiesis recovery when transplanted BM-MSCs in the femur of Balb/c mice. For the mechanism of action, we found that IL-12 receptor beta 1 (IL-12Rβ1) expression of irradiated BM-MSCs was upregulated rapidly, coincidentally consistent with early use of IL-12 induced osteogenic differentiation enhancement. IL-12Rβ1 and tyrosine kinase 2 gene (Tyk2) silencing experiments and phosphotyrosine of signal transducer and activator of transcription 3 (p-STAT3) suppression experiments indicated the IL-12Rβ1/TYK2/STAT3 signaling was essential in IL-12-induced osteogenic differentiation enhancement of BM-MSCs.ConclusionThese findings suggested that IL-12 may exert BM-MSCs-based hematopoietic recovery by repairing osteogenic differentiation abilities damages through IL-12Rβ1/TYK2/STAT3 signaling pathway post-irradiation.

scholarly journals Thrombin-activated platelet rich plasma (PRP) enhanced osteogenic differentiation of bone marrow mesenchymal stem cells by activing autophagy through miR-140-3p/SPRED2 axis

2020 ◽  
Author(s):  
Shuting Jiang ◽  
Hongyan Liu ◽  
Weiyan Zhu ◽  
Hui Yan ◽  
Beizhan Yan
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Alizarin Red ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

Abstract Background Mesenchymal stem cells transplantation gradually become a potential treatment for bone defect in clinic practice. This study aimed to investigate the molecular mechanism of PRP and autophagy for osteogenic differentiation in bone marrow mesenchymal stem cells (BMSCs). Methods Thrombin activated PRP was prepared and the BMSCs were treated with activated PRP with different concentration and transfected with miR-140-3p vector (mimics or inhibitor), si-SPRED2 or co-transfected with miR-140-3p inhibitor and si-SPRED2, respectively. qRT-PCR and Western blotting were used to determine the mRNA expression and protein expression. A luciferase reporter assay was conducted to identified the targeting relationship between iR-140-3p and SPRED2 Subsequently, cell proliferation was detected by MTT and ALP activity was also determined. Alizarin red staining was used for the evaluating the formation of calcium nodules. Results MiR-140-3p expression was found to be inhibited by PRP in a dose-dependent manner, besides, cell proliferation, ALP activity, the expression of COL-I, OPN, Runx2 and OCN, and the formation of calcium nodules related to osteogenic differentiation were enhanced by PRP. Subsequently, we found that PRP activated autophagy and up-regulated SPRED2 expression in BMSCs through suppressing miR-140-3p expression. Moreover, we confirmed that miR-140-3p targeted SPRED2 and negatively regulation its expression. Finally, the findings showed that inhibition of miR-140-3p enhanced cell proliferation, osteogenic differentiation and autophagy of BMSCs by negatively regulating SPRED2 expression. Conclusion Thrombin activated PRP accelerated osteogenic differentiation of BMSCs by activing autophagy through miR-140-3p/SPRED2 axis.

Effect of Chordin-Like 1 on Differentiation of Bone Marrow Mesenchymal Stem Cells Through Wnt5/TGF-β Signaling Pathway

2019 ◽  
Vol 9 (9) ◽  
pp. 1304-1310
Author(s):  
Qing Yang ◽  
Lei Wu ◽  
Yang Liu ◽  
Bing Yuan
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Real Time Pcr ◽  
Caspase 3 ◽  
Control Group ◽  
Alp Activity ◽  
Marrow Mesenchymal Stem Cells

Chordin-like 1 (CHRDL1) functions in multiple tissues and organs. However, whether CHRDL1 affects bone marrow mesenchymal stem cells (BMSCs) differentiation remain unclear. Rat BMSCs were isolated and divided into control group, CHRDL1 group and CHRDL1 siRNA group followed by analysis of CHRDL1 level by real time PCR and ELISA, cell proliferation by MTT assay, Caspase 3 activity, ALP activity, expression of o Runx2, OC and PPARγ2 by Real time PCR, TGF-β secretion by ELIS, and Wnt5 protein expression by Western blot. CHRDL1 expression was significantly increased in CHRDL1 group, along with significantly promoted cell proliferation, decreased Caspase 3 activity, increased ALP activity and expression of Runx2 and OC, decreased PPARγ2 expression, increased TGF-β secretion and Wnt5 expression compared to control group (P < 0.05). However, CHRDL1 siRNA transfection significantly decreased CHRDL1 expression, inhibited cell proliferation, increased Caspase 3 activity, decreased ALP activity and Runx2 and OC expression, increased PPARγ2 expression, decreased TGF-β secretion and Wnt5 expression. (P < 0.05). Down-regulation of CHRDL1 expression in BMSCs promotes Wnt5/TGF-β signaling transduction, which in turn increases BMSCs proliferation and osteogenic differentiation. Up-regulation of CHRDL1 expression in BMSCs inhibited the activation of Wnt5/TGF-β signaling pathway, promoted BMSCs apoptosis, and inhibited BMSCs proliferation and osteogenic differentiation.

Bmi1 Induces Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Under Inflammatory Environment

2019 ◽  
Vol 9 (12) ◽  
pp. 1783-1789
Author(s):  
Chungang Dong ◽  
Junyu Wei
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Proliferation ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Real Time Pcr ◽  
Alp Activity ◽  
Bmi1 Expression ◽  
Tnf Α ◽  
Marrow Mesenchymal Stem Cells

Bmi1 is a polycomb histone that regulates stem cells, but the role and mechanism of Bmi1 in bone marrow mesenchymal stem cells (BMSCs) differentiation has not been elucidated. Rat BMSCs were cultured in vitro and randomly divided into control group and inflammation group (treated with LPS). Bmi1 and Bmi1 siRNA were transfected into inflammatory BMSCs, followed by analysis of Bmi1 expression by Real time PCR, cell proliferation by MTT assay, Caspase3 activity, ALP activity, expression of Runx2, OP and PPARγ 2 by Real time PCR, as well as secretion of TNF-α and IL-1β by ELISA. In inflammatory environment, Bmi1 expression was significantly decreased, cell proliferation was significantly inhibited, along with increased Caspase3 activity, decreased ALP activity and the expression of Runx2 and OP, increased PPAR 2 expression and secretion of TNF-α and IL-1β (P < 0 05). Transfection of Bmi1 siRNA into inflammatory BMSCs further significantly aggravated the above changes (P < 0 05). Bmi1 plasmid transfected into inflammatory BMSCs significantly promoted Bmi1 expression and cell proliferation, decreased Caspase3 activity, increased ALP activity and expression of Runx2 and OP, decreased PPAR γ2 expression and TNF-α and IL-1β secretion (P < 0 05). Bmi1 expression is reduced in BMSCs under inflammation. Up-regulation of Bmi1 can inhibit the secretion of inflammatory factors, regulate the proliferation and apoptosis of BMSCs, and promote the proliferation and osteogenic differentiation of BMSCs.

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