Mechanical Stretch Promotes the Osteogenic Differentiation of Bone Mesenchymal Stem Cells Induced by Erythropoietin

Introduction. The effects of erythropoietin (EPO) on the behaviors of bone marrow mesenchymal stem cells (BMSCs) subjected to mechanical stretch remain unclear. This study was therefore aimed at establishing the dose-response effect of EPO stimulation on rat BMSCs and investigating the effects of mechanical stretch combined with EPO on the proliferation and osteogenic differentiation of BMSCs. Material and Methods. The proliferation and osteogenic differentiation of rat BMSCs were examined and compared using EPO with different concentrations. Thereafter, BMSCs were subjected to 10% elongation using a Flexcell strain unit, combined with 20 IU/ml EPO. The proliferation of BMSCs was detected by Cell Counting Kit-8, colony formation assay, and cell cycle assay; meanwhile, the mRNA expression levels of Ets-1, C-myc, Ccnd1, and C-fos were detected by reverse transcription and real-time quantitative PCR (qPCR). The osteogenic differentiation of BMSCs was detected by alkaline phosphatase (ALP) staining, and the mRNA expression levels of ALP, OCN, COL, and Runx2 were detected by qPCR. The role of the extracellular signal-regulated kinases 1/2 (ERK1/2) in the osteogenesis of BMSCs stimulated by mechanical stretch combined with 20 IU/ml EPO was examined by Western blot. Results. Our results showed that effects of EPO on BMSCs included a dose-response relationship, with the 20 IU/ml EPO yielding the largest. Mechanical stretch combined with 20 IU/ml EPO promoted proliferation and osteogenic differentiation of BMSCs. The increase in ALP, mineral deposition, and osteoblastic genes induced by the mechanical stretch–EPO combination was inhibited by U0126, an ERK1/2 inhibitor. Conclusion. EPO was able to promote the proliferation and osteogenic differentiation of BMSCs, and these effects were enhanced when combined with mechanical stretch. The underlying mechanism may be related to the activation of the ERK1/2 signaling pathway.

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Fak silencing impairs osteogenic differentiation of bone mesenchymal stem cells induced by uniaxial mechanical stretch

Journal of Dental Sciences ◽

10.1016/j.jds.2019.03.001 ◽

2019 ◽

Vol 14 (3) ◽

pp. 225-233 ◽

Cited By ~ 2

Author(s):

Pei Hu ◽

Xiaowen Zhu ◽

Chuang Zhao ◽

Jing Hu ◽

En Luo ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Mechanical Stretch ◽

Bone Mesenchymal Stem Cells

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Knockdown of has_circ_0010452 Promotes Proliferation and Osteogenic Differentiation via Up-Regulating miR-543 in Human Bone Marrow Mesenchymal Stem Cells

Journal of Biomaterials and Tissue Engineering ◽

10.1166/jbt.2022.2966 ◽

2022 ◽

Vol 12 (4) ◽

pp. 770-777

Author(s):

Siyuan Chen ◽

Weixiong Guo ◽

Jinsong Wei ◽

Han Lin ◽

Fengyan Guo

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Human Bone ◽

Human Bone Marrow ◽

Cell Counting ◽

Luciferase Reporter ◽

Expression Levels ◽

Marrow Mesenchymal Stem Cells

Objective: The aim of this study was to explore the role of has_circ_0010452 in the progression of osteoporosis (OP) targeting miR-543, as well as their functions in regulating proliferation and osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs). Methods: The expression levels of circ_0010452 and miR-543 in hBMSCs at different time points of osteogenic differentiation were determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). After transfection of circ_0010452 siRNA or miR-543 inhibitor in hBMSCs, the relative expression levels of osteogenic marker proteins, including oat spelt xylan (OSX), osteocalcin (OCN) and collagen I (Col-1), were determined by western blot. Cell proliferation of hBMSCs was valued by Cell Counting Kit 8 (CCK-8) assay. Dual-Luciferase reporter gene assay was performed to verify the relationship between circ_0010452 and miR-543. Subsequently, the regulatory effects of circ_0010452 and miR-543 on osteogenic differentiation and the capability of mineralization were evaluated by alkaline phosphatase (ALP) determination and alizarin red staining, respectively. Results: The expression of circ_0010452 decreased gradually and miR-543 increased in hBMSCs with the prolongation of osteogenic differentiation. circ_0010452 could bind to miR-543, which was negatively regulated by miR-543 in hBMSCs. Moreover, knockdown of circ_0010452 inhibited proliferation and osteogenic differentiation by upregulating miR-543, as well as upregulating expressions of OSX, OCN and Col-1. Furthermore, knockdown of circ_0010452 markedly promoted the capability of mineralization of hBMSCs, which was further reversed by transfection of miR-543 inhibitor. The knockdown of miR-543 partially reversed the inhibitory effect of circ_0010452 on the osteogenesis of hBMSCs. Conclusions: Silence of circ_0010452 promotes the development of OP via binding to miR-543 regulating proliferation and osteogenic differentiation of hBMSCs, thus promoting the progression of osteoporosis.

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Hsa_circ_0066523 promotes the proliferation and osteogenic differentiation of bone mesenchymal stem cells by repressing PTEN

Bone and Joint Research ◽

10.1302/2046-3758.108.bjr-2020-0127.r2 ◽

2021 ◽

Vol 10 (8) ◽

pp. 526-535

Author(s):

Wei Xin ◽

Shuai Yuan ◽

Bo Wang ◽

Qirong Qian ◽

Yi Chen

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Cell Counting ◽

Luciferase Reporter ◽

Akt Pathway ◽

Bone Mesenchymal Stem Cells ◽

Alizarin Red ◽

Proliferation And Differentiation ◽

The Impact

Aims Circular RNAs (circRNAs) are a novel type of non-coding RNA that plays major roles in the development of diverse diseases including osteonecrosis of the femoral head (ONFH). Here, we explored the impact of hsa_circ_0066523 derived from forkhead box P1 (FOXP1) (also called circFOXP1) on bone mesenchymal stem cells (BMSCs), which is important for ONFH development. Methods RNA or protein expression in BMSCs was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) or western blot, respectively. Cell Counting Kit 8 (CCK8) and 5-ethynyl-2’-deoxyuridine (EdU) were used to analyze cell proliferation. Alkaline phosphatase (ALP) activity, ALP staining, and Alizarin Red S staining were employed to evaluate the osteoblastic differentiation. Chromatin immunoprecipitation (ChIP), luciferase reporter, RNA pull down, and RNA immunoprecipitation (RIP) assays were combined for exploring molecular associations. Results Circ_0066523 was upregulated in osteogenic induction process of BMSCs. Silencing circ_0066523 restrained the proliferation and osteogenic differentiation of BMSCs. Mechanistically, circ_0066523 activated phosphatidylinositol-4,5-bisphosphate 3-kinase / AKT serine/threonine kinase 1 (PI3K/AKT) pathway via recruiting lysine demethylase 5B (KDM5B) to epigenetically repress the transcription of phosphatase and tensin homolog (PTEN). Functionally, AKT signalling pathway agonist or PTEN knockdown counteracted the effects of silenced circ_0066523 on BMSC proliferation and differentiation. Conclusion Circ_0066523 promotes the proliferation and differentiation of BMSCs by epigenetically repressing PTEN and therefore activating AKT pathway. This finding might open new avenues for the identification of therapeutic targets for osteoblast differentiation related diseases such as ONFH. Cite this article: Bone Joint Res 2021;10(8):526–535.

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The 3D-Printed PLGA Scaffolds Loaded with Bone Marrow-Derived Mesenchymal Stem Cells Augment the Healing of Rotator Cuff Repair in the Rabbits

Cell Transplantation ◽

10.1177/0963689720973647 ◽

2020 ◽

Vol 29 ◽

pp. 096368972097364

Author(s):

Peng Chen ◽

Lei Cui ◽

Sai Chuen Fu ◽

Li Shen ◽

Wentao Zhang ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Rotator Cuff ◽

Mrna Expression ◽

Rotator Cuff Repair ◽

Cell Counting ◽

Expression Levels ◽

3D Printed ◽

Cuff Repair

The healing of tendon–bone in the rotator cuff is featured by the formation of the scar tissues in the interface after repair. This study aimed to determine if the 3D-printed poly lactic-co-glycolic acid (PLGA) scaffolds loaded with bone marrow-derived mesenchymal stem cells (BMSCs) could augment the rotator cuff repair in the rabbits. PLGA scaffolds were generated by the 3D-printed technology; Cell Counting Kit-8 assay evaluated the proliferation of BMSCs; the mRNA and protein expression levels were assessed by quantitative real-time polymerase chain reaction and western blot, respectively; immunohistology evaluated the rotator cuff repair; biomechanical characteristics of the repaired tissues were also assessed. 3D-printed PLGA scaffolds showed good biocompatibility without affecting the proliferative ability of BMSCs. BMSCs–PLGA scaffolds implantation enhanced the cell infiltration into the tendon-bone injunction at 4 weeks after implantation and improved the histology score in the tendon tissues after implantation. The mRNA expression levels of collagen I, III, tenascin, and biglycan were significantly higher in the scaffolds + BMSCs group at 4 weeks post-implantation than that in the scaffolds group. At 8 and 12 weeks after implantation, the biglycan mRNA expression level in the BMSCs–PLGA scaffolds group was significantly lower than that in the scaffolds group. BMSCs–PLGA scaffolds implantation enhanced collagen formation and increased collagen dimeter in the tendon–bone interface. The biomechanical analysis showed that BMSCs–PLGA scaffolds implantation improved the biomechanical properties of the regenerated tendon. The combination of 3D-printed PLGA scaffolds with BMSCs can augment the tendon–bone healing in the rabbit rotator cuff repair model.

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Delivery of miRNA-29b Using R9-LK15, a Novel Cell-Penetrating Peptide, Promotes Osteogenic Differentiation of Bone Mesenchymal Stem Cells

BioMed Research International ◽

10.1155/2019/3032158 ◽

2019 ◽

Vol 2019 ◽

pp. 1-12 ◽

Cited By ~ 4

Author(s):

Qiuling Liu ◽

Zhen Lin ◽

Yi Liu ◽

Jiang Du ◽

Hongsheng Lin ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Transfection Efficiency ◽

Cell Counting ◽

Cell Penetrating Peptide ◽

Bone Mesenchymal Stem Cells ◽

Matrix Mineralization ◽

Novel Mirna ◽

Cell Penetrating

Delivery of osteogenesis-promoting microRNAs (miRNAs) is a promising approach to enhance bone regeneration. In this study, we generated nanocomplexes comprising the novel cell-penetrating peptide R9-LK15 and miR-29b and investigated their effects on osteogenic differentiation of bone mesenchymal stem cells (BMSCs). R9-LK15/miR-29b nanocomplexes were prepared and characterized. The transfection efficiency, cell viability, and osteogenic differentiation were investigated. The results showed that R9-LK15 maintained the stability of miR-29b in serum for up to 24 h. Moreover, R9-LK15 efficiently delivered miR-29b into BMSCs; the transfection efficiency was ~10-fold higher than that achieved using Lipofectamine 2000. The Cell Counting Kit-8 assay showed that R9-LK15 and R9-LK15/miR-29b nanocomplexes had negligible cytotoxic effects on BMSCs. Delivery of R9-LK15/miR-29b nanocomplexes promoted osteogenic differentiation of BMSCs and extracellular matrix mineralization by upregulating alkaline phosphatase expression and downregulating histone deacetylase-4 expression. In general, we developed a novel miRNA delivery system that has a high transfection efficiency and promotes osteogenic differentiation.

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Effects of Placenta-Derived Mesenchymal Stem Cells on the Particulate Matter-Induced Damages in Human Middle Ear Epithelial Cells

Stem Cells International ◽

10.1155/2019/4357684 ◽

2019 ◽

Vol 2019 ◽

pp. 1-7

Author(s):

So Young Kim ◽

Seung Ha Oh ◽

Jun Ho Lee ◽

Myung-Whan Suh ◽

Moo Kyun Park

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Particulate Matter ◽

Epithelial Cells ◽

Mrna Expression ◽

Middle Ear ◽

Inflammatory Genes ◽

Expression Levels ◽

Human Middle Ear ◽

Mrna Expression Levels

This study was aimed at investigating the effects of placenta-derived mesenchymal stem cells (PL-MSCs) on particulate matter- (PM-) exposed human middle ear epithelial cells (HMEECs). HMEECs were treated with 300 μg/ml PM for 24 hours. The PL-MSCs were cocultured with PM-treated HMEECs. Cells were harvested on days 0, 1, and 4, and the expression of the inflammatory genes TNFα, COX2, IL1β, IL6, and MUC5B in HMEECs and anti-inflammatory genes PTGES, TGFβ, and VEGF in PL-MSCs was examined by qRT-PCR. The culture media were collected to measure the secreted PGE2 level using an enzyme-linked immunosorbent assay. The mRNA expression of TNFα, COX2, IL1β, IL6, and MUC5B in HMEECs increased following PM treatment. PM-treated HMEECs cocultured with PL-MSCs showed alleviated inflammatory reactions represented by lower mRNA expression levels of MUC5B, TNFα, IL1β, and IL6 compared to monocultured PM-treated HMEECs. The mRNA expression levels of PGE2, TGFβ, and VEGF were elevated in cocultured PL-MSCs compared to those of control PL-MSCs. The medium of PM-treated HMEECs cocultured with PL-MSCs exhibited increased PGE2 levels. The increased inflammatory response in PM-treated HMEECs was reversed using PL-MSCs. The PGE2, TGFβ, and VEGF were the mediators of the anti-inflammatory effects of PL-MSCs.

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TSG Targeting KDM5A affects osteogenic differentiation of bone mesenchymal stem cells induced by bone morphogenetic protein 2

10.1101/2020.07.25.221424 ◽

2020 ◽

Author(s):

Min Wei ◽

Yi Jiang ◽

Yuanqing Huang

Keyword(s):

Stem Cells ◽

Alkaline Phosphatase ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Inhibitory Effect ◽

Bone Morphogenetic Protein 2 ◽

Nodule Formation ◽

Mechanism Study ◽

Bone Mesenchymal Stem Cells ◽

Expression Levels

AbstractTo investigate the effect of 2, 3, 5, 4’-tetrahydroxystilbene-2-O-β-D-glucoside (TSG) on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSC) and its molecular mechanism. The effects of TSG on alkaline phosphatase positive cloning and mineralized nodule formation were also detected. Total mRNA and protein were extracted and effects of TSG on the expression levels of osteopontin (OPN), osteocalcin (OCN), Runt-related transcription factor 2 (Runx2), Osterix and Col1a1 were detected by real-time fluorescence quantitative PCR. Western Blotting was used to detect the inhibitory effect of TSG on KDM5A. BMSCs were transfected with Small interfering RNA (siRNA) targeting KDM5A (si-KMD5A) and pcDNA3.1 KMD5A. TSG significantly increased the activity of ALP and the number of alkaline phosphatase clones and calcified nodule formation. The OPN, OCN, Runx2 and Osterix expression levels were significantly increased among the osteoblasts after TSG treatment. The mechanism study showed that the effect of TSG is realized by inhibiting KDM5A. KDM5A signaling may be involved in the regulation of osteogenic differentiation of rBMSC. TSG can promote osteogenic differentiation and maturation of rBMSC at 0.1-50 μmol / L. The mechanism of action was realized by inhibiting the expression of KDM5A.

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