Development of a simvastatin loaded injectable porous scaffold in situ formed by phase inversion method for bone tissue regeneration

2018 ◽  
Vol 42 (2) ◽  
pp. 72-79 ◽  
Author(s):  
Mina Hajihasani Biouki ◽  
Hamid Mobedi ◽  
Akbar Karkhaneh ◽  
Morteza Daliri Joupari
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Osteogenic Differentiation ◽  
Glycolic Acid ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Injectable Scaffolds ◽  
Interconnected Pores

Introduction: The use of injectable scaffolds as a minimally invasive method is a good choice in tissue engineering applications. A critical parameter for the tissue engineering scaffolds is a suitable morphology with interconnected pores. We present the development of a simvastatin loaded scaffold that forms in situ and provides the porous structure with interconnected pores. Methods: The formulation of these scaffolds includes a polymeric solution of poly lactic-co-glycolic acid (25 wt%) in N-methyl-2-pyrrolidone containing 6 wt% deionized water and porogen (mannitol, four times the weight of the polymer). We have grafted simvastatin to poly lactic-co-glycolic acid by the esterification reactions. Simvastatin or simvastatin-grafted poly lactic-co-glycolic acid in different levels was added to polymer solution and finally the solution was injected into phosphate buffered saline. The simvastatin-grafted poly lactic-co-glycolic acid was characterized by attenuated total reflection Fourier-transform infra-red and 1H-nuclear magnetic resonance spectroscopy. The morphology, porosity, and biocompatibility of the scaffolds were evaluated. The in vitro simvastatin release from the various formulations was studied. Osteogenic differentiation of the adipose-derived stem cells was investigated using alkaline phosphatase activity assay and cell mineralization was evaluated using Alizarin red staining. Results: The morphology results showed the resultant scaffold was porous with the interconnected pores. The scaffolds presented 91% porosity. Non-toxic doses of simvastatin in the scaffolds were determined by methyl-thiazolyl diphenyl-tetrazolium bromide assay. The released simvastatin from the scaffolds continues over 80 days. Alkaline phosphatase activity and Alizarin red results indicated that cell osteogenic differentiation is promoted. Conclusion: The results demonstrated that release of simvastatin from the injectable scaffolds can have positive effects on osteogenic differentiation of the adipose-derived stem cells.

scholarly journals Composite Hydrogel of Methacrylated Hyaluronic Acid and Fragmented Polycaprolactone Nanofiber for Osteogenic Differentiation of Adipose-Derived Stem Cells

Pharmaceutics ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 902
Author(s):  
Madhumita Patel ◽  
Won-Gun Koh
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Hyaluronic Acid ◽  
Bone Tissue Engineering ◽  
Osteogenic Differentiation ◽  
Bone Tissue ◽  
Composite Hydrogel ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Composite Hydrogels

Composite hydrogels with electrospun nanofibers (NFs) have recently been used to mimic the native extracellular matrix. In this study, composite hydrogels of methacrylated hyaluronic acid containing fragmented polycaprolactone NFs were used for bone tissue engineering. The composite (NF/hydrogel) was crosslinked under ultraviolet (UV) light. The incorporation of fragmented polycaprolactone NFs increased the compression modulus from 1762.5 to 3122.5 Pa. Subsequently, adipose-derived stem cells incorporated into the composite hydrogel exhibited a more stretched and elongated morphology and osteogenic differentiation in the absence of external factors. The mRNA expressions of osteogenic biomarkers, including collagen 1 (Col1), alkaline phosphatase, and runt-related transcription factor 2, were 3–5-fold higher in the composite hydrogel than in the hydrogel alone. In addition, results of the protein expression of Col1 and alizarin red staining confirmed osteogenic differentiation. These findings suggest that our composite hydrogel provides a suitable microenvironment for bone tissue engineering.

scholarly journals Azithromycin Promotes the Osteogenic Differentiation of Human Periodontal Ligament Stem Cells after Stimulation with TNF-α

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Tingting Meng ◽  
Ying Zhou ◽  
Jingkun Li ◽  
Meilin Hu ◽  
Xiaomeng Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Osteogenic Differentiation ◽  
Alkaline Phosphatase Activity ◽  
Periodontal Ligament ◽  
Caspase 3 ◽  
Caspase 8 ◽  
Periodontal Ligament Stem Cells ◽  
Alizarin Red ◽  
Induced Apoptosis

Background and Objective. This study investigated the effects and underlying mechanisms of azithromycin (AZM) treatment on the osteogenic differentiation of human periodontal ligament stem cells (PDLSCs) after their stimulation with TNF-α in vitro. Methods. PDLSCs were isolated from periodontal ligaments from extracted teeth, and MTS assay was used to evaluate whether AZM and TNF-α had toxic effects on PDLSCs viability and proliferation. After stimulating PDLSCs with TNF-α and AZM, we analyzed alkaline phosphatase staining, alkaline phosphatase activity, and alizarin red staining to detect osteogenic differentiation. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis was performed to detect the mRNA expression of osteogenic-related genes, including RUNX2, OCN, and BSP. Western blotting was used to measure the NF-κB signaling pathway proteins p65, phosphorylated p65, IκB-α, phosphorylated IκB-α, and β-catenin as well as the apoptosis-related proteins caspase-8 and caspase-3. Annexin V assay was used to detect PDLSCs apoptosis. Results. TNF-α stimulation of PDLSCs decreased alkaline phosphatase and alizarin red staining, alkaline phosphatase activity, and mRNA expression of RUNX2, OCN, and BSP in osteogenic-conditioned medium. AZM enhanced the osteogenic differentiation of PDLSCs that were stimulated with TNF-α. Western blot analysis showed that β-catenin, phosphorated p65, and phosphorylated IκB-α protein expression decreased in PDLSCs treated with AZM. In addition, pretreatment of PDLSCs with AZM (10 μg/ml, 20 μg/ml) prevented TNF-α-induced apoptosis by decreasing caspase-8 and caspase-3 expression. Conclusions. Our results showed that AZM promotes PDLSCs osteogenic differentiation in an inflammatory microenvironment by inhibiting the WNT and NF-κB signaling pathways and by suppressing TNF-α-induced apoptosis. This suggests that AZM has potential as a clinical therapeutic for periodontitis.

scholarly journals Yunnan Baiyao Conditioned Medium Promotes the Odonto/Osteogenic Capacity of Stem Cells from Apical Papilla via Nuclear Factor Kappa B Signaling Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Xiyao Pang ◽  
Yanqiu Wang ◽  
Jintao Wu ◽  
Zhou Zhou ◽  
Tao Xu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Osteogenic Differentiation ◽  
Conditioned Medium ◽  
Signaling Pathway ◽  
Cell Counting ◽  
Dependent Manner ◽  
Alizarin Red ◽  
Apical Papilla ◽  
Yunnan Baiyao

Yunnan Baiyao is a traditional Chinese herbal remedy that has long been used for its characteristics of wound healing, bone regeneration, and anti-inflammation. However, the effects of Yunnan Baiyao on the odonto/osteogenic differentiation of stem cells from apical papilla (SCAPs) and the potential mechanisms remain unclear. The aim of this study was to investigate the odonto/osteogenic differentiation effects of Yunnan Baiyao on SCAPs and the underlying mechanisms involved. SCAPs were isolated and cocultured with Yunnan Baiyao conditioned media. The proliferation ability was determined by cell counting kit 8 and flow cytometry. The differentiation capacity and the involvement of NF-κB pathway were investigated by alkaline phosphatase assay, alizarin red staining, immunofluorescence assay, real-time RT-PCR, and western blot analyses. Yunnan Baiyao conditioned medium at the concentration of 50 μg/mL upregulated alkaline phosphatase activity, induced more mineralized nodules, and increased the expression of odonto/osteogenic genes/proteins (e.g., OCN/OCN, OPN/OPN, OSX/OSX, RUNX2/RUNX2, ALP/ALP, COL-I/COL-I, DMP1, DSP/DSPP) of SCAPs. In addition, the expression of cytoplasmic phos-IκBα, phos-P65, and nuclear P65 was significantly increased in Yunnan Baiyao conditioned medium treated SCAPs in a time-dependent manner. Conversely, the differentiation of Yunnan Baiyao conditioned medium treated SCAPs was obviously inhibited when these stem cells were cocultured with the specific NF-κB inhibitor BMS345541. Yunnan Baiyao can promote the odonto/osteogenic differentiation of SCAPs via the NF-κB signaling pathway.

scholarly journals TRIM16 Positively Regulates Osteogenic Differentiation of Human Periodontal Ligament Stem Cells by Modulating the Ubiquitination and Degradation of RUNX2

2020 ◽  
Author(s):  
Yi Zhao ◽  
Qiaoli Zhai ◽  
Hong Liu ◽  
Xun Xi ◽  
Shuai Chen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Alkaline Phosphatase ◽  
Bone Regeneration ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Osteogenic Potential ◽  
Common Disease ◽  
Periodontal Ligament Stem Cells ◽  
Alizarin Red ◽  
Periodontal Tissues

Abstract BackgroundPeriodontal disease is a common disease that compromises the integrity of tooth-supporting tissues. Bone regeneration is the ultimate goal of periodontal therapies, in which osteogenic differentiation of human periodontal ligament stem cells plays a critical role. The tripartite motif (TRIM)16 is downregulated in periodontal tissues of patients with periodontitis and involved in osteogenic differentiation of human bone marrow mesenchymal stem cells(hBMSCs).However, the role of TRIM16 in the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) is largely unknown.MethodshPDLSCs were isolated and identified by immunophenotype assays using flow cytometry. Overexpression plasmids and specific short-hairpin RNAs (shRNAs) were constructed to manipulate the expression of target molecules. Alkaline phosphatase (ALP) staining, alizarin red staining (ARS) and enzyme‐linked immunosorbent assays (ELISA) were used to evaluate osteogenic potential capacity. Reverse transcription quantitative PCR (RT-qPCR) and Western blot analysis were performed to determine the expression of osteogenic-related markers and activation of relevant signaling pathways. Co-immunoprecipitation assays were performed to confirm the interactions between proteins and the ubiquitination of RUNX2. A LC-MS/MS analysis was performed to explore the different expression proteins in present of TRIM16.ResultsTRIM16 significantly promoted alkaline phosphatase activity and mineralized nodule formation, and positively regulated the osteogenic differentiation of hPDLSCs by enhancing protein expression of RUNX2, COL1A1 and OCN. Mechanistically, TRIM16 serves as a pivotal factor that stabilizes RUNX2 protein levels by decreasing CHIP-mediated K48-linked ubiquitination degradation of the RUNX2 protein. Besides, TRIM16 significantly increased expression of COL1A1 via activation of p38MAPK/RUNX2.ConclusionThis study identified a novel mechanism of TRIM16 in regulating stability of the RUNX2 protein, which may promote the osteogenic differentiation of hPDLSCs. TRIM16 may be a potential target of stem cell based-bone regeneration for periodontal therapies.

scholarly journals Differentiation of Rat Adipose-Derived Stem Cells into Parathyroid-Like Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-6
Author(s):  
Ping Zhang ◽  
Hao Zhang ◽  
Wenwu Dong ◽  
Zhihong Wang ◽  
Yuan Qin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Parathyroid Hormone ◽  
Osteogenic Differentiation ◽  
Hormone Secretion ◽  
Activin A ◽  
High Price ◽  
Dependent Manner ◽  
Adipose Derived Stem Cells ◽  
Alizarin Red ◽  
Postoperative Hypoparathyroidism

Background. The current treatment for postoperative hypoparathyroidism has shortcomings, such as repeated blood monitoring for dosage adjustment, uncertain long-term efficacy, and the high price of recombinant parathyroid hormone therapy. Adipose-derived stem cells can undergo adipogenic and osteogenic differentiation in vitro and are considered a novel source of parathyroid-like cells, but the idea lacks theoretical basis and feasibility. We aimed at establishing a protocol for differentiating adipose-derived stem cells into parathyroid-like cells for treating hypoparathyroidism. Materials/Methods. Adipose-derived stem cells were isolated and purified from the inguinal adipose tissue of Sprague Dawley rats. Adipogenic differentiation and osteogenic differentiation of the cells were identified by oil red O and alizarin red S staining, respectively. The adipose-derived stem cells were stimulated by sonic hedgehog (SHH) and activin A. The differentiation of the adipose-derived stem cells to parathyroid-like cells was confirmed by the detection of parathyroid hormone and the related parathyroid markers. Results. Adipose-derived stem cells were successfully isolated and purified from the rat adipocytes. The adipogenic and osteogenic differentiation capabilities of the adipose-derived stem cells were determined. SHH and activin A stimulated parathyroid hormone secretion by the adipose-derived stem cells and significantly increased the expression of calcium-sensing receptor (CaSR), parathyroid hormone, and glial cells missing homolog 2 (GCM2) in the cells in a time- and concentration-dependent manner. Conclusion. We successfully differentiated rat adipose-derived stem cells into parathyroid-like cells, which will pave a new route to curing hypoparathyroidism.

286 OSTEOGENIC DIFFERENTIATION IN VITRO OF PORCINE ADULT MESENCHYMAL STEM CELLS

2008 ◽  
Vol 20 (1) ◽  
pp. 223
Author(s):  
A. Lima ◽  
E. Monaco ◽  
S. Wilson ◽  
D. Kim ◽  
C. Feltrin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Adipose Tissue ◽  
Osteogenic Differentiation ◽  
Subcutaneous Adipose Tissue ◽  
Cell Types ◽  
Alizarin Red ◽  
Subcutaneous Adipose

The quantity and accessibility of subcutaneous adipose tissue in humans make it an attractive alternative to bone marrow as a source of adult stem cells for therapeutic purposes. However, before such a cell source substitution can be proposed, the properties of stem cells derived from adipose (ADSCs) and bone marrow (MSCs) and their differentiated progeny must be compared in an animal model that adequately simulates the structure and physiology of humans. The objective of this work was to induce adult porcine stem cells isolated from subcutaneous adipose tissue and bone marrow to differentiate in vitro along the osteoblastic lineage and to compare their morphological, phenotypic, and genotypic properties. MSCs and ADSCs were isolated respectively from femurs and subcutaneous adipose tissue of adult pigs and cultured in vitro using DMEM supplemented with 10% fetal bovine serum (FBS), 1% penicillin G-streptomycin, and 5.6 mg L–1 amphotericin B. After 3 passages, cells were differentiated along the osteogenic lineage using lineage-specific inducing medium. Osteogenic medium contained 100 nm dexamethasone, 10 mm β-glycerophosphate, and 0.005 mm ascorbic acid-2-phosphate. Osteogenic cultures were incubated for 4 weeks in 95% air and 5% CO2 at 39�C. Spent medium was replaced with fresh medium every 3 days. Histological staining with alkaline phosphatase, Von Kossa, and alizarin red S were performed at 0, 2, 4, 7, 14, 21, and 28 days of differentiation (dd). At the same time points, RNA was extracted. qPCR was performed on COL1A1, BGLAP, SPARC, and SPP1. As internal control, the geometrical mean of GTF2H, NUBP, and PPP2C was used. Relative mRNA abundance between cell types was calculated using 1/efficiencydCT. The osteogenic differentiation of both MSCs and ADScs was confirmed by the organization of the cells in nodules and by alkaline phosphatase-, Von Kossa-, and alizarin red S-positive staining. The percent relative abundance of the 4 genes in both cell types was COL1A1 (ca. 50) > SPARC (ca. 45) > SPP1 (ca. 5) > BGLAP ( < 0.1). Cell types showed similar mRNA abundance for COL1A1 and SPARC while SPP1 and BGLAP were, respectively, 10- and 19-fold higher in MSCs than in ADSCs. All of the genes had the same pattern among tissues during differentiation except for SPP1, which showed a >10-fold increase at 14 v. 0 dd only for MSCs. Adipose-derived stem cells demonstrated a clear osteogenic differentiation and similar expression and pattern of the two osteogenic genes most abundant in MSCs (COL1A1 and SPARC). However, the higher abundance of SPP1 and BGLAP and the different behavior of SPP1 in MSCs suggest a different transcription profile between the two cell types. From these preliminary results, adipose tissue can be a practical alternative source for stem cells in future human clinical applications.

178 Differential Behavior of Porcine Mesenchymal Stem Cells from Bone Marrow and Adipose During Osteogenic Differentiation on a Glycosaminoglycan Hydrogel Scaffold for Bone and Cartilage Tissue Engineering

2018 ◽  
Vol 30 (1) ◽  
pp. 229 ◽  
Author(s):  
S. A. Womack ◽  
D. J. Milner ◽  
D. W. Weisgerber ◽  
B. A. Harley ◽  
M. B. Wheeler
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Cartilage Tissue Engineering ◽  
Cartilage Tissue ◽  
Scaffold Material ◽  
Alizarin Red ◽  
And Cartilage

The pig is an ideal species for use in tissue engineering studies targeted towards repair of bone and cartilage defects. Novel collagen-glycosaminoglycan hydrogel (CG) scaffolds have shown promise for supporting bone and cartilage growth from mesenchymal stem cells. In order to determine the suitability of these scaffolds for use in porcine model systems for bone and cartilage tissue engineering, we have begun to investigate the behaviour of porcine mesenchymal stem cells on this material. The purpose of this study was to determine whether mesenchymal stem cells from adipose (ASC) and bone marrow (BMSC) form bone on a CG scaffold material. Primary BMSC and ASC from 6-month-old Yorkshire pigs were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) with 10% fetal bovine serum. The ASC and BMSC were then trypsinized at passage 4 or 5 and used to seed ~4-mm-diameter CG scaffolds with 2 million cells/scaffold. Scaffolds were seeded by suspending the cells in medium that had been equilibrated for 30 min, and then placing the CG scaffold into the medium. This method of seeding was determined to be most effective in previous experiments. Scaffolds were then cultured for 7 days in DMEM followed by 21 days in osteogenic media. At the conclusion of the incubation period, the diameter of the scaffolds was measured, and they were fixed with 4% paraformaldehyde and cryosectioned. Then, 10-µm sections were stained with Alizarin Red to assay for mineralization, a hallmark of osteogenic differentiation. Both ASC- and BMSC-loaded scaffolds showed Alizarin Red staining throughout the section after incubation, demonstrating that both undergo osteogenesis on the scaffold material (n = 4). During osteogenic differentiation, scaffolds seeded with both ASC and BMSC showed a decrease in diameter. Unseeded scaffolds showed no decrease in size when in media. The BMSC scaffolds demonstrated a more extensive decrease in size than ASC. The average diameter of ASC loaded scaffolds after differentiation was 2.49 ± 0.39 mm, and that of BMSC-loaded scaffolds was 1.47 ± 0 0.18 mm (n = 3, P < 0.05, Student’s t-test). This suggests a differential ability of ASC and BMSC to break down and metabolize the scaffold matrix, and may indicate that one cell type may be preferable to the other for repairing osteogenic defects using these scaffolds. Current experiments underway will analyse expression of matrix-degrading enzymes to determine the source of the difference between cell types in scaffold shrinkage during differentiation. We will also quantify mineralization in ASC- v. BMSC-loaded scaffolds and assay gene expression of osteogenic markers to determine if there is a difference in osteogenic potential between sources of mesenchymal stem cells on these scaffolds.

Osteogenic differentiation of adipose-derived stem cells in mesoporous SBA-16 and SBA-16 hydroxyapatite scaffolds

RSC Advances ◽  
2015 ◽  
Vol 5 (67) ◽  
pp. 54551-54562 ◽  
Author(s):  
Gracielle F. Andrade ◽  
Juliana L. Carvalho ◽  
Armando S. C. Júnior ◽  
Alfredo M. Goes ◽  
Edésia M. B. Sousa
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Bone Tissue Engineering ◽  
Osteogenic Differentiation ◽  
Bone Tissue ◽  
Adipose Derived Stem Cells ◽  
Engineering Applications

Adipose-derived stem cells (ASCs) are currently a point of focus for bone tissue engineering applications.

scholarly journals Impact of Age on Human Adipose Stem Cells for Bone Tissue Engineering

2017 ◽  
Vol 26 (9) ◽  
pp. 1496-1504 ◽  
Author(s):  
Denis Dufrane
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Bone Tissue Engineering ◽  
Osteogenic Differentiation ◽  
Bone Tissue ◽  
Pathological Condition ◽  
Growth Potential ◽  
Matrix Mineralization

Bone nonunion is a pathological condition in which all bone healing processes have stopped, resulting in abnormal mobility between 2 bone segments. The incidence of bone-related injuries will increase in an aging population, leading to such injuries reaching epidemic proportions. Tissue engineering and cell therapy using mesenchymal stem cells (MSCs) have raised the possibility of implanting living tissue for bone reconstruction. Bone marrow was first proposed as the source of stem cells for bone regeneration. However, as the quantity of MSCs in the bone marrow decreases, the capacity of osteogenic differentiation of bone marrow stem cells is also impaired by the donor’s age in terms of reduced MSC replicative capacity; an increased number of apoptotic cells; formation of colonies positive for alkaline phosphatase; and decreases in the availability, growth potential, and temporal mobilization of MSCs for bone formation in case of fracture. Adipose-derived stem cells (ASCs) demonstrate several advantages over those from bone marrow, including a less invasive harvesting procedure, a higher number of stem cell progenitors from an equivalent amount of tissue harvested, increased proliferation and differentiation capacities, and better angiogenic and osteogenic properties in vivo. Subcutaneous native adipose tissue was not affected by the donor’s age in terms of cellular senescence and yield of ASC isolation. In addition, a constant mRNA level of osteocalcin and alkaline phosphatase with a similar level of matrix mineralization of ASCs remained unaffected by donor age after osteogenic differentiation. The secretome of ASCs was also unaffected by age when aiming to promote angiogenesis by vascular endothelial growth factor (VEGF) release in hypoxic conditions. Therefore, the use of adipose cells for bone tissue engineering is not limited by the donor’s age from the isolation of stem cells up to the manufacturing of a complex osteogenic graft.

Stimulated Osteogenic Differentiation of Human Mesenchymal Stem Cells by Reduced Graphene Oxide

2015 ◽  
Vol 15 (10) ◽  
pp. 7966-7970 ◽  
Author(s):  
Linhua Jin ◽  
Jong Ho Lee ◽  
Oh Seong Jin ◽  
Yong Cheol Shin ◽  
Min Jeong Kim ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Graphene Oxide ◽  
Osteogenic Differentiation ◽  
Reduced Graphene Oxide ◽  
Human Mesenchymal Stem Cells ◽  
Water Soluble ◽  
Alizarin Red ◽  
Reduced Graphene

Osteoprogenitor cells play a significant role in the growth or repair of bones, and have great potential as cell sources for regenerative medicine and bone tissue engineering, but control of their specific differentiation into bone cells remains a challenge. Graphene-based nanomaterials are attractive candidates for biomedical applications as substrates for stem cell (SC) differentiation, scaffolds in tissue engineering, and components of implant devices owing to their biocompatible, transferable and implantable properties. This study examined the enhanced osteogenic differentiation of human mesenchymal stem cells (hMSCs) by reduced graphene oxide (rGO) nanoparticles (NPs), and rGO NPs was prepared by reducing graphene oxide (GO) with a hydrazine treatment followed by annealing in argon and hydrogen. The cytotoxicity profile of each particle was examined using a water-soluble tetrazolium-8 (WST-8) assay. At different time-points, a WST-8 assay, alkaline phosphatase (ALP) activity assay and alizarin red S (ARS) staining were used to determine the effects of rGO NPs on proliferation, differentiation and mineralization, respectively. The results suggest that graphene-based materials have potential as a platform for stem cells culture and biomedicalapplications.

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