scholarly journals The composite sandwich structure of dNCPs polyelectrolyte multilayers induced the osteogenic differentiation of PDLSCs in vitro

2020 ◽  
Vol 18 ◽  
pp. 228080002094271
Author(s):  
Jing Chen ◽  
Wenxing Li ◽  
Qiang Li ◽  
Yuhui Wang ◽  
Bingjiao Zhao ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Sandwich Structure ◽  
Polyelectrolyte Multilayers ◽  
Osteogenic Potential ◽  
Fiber Morphology ◽  
Periodontal Ligament Stem Cells ◽  
Composite Sandwich ◽  
Coated Membranes ◽  
Experimental Group

This study reported about the fabrication of dentin non-collagenous proteins (dNCPs) polyelectrolyte multilayers and evaluated its osteogenic potential. The composite sandwich structure of dNCPs polyelectrolyte multilayers was generated on the surface of polycaprolactone electrospinning membranes by the Layer-by-Layer self-assembly technique. The dNCPs-coated membranes comprised the experimental group and the non-coated membranes acted as the control. Nanofiber morphologies of both membranes were observed under scanning electron microscope. The release of dNCPs was evaluated by ELISA kit. Periodontal ligament stem cells (PDLSCs) were seeded on both membranes. The morphology changes and proliferation of cells were tested. The expressions of osteogenic-related genes and proteins were evaluated by RT-PCR, alkaline phosphatase (ALP) activity assay, and immunofluorescence staining. dNCPs-coated membranes displayed significantly different fiber morphology than the non-coated membranes. A stable release of dentin phosphoprotein was maintained from day 4 to day 15 in the experimental group. Cells on dNCPs-coated membranes were found to have cuboidal or polygonal shapes. The proliferative rate of cells was significantly lower in the experimental group from day 4 to day 9 ( p<0.05). However, cells on the dNCPs-coated membranes demonstrated a significantly higher ALP content and expression levels of osteogenic gene and proteins than the controls ( p<0.05). These results indicated that dNCPs polyelectrolyte multilayers could induce the osteogenic differentiation of PDLSCs in vitro.

scholarly journals Effects of Naringin on Proliferation and Osteogenic Differentiation of Human Periodontal Ligament Stem Cells In Vitro and In Vivo

2015 ◽  
Vol 2015 ◽  
pp. 1-9 ◽  
Author(s):  
Lihua Yin ◽  
Wenxiao Cheng ◽  
Zishun Qin ◽  
Hongdou Yu ◽  
Zhanhai Yu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Trabecular Structure ◽  
Control Group ◽  
Periodontal Ligament Stem Cells ◽  
Expression Levels ◽  
Proliferation And Differentiation

This study is to explore the osteogenesis potential of the human periodontal ligament stem cells (hPDLSCs) induced by naringin in vitro and in vitro. The results confirmed that 1 μM naringin performs the best effect and a collection of bone-related genes (RUNX2,COL1A2, OPN, and OCN) had significantly higher expression levels compared to the control group. Furthermore, a typical trabecular structure was observed in vivo, surrounded by a large amount of osteoblasts. These results demonstrated that naringin, at a concentration of 1 μM, can efficiently promote the proliferation and differentiation of hPDLSCs both in vitro and in vivo.

scholarly journals Combination of Human Amniotic Fluid Derived-Mesenchymal Stem Cells and Nano-hydroxyapatite Scaffold Enhances Bone Regeneration

2019 ◽  
Vol 7 (17) ◽  
pp. 2739-2750 ◽  
Author(s):  
Eman E. A. Mohammed ◽  
Hanan Beherei ◽  
Mohamed El-Zawahry ◽  
Abdel Razik Farrag ◽  
Naglaa Kholoussi ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Bone Healing ◽  
Therapeutic Potential ◽  
Osteogenic Potential ◽  
3D Scaffold ◽  
Human Amniotic Fluid ◽  
Post Transplantation ◽  
Defect Site ◽  
Tibia Defect

BACKGROUND: Human amniotic fluid-derived stem cells (hAF-MSCs) have a high proliferative capacity and osteogenic differentiation potential in vitro. The combination of hAF-MSCs with three-dimensional (3D) scaffold has a promising therapeutic potential in bone tissue engineering and regenerative medicine. Selection of an appropriate scaffold material has a crucial role in a cell supporting and osteoinductivity to induce new bone formation in vivo. AIM: This study aimed to investigate and evaluate the osteogenic potential of the 2nd-trimester hAF-MSCs in combination with the 3D scaffold, 30% Nano-hydroxyapatite chitosan, as a therapeutic application for bone healing in the induced tibia defect in the rabbit. SUBJECT AND METHODS: hAF-MSCs proliferation and culture expansion was done in vitro, and osteogenic differentiation characterisation was performed by Alizarin Red staining after 14 & 28 days. Expression of the surface markers of hAF-MSCs was assessed using Flow Cytometer with the following fluorescein-labelled antibodies: CD34-PE, CD73-APC, CD90-FITC, and HLA-DR-FITC. Ten rabbits were used as an animal model with an induced defect in the tibia to evaluate the therapeutic potential of osteogenic differentiation of hAF-MSCs seeded on 3D scaffold, 30% Nano-hydroxyapatite chitosan. The osteogenic differentiated hAF-MSCs/scaffold composite system applied and fitted in the defect region and non-seeded scaffold was used as control. The histopathological investigation was performed at 2, 3, & 4 weak post-transplantation and scanning electron microscope (SEM) was assessed at 2 & 4 weeks post-transplantation to evaluate the bone healing potential in the rabbit tibia defect. RESULTS: Culture and expansion of 2nd-trimester hAF-MSCs presented high proliferative and osteogenic potential in vitro. Histopathological examination for the transplanted hAF-MSCs seeded on the 3D scaffold, 30% Nano-hydroxyapatite chitosan, demonstrated new bone formation in the defect site at 2 & 3 weeks post-transplantation as compared to the control (non-seeded scaffold). Interestingly, the scaffold accelerated the osteogenic differentiation of AF-MSCs and showed complete bone healing of the defect site as compared to the control (non-seeded scaffold) at 4 weeks post-transplantation. Furthermore, the SEM analysis confirmed these findings. CONCLUSION: The combination of the 2nd-trimester hAF-MSCs and 3D scaffold, 30% Nano-hydroxyapatite chitosan, have a therapeutic perspective for large bone defect and could be used effectively in bone tissue engineering and regenerative medicine.

scholarly journals Effect of puerarin on osteogenic differentiation of human periodontal ligament stem cells

2019 ◽  
Vol 48 (4) ◽  
pp. 030006051985164
Author(s):  
Jun Li ◽  
Youjian Peng
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Calcium Deposition ◽  
Pcr Analysis ◽  
Cell Surface Markers ◽  
Periodontal Ligament Stem Cells ◽  
Alizarin Red ◽  
Alp Activity ◽  
Experimental Group

Objective To investigate the effects of the flavonoid, puerarin, on osteogenic differentiation of human periodontal ligament stem cells (PDLSCs). Methods Human PDLSCs were isolated from patients undergoing orthodontic treatment, and the cell surface markers CD146, CD34, CD45, and STRO-1 were identified by immunofluorescence. Cell proliferation was detected by MTT assay; alkaline phosphatase (ALP) activity was measured, and calcium deposition was detected by alizarin red staining. PCR was then used to detect the distributions of COL-I, OPN, Runx2, and OCN, genes related to osteogenic differentiation. Results Staining was positive for cytokines CD146, CD34, CD45, and STRO-1 in the experimental group; staining was also positive for silk protein, but negative for keratin. After 7 days of culture, exposure to puerarin significantly promoted the level of intracellular ALP; increased puerarin concentration led to increased intracellular ALP. Red mineralized nodules appeared upon exposure to puerarin and the number of nodules was concentration-dependent. PCR analysis revealed that COL-I, OPN, Runx2, and OCN expression levels increased as puerarin concentration increased. Conclusions Exposure to puerarin can promote proliferation and ALP activity in human PDLSCs, thus promoting both molecular and osteogenic differentiation; these findings may provide a theoretical basis for the clinical treatment of periodontal disease with puerarin.

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 MiR-144-5p, an exosomal miRNA from bone marrow-derived macrophage in type 2 diabetes, impairs bone fracture healing via targeting Smad1

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Dong Zhang ◽  
Yifan Wu ◽  
Zonghuan Li ◽  
Hairen Chen ◽  
Siyuan Huang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Fracture Healing ◽  
Bone Regeneration ◽  
Osteogenic Differentiation ◽  
Fracture Repair ◽  
Osteogenic Potential ◽  
Patients With Diabetes ◽  
The Impact

Abstract Background Patients with diabetes have an increased risk of nonunion and delayed union of fractures. Macrophages have been shown as a key player in diabetic complications. However, it remains obscure how diabetic milieu affects macrophage-derived exosomes and its implications on osteogenic differentiation of BMSCs. In this study, we aim to define the impact of diabetic milieu on macrophage-derived exosomes, role of extracellular vesicles in intercellular communication with BMSCs, and subsequent effects on osteogenic differentiation and fracture repair. Results The osteogenic potential and the ability of fracture repair of exosomes derived from diabetic bone marrow-derived macrophages (dBMDM-exos) were revealed to be lower, as compared with non-diabetic bone marrow-derived macrophages (nBMDM-exos) in vitro and in vivo. Interestingly, miR-144-5p levels were sharply elevated in dBMDM-exos and it could be transferred into BMSCs to regulate bone regeneration by targeting Smad1. In addition, the adverse effects of dBMDM-exos on the osteogenic potential and the ability of fracture repair were reversed through the suppression of miR-144-5p inhibition in vitro and vivo. Conclusions The results demonstrated an important role of exosomal miR-144-5p in bone regeneration, offering insight into developing new strategy for the improvement of fracture healing in patients with diabetes mellitus. Graphic Abstract

scholarly journals Mesenchymal Stromal Cell Differentiation for Generating Cartilage and Bone-Like Tissues In Vitro

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 2165
Author(s):  
Graziana Monaco ◽  
Yann D. Ladner ◽  
Alicia J. El Haj ◽  
Nicholas R. Forsyth ◽  
Mauro Alini ◽  
...  
Keyword(s):  
Gene Expression ◽  
Growth Factors ◽  
Osteogenic Differentiation ◽  
Type I Collagen ◽  
Specific Growth ◽  
Culture Media ◽  
Culture Conditions ◽  
Osteogenic Potential ◽  
Type I

In the field of tissue engineering, progress has been made towards the development of new treatments for cartilage and bone defects. However, in vitro culture conditions for human bone marrow mesenchymal stromal cells (hBMSCs) have not yet been fully defined. To improve our understanding of cartilage and bone in vitro differentiation, we investigated the effect of culture conditions on hBMSC differentiation. We hypothesized that the use of two different culture media including specific growth factors, TGFβ1 or BMP2, as well as low (2% O2) or high (20% O2) oxygen tension, would improve the chondrogenic and osteogenic potential, respectively. Chondrogenic and osteogenic differentiation of hBMSCs isolated from multiple donors and expanded under the same conditions were directly compared. Chondrogenic groups showed a notable upregulation of chondrogenic markers compared with osteogenic groups. Greater sGAG production and deposition, and collagen type II and I accumulation occurred for chondrogenic groups. Chondrogenesis at 2% O2 significantly reduced ALP gene expression and reduced type I collagen deposition, producing a more stable and less hypertrophic chondrogenic phenotype. An O2 tension of 2% did not inhibit osteogenic differentiation at the protein level but reduced ALP and OC gene expression. An upregulation of ALP and OC occurred during osteogenesis in BMP2 containing media under 20% O2; BMP2 free osteogenic media downregulated ALP and also led to higher sGAG release. A higher mineralization was observed in the presence of BMP2 during osteogenesis. This study demonstrates how the modulation of O2 tension, combined with tissue-specific growth factors and media composition can be tailored in vitro to promote chondral or endochondral differentiation while using the same donor cell population.

scholarly journals The potential of human-derived periodontal ligament stem cells to osteogenic differentiation: An In vitro investigation

2014 ◽  
Vol 2 (4) ◽  
pp. 18-23
Author(s):  
Behzad Houshmand ◽  
Omolbanin Amjadi ◽  
Alireza Rafiei ◽  
Mohammadali Rouzegar ◽  
Mohammadreza Abrishami ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Periodontal Ligament Stem Cells ◽  
In Vitro Investigation

Abstract 525: The Role of FHL2 in Mechanically Regulated Valve Interstitial Cell Osteogenic Differentiation

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Alan Y Lam ◽  
Jan-Hung Chen ◽  
Craig A Simmons
Keyword(s):  
Osteogenic Differentiation ◽  
Nuclear Translocation ◽  
Dominant Negative ◽  
Osteogenic Potential ◽  
Matrix Stiffness ◽  
Calcific Aortic Valve Disease ◽  
Regulatory Relationship ◽  
Sirna Knockdown

Introduction: Lesions in calcific aortic valve disease (CAVD) often include osteoblasts and occur preferentially in the natively stiffer fibrosa side of the valve, suggesting roles for osteogenesis and matrix stiffness in CAVD. Valve interstitial cells (VICs) have osteogenic potential that is modulated by matrix stiffness in vitro . Osteogenesis of mesenchymal stem cells is mediated by FHL2 and modulated by matrix stiffness via RhoA. However, the connection between FHL2 and RhoA and their roles in mechanically-regulated osteogenic differentiation of VICs and CAVD have not been established. Hypothesis: Matrix stiffness regulates RhoA-dependent FHL2 expression to direct VIC osteogenesis. Methods/Results: Primary porcine aortic VICs were grown on collagen-coated polyacrylamide gels of varying elastic moduli in vitro . With increasing substrate stiffness, VICs demonstrated increased RhoA activation by ELISA and increased FHL2 nuclear translocation by immunofluorescence and western blotting. Notably, significant increases were seen on 22 kPa substrates, a stiffness unique to the disease-prone fibrosa, compared to 11 kPa substrates (p<0.05). We investigated the regulatory relationship between RhoA and FHL2 in VICs by controlling RhoA levels with adenoviruses. With constitutively active RhoA, FHL2 nuclear translocation increased (p<0.06); in contrast, with dominant negative RhoA, FHL2 nuclear translocation decreased (p<0.05). The effect of RhoA levels on stiffness-dependent FHL2 activity was verified by knocking down RhoA levels pharmacologically (C3 toxin) which abrogated stiffness-dependent FHL2 nuclear translocation. Alkaline phosphatase activity as a marker of osteogenesis was highest on substrates with fibrosa-like stiffness and was reduced by FHL2 siRNA knockdown. Conclusions: Together, these results support the hypothesis that mechanically-regulated VIC osteogenic differentiation in vitro is mediated by FHL2 operating downstream of RhoA. New insights into the mechanotransduction mechanisms that regulate VIC osteogenesis may lead to novel strategies for treating CAVD.

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