scholarly journals The Role of Pannexin3-Modified Human Dental Pulp-Derived Mesenchymal Stromal Cells in Repairing Rat Cranial Critical-Sized Bone Defects

2017 ◽  
Vol 44 (6) ◽  
pp. 2174-2188 ◽  
Author(s):  
Fangfang Song ◽  
Hualing Sun ◽  
Liyuan Huang ◽  
Dongjie Fu ◽  
Cui Huang
Keyword(s):  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Underlying Mechanism ◽  
Bone Defects ◽  
Dependent Manner ◽  
Alizarin Red ◽  
Human Dental Pulp

Background/Aims: Human dental pulp-derived mesenchymal stromal cells (hDPSCs) are promising seed cells for tissue engineering due to their easy accessibility and multi-lineage differentiation. Pannexin3 (Panx3) plays crucial roles during bone development and differentiation. The aim of the present study was to investigate the effect of Panx3 on osteogenesis of hDPSCs and the underlying mechanism. Methods: Utilizing qRT-PCR, Western blot, and immunohistochemistry, we explored the change of Panx3 during osteogenic differentiation of hDPSCs. Next, hDPSCs with loss (Panx3 knockdown) and gain (Panx3 overexpression) of Panx3 function were developed to investigate the effects of Panx3 on osteogenic differentiation of hDPSC and the underlying mechanism. Finally, a commercial β-TCP scaffold carrying Panx3-modified hDPSCs was utilized to evaluate bone defect repair. Results: Panx3 was upregulated during osteogenic differentiation in a time-dependent manner. Panx3 overexpression promoted osteogenic differentiation of hDPSCs, whereas depletion of Panx3 resulted in a decline of differentiation, evidenced by upregulated expression of mineralization-related markers, increased alkaline phosphatase (ALP) activity, and enhanced ALP and Alizarin red staining. Panx3 was found to interact with the Wnt/β-catenin signaling pathway, forming a negative feedback loop. However, Wnt/β-catenin did not contribute to enhancement of osteogenic differentiation as observed in Panx3 overexpression. Moreover, Panx3 promoted osteogenic differentiation of hDPSCs via increasing ERK signaling pathway. Micro-CT and histological staining results showed that Panx3-modified hDPSCs significantly improved ossification of critical-sized bone defects. Conclusion: These findings suggest that Panx3 is a crucial modulator of hDPSCs differentiation.

scholarly journals circAKT3 positively regulates osteogenic differentiation of human dental pulp stromal cells via miR-206/CX43 axis

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Bo Zhang ◽  
Sibei Huo ◽  
Xiao Cen ◽  
Xuefeng Pan ◽  
Xinqi Huang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Alizarin Red ◽  
Cx43 Expression ◽  
Human Dental Pulp

Abstract Background Human dental pulp stromal cells (hDPSCs) are promising sources of mesenchymal stem cells (MSCs) for bone tissue regeneration. Circular RNAs (circRNAs) have been demonstrated to play critical roles in stem cell osteogenic differentiation. Herein, we aimed to investigate the role of circAKT3 during osteogenesis of hDPSCs and the underlying mechanisms of its function. Methods We performed circRNA sequencing to investigate the expression profiles of circular RNAs during osteogenesis of hDPSCs. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to detect the expression pattern of circAKT3 and miR-206 in hDPSCs during osteogenesis. We knocked down circAKT3 and interfered the expression of miR-206 to verify their regulatory role in hDPSC osteogenesis. We detected hDPSCs mineralization by alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining and used dual-luciferase reporter assay to validate the direct binding between circAKT3 and miR-206. To investigate in vivo mineralization, we performed subcutaneous transplantation in nude mice and used hematoxylin and eosin, Masson’s trichrome, and immunohistochemistry staining. Results Totally, 86 circRNAs were differentially expressed during hDPSC osteogenesis, in which 29 were downregulated while 57 were upregulated. circAKT3 was upregulated while miR-206 was downregulated during hDPSC osteogenesis. Knockdown of circAKT3 inhibited ALP/ARS staining and expression levels of osteogenic genes. circAKT3 directly interacted with miR-206, and the latter one suppressed osteogenesis of hDPSCs. Silencing miR-206 partially reversed the inhibitory effect of circAKT3 knockdown on osteogenesis. Connexin 43 (CX43), which positively regulates osteogenesis of stem cells, was predicted as a target of miR-206, and overexpression or knockdown of miR-206 could correspondingly decrease and increase the expression of CX43. In vivo study showed knockdown of circAKT3 suppressed the formation of mineralized nodules and expression of osteogenic proteins. Conclusion During osteogenesis of hDPSCs, circAKT3 could function as a positive regulator by directly sponging miR-206 and arresting the inhibitive effect of miR-206 on CX43 expression.

scholarly journals The Selective Histone Deacetylase Inhibitor MI192 Enhances the Osteogenic Differentiation Efficacy of Human Dental Pulp Stromal Cells

2021 ◽  
Vol 22 (10) ◽  
pp. 5224
Author(s):  
Kenny Man ◽  
Liam Lawlor ◽  
Lin-Hua Jiang ◽  
Xuebin B. Yang
Keyword(s):  
Alkaline Phosphatase ◽  
Osteogenic Differentiation ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Selective Inhibitor ◽  
Epigenetic Reprogramming ◽  
Type I ◽  
Human Dental Pulp ◽  
Pre Treatment ◽  
Dose Dependent

The use of human dental pulp stromal cells (hDPSCs) has gained increasing attention as an alternative stem cell source for bone tissue engineering. The modification of the cells’ epigenetics has been found to play an important role in regulating differentiation, with the inhibition of histone deacetylases 3 (HDAC3) being linked to increased osteogenic differentiation. This study aimed to induce epigenetic reprogramming using the HDAC2 and 3 selective inhibitor, MI192 to promote hDPSCs osteogenic capacity for bone regeneration. MI192 treatment caused a time–dose-dependent change in hDPSC morphology and reduction in viability. Additionally, MI192 successfully augmented hDPSC epigenetic functionality, which resulted in increased histone acetylation and cell cycle arrest at the G2/M phase. MI192 pre-treatment exhibited a dose-dependent effect on hDPSCs alkaline phosphatase activity. Quantitative PCR and In-Cell Western further demonstrated that MI192 pre-treatment significantly upregulated hDPSCs osteoblast-related gene and protein expression (alkaline phosphatase, bone morphogenic protein 2, type I collagen and osteocalcin) during osteogenic differentiation. Importantly, MI192 pre-treatment significantly increased hDPSCs extracellular matrix collagen production and mineralisation. As such, for the first time, our findings show that epigenetic reprogramming with the HDAC2 and 3 selective inhibitor MI192 accelerates the osteogenic differentiation of hDPSCs, demonstrating the considerable utility of this MSCs engineering approach for bone augmentation strategies.

scholarly journals The oncometabolite R-2-hydroxyglutarate dysregulates the differentiation of human mesenchymal stromal cells via inducing DNA hypermethylation

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Lizhen Liu ◽  
Kaimin Hu ◽  
Jingjing Feng ◽  
Huafang Wang ◽  
Shan Fu ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Chondrogenic Differentiation ◽  
Cell Counting ◽  
Human Mscs ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Dna Hypermethylation ◽  
Cartilaginous Tumors

Abstract Background Isocitrate dehydrogenase (IDH1/2) gene mutations are the most frequently observed mutations in cartilaginous tumors. The mutant IDH causes elevation in the levels of R-enantiomer of 2-hydroxylglutarate (R-2HG). Mesenchymal stromal cells (MSCs) are reasonable precursor cell candidates of cartilaginous tumors. This study aimed to investigate the effect of oncometabolite R-2HG on MSCs. Methods Human bone marrow MSCs treated with or without R-2HG at concentrations 0.1 to 1.5 mM were used for experiments. Cell Counting Kit-8 was used to detect the proliferation of MSCs. To determine the effects of R-2HG on MSC differentiation, cells were cultured in osteogenic, chondrogenic and adipogenic medium. Specific staining approaches were performed and differentiation-related genes were quantified. Furthermore, DNA methylation status was explored by Illumina array-based arrays. Real-time PCR was applied to examine the signaling component mRNAs involved in. Results R-2HG showed no influence on the proliferation of human MSCs. R-2HG blocked osteogenic differentiation, whereas promoted adipogenic differentiation of MSCs in a dose-dependent manner. R-2HG inhibited chondrogenic differentiation of MSCs, but increased the expression of genes related to chondrocyte hypertrophy in a lower concentration (1.0 mM). Moreover, R-2HG induced a pronounced DNA hypermethylation state of MSC. R-2HG also improved promotor methylation of lineage-specific genes during osteogenic and chondrogenic differentiation. In addition, R-2HG induced hypermethylation and decreased the mRNA levels of SHH, GLI1and GLI2, indicating Sonic Hedgehog (Shh) signaling inhibition. Conclusions The oncometabolite R-2HG dysregulated the chondrogenic and osteogenic differentiation of MSCs possibly via induction of DNA hypermethylation, improving the role of R-2HG in cartilaginous tumor development.

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 An Inhibitory Role of Osthole in Rat MSCs Osteogenic Differentiation and Proliferation via Wnt/β-Catenin and Erk1/2-MAPK Pathways

2016 ◽  
Vol 38 (6) ◽  
pp. 2375-2388 ◽  
Author(s):  
Hongyang Hu ◽  
Min Chen ◽  
Guangzu Dai ◽  
Guoqing Du ◽  
Xuezong Wang ◽  
...  
Keyword(s):  
Osteogenic Differentiation ◽  
Signaling Pathways ◽  
Western Blotting ◽  
Underlying Mechanism ◽  
Mapk Signaling ◽  
Cell Counting ◽  
Nuclear Antigen ◽  
Dependent Manner ◽  
Alizarin Red ◽  
Mapk Signaling Pathways

Background/Aims: Bone marrow-derived mesenchymal stem cells (MSCs) are responsible for new bone formation during adulthood. Accumulating evidences showed that Osthole promotes the osteogenic differentiation in primary osteoblasts. The aim of this study was to investigate whether Osthole exhibits a potential to stimulate the osteogenic differentiation of MSCs and the underlying mechanism. Methods: MSCs were treated with a gradient concentration of Osthole (6.25 µM, 12.5 µM, and 25 µM). Cell proliferation was assessed by western blotting with the proliferating cell nuclear antigen (PCNA) and Cyclin D1 antibodies, fluorescence activated cell sorting (FACS), and cell counting kit 8 (CCK8). MSCs were cultured in osteogenesis-induced medium for one or two weeks. The osteogenic differentiation of MSCs was estimated by Alkaline Phosphatase (ALP) staining, Alizarin red staining, Calcium influx, and quantitative PCR (qPCR). The underlying mechanism of Osthole-induced osteogenesis was further evaluated by western blotting with antibodies in Wnt/β-catenin, PI3K/Akt, BMPs/smad1/5/8, and MAPK signaling pathways. Results: Osthole inhibited proliferation of rat MSCs in a dose-dependent manner. Osthole suppressed osteogenic differentiation of rat MSCs by down-regulating the activities of Wnt/β-catenin and Erk1/2-MAPK signaling. Conclusions: Osthole inhibits the proliferation and osteogenic differentiation of rat MSCs, which might be mediated through blocking the Wnt/β-catenin and Erk1/2-MAPK signaling pathways.

scholarly journals Effects of TGF-β1 Overexpression on Biological Characteristics of Human Dental Pulp-derived Mesenchymal Stromal Cells

2019 ◽  
Vol 12 (1) ◽  
pp. 170-182 ◽  
Author(s):  
Hasan Salkın ◽  
Zeynep Burçin Gönen ◽  
Ergül Ergen ◽  
Dilek Bahar ◽  
Mustafa Çetin
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Biological Characteristics ◽  
Human Dental Pulp ◽  
Tgf Β1

Osteogenic Differentiation of Mesenchymal Stromal Cells: A Comparative Analysis Between Human Subcutaneous Adipose Tissue and Dental Pulp

2017 ◽  
Vol 26 (11) ◽  
pp. 843-855 ◽  
Author(s):  
Iolanda D'Alimonte ◽  
Filiberto Mastrangelo ◽  
Patricia Giuliani ◽  
Laura Pierdomenico ◽  
Marco Marchisio ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Comparative Analysis ◽  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Subcutaneous Adipose Tissue ◽  
Subcutaneous Adipose

scholarly journals Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

2021 ◽  
Vol 8 (11) ◽  
pp. 165
Author(s):  
Masanori Tsubosaka ◽  
Masahiro Maruyama ◽  
Elijah Ejun Huang ◽  
Ning Zhang ◽  
Takeshi Utsunomiya ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Osteogenic Differentiation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Early Stage ◽  
Genetically Modified ◽  
Interleukin 4 ◽  
Alizarin Red ◽  
Cell Based Therapy

The use of genetically modified (GM) mesenchymal stromal cells (MSCs) and preconditioned MSCs (pMSCs) may provide further opportunities to improve the outcome of core decompression (CD) for the treatment of early-stage osteonecrosis of the femoral head (ONFH). GM interleukin-4 (IL4) over-expressing MSCs (IL4-MSCs), platelet-derived growth factor (PDGF)-BB over-expressing MSCs (PDGF-BB-MSCs), and IL4-PDGF-BB co-over-expressing MSCs (IL4-PDGF-BB-MSCs) and their respective pMSCs were used in this in vitro study and compared with respect to cell proliferation and osteogenic differentiation. IL4-MSCs, PDGF-BB-MSCs, IL4-PDGF-BB-MSCs, and each pMSC treatment significantly increased cell proliferation compared to the MSC group alone. The percentage of Alizarin red-stained area in the IL4-MSC and IL4-pMSC groups was significantly lower than in the MSC group. However, the percentage of Alizarin red-stained area in the PDGF-BB-MSC group was significantly higher than in the MSC and PDGF-BB-pMSC groups. The percentage of Alizarin red-stained area in the IL4-PDGF-BB-pMSC was significantly higher than in the IL4-PDGF-BB-MSC group. There were no significant differences in the percentage of Alizarin red-stained area between the MSC and IL4-PDGF-BB-pMSC groups. The use of PDGF-BB-MSCs or IL4-PDGF-BB-pMSCs increased cell proliferation. Furthermore, PDGF-BB-MSCs promoted osteogenic differentiation. The addition of GM MSCs may provide a useful supplementary cell-based therapy to CD for treatment of ONFH.

scholarly journals Functional differences in mesenchymal stromal cells from human dental pulp and periodontal ligament

2014 ◽  
Vol 18 (2) ◽  
pp. 344-354 ◽  
Author(s):  
Anoop Babu Vasandan ◽  
Shilpa Rani Shankar ◽  
Priya Prasad ◽  
Vulugundam Sowmya Jahnavi ◽  
Ramesh Ramachandra Bhonde ◽  
...  
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Dental Pulp ◽  
Periodontal Ligament ◽  
Functional Differences ◽  
Human Dental Pulp
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