PP2A in LepR+ mesenchymal stem cells contributes to embryonic and postnatal endochondral ossification through Runx2 dephosphorylation

AbstractIt has not been well studied which cells and related mechanisms contribute to endochondral ossification. Here, we fate mapped the leptin receptor-expressing (LepR+) mesenchymal stem cells (MSCs) in different embryonic and adult extremities using Lepr-cre; tdTomato mice and investigated the underling mechanism using Lepr-cre; Ppp2r1afl/fl mice. Tomato+ cells appear in the primary and secondary ossification centers and express the hypertrophic markers. Ppp2r1a deletion in LepR+ MSCs reduces the expression of Runx2, Osterix, alkaline phosphatase, collagen X, and MMP13, but increases that of the mature adipocyte marker perilipin, thereby reducing trabecular bone density and enhancing fat content. Mechanistically, PP2A dephosphorylates Runx2 and BRD4, thereby playing a major role in positively and negatively regulating osteogenesis and adipogenesis, respectively. Our data identify LepR+ MSC as the cell origin of endochondral ossification during embryonic and postnatal bone growth and suggest that PP2A is a therapeutic target in the treatment of dysregulated bone formation.

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Clumps of Mesenchymal Stem Cells/Extracellular Matrix Complexes Generated with Xeno-Free Chondro-Inductive Medium induce Bone Regeneration via Endochondral Ossification

Biomedicines ◽

10.3390/biomedicines9101408 ◽

2021 ◽

Vol 9 (10) ◽

pp. 1408

Author(s):

Susumu Horikoshi ◽

Mikihito Kajiya ◽

Souta Motoike ◽

Mai Yoshino ◽

Shin Morimoto ◽

...

Keyword(s):

Stem Cells ◽

Extracellular Matrix ◽

Mesenchymal Stem Cells ◽

Bone Formation ◽

Bone Regeneration ◽

Endochondral Ossification ◽

Healing Process ◽

Early Period ◽

Cartilage Matrix

Three-dimensional clumps of mesenchymal stem cells (MSCs)/extracellular matrix (ECM) complexes (C-MSCs) can be transplanted into tissue defect site with no artificial scaffold. Importantly, most bone formation in the developing process or fracture healing proceeds via endochondral ossification. Accordingly, this present study investigated whether C-MSCs generated with chondro-inductive medium (CIM) can induce successful bone regeneration and assessed its healing process. Human bone marrow-derived MSCs were cultured with xeno-free/serum-free (XF) growth medium. To obtain C-MSCs, confluent cells that had formed on the cellular sheet were scratched using a micropipette tip and then torn off. The sheet was rolled to make a round clump of cells. The cell clumps, i.e., C-MSCs, were maintained in XF-CIM. C-MSCs generated with XF-CIM showed enlarged round cells, cartilage matrix, and hypertrophic chondrocytes genes elevation in vitro. Transplantation of C-MSCs generated with XF-CIM induced successful bone regeneration in the SCID mouse calvaria defect model. Immunofluorescence staining for human-specific vimentin demonstrated that donor human and host mouse cells cooperatively contributed the bone formation. Besides, the replacement of the cartilage matrix into bone was observed in the early period. These findings suggested that cartilaginous C-MSCs generated with XF-CIM can induce bone regeneration via endochondral ossification.

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Osteogenic differentiation of adipose tissue-derived mesenchymal stem cells cultured with different concentrations of prolactin

Arquivo Brasileiro de Medicina Veterinária e Zootecnia ◽

10.1590/1678-4162-9364 ◽

2017 ◽

Vol 69 (6) ◽

pp. 1573-1580

Author(s):

K.P. Oliveira ◽

A.M.S. Reis ◽

A.P. Silva ◽

C.L.R. Silva ◽

A.M. Goes ◽

...

Keyword(s):

Stem Cells ◽

Alkaline Phosphatase ◽

Mesenchymal Stem Cells ◽

Adipose Tissue ◽

Osteogenic Differentiation ◽

Bone Sialoprotein ◽

Collagen I ◽

Female Rats ◽

Osteogenic Potential ◽

Vitro Effect

ABSTRACT The objective was to evaluate the in vitro effect of prolactin in osteogenic potential of adipose tissue-derived mesenchymal stem cells (ADSCs) in female rats. ADSCs were cultured in osteogenic medium with and without the addition of prolactin and distributed into three groups: 1) ADSCs (control), 2) ADSCs with addition of 100ng/mL of prolactin and 3) ADSCs with addition of 300ng/mL of prolactin. At 21 days of differentiation, the tests of MTT conversion into formazan crystals, percentage of mineralized nodules and cells per field and quantification of genic transcript for alkaline phosphatase, osteopontin, osteocalcin, bone sialoprotein, BMP-2 and collagen I by real-time RT-PCR were made. The addition of prolactin reduced the conversion of MTT in group 3 and increased the percentage of cells per field in the groups 2 and 3, however without significantly increasing the percentage of mineralized nodules and the expression of alkaline phosphatase, osteopontin, osteocalcin, bone sialoprotein, BMP-2 and collagen I. In conclusion, the addition of prolactin in concentrations of 100ng/mL and 300ng/mL does not change the osteogenic differentiation to the ADSCs of female rats despite increase in the cellularity of the culture.

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Micro-Computed Tomography Analysis on Administration of Mesenchymal Stem Cells - Bovine Teeth Scaffold Composites for Alveolar Bone Tissue Engineering

Journal of Biomimetics Biomaterials and Biomedical Engineering ◽

10.4028/www.scientific.net/jbbbe.52.86 ◽

2021 ◽

Vol 52 ◽

pp. 86-96

Author(s):

Desi Sandra Sari ◽

Fourier Dzar Eljabbar Latief ◽

Ferdiansyah ◽

Ketut Sudiana ◽

Fedik Abdul Rantam

Keyword(s):

Computed Tomography ◽

Tissue Engineering ◽

Stem Cells ◽

Mesenchymal Stem Cells ◽

Bone Regeneration ◽

Bone Growth ◽

Alveolar Bone ◽

Micro Computed Tomography ◽

Freeze Dried ◽

Significant Difference

The tissue engineering approach for periodontal tissue regeneration using a combination of stem cells and scaffold has been vastly developed. Mesenchymal Stem Cells (MSCs) seeded with Bovine Teeth Scaffold (BTSc) can repair alveolar bone damage in periodontitis cases. The alveolar bone regeneration process was analyzed by micro-computed tomography (µ-CT) to observe the structure of bone growth and to visualize the scaffold in 3-Dimensional (3D). The purpose of this study is to analyze alveolar bone regeneration by µ-CT following the combination of MSCs and bovine teeth scaffold (MSCs-BTSc) implantation in the Wistar rat periodontitis model. Methods. MSCs were cultured from adipose-derived mesenchymal stem cells of rats. BTSc was taken from bovine teeth and freeze-dried with a particle size of 150-355 µm. MSCs were seeded on BTSc for 24 hours and transplanted in a rat model of periodontitis. Thirty-five Wistar rats were made as periodontitis models with LPS induction from P. gingivalis injected to the buccal section of interproximal gingiva between the first and the second mandibular right-molar teeth for six weeks. There were seven groups (control group, BTSc group on day 7, BTSc group on day 14, BTSc group on day 28, MSCs-BTSc group on day 7, MSCs-BTSc group on day 14, MSCs-BTSc group on day 28). The mandibular alveolar bone was analyzed and visualized in 3D with µ-CT to observe any new bone growth. Statistical Analysis. Group data were subjected to the Kruskal Wallis test followed by the Mann-Whitney (p <0.05). The µ-CT qualitative analysis shows a fibrous structure, which indicates the existence of new bone regeneration. Quantitative analysis of the periodontitis model showed a significant difference between the control model and the model with the alveolar bone resorption (p <0.05). The bone volume and density measurements revealed that the MSCs-BTSc group on day 28 formed new bone compared to other groups (p <0.05). Administration of MSCs-BTSc combination has the potential to form new alveolar bone.

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3D bioprinting of mesenchymal stem cells and endothelial cells in an alginate-gelatin-based bioink

Journal of 3D Printing in Medicine ◽

10.2217/3dp-2020-0026 ◽

2021 ◽

Author(s):

Sindhuja D Eswaramoorthy ◽

Nandini Dhiman ◽

Akshay Joshi ◽

Subha N Rath

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Alkaline Phosphatase ◽

Endothelial Cells ◽

Mesenchymal Stem Cells ◽

Osteogenic Differentiation ◽

Alkaline Phosphatase Activity ◽

Histochemical Staining ◽

Vital Role ◽

3D Bioprinting

Aim: Bioink is one of the essential factors in 3D bioprinting that determines the fate of cells, in our case, umbilical cord-derived mesenchymal stem cells (UMSC). The aim was to determine if the presence of the osteoinductive factors in the bioink enhances osteodifferentiation as compared with adding them postprinting and if the UMSC and endothelial cells (EC) coculture result in better osteodifferentiation. Materials & methods: Alginate-gelatin along with UMSC–EC were bioprinted using an extrusion 3D bioprinter. Results & conclusion: The UMSC–EC interaction, as well as intrinsic addition of the differentiation components in the bioink, were observed to play a vital role in increasing the osteogenic differentiation as shown by the histochemical staining, alkaline phosphatase activity and gene expression of osteogenic markers.

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Mesenchymal Stem Cells Coated with Synthetic Bone-Targeting Polymers Enhance Osteoporotic Bone Fracture Regeneration

Bioengineering ◽

10.3390/bioengineering7040125 ◽

2020 ◽

Vol 7 (4) ◽

pp. 125

Author(s):

Yuliya Safarova (Yantsen) ◽

Farkhad Olzhayev ◽

Bauyrzhan Umbayev ◽

Andrey Tsoy ◽

Gonzalo Hortelano ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Bone Density ◽

Active Sites ◽

Bone Fractures ◽

Bone Fragility ◽

Female Rats ◽

Osteoporotic Bone ◽

Osteoblast Progenitor ◽

Osteoclastic Activity

Osteoporosis is a progressive skeletal disease characterized by reduced bone density leading to bone fragility and an elevated risk of bone fractures. In osteoporotic conditions, decrease in bone density happens due to the augmented osteoclastic activity and the reduced number of osteoblast progenitor cells (mesenchymal stem cells, MSCs). We investigated a new method of cell therapy with membrane-engineered MSCs to restore the osteoblast progenitor pool and to inhibit osteoclastic activity in the fractured osteoporotic bones. The primary active sites of the polymer are the N-hydroxysuccinimide and bisphosphonate groups that allow the polymer to covalently bind to the MSCs’ plasma membrane, target hydroxyapatite molecules on the bone surface and inhibit osteolysis. The therapeutic utility of the membrane-engineered MSCs was investigated in female rats with induced estrogen-dependent osteoporosis and ulnar fractures. The analysis of the bone density dynamics showed a 27.4% and 21.5% increase in bone density at 4 and 24 weeks after the osteotomy of the ulna in animals that received four transplantations of polymer-modified MSCs. The results of the intravital observations were confirmed by the post-mortem analysis of histological slices of the fracture zones. Therefore, this combined approach that involves polymer and cell transplantation shows promise and warrants further bio-safety and clinical exploration.

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Do Autologous Mesenchymal Stem Cells Augment Bone Growth and Contact to Massive Bone Tumor Implants?

Tissue Engineering ◽

10.1089/ten.2006.12.ft-171 ◽

2006 ◽

Vol 0 (0) ◽

pp. 060713121012001

Author(s):

Priya Kalia ◽

Gordon W. Blunn ◽

Jemima Miller ◽

Aprajit Bhalla ◽

Michael Wiseman ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Bone Tumor ◽

Bone Growth ◽

Autologous Mesenchymal Stem Cells ◽

Massive Bone

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Bone Growth Capacity of Human Umbilical Cord Mesenchymal Stem Cells and BMP-2 Seeded Into Hydroxyapatite/Chitosan/Gelatin Scaffold in Alveolar Cleft Defects: An Experimental Study in Goat

The Cleft Palate-Craniofacial Journal ◽

10.1177/1055665620962360 ◽

2020 ◽

pp. 105566562096236

Author(s):

Kristaninta Bangun ◽

Chaula L. Sukasah ◽

Ismail H. Dilogo ◽

Decky J. Indrani ◽

Nurjati Chairani Siregar ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Bone Growth ◽

Alveolar Bone ◽

Operating Time ◽

Inflammatory Cells ◽

Capra Hircus ◽

Functional Score ◽

Human Umbilical Cord ◽

Group Design

Objective: To evaluate bone regeneration in alveolar defects treated with human umbilical cord–derived mesenchymal stem cells (hUCMSCs), hydroxyapatite/chitosan/gelatin (HA/CS/Gel) scaffold, and bone morphogenic protein-2 (BMP-2) in Capra hircus models. Design: Randomized posttest-only control group design. Setting: Animal Hospital at Bogor Agricultural Institute. Participants: Healthy and equally treated 24 female Capra hircus/goats. Intervention: Animals were randomly assigned to 3 experimental group design (iliac crest alveolar bone graft/ICABG [control], HA/Cs/Gel+BMP-2 [ Novosys], and HA/Cs/Gel+BMP-2+UCMSCs). Graft materials were implanted in surgically made alveolar defects. Main Outcome Measures: Postoperative functional score and operating time were assessed. New bone growth, bone density, inflammatory cells recruitment, and neoangiogenesis were evaluated based on radiological and histological approach at 2 time points, week 4 and 12. Statistical analysis was done between treatment groups. Results: Operating time was 34% faster and functional score 94.5% more superior in HA/Cs/Gel+BMP-2+hUCMSC group. Bone growth capacity in HA/Cs/Gel+BMP-2+UCMSCs mimicked ICABG, but ICABG showed possibility of bone loss between week 4 and 12. The HA/Cs/Gel+BMP-2+UCMSCs showed early bone repopulation and unseen inflammatory cells and angiogenesis on week 12. Discussion and Conclusion: The HA/Cs/Gel+BMP-2+hUCMSCs were superior in enhancing new bone growth without donor site morbidity compared to ICABG. The presence of hUCMSCs in tissue-engineered alveolar bone graft (ABG), supported with paracrine activity of the resident stem cells, initiated earlier new bone repopulation, and completed faster bone regeneration. The HA/Cs/Gel scaffold seeded with UCMSCs+BMP-2 is a safe substitute of ICABG to close alveolar bone defects suitable for patients with cleft lip, alveolus, and palate.

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