Engineered scaffolds based on mesenchymal stem cells/preosteoclasts extracellular matrix promote bone regeneration

Recently, extracellular matrix-based tissue-engineered bone is a promising approach to repairing bone defects, and the seed cells are mostly mesenchymal stem cells. However, bone remodelling is a complex biological process, in which osteoclasts perform bone resorption and osteoblasts dominate bone formation. The interaction and coupling of these two kinds of cells is the key to bone repair. Therefore, the extracellular matrix secreted by the mesenchymal stem cells alone cannot mimic a complex bone regeneration microenvironment, and the addition of extracellular matrix by preosteoclasts may contribute as an effective strategy for bone regeneration. Here, we established the mesenchymal stem cell/preosteoclast extracellular matrix -based tissue-engineered bones and demonstrated that engineered-scaffolds based on mesenchymal stem cell/ preosteoclast extracellular matrix significantly enhanced osteogenesis in a 3 mm rat femur defect model compared with mesenchymal stem cell alone. The bioactive proteins released from the mesenchymal stem cell/ preosteoclast extracellular matrix based tissue-engineered bones also promoted the migration, adhesion, and osteogenic differentiation of mesenchymal stem cells in vitro. As for the mechanisms, the iTRAQ-labeled mass spectrometry was performed, and 608 differentially expressed proteins were found, including the IGFBP5 and CXCL12. Through in vitro studies, we proved that CXCL12 and IGFBP5 proteins, mainly released from the preosteoclasts, contributed to mesenchymal stem cells migration and osteogenic differentiation, respectively. Overall, our research, for the first time, introduce pre-osteoclast into the tissue engineering of bone and optimize the strategy of constructing extracellular matrix–based tissue-engineered bone using different cells to simulate the natural bone regeneration environment, which provides new sight for bone tissue engineering.

Download Full-text

Osteogenic Differentiation Potential of Human Bone Marrow and Amniotic Fluid-Derived Mesenchymal Stem Cells in Vitro & in Vivo

Open Access Macedonian Journal of Medical Sciences ◽

10.3889/oamjms.2019.124 ◽

2019 ◽

Vol 7 (4) ◽

pp. 507-515 ◽

Cited By ~ 4

Author(s):

Eman E. A. Mohammed ◽

Mohamed El-Zawahry ◽

Abdel Razik H. Farrag ◽

Nahla N. Abdel Aziz ◽

Wessam Sharaf-ElDin ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Amniotic Fluid ◽

Bone Regeneration ◽

Osteogenic Differentiation ◽

Differentiation Potential

BACKGROUND: Cell therapies offer a promising potential in promoting bone regeneration. Stem cell therapy presents attractive care modality in treating degenerative conditions or tissue injuries. The rationale behind this is both the expansion potential of stem cells into a large cell population size and its differentiation abilities into a wide variety of tissue types, when given the proper stimuli. A progenitor stem cell is a promising source of cell therapy in regenerative medicine and bone tissue engineering. AIM: This study aimed to compare the osteogenic differentiation and regenerative potentials of human mesenchymal stem cells derived from human bone marrow (hBM-MSCs) or amniotic fluid (hAF-MSCs), both in vitro and in vivo studies. SUBJECTS AND METHODS: Human MSCs, used in this study, were successfully isolated from two human sources; the bone marrow (BM) and amniotic fluid (AF) collected at the gestational ages of second or third trimesters. RESULTS: The stem cells derived from amniotic fluid seemed to be the most promising type of progenitor cells for clinical applications. In a pre-clinical experiment, attempting to explore the therapeutic application of MSCs in bone regeneration, Rat lumbar spines defects were surgically created and treated with undifferentiated and osteogenically differentiated MSCs, derived from BM and second trimester AF. Cells were loaded on gel-foam scaffolds, inserted and fixed in the area of the surgical defect. X-Ray radiography follows up, and histopathological analysis was done three-four months post- operation. The transplantation of AF-MSCs or BM-MSCs into induced bony defects showed promising results. The AF-MSCs are offering a better healing effect increasing the likelihood of achieving successful spinal fusion. Some bone changes were observed in rats transplanted with osteoblasts differentiated cells but not in rats transplanted with undifferentiated MSCs. Longer observational periods are required to evaluate a true bone formation. The findings of this study suggested that the different sources; hBM-MSCs or hAF-MSCs exhibited remarkably different signature regarding the cell morphology, proliferation capacity and osteogenic differentiation potential CONCLUSIONS: AF-MSCs have a better performance in vivo bone healing than that of BM-MSCs. Hence, AF derived MSCs is highly recommended as an alternative source to BM-MSCs in bone regeneration and spine fusion surgeries. Moreover, the usage of gel-foam as a scaffold proved as an efficient cell carrier that showed bio-compatibility with cells, bio-degradability and osteoinductivity in vivo.

Download Full-text

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.

Download Full-text

Emilin-2 is a component of bone marrow extracellular matrix regulating mesenchymal stem cell differentiation and hematopoietic progenitors

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02674-2 ◽

2022 ◽

Vol 13 (1) ◽

Author(s):

Francesco Da Ros ◽

Luca Persano ◽

Dario Bizzotto ◽

Mariagrazia Michieli ◽

Paola Braghetta ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Extracellular Matrix ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Adipogenic Differentiation ◽

Stem Cell Differentiation ◽

Dependent Manner ◽

Hematopoietic Stem

Abstract Background Dissection of mechanisms involved in the regulation of bone marrow microenvironment through cell–cell and cell–matrix contacts is essential for the detailed understanding of processes underlying bone marrow activities both under physiological conditions and in hematologic malignancies. Here we describe Emilin-2 as an abundant extracellular matrix component of bone marrow stroma. Methods Immunodetection of Emilin-2 was performed in bone marrow sections of mice from 30 days to 6 months of age. Emilin-2 expression was monitored in vitro in primary and mesenchymal stem cell lines under undifferentiated and adipogenic conditions. Hematopoietic stem cells and progenitors in bone marrow of 3- to 10-month-old wild-type and Emilin-2 null mice were analyzed by flow cytometry. Results Emilin-2 is deposited in bone marrow extracellular matrix in an age-dependent manner, forming a meshwork that extends from compact bone boundaries to the central trabecular regions. Emilin-2 is expressed and secreted by both primary and immortalized bone marrow mesenchymal stem cells, exerting an inhibitory action in adipogenic differentiation. In vivo Emilin-2 deficiency impairs the frequency of hematopoietic stem/progenitor cells in bone marrow during aging. Conclusion Our data provide new insights in the contribution of bone marrow extracellular matrix microenvironment in the regulation of stem cell niches and hematopoietic progenitor differentiation.

Download Full-text

Human Adipose-Derived Mesenchymal Stem Cells-Incorporated Silk Fibroin as a Potential Bio-Scaffold in Guiding Bone Regeneration

Polymers ◽

10.3390/polym12040853 ◽

2020 ◽

Vol 12 (4) ◽

pp. 853 ◽

Cited By ~ 1

Author(s):

Dewi Sartika ◽

Chih-Hsin Wang ◽

Ding-Han Wang ◽

Juin-Hong Cherng ◽

Shu-Jen Chang ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Bone Regeneration ◽

Osteogenic Differentiation ◽

Silk Fibroin ◽

Bone Repair ◽

Matrix Deposition ◽

Calvarial Defects

Recently, stem cell-based bone tissue engineering (BTE) has been recognized as a preferable and clinically significant strategy for bone repair. In this study, a pure 3D silk fibroin (SF) scaffold was fabricated as a BTE material using a lyophilization method. We aimed to investigate the efficacy of the SF scaffold with and without seeded human adipose-derived mesenchymal stem cells (hASCs) in facilitating bone regeneration. The effectiveness of the SF-hASCs scaffold was evaluated based on physical characterization, biocompatibility, osteogenic differentiation in vitro, and bone regeneration in critical rat calvarial defects in vivo. The SF scaffold demonstrated superior biocompatibility and significantly promoted osteogenic differentiation of hASCs in vitro. At six and twelve weeks postimplantation, micro-CT showed no statistical difference in new bone formation amongst all groups. However, histological staining results revealed that the SF-hASCs scaffold exhibited a better bone extracellular matrix deposition in the defect regions compared to other groups. Immunohistochemical staining confirmed this result; expression of osteoblast-related genes (BMP-2, COL1a1, and OCN) with the SF-hASCs scaffold treatment was remarkably positive, indicating their ability to achieve effective bone remodeling. Thus, these findings demonstrate that SF can serve as a potential carrier for stem cells, to be used as an osteoconductive bioscaffold for BTE applications.

Download Full-text

Rapid Rapamycin-Only Induced Osteogenic Differentiation of Blood-Derived Stem Cells and Their Adhesion to Natural and Artificial Scaffolds

Stem Cells International ◽

10.1155/2017/2976541 ◽

2017 ◽

Vol 2017 ◽

pp. 1-12

Author(s):

Carpentieri Arianna ◽

Cozzoli Eliana ◽

Acri Flavio ◽

Ranalli Marco ◽

Diedenhofen Giacomo ◽

...

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Stem Cell ◽

Osteogenic Differentiation ◽

Molecular Mechanisms ◽

Maxillofacial Surgery ◽

Ongoing Research ◽

Medicine Research ◽

Cell Based Therapy

Stem cells are a centerpiece of regenerative medicine research, and the recent development of adult stem cell-based therapy systems has vigorously expanded the scope and depth of this scientific field. The regeneration of damaged and/or degraded bone tissue in orthopedic, dental, or maxillofacial surgery is one of the main areas where stem cells and their regenerative potential could be used successfully, requiring tissue engineering solutions incorporating an ideal stem cell type paired with the correct mechanical support. Our contribution to this ongoing research provides a new model of in vitro osteogenic differentiation using blood-derived stem cells (BDSCs) and rapamycin, visibly expressing typical osteogenic markers within ten days of treatment. In depth imaging studies allowed us to observe the adhesion, proliferation, and differentiation of BDSCs to both titanium and bone scaffolds. We demonstrate that BDSCs can differentiate towards the osteogenic lineage rapidly, while readily adhering to the scaffolds we exposed them to. Our results show that our model can be a valid tool to study the molecular mechanisms of osteogenesis while tailoring tissue engineering solutions to these new insights.

Download Full-text

Osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells encapsulated in Thai silk fibroin/collagen hydrogel: a pilot study in vitro

Asian Biomedicine ◽

10.1515/abm-2019-0030 ◽

2019 ◽

Vol 12 (6) ◽

pp. 273-279 ◽

Cited By ~ 1

Author(s):

Jirun Apinun ◽

Sittisak Honsawek ◽

Somsak Kuptniratsaikul ◽

Jutarat Jamkratoke ◽

Sorada Kanokpanont

Keyword(s):

Tissue Engineering ◽

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Pilot Study ◽

Osteogenic Differentiation ◽

Collagen Hydrogel ◽

Collagen Hydrogels ◽

Rat Bone

AbstractBackgroundSilk fibroin (SF) can be processed into a hydrogel. SF/collagen hydrogel may be a suitable biomaterial for bone tissue engineering.ObjectivesTo investigate in vitro biocompatibility and osteogenic potential of encapsulated rat bone marrow-derived mesenchymal stem cells (rat MSCs) in an injectable Thai SF/collagen hydrogel induced by oleic acid–poloxamer 188 surfactant mixture in an in vitro pilot study.MethodsRat MSCs were encapsulated in 3 groups of hydrogel scaffolds (SF, SF with 0.05% collagen [SF/0.05C], and SF with 0.1% collagen [SF/0.1C]) and cultured in a growth medium and an osteogenic induction medium. DNA, alkaline phosphatase (ALP) activity, and calcium were assayed at periodically for up to 5 weeks. After 6 weeks of culture the cells were analyzed by scanning electron microscopy and energy dispersive spectroscopy.ResultsAlthough SF hydrogel with collagen seems to have less efficiency to encapsulate rat MSCs, their plateau phase growth in all hydrogels was comparable. Inability to maintain cell viability as cell populations declined over 1–5 days was observed. Cell numbers then plateaued and were maintained until day 14 of culture. ALP activity and calcium content of rat MSCs in SF/collagen hydrogels were highest at day 21. An enhancing effect of collagen combined with the hydrogel was observed for proliferation and matrix formation; however, benefits of the combination on osteogenic differentiation and biomineralization are as yet unclear.ConclusionRat MSCs in SF and SF/collagen hydrogels showed osteogenic differentiation. Accordingly, these hydrogels may serve as promising scaffolds for bone tissue engineering.

Download Full-text

204 ISOLATION, CHARACTERIZATION, AND DIFFERENTIATION OF ADIPOSE TISSUE DERIVED MESENCHYMAL STEM CELLS: AN AUTOLOGOUS TRANSPLANTATION TO PATIENTS

Reproduction Fertility and Development ◽

10.1071/rdv26n1ab204 ◽

2014 ◽

Vol 26 (1) ◽

pp. 216 ◽

Cited By ~ 1

Author(s):

H. N. Malik ◽

A. Dubey ◽

D. K. Singhal ◽

S. Saugandhika ◽

S. Boeteng ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Adipose Tissue ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Hip Dysplasia ◽

Autologous Transplantation ◽

Embryonic Stem ◽

Patient Specific

Adult stem cells derived from all possible sources of livestock serve as the best possible alternative to embryonic stem cells. The discovery of mesenchymal stem cells has provided the new horizon to stem cell therapy. Adipose tissue derived mesenchymal stem cell (ADSCs), an easy source of adult stem cell has created a lot of interest among researchers as patient specific treatment and autologous transplantation in animals is becoming a viable option. The proposed study was carried out for 1) isolation of ADSCs from dogs, suffering from hip dysplasia or from paraplegia, 2) ADSC characterisation and in vitro differentiation ability into osteocytes, chondrocytes, adipocytes and neurocytes specific cells. Adipose tissues were collected from belly/umbilical cord region. ADSCs were isolated by enzymatic digestion method followed by enriching through a 41 μm filter. Filtered cells were then resuspended in cell culture flasks containing growth enriching medium and cultured in 5% CO2 in air at 37°C for 5 days. ADSCs were characterised by amplification of mesenchymal stem cell specific markers i.e. CD29, CD44, CD90, and CD166 and by immunocytochemistry of mesenchymal stem cell specific protein i.e. CD44 and CD90. ADSCs were further in vitro differentiated. ADSCs derived osteocytes, chondrocytes, and adipocytes were validated through the amplification of specific markers of osteocytes (Osteopontin, Collagen I); chondrocytes (Aggrecan and Collagen II) and adipocytes (LPL, PPARα, PPARγ). Dog ADSCs were further autogenic transplanted into hip dysplasia and paraplegic patients. These patients recovered well one month from transplantation and were able to move freely. It may be concluded that these findings may have implications for defining the physiological roles of ADSCs in arthritis; orthopaedic ailments, joint regeneration, neuronal disorders, and several other applications leading to novel therapeutic opportunities.

Download Full-text

Osteogenic Response of Human Mesenchymal Stem Cells Analysed Using Combined Intracellular and Extracellular Metabolomic Monitoring

Cellular Physiology and Biochemistry ◽

10.33594/000000377 ◽

2021 ◽

Vol 55 (3) ◽

pp. 311-326

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Osteogenic Differentiation ◽

Bone Microarchitecture ◽

Human Mesenchymal Stem Cells ◽

Stem Cell Differentiation ◽

Extracellular Metabolite ◽

Osteogenic Response

Background/Aims: The skeleton is a metabolically active organ undergoing continuous remodelling initiated by mesenchymal progenitors present in bone and bone marrow. Under certain pathological conditions this remodelling balance shifts towards increased resorption resulting in weaker bone microarchitecture, and there is consequently a therapeutic need to identify pathways that could inversely enhance bone formation from stem cells. Metabolomics approaches recently applied to stem cell characterisation could help identify new biochemical markers involved in osteogenic differentiation. Methods: Combined intra- and extracellular metabolite profiling was performed by liquid chromatography-mass spectrometry (LC-MS) on human mesenchymal stem cells (MSCs) undergoing osteogenic differentiation in vitro. Using a combination of univariate and multivariate analyses, changes in metabolite and nutrient concentration were monitored in cultures under osteogenic treatment over 10 days. Results: A subset of differentially detected compounds was identified in differentiating cells, suggesting a direct link to metabolic processes involved in osteogenic response. Conclusion: These results highlight new metabolite candidates as potential biomarkers to monitor stem cell differentiation towards the bone lineage.

Download Full-text

Bio-functionalization and in-vitro evaluation of titanium surface with recombinant fibronectin and elastin fragment in human mesenchymal stem cell

PLoS ONE ◽

10.1371/journal.pone.0260760 ◽

2021 ◽

Vol 16 (12) ◽

pp. e0260760

Author(s):

Bo-Hyun Park ◽

Eui-Seung Jeong ◽

Sujin Lee ◽

Jun-Hyeog Jang

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Bone Regeneration ◽

Osteogenic Differentiation ◽

Cellular Response ◽

Titanium Surface ◽

Stem Cell Differentiation ◽

Binding Motif ◽

Type I

Titanium is a biomaterial that meets a number of important requirements, including excellent mechanical and chemical properties, but has low bioactivity. To improve cellular response onto titanium surfaces and hence its osseointegration, the titanium surface was bio-functionalized to mimic an extracellular matrix (ECM)-like microenvironment that positively influences the behavior of stem cells. In this respect, fibronectin and elastin are important components of the ECM that regulate stem cell differentiation by supporting the biological microenvironment. However, each native ECM is unsuitable due to its high production cost and immunogenicity. To overcome these problems, a recombinant chimeric fibronectin type III9-10 and elastin-like peptide fragments (FN9-10ELP) was developed herein and applied to the bio-functionalized of the titanium surface. An evaluation of the biological activity and cellular responses with respect to bone regeneration indicated a 4-week sustainability on the FN9-10ELP functionalized titanium surface without an initial burst effect. In particular, the adhesion and proliferation of human mesenchymal stem cells (hMSCs) was significantly increased on the FN9-10ELP coated titanium compared to that observed on the non-coated titanium. The FN9-10ELP coated titanium induced osteogenic differentiation such as the alkaline phosphatase (ALP) activity and mineralization activity. In addition, expressions of osteogenesis-related genes such as a collagen type I (Col I), Runt-related transcription factor 2 (RUNX2), osteopontin (OPN), osteocalcin (OCN), bone sialo protein (BSP), and PDZ-binding motif (TAZ) were further increased. Thus, in vitro the FN9-10ELP functionalization titanium not only sustained bioactivity but also induced osteogenic differentiation of hMSCs to improve bone regeneration.

Download Full-text

ID: 1032 Differentiation Potential and Characteristics of Mesenchymal Stem Cells Isolated from Human Umbilical Cord membrane to Hepatocyte-like Cells

Biomedical Research and Therapy ◽

10.15419/bmrat.v4is.308 ◽

2017 ◽

Vol 4 (S) ◽

pp. 107

Author(s):

Trung Kien Do ◽

Van Hanh Nguyen ◽

Huu Duc Nguyen ◽

Chu Hoang Ha

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Umbilical Cord ◽

Gene Transfection ◽

Human Umbilical Cord ◽

Specific Marker ◽

Differentiation Potential

Recent studies indicated that Mesenchymal stem cell has become a potential objective for therapy. In this study, umbilical cord cells were isolated and analyzed the expression of mesenchymal stem cells specific markers then they were differentiated into hepatocyte-like cells by DMSO and Gene transfection. Umbilical cord mesenchymal stem cell (MSC) was isolated by explant culture in media DMEM/F12, complementing with growth factors EGF, FGF and IST. After that, they were exposured to DMSO with three concentrations: 0.01%, 0.1%, 1% and another group was transfection with HNF4α by Lipofectamin LX plus. The cells were analyzed at 1, 2, 3, and 4 weeks after treatment. The cells isolation was shown the positive with markers CD73, CD34, CD86, CD90, CD105, eras, Oct 1, GATA, and negative with markers HNF4α, Alb and G6P. In group 0,1% DMSO treatment, after 3 weeks the cells were positive with markers HNF4α but it was also negative with markers Alb and G6P. In the transfection group, the cell expresses HNF4α at three weeks after treatment. Although our results exposure that the umbilical cord mesenchymal stem cells expressed hepatic specific marker after DMSO induced and DNA treatment. So it will be necessary to optimize research conditional and investigate the hepatic functions of these cells in a longer in vitro culture.

Download Full-text