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Ran C, Liu T, Bao Y, Wang W, Xue D, Yin G, Zhang X, Zhao D. Proteoglycans Enhance the Therapeutic Effect of BMSC Transplantation on Osteoarthritis. Bioengineering (Basel) 2024; 11:1167. [PMID: 39593826 PMCID: PMC11592059 DOI: 10.3390/bioengineering11111167] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2024] [Revised: 11/01/2024] [Accepted: 11/08/2024] [Indexed: 11/28/2024] Open
Abstract
BACKGROUND The injection of bone mesenchymal stem cells (BMSCs) for osteoarthritis (OA) treatment fails to address the disrupted extracellular microenvironment, limiting the differentiation and paracrine functions of BMSCs and resulting in suboptimal therapeutic outcomes. Proteoglycans (PGs) promote cell differentiation, tissue repair, and microenvironment remodeling. This study investigated the potential of combining PGs with BMSCs to increase the efficacy of OA treatment. METHODS We evaluated the effects of PG on BMSC and chondrocyte functions by adding various PG concentrations to the culture media. Additionally, a Transwell system was used to assess the impact of PG on the communication between BMSCs and chondrocytes. The results of the in vitro experiment were verified by tissue staining and immunohistochemistry following the treatment of OA model rats. RESULTS Our findings indicate that PG effectively induces Col II expression in BMSCs and enhances the paracrine secretion of TGF-β1, thereby activating the TGF-β signaling pathway in chondrocytes and increasing PRG4 gene expression. Compared with the other groups, the BMSC/PG treatment group presented a smoother articular surface and more robust extracellular matrix than the other groups in vivo, with significantly increased expression and distribution of Smad2/3 and PRG4. CONCLUSIONS PG enhances BMSC differentiation into chondrocytes and stimulates paracrine TGF-β1 secretion. Proteoglycans not only promote chondrocyte differentiation and paracrine TGF-β1 signaling in BMSCs but also increase the sensitivity of chondrocytes to TGF-β1 secreted from BMSCs, leading to PRG4 expression through the TGFR/Smad2/3 pathway. Proteoglycans can enhance the therapeutic effect of BMSC treatment on OA and have the potential to delay the degeneration of OA cartilage.
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Affiliation(s)
- Chunxiao Ran
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Tianhao Liu
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Yongming Bao
- Department of Life Science and Biotechnology, Dalian University of Technology, Dalian 116024, China;
| | - Weidan Wang
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Dongling Xue
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Guangxiao Yin
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Xiuzhi Zhang
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
| | - Dewei Zhao
- Department of Orthopedics, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China; (C.R.); (W.W.); (D.X.); (G.Y.)
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Roncada T, Blunn G, Roldo M. Collagen and Alginate Hydrogels Support Chondrocytes Redifferentiation In Vitro without Supplementation of Exogenous Growth Factors. ACS OMEGA 2024; 9:21388-21400. [PMID: 38764657 PMCID: PMC11097186 DOI: 10.1021/acsomega.4c01675] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 02/21/2024] [Revised: 04/24/2024] [Accepted: 04/26/2024] [Indexed: 05/21/2024]
Abstract
Focal cartilage defects are a prevalent knee problem affecting people of all ages. Articular cartilage (AC) possesses limited healing potential, and osteochondral defects can lead to pain and long-term complications such as osteoarthritis. Autologous chondrocyte implantation (ACI) has been a successful surgical approach for repairing osteochondral defects over the past two decades. However, a major drawback of ACI is the dedifferentiation of chondrocytes during their in vitro expansion. In this study, we isolated ovine chondrocytes and cultured them in a two-dimensional environment for ACI procedures. We hypothesized that 3D scaffolds would support the cells' redifferentiation without the need for growth factors so we encapsulated them into soft collagen and alginate (col/alg) hydrogels. Chondrocytes embedded into the hydrogels were viable and proliferated. After 7 days, they regained their original rounded morphology (aspect ratio 1.08) and started to aggregate. Gene expression studies showed an upregulation of COL2A1, FOXO3A, FOXO1, ACAN, and COL6A1 (37, 1.13, 22, 1123, and 1.08-fold change expression, respectively) as early as day one. At 21 days, chondrocytes had extensively colonized the hydrogel, forming large cell clusters. They started to replace the degrading scaffold by depositing collagen II and aggrecan, but with limited collagen type I deposition. This approach allows us to overcome the limitations of current approaches such as the dedifferentiation occurring in 2D in vitro expansion and the necrotic formation in spheroids. Further studies are warranted to assess long-term ECM deposition and integration with native cartilage. Though limitations exist, this study suggests a promising avenue for cartilage repair with col/alg hydrogel scaffolds.
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Affiliation(s)
- Tosca Roncada
- School
of Pharmacy and Biomedical Sciences, University
of Portsmouth, St Michael’s
Building, White Swan Road, Portsmouth PO1 2DT, U.K.
| | - Gordon Blunn
- School
of Pharmacy and Biomedical Sciences, University
of Portsmouth, St Michael’s
Building, White Swan Road, Portsmouth PO1 2DT, U.K.
| | - Marta Roldo
- School
of Pharmacy and Biomedical Sciences, University
of Portsmouth, St Michael’s
Building, White Swan Road, Portsmouth PO1 2DT, U.K.
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Majumder N, Seit S, Bhabesh NS, Ghosh S. An Advanced Bioconjugation Strategy for Covalent Tethering of TGFβ3 with Silk Fibroin Matrices and its Implications in the Chondrogenesis Profile of Human BMSCs and Human Chondrocytes: A Paradigm Shift in Cartilage Tissue Engineering. Adv Healthc Mater 2024; 13:e2303513. [PMID: 38291832 DOI: 10.1002/adhm.202303513] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2023] [Revised: 01/25/2024] [Indexed: 02/01/2024]
Abstract
The transforming growth factor-β class of cytokines plays a significant role in articular cartilage formation from mesenchymal condensation to chondrogenic differentiation. However, their exogenous addition to the chondrogenic media makes the protocol expensive. It reduces the bioavailability of the cytokine to the cells owing to their burst release. The present study demonstrates an advanced bioconjugation strategy to conjugate transforming growth factor-β3 (TGFβ3) with silk fibroin matrix covalently via a cyanuric chloride coupling reaction. The tethering and change in secondary conformation are confirmed using various spectroscopic analyses. To assess the functionality of the chemically modified silk matrix, human bone marrow-derived mesenchymal stem cells (hBMSCs) and chondrocytes are cultured for 28 days in a chondrogenic differentiation medium. Gene expression and histological analysis reveal enhanced expression of chondrogenic markers with intense Safranin-O and Alcian Blue staining in TGFβ3 conjugated silk matrices than where TGFβ3 is exogenously added to the media for both hBMSCs and chondrocytes. Therefore, this study successfully recapitulates the native niche of TGFβ3 and the role of the silk as a growth factor stabilizer. When cultured over TGFβ3 conjugated silk matrices, hBMSCs display increased proteoglycan secretion and maximum chondrogenic trait with attenuation of chondrocyte hypertrophy over human chondrocytes.
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Affiliation(s)
- Nilotpal Majumder
- Department of Textile and Fibre Engineering, Indian Institute of Technology Delhi, New Delhi, 110016, India
| | - Sinchan Seit
- Department of Textile and Fibre Engineering, Indian Institute of Technology Delhi, New Delhi, 110016, India
| | - Neel Sarovar Bhabesh
- International Centre for Genetic Engineering and Biotechnology (ICGEB), Transcription Regulation group, New Delhi, 110067, India
| | - Sourabh Ghosh
- Department of Textile and Fibre Engineering, Indian Institute of Technology Delhi, New Delhi, 110016, India
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Al Maruf DSA, Xin H, Cheng K, Garcia AG, Mohseni-Dargah M, Ben-Sefer E, Tomaskovic-Crook E, Crook JM, Clark JR. Bioengineered cartilaginous grafts for repairing segmental mandibular defects. J Tissue Eng 2024; 15. [DOI: 10.1177/20417314241267017] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2025] Open
Abstract
Reconstructing critical-sized craniofacial bone defects is a global healthcare challenge. Current methods, like autologous bone transplantation, face limitations. Bone tissue engineering offers an alternative to autologous bone, with traditional approaches focusing on stimulating osteogenesis via the intramembranous ossification (IMO) pathway. However, IMO falls short in addressing larger defects, particularly in clinical scenarios where there is insufficient vascularisation. This review explores redirecting bone regeneration through endochondral ossification (ECO), a process observed in long bone healing stimulated by hypoxic conditions. Despite its promise, gaps exist in applying ECO to bone tissue engineering experiments, requiring the elucidation of key aspects such as cell sources, biomaterials and priming protocols. This review discusses various scaffold biomaterials and cellular sources for chondrogenesis and hypertrophic chondrocyte priming, mirroring the ECO pathway. The review highlights challenges in current endochondral priming and proposes alternative approaches. Emphasis is on segmental mandibular defect repair, offering insights for future research and clinical application. This concise review aims to advance bone tissue engineering by addressing critical gaps in ECO strategies.
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Affiliation(s)
- D S Abdullah Al Maruf
- Integrated Prosthetics and Reconstruction, Department of Head and Neck Surgery, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
- Central Clinical School, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
| | - Hai Xin
- Central Clinical School, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
| | - Kai Cheng
- Royal Prince Alfred Institute of Academic Surgery, Sydney Local Health District, Camperdown, NSW, Australia
| | - Alejandro Garcia Garcia
- Cell, Tissue and Organ Engineering Laboratory, Biomedical Centre (BMC), Department of Clinical Sciences Lund, Stem Cell Centre, Lund University, Lund, Sweden
| | - Masoud Mohseni-Dargah
- Integrated Prosthetics and Reconstruction, Department of Head and Neck Surgery, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
| | - Eitan Ben-Sefer
- Integrated Prosthetics and Reconstruction, Department of Head and Neck Surgery, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
- Arto Hardy Biomedical Innovation Hub, Chris O`Brien Lifehouse, Camperdown, NSW, Australia
- School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
| | - Eva Tomaskovic-Crook
- Arto Hardy Biomedical Innovation Hub, Chris O`Brien Lifehouse, Camperdown, NSW, Australia
- School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
- Sarcoma and Surgical Research Centre, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
- ARC Centre of Excellence for Electromaterials Science, The University of Wollongong, Wollongong, NSW, Australia
- Intelligent Polymer Research Institute, AIIM Facility, The University of Wollongong, Wollongong, NSW, Australia
| | - Jeremy Micah Crook
- Arto Hardy Biomedical Innovation Hub, Chris O`Brien Lifehouse, Camperdown, NSW, Australia
- School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
- Sarcoma and Surgical Research Centre, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
- ARC Centre of Excellence for Electromaterials Science, The University of Wollongong, Wollongong, NSW, Australia
- Intelligent Polymer Research Institute, AIIM Facility, The University of Wollongong, Wollongong, NSW, Australia
| | - Jonathan Robert Clark
- Integrated Prosthetics and Reconstruction, Department of Head and Neck Surgery, Chris O’Brien Lifehouse, Camperdown, NSW, Australia
- Central Clinical School, Faculty of Medicine and Health, The University of Sydney, Camperdown, NSW, Australia
- Royal Prince Alfred Institute of Academic Surgery, Sydney Local Health District, Camperdown, NSW, Australia
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Strauß S, Diemer M, Bucan V, Kuhbier JW, Asendorf T, Vogt PM, Schlottmann F. Spider silk enhanced tissue engineering of cartilage tissue: Approach of a novel bioreactor model using adipose derived stromal cells. J Appl Biomater Funct Mater 2024; 22:22808000241226656. [PMID: 38253568 DOI: 10.1177/22808000241226656] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2024] Open
Abstract
Human cartilage tissue remains a challenge for the development of therapeutic options due to its poor vascularization and reduced regenerative capacities. There are a variety of research approaches dealing with cartilage tissue engineering. In addition to different biomaterials, numerous cell populations have been investigated in bioreactor-supported experimental setups to improve cartilage tissue engineering. The concept of the present study was to investigate spider silk cocoons as scaffold seeded with adipose-derived stromal cells (ASC) in a custom-made bioreactor model using cyclic axial compression to engineer cartilage-like tissue. For chemical induction of differentiation, BMP-7 and TGF-β2 were added and changes in cell morphology and de-novo tissue formation were investigated using histological staining to verify chondrogenic differentiation. By seeding spider silk cocoons with ASC, a high colonization density and cell proliferation could be achieved. Mechanical induction of differentiation using a newly established bioreactor model led to a more roundish cell phenotype and new extracellular matrix formation, indicating a chondrogenic differentiation. The addition of BMP-7 and TGF-β2 enhanced the expression of cartilage specific markers in immunohistochemical staining. Overall, the present study can be seen as pilot study and valuable complementation to the published literature.
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Affiliation(s)
- Sarah Strauß
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
| | - Maximilian Diemer
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
| | - Vesna Bucan
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
| | - Jörn W Kuhbier
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
- Department of Plastic, Aesthetic and Hand Surgery, Helios Klinikum Hildesheim, Hildesheim, Germany
| | - Tomke Asendorf
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
| | - Peter M Vogt
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
| | - Frederik Schlottmann
- Department of Plastic, Aesthetic, Hand and Reconstructive Surgery, Hannover Medical School, Hannover, Germany
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Chondrocyte Hypertrophy in Osteoarthritis: Mechanistic Studies and Models for the Identification of New Therapeutic Strategies. Cells 2022; 11:cells11244034. [PMID: 36552796 PMCID: PMC9777397 DOI: 10.3390/cells11244034] [Citation(s) in RCA: 31] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Accepted: 12/08/2022] [Indexed: 12/16/2022] Open
Abstract
Articular cartilage shows limited self-healing ability owing to its low cellularity and avascularity. Untreated cartilage defects display an increased propensity to degenerate, leading to osteoarthritis (OA). During OA progression, articular chondrocytes are subjected to significant alterations in gene expression and phenotype, including a shift towards a hypertrophic-like state (with the expression of collagen type X, matrix metalloproteinases-13, and alkaline phosphatase) analogous to what eventuates during endochondral ossification. Present OA management strategies focus, however, exclusively on cartilage inflammation and degradation. A better understanding of the hypertrophic chondrocyte phenotype in OA might give new insights into its pathogenesis, suggesting potential disease-modifying therapeutic approaches. Recent developments in the field of cellular/molecular biology and tissue engineering proceeded in the direction of contrasting the onset of this hypertrophic phenotype, but knowledge gaps in the cause-effect of these processes are still present. In this review we will highlight the possible advantages and drawbacks of using this approach as a therapeutic strategy while focusing on the experimental models necessary for a better understanding of the phenomenon. Specifically, we will discuss in brief the cellular signaling pathways associated with the onset of a hypertrophic phenotype in chondrocytes during the progression of OA and will analyze in depth the advantages and disadvantages of various models that have been used to mimic it. Afterwards, we will present the strategies developed and proposed to impede chondrocyte hypertrophy and cartilage matrix mineralization/calcification. Finally, we will examine the future perspectives of OA therapeutic strategies.
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7
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Sun H, Zhou X, Zhang Y, Zhang L, Yu X, Ye Z, Laurencin CT. Bone Implants (Bone Regeneration and Bone Cancer Treatments). BIOFABRICATION FOR ORTHOPEDICS 2022:265-321. [DOI: 10.1002/9783527831371.ch10] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/06/2025]
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Staubli F, Stoddart MJ, D'Este M, Schwab A. Pre-culture of human mesenchymal stromal cells in spheroids facilitates chondrogenesis at a low total cell count upon embedding in biomaterials to generate cartilage microtissues. Acta Biomater 2022; 143:253-265. [PMID: 35240315 DOI: 10.1016/j.actbio.2022.02.038] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/08/2021] [Revised: 02/21/2022] [Accepted: 02/24/2022] [Indexed: 12/29/2022]
Abstract
Material-assisted cartilage tissue engineering has limited application in cartilage treatment due to hypertrophic tissue formation and high cell counts required. This study aimed at investigating the potential of human mesenchymal stromal cell (hMSC) spheroids embedded in biomaterials to study the effect of biomaterial composition on cell differentiation. Pre-cultured (3 days, chondrogenic differentiation media) spheroids (250 cells/spheroid) were embedded in tyramine-modified hyaluronic acid (THA) and collagen type I (Col) composite hydrogels (four combinations of THA (12.5 vs 16.7 mg/ml) and Col (2.5 vs 1.7 mg/ml) content) at a cell density of 5 × 106 cells/ml (2 × 104 spheroids/ml). Macropellets derived from single hMSCs (2.5 × 105 cells, ScMP) or hMSC spheroids (2.5 × 105 cells, 103 spheroids, SpMP) served as control. hMSC differentiation was analyzed using glycosaminoglycan (GAG) quantification, gene expression analysis and (immuno-)histology. Embedding of hMSC spheroids in THA-Col induced chondrogenic differentiation marked by upregulation of aggrecan (ACAN) and COL2A1, and the production of GAGs . Lower THA led to more pronounced chondrogenic phenotype compared to higher THA content. Col content had no significant influence on hMSC chondrogenesis. Pellet cultures showed an upregulation in chondrogenic-associated genes and production of GAGs with less upregulation of hypertrophic-associated genes in SpMP culture compared to ScMP group. This study presents hMSC pre-culture in spheroids as promising approach to study chondrogenic differentiation after biomaterial encapsulation at low total cell count (5 × 106/ml) without compromising chondrogenic matrix production. This approach can be applied to assemble microtissues in biomaterials to generate large cartilage construct. STATEMENT OF SIGNIFICANCE: In vitro studies investigating the chondrogenic potential of biomaterials are limited due to the low cell-cell contact of encapsulated single cells. Here, we introduce the use of pre-cultured hMSC spheroids to study chondrogenesis upon encapsulation in a biomaterial. The use of spheroids takes advantage of the high cell-cell contact within each spheroid being critical in the early chondrogenesis of hMSCs. At a low seeding density of 5·106 cells/ml (2 × 104 spheroids/ml) we demonstrated hMSC chondrogenesis and cartilaginous matrix deposition. Our results indicate that the pre-culture might have a beneficial effect on hypertrophic gene expression without compromising chondrogenic differentiation. This approach has shown potential to assemble microtissues (here spheroids) in biomaterials to generate large cartilage constructs and to study the effect of biomaterial composition on cell alignment and migration.
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Ferrao Blanco MN, Bastiaansen-Jenniskens YM, Chambers MG, Pitsillides AA, Narcisi R, van Osch GJ. Effect of Inflammatory Signaling on Human Articular Chondrocyte Hypertrophy: Potential Involvement of Tissue Repair Macrophages. Cartilage 2021; 13:168S-174S. [PMID: 34165367 PMCID: PMC8739598 DOI: 10.1177/19476035211021907] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
OBJECTIVE In osteoarthritis, chondrocytes tend to acquire a hypertrophic phenotype, which contributes to the modification of the extracellular matrix, resulting in permanent cartilage changes. In mouse chondrocytes, pro-inflammatory macrophages and pro-inflammatory cytokines have been shown to stimulate hypertrophy via the activation of the nuclear factor kappa B (NF-κB) pathway. Whether or not this also occurs in human chondrocytes remains unclear. We therefore aimed to investigate whether hypertrophy-like responses in human cartilage are driven mainly by intrinsic inflammatory signaling or shaped by specific macrophage populations. DESIGN Human articular chondrocytes were cultured with pro-inflammatory cytokines or medium conditioned by defined macrophage subsets. Furthermore, the effect of inhibition of NF-κB-dependent gene expression was evaluated using the NF-κB inhibitor SC-514. Hypertrophy was assessed by measuring the transcription level of alkaline phosphatase (ALPL), type X collagen (COL10A1), Indian hedgehog (IHH), and runt-related transcription factor 2 (RUNX2). RESULTS The expression of hypertrophic genes was not promoted in human chondrocytes by pro-inflammatory cytokines neither pro-inflammatory M(IFNγ + TNFα) macrophages. Inhibition of the NF-κB-dependent gene expression did not affect human articular chondrocyte hypertrophy. However, tissue repair M(IL4) macrophages induced hypertrophy by promoting the expression of COL10A1, RUNX2, and IHH. CONCLUSION Intrinsic inflammatory signaling activation is not involved in the hypertrophic shift observed in human articular chondrocytes cultured in vitro. However, tissue repair macrophages may contribute to the onset of this detrimental phenotype in human osteoarthritic cartilage, given the effect observed in our experimental models.
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Affiliation(s)
- Mauricio N. Ferrao Blanco
- Department of Orthopaedics and
Sports Medicine, Erasmus MC, University Medical Center Rotterdam, Rotterdam,
The Netherlands
| | | | - Mark G. Chambers
- Lilly Research Laboratories, Eli
Lilly Pharmaceuticals, Indianapolis, IN, USA
| | | | - Roberto Narcisi
- Department of Orthopaedics and
Sports Medicine, Erasmus MC, University Medical Center Rotterdam, Rotterdam,
The Netherlands
| | - Gerjo J.V.M. van Osch
- Department of Orthopaedics and
Sports Medicine, Erasmus MC, University Medical Center Rotterdam, Rotterdam,
The Netherlands,Department of
Otorhinolaryngology, Erasmus MC, University Medical Center Rotterdam,
Rotterdam, The Netherlands,Department of Biomechanical
Engineering, TU Delft, Delft, The Netherlands,Gerjo J.V.M. van Osch, Erasmus MC,
University Medical Center Rotterdam, Wytemaweg 80, Room Ee 16.51b,
Rotterdam, 3015 CN, The Netherlands.
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Wilken F, Slotta-Huspenina J, Laux F, Blanke F, Schauwecker J, Vogt S, Gollwitzer H. Autologous Chondrocyte Transplantation in Femoroacetabular Impingement Syndrome: Growth and Redifferentiation Potential of Chondrocytes Harvested from the Femur in Cam-Type Deformities. Cartilage 2021; 12:377-386. [PMID: 30862178 PMCID: PMC8236656 DOI: 10.1177/1947603519833138] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/10/2023] Open
Abstract
OBJECTIVE Cam-type femoroacetabular impingement (FAI) syndrome is one of the most frequent reasons for cartilage damage in the hip. Autologous chondrocyte transplantation has proven high success rates in the treatment of focal chondral defects; however, harvesting of chondrocytes in the hip has been reported but not specifically from the region of femoral cam lesions. Therefore, the goal of this study was to analyze the growth and redifferentiation potential of cartilage samples harvested from the cam deformities in patients with FAI. DESIGN Cartilage samples were gained from 15 patients with cam-type FAI undergoing arthroscopic femoral cam resection. Healthy (hyaline cartilage of the hip and knee joint, n = 12) and arthritic control groups (degenerative changes in cartilage of the hip joint, n = 8) were also analyzed. Chondrocytes were initially cultured under monolayer, and subsequently under pellet conditions. A comparative representation of the groups was performed by Mankin score classification, immunohistochemistry (IHC) (Col1, Col2, aggrecan), and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) (Col1, Col2, Col10, Sox9, RunX2). RESULTS Mankin score of FAI-samples (4.1±3.1, Range 0-10) showed a wide variation but was significant lower (P = 0.0244) when compared with the arthritic control (7.5 ± 2.7, range 4-12). IHC showed an increased deposition of Col2 (P = 0.0002) and aggrecan (P = 0.0261) after pellet culture compared with deposition after monolayer culture in all groups. In qRT-PCR, FAI samples showed after pellet culture increased Col2 (P = 0.0050) and Col10 expression (P = 0.0006) and also Mankin score correlated increasing gene-expression of Col10 (r = 0.8108, P = 0.0341) and RunX2 (r = 0.8829, P = 0.123). CONCLUSIONS Cartilage samples of patients with cam-type FAI showed sufficient but heterogeneous composition relating to histological quality and chondrogenic potential. However, harvesting of chondrocytes from the cam lesion might be a valid option especially if a cartilage lesion is noted in a diagnostic arthroscopy and individual preexisting stage of cartilage degeneration and appropriate pellet-culturing conditions are considered.
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Affiliation(s)
- Frauke Wilken
- Clinic of Orthopaedics and Orthopaedic Sports Medicine, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany,Department of Orthopaedic Sports Medicine, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany
| | - Julia Slotta-Huspenina
- Institute of Pathology, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany
| | - Florian Laux
- Clinic of Orthopaedics and Orthopaedic Sports Medicine, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany
| | - Fabian Blanke
- Department of Orthopaedic Sports Medicine, Hessing Stiftung, Augsburg, Germany
| | - Johannes Schauwecker
- Clinic of Orthopaedics and Orthopaedic Sports Medicine, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany
| | - Stephan Vogt
- Department of Orthopaedic Sports Medicine, Klinikum Rechts der Isar, Technical University of Munich, Munich, Germany,Department of Orthopaedic Sports Medicine, Hessing Stiftung, Augsburg, Germany,Stephan Vogt, Department of Orthopaedic Sports Medicine, Hessing Stiftung, Hessingstraße 17, 86199 Augsburg, Germany.
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11
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Fu R, Liu C, Yan Y, Li Q, Huang RL. Bone defect reconstruction via endochondral ossification: A developmental engineering strategy. J Tissue Eng 2021; 12:20417314211004211. [PMID: 33868628 PMCID: PMC8020769 DOI: 10.1177/20417314211004211] [Citation(s) in RCA: 30] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2021] [Accepted: 03/03/2021] [Indexed: 02/05/2023] Open
Abstract
Traditional bone tissue engineering (BTE) strategies induce direct bone-like matrix formation by mimicking the embryological process of intramembranous ossification. However, the clinical translation of these clinical strategies for bone repair is hampered by limited vascularization and poor bone regeneration after implantation in vivo. An alternative strategy for overcoming these drawbacks is engineering cartilaginous constructs by recapitulating the embryonic processes of endochondral ossification (ECO); these constructs have shown a unique ability to survive under hypoxic conditions as well as induce neovascularization and ossification. Such developmentally engineered constructs can act as transient biomimetic templates to facilitate bone regeneration in critical-sized defects. This review introduces the concept and mechanism of developmental BTE, explores the routes of endochondral bone graft engineering, highlights the current state of the art in large bone defect reconstruction via ECO-based strategies, and offers perspectives on the challenges and future directions of translating current knowledge from the bench to the bedside.
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Affiliation(s)
- Rao Fu
- Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Chuanqi Liu
- Department of Plastic and Burn Surgery, West China Hospital, Sichuan University, Chengdu, China
| | - Yuxin Yan
- Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Qingfeng Li
- Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Ru-Lin Huang
- Department of Plastic and Reconstructive Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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12
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Lo Monaco M, Gervois P, Beaumont J, Clegg P, Bronckaers A, Vandeweerd JM, Lambrichts I. Therapeutic Potential of Dental Pulp Stem Cells and Leukocyte- and Platelet-Rich Fibrin for Osteoarthritis. Cells 2020; 9:cells9040980. [PMID: 32326610 PMCID: PMC7227024 DOI: 10.3390/cells9040980] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2020] [Revised: 04/07/2020] [Accepted: 04/13/2020] [Indexed: 12/18/2022] Open
Abstract
Osteoarthritis (OA) is a degenerative and inflammatory joint disorder with cartilage loss. Dental pulp stem cells (DPSCs) can undergo chondrogenic differentiation and secrete growth factors associated with tissue repair and immunomodulation. Leukocyte- and platelet-rich fibrin (L-PRF) emerges in regenerative medicine because of its growth factor content and fibrin matrix. This study evaluates the therapeutic application of DPSCs and L-PRF in OA via immunomodulation and cartilage regeneration. Chondrogenic differentiation of DPSCs, with or without L-PRF exudate (ex) and conditioned medium (CM), and of bone marrow-mesenchymal stem cells was compared. These cells showed differential chondrogenesis. L-PRF was unable to increase cartilage-associated components. Immature murine articular chondrocytes (iMACs) were cultured with L-PRF ex, L-PRF CM, or DPSC CM. L-PRF CM had pro-survival and proliferative effects on unstimulated and cytokine-stimulated iMACs. L-PRF CM stimulated the release of IL-6 and PGE2, and increased MMP-13, TIMP-1 and IL-6 mRNA levels in cytokine-stimulated iMACs. DPSC CM increased the survival and proliferation of unstimulated iMACs. In cytokine-stimulated iMACs, DPSC CM increased TIMP-1 gene expression, whereas it inhibited nitrite release in 3D culture. We showed promising effects of DPSCs in an in vitro OA model, as they undergo chondrogenesis in vitro, stimulate the survival of chondrocytes and have immunomodulatory effects.
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Affiliation(s)
- Melissa Lo Monaco
- Cardio & Organ Systems (COST), Biomedical Research Institute (BIOMED), Hasselt University, 3590 Diepenbeek, Belgium; (P.G.); (J.B.); (A.B.); (I.L.)
- Department of Veterinary Medicine, Integrated Veterinary Research Unit (IVRU) - Namur Research Institute for Life Science (NARILIS), University of Namur, 5000 Namur, Belgium;
- Correspondence: ; Tel.: +32-(0)-26-92-09
| | - Pascal Gervois
- Cardio & Organ Systems (COST), Biomedical Research Institute (BIOMED), Hasselt University, 3590 Diepenbeek, Belgium; (P.G.); (J.B.); (A.B.); (I.L.)
| | - Joel Beaumont
- Cardio & Organ Systems (COST), Biomedical Research Institute (BIOMED), Hasselt University, 3590 Diepenbeek, Belgium; (P.G.); (J.B.); (A.B.); (I.L.)
- Maastricht Radiation Oncology (MaastRO) Lab, GROW—School for Oncology and Developmental Biology, Maastricht University, 6229ER Maastricht, The Netherlands
| | - Peter Clegg
- Department of Musculoskeletal and Ageing Sciences, Institute of Lifecourse and Medical Sciences, University of Liverpool, L7 8TX Liverpool, UK;
| | - Annelies Bronckaers
- Cardio & Organ Systems (COST), Biomedical Research Institute (BIOMED), Hasselt University, 3590 Diepenbeek, Belgium; (P.G.); (J.B.); (A.B.); (I.L.)
| | - Jean-Michel Vandeweerd
- Department of Veterinary Medicine, Integrated Veterinary Research Unit (IVRU) - Namur Research Institute for Life Science (NARILIS), University of Namur, 5000 Namur, Belgium;
| | - Ivo Lambrichts
- Cardio & Organ Systems (COST), Biomedical Research Institute (BIOMED), Hasselt University, 3590 Diepenbeek, Belgium; (P.G.); (J.B.); (A.B.); (I.L.)
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13
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Vieira JS, Cunha EJ, de Souza JF, Chaves LHK, de Souza JL, Giovanini AF. Alendronate disturbs femoral growth due to changes during immunolocalization of transforming growth factor-β1 and bone morphogenetic protein-2 in epiphyseal plate. World J Exp Med 2020; 10:1-9. [PMID: 31942441 PMCID: PMC6960019 DOI: 10.5493/wjem.v10.i1.1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/05/2019] [Revised: 11/26/2019] [Accepted: 12/15/2019] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND The epiphyseal growth plate is an important anatomical segment localized on the ends of a long bone. Despite the abovementioned atractive reasons for alendronate’s use, few data on the effect of alendronate during epiphyseal growth exist.
AIM Verify the effect of alendronate on the growth epiphyseal plate, and compare its effect with the size of the femur during the double-staining of the immunolocalization of transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP2) in endochondral ossifing in specimens that have received alendronate.
METHODS Forty newborn rats were randomly divided into two groups: a control group (were given applications of 1 mg/kg physiologic saline) and a group that received Alendronate (a dose of 2.5 mg/kg). These groups were then divided into two subgroups for euthanasia in two and 12 d of life. After euthanasia, the femurs were removed, and the femoral bones were measured linearly between the apex of the greater trochanter until the lower intercondylar midlle face to verify the probable bone growth between 3 and 12 d in control and alednroanto treated rats. Posteriorly, the surgical pieces were also sent to the histopathology laboratory to produce histological slides. The obtained slides were stained with hematoxylin and eosin to measure each of the cartilage zones in endochondral development. and other slides were immunohistochemically tested for anti- TGF-β1 and BMP-2 antibodies to investigate the immunolocalization of these proteins in the epiphyseal plaque area.
RESULTS On the third day, some diferences between the control group and specimens treated with alendronate were verified. Macroscopiccaly, we found similarities in size between the femoral bones when we compared the control group with the specimens that received alendronate. On the 12th day, the bone size of the mice receiving the drug was significantly smaller than those of the control group. These results coincide with changes in the TGF-β1 and BMP-2 expression. In the specimens that received alendronate, the TGF-β1 was expressed in some sites of trabecular bone that was neoformed, peripherally to the bone marrow area. The BMP-2 was also positive in proliferative chondrocytes and hypertrofic chondrocytes. On the 12th day, all layers of chondrocytes exhibited positivity for BMP-2 in the specimens that received alendronate. In the interface between the trabecular bone and cartilage, an area of disorganized bone deposition was evident. Neoformed bone also appeared to be different at 12 d. In the control group, BMP-2 was positive in an intense area of bone trabeculae, whereas the alendronate-treated group showed TGF-β1 positive trabeculae and a greater bone area.
CONCLUSION Alendronate alters the immunolocalization of TGF-β1 and BMP-2 simultaneously, a condition that changes the usual histological aspects of the cartilage zone and impairs epiphysis growth and femur growth.
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14
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Alteration of the Total Cellular Glycome during Late Differentiation of Chondrocytes. Int J Mol Sci 2019; 20:ijms20143546. [PMID: 31331074 PMCID: PMC6678350 DOI: 10.3390/ijms20143546] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2019] [Revised: 07/16/2019] [Accepted: 07/17/2019] [Indexed: 12/24/2022] Open
Abstract
In normal articular cartilage, chondrocytes do not readily proliferate or terminally differentiate, and exhibit a low level of metabolism. Hypertrophy-like changes of chondrocytes have been proposed to play a role in the pathogenesis of osteoarthritis by inducing protease-mediated cartilage degradation and calcification; however, the molecular mechanisms underlying these changes are unclear. Glycans are located on the outermost cell surface. Dynamic cellular differentiation can be monitored and quantitatively characterized by profiling the glycan structures of total cellular glycoproteins. This study aimed to clarify the alterations in glycans upon late differentiation of chondrocytes, during which hypertrophy-like changes occur. Primary mouse chondrocytes were differentiated using an insulin-induced chondro-osteogenic differentiation model. Comprehensive glycomics, including N-glycans, O-glycans, free oligosaccharides, glycosaminoglycan, and glycosphingolipid, were analyzed for the chondrocytes after 0-, 10- and 20-days cultivation. The comparison and clustering of the alteration of glycans upon hypertrophy-like changes of primary chondrocytes were performed. Comprehensive glycomic analyses provided complementary alterations in the levels of various glycans derived from glycoconjugates during hypertrophic differentiation. In addition, expression of genes related to glycan biosynthesis and metabolic processes was significantly correlated with glycan alterations. Our results indicate that total cellular glycan alterations are closely associated with chondrocyte hypertrophy and help to describe the glycophenotype by chondrocytes and their hypertrophic differentiation. our results will assist the identification of diagnostic and differentiation biomarkers in the future.
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15
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Jonitz-Heincke A, Klinder A, Boy D, Salamon A, Hansmann D, Pasold J, Buettner A, Bader R. In Vitro Analysis of the Differentiation Capacity of Postmortally Isolated Human Chondrocytes Influenced by Different Growth Factors and Oxygen Levels. Cartilage 2019; 10:111-119. [PMID: 28715962 PMCID: PMC6376569 DOI: 10.1177/1947603517719318] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
OBJECTIVE In the present in vitro study, we analyzed the chondrogenic differentiation capacity of human chondrocytes postmortally isolated from unaffected knee cartilage by the addition of transforming growth factor-β1 (TGF-β1) and/or insulin-like growth factor-1 (IGF-1) and different oxygen levels. DESIGN After 14 and 35 days, DNA concentrations and protein contents of Col1, Col2, aggrecan as well as glycosaminoglycans (GAGs) of chondrocytes cultivated as pellet cultures were analyzed. Additionally, expression rates of mesenchymal stem cell (MSC)-associated differentiation markers were assessed in monolayer cultures. RESULTS All cultivated chondrocytes were found to be CD29+/CD44+/CD105+/CD166+. Chondrocytic pellets stimulated with TGF-β1 showed enhanced synthesis rates of hyaline cartilage markers and reduced expression of the non-hyaline cartilage marker Col1 under hypoxic culture conditions. CONCLUSIONS Our results underline the substantial chondrogenic potential of human chondrocytes postmortally isolated from unaffected articular knee cartilage especially in case of TGF-β1 administration.
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Affiliation(s)
- Anika Jonitz-Heincke
- Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Rostock, Germany,Anika Jonitz-Heincke, Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Doberaner Strasse 142, 18057 Rostock, Germany.
| | - Annett Klinder
- Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Rostock, Germany
| | - Diana Boy
- Institute of Forensic Medicine, University Medical Center Rostock, Rostock, Germany
| | - Achim Salamon
- Department of Cell Biology, University Medical Center Rostock, Rostock, Germany
| | - Doris Hansmann
- Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Rostock, Germany
| | - Juliane Pasold
- Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Rostock, Germany
| | - Andreas Buettner
- Institute of Forensic Medicine, University Medical Center Rostock, Rostock, Germany
| | - Rainer Bader
- Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University Medical Center Rostock, Rostock, Germany
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16
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Bwalya EC, Wijekoon HS, Fang J, Kim S, Hosoya K, Okumura M. Independent chondrogenic potential of canine bone marrow-derived mesenchymal stem cells in monolayer expansion cultures decreases in a passage-dependent pattern. J Vet Med Sci 2018; 80:1681-1687. [PMID: 30210068 PMCID: PMC6261819 DOI: 10.1292/jvms.18-0202] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marrow-derived mesenchymal stem cells (BMSC), independent and spontaneous chondrogenesis has
been previously demonstrated in adult horses, bovine calves and adult human BMSC. Surprisingly, adult canine BMSC under similar culture conditions previously failed to demonstrate
chondrogenesis. The present study evaluated independent chondrogenic potential of BMSC sourced from three young dogs in the absence of known chondroinductive factors. BMSC were culture
expanded in 10% DMEM up to third passage (P3). At each passage, the phenotype of BMSC was evaluated by RT-PCR gel electrophoresis and qPCR. BMSC exhibited a chondrogenic phenotype in the
absence of dexamethasone and TGF-β1 as verified by the expression of Sox-9, type II collagen and aggrecan. Sox-9 was
significantly downregulated (P<0.05) from P1−P3 compared to P0 while type II and X collagen, and aggrecan were
significantly downregulated at P3 compared to P0. There was a significant (P<0.01) negative correlation between passaging and Sox-9, type II
collagen and aggrecan gene expression. These results indicate that independent chondrogenic potential and phenotype retention of BMSC decreases in a
passage-dependent pattern. Therefore, caution should be exercised for future experiments evaluating the chondrogenic potential of BMSC after extensive expansion cultures in 10% DMEM.
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Affiliation(s)
- Eugene C Bwalya
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
| | - Hm Suranji Wijekoon
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
| | - Jing Fang
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
| | - Sangho Kim
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
| | - Kenji Hosoya
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
| | - Masahiro Okumura
- Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita 18 Nishi 9, Kita-ku, Sapporo, Hokkaido 060-0818, Japan
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17
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Perez JR, Kouroupis D, Li DJ, Best TM, Kaplan L, Correa D. Tissue Engineering and Cell-Based Therapies for Fractures and Bone Defects. Front Bioeng Biotechnol 2018; 6:105. [PMID: 30109228 PMCID: PMC6079270 DOI: 10.3389/fbioe.2018.00105] [Citation(s) in RCA: 237] [Impact Index Per Article: 33.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2017] [Accepted: 07/09/2018] [Indexed: 12/25/2022] Open
Abstract
Bone fractures and segmental bone defects are a significant source of patient morbidity and place a staggering economic burden on the healthcare system. The annual cost of treating bone defects in the US has been estimated to be $5 billion, while enormous costs are spent on bone grafts for bone injuries, tumors, and other pathologies associated with defective fracture healing. Autologous bone grafts represent the gold standard for the treatment of bone defects. However, they are associated with variable clinical outcomes, postsurgical morbidity, especially at the donor site, and increased surgical costs. In an effort to circumvent these limitations, tissue engineering and cell-based therapies have been proposed as alternatives to induce and promote bone repair. This review focuses on the recent advances in bone tissue engineering (BTE), specifically looking at its role in treating delayed fracture healing (non-unions) and the resulting segmental bone defects. Herein we discuss: (1) the processes of endochondral and intramembranous bone formation; (2) the role of stem cells, looking specifically at mesenchymal (MSC), embryonic (ESC), and induced pluripotent (iPSC) stem cells as viable building blocks to engineer bone implants; (3) the biomaterials used to direct tissue growth, with a focus on ceramic, biodegradable polymers, and composite materials; (4) the growth factors and molecular signals used to induce differentiation of stem cells into the osteoblastic lineage, which ultimately leads to active bone formation; and (5) the mechanical stimulation protocols used to maintain the integrity of the bone repair and their role in successful cell engraftment. Finally, a couple clinical scenarios are presented (non-unions and avascular necrosis—AVN), to illustrate how novel cell-based therapy approaches can be used. A thorough understanding of tissue engineering and cell-based therapies may allow for better incorporation of these potential therapeutic approaches in bone defects allowing for proper bone repair and regeneration.
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Affiliation(s)
- Jose R Perez
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States
| | - Dimitrios Kouroupis
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States.,Diabetes Research Institute & Cell Transplant Center, Miller School of Medicine, University of Miami, Miami, FL, United States
| | - Deborah J Li
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States
| | - Thomas M Best
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States
| | - Lee Kaplan
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States
| | - Diego Correa
- Department of Orthopedics, UHealth Sports Medicine Institute, Miller School of Medicine, University of Miami, Miami, FL, United States.,Diabetes Research Institute & Cell Transplant Center, Miller School of Medicine, University of Miami, Miami, FL, United States
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18
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Sun Y, Yan L, Chen S, Pei M. Functionality of decellularized matrix in cartilage regeneration: A comparison of tissue versus cell sources. Acta Biomater 2018; 74:56-73. [PMID: 29702288 PMCID: PMC7307012 DOI: 10.1016/j.actbio.2018.04.048] [Citation(s) in RCA: 59] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2018] [Revised: 04/20/2018] [Accepted: 04/23/2018] [Indexed: 01/12/2023]
Abstract
Increasing evidence indicates that decellularized extracellular matrices (dECMs) derived from cartilage tissues (T-dECMs) or chondrocytes/stem cells (C-dECMs) can support proliferation and chondrogenic differentiation of cartilage-forming cells. However, few review papers compare the differences between these dECMs when they serve as substrates for cartilage regeneration. In this review, after an introduction of cartilage immunogenicity and decellularization methods to prepare T-dECMs and C-dECMs, a comprehensive comparison focuses on the effects of T-dECMs and C-dECMs on proliferation and chondrogenic differentiation of chondrocytes/stem cells in vitro and in vivo. Key factors within dECMs, consisting of microarchitecture characteristics and micromechanical properties as well as retained insoluble and soluble matrix components, are discussed in-depth for potential mechanisms underlying the functionality of these dECMs in regulating chondrogenesis. With this information, we hope to benefit dECM based cartilage engineering and tissue regeneration for future clinical application. STATEMENT OF SIGNIFICANCE The use of decellularized extracellular matrix (dECM) is becoming a promising approach for tissue engineering and regeneration. Compared to dECM derived from cartilage tissue, recently reported dECM from cell sources exhibits a distinct role in cell based cartilage regeneration. In this review paper, for the first time, tissue and cell based dECMs are comprehensively compared for their functionality in cartilage regeneration. This information is expected to provide an update for dECM based cartilage regeneration.
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Affiliation(s)
- Yu Sun
- Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA; Department of Orthopaedics, Orthopaedics Institute, Subei People's Hospital of Jiangsu Province, Yangzhou, Jiangsu 225001, China
| | - Lianqi Yan
- Department of Orthopaedics, Orthopaedics Institute, Subei People's Hospital of Jiangsu Province, Yangzhou, Jiangsu 225001, China
| | - Song Chen
- Department of Orthopaedics, Chengdu Military General Hospital, Chengdu, Sichuan 610083, China
| | - Ming Pei
- Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA; Exercise Physiology, West Virginia University, Morgantown, WV 26506, USA; WVU Cancer Institute, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, WV 26506, USA.
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19
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Gasparini S, Villa F, Molfetta L, Repaci E, Castagnola P, Quarto R, Giannoni P. Exposure to reversine affects the chondrocyte morphology and phenotype in vitro. J Tissue Eng Regen Med 2017; 12:e1337-e1348. [PMID: 28714568 DOI: 10.1002/term.2515] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2016] [Revised: 07/05/2017] [Accepted: 07/11/2017] [Indexed: 11/11/2022]
Abstract
Articular chondrocytes derived from osteoarthritic tissues (OA HAC) show a severely reduced chondrogenic commitment. This impairment undermines their use for tissue-engineered cartilage repair, which relies on cell proliferation and growth to meet therapeutic needs, but also on efficient cell plasticity to recover the chondrogenic phenotype. Reversine (Rev), a 2,6-disubstituted purine inhibitor of spindle-assembly checkpoints, was described to convert differentiated mesenchymal cells to their undifferentiated precursors. We hypothesized that Rev exposure could divert OA HAC to more plastic cells, re-boosting their subsequent commitment. HAC were enzymatically released from OA cartilage specimens, expanded for 2 weeks and treated with 5 μm Rev in dimethylsulphoxide (DMSO) or with DMSO alone for 6 days. Cell growth was assessed using the AlamarBlueTM assay. Cytoskeletal structure, endoproliferation and caspase-3-immunopositivity were assayed by epifluorescence microscopy. The OA HAC chondrogenic performance was evaluated by quantitative reverse transcription-polymerase chain reaction (RT-PCR) for glyceraldehyde-3-phosphate dehydrogenase, Sox9, Aggrecan (Agg), type II collagen (Col2), Ki67, cyclinD1, transforming growth factor-β1 (TGF-β1), -2 and -3, interleukin-1β (IL-1β) and -6 , SMAD3 and -7, and vascular endothelial growth factor. Rev-treated OA HAC recovered polygonal morphology and reduced Ki67 expression and proliferation. Cell-cycle impairment accounted for altered cytoskeletal organization, endoproliferation and apoptosis, whereas a compensatory mechanism sustained the increased cyclinD1 transcript levels. Sox9, Agg and TGFs were overexpressed, but not Col2. IL transcripts were massively downregulated. These events were dose-related and transient. Overall, in spite of a higher Rev-induced transcriptional activity for extracellular matrix components and in spite of a Rev-treated cell phenotype closer to that of the three-dimensional native articular chondrocyte, Rev effects seem unleashed from a full regained chondrogenic potential.
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Affiliation(s)
- S Gasparini
- Stem Cell Laboratory, Department of Experimental Medicine (Di.Me.S.), University of Genova, Advanced Biotechnology Centre, Genova, Italy
| | - F Villa
- Stem Cell Laboratory, Department of Experimental Medicine (Di.Me.S.), University of Genova, Advanced Biotechnology Centre, Genova, Italy
| | - L Molfetta
- Orthopedic Unit, Department of Neuroscience, Rehabilitation, Ophthalmology, Genetics and Maternal-Infant Sciences (Di.N.O.G.M.I), University of Genova, Genova, Italy
| | - E Repaci
- Stem Cell Laboratory, Department of Experimental Medicine (Di.Me.S.), University of Genova, Advanced Biotechnology Centre, Genova, Italy
| | | | - R Quarto
- Stem Cell Laboratory, Department of Experimental Medicine (Di.Me.S.), University of Genova, Advanced Biotechnology Centre, Genova, Italy
| | - P Giannoni
- Stem Cell Laboratory, Department of Experimental Medicine (Di.Me.S.), University of Genova, Advanced Biotechnology Centre, Genova, Italy
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20
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Jia Y, Yue Y, Hu DN, Chen JL, Zhou JB. Human aqueous humor levels of transforming growth factor-β2: Association with matrix metalloproteinases/tissue inhibitors of matrix metalloproteinases. Biomed Rep 2017; 7:573-578. [PMID: 29188062 DOI: 10.3892/br.2017.1004] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2017] [Accepted: 09/29/2017] [Indexed: 01/04/2023] Open
Abstract
The present study aims to investigate the association of transforming growth factor-β2 (TGF-β2) and matrix metalloproteinases (MMPs), MMP-2 and MMP-3, and tissue inhibitors of matrix metalloproteinases (TIMPs), TIMP-1, TIMP-2 and TIMP-3 in the aqueous humor of patients with high myopia or cataracts. The levels of TGF-β2 and MMPs/TIMPs were measured with the Luminex xMAP Technology using commercially available Milliplex xMAP kits. The association between TGF-β2 and MMPs/TIMPs levels was analyzed using the Spearmans correlation test. The levels of TGF-β2 were identified to be positively correlated with the levels of TIMP-1 and TIMP-3 (TIMP-1: r=0.334; P=0.007; TIMP-3: r=0.309; P=0.012). The levels of MMP-2, MMP-3 and TIMP-2 did not significantly correlate with TGF-β2 levels (P>0.05). A positive correlation was identified between TGF-β2 and TIMPs in the aqueous humor of human eyes with elongated axial length. It appears that TGF-β2 stimulates the expression of TIMPs as a compensatory reaction to the development of high myopia.
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Affiliation(s)
- Yan Jia
- Department of Ophthalmology, Shanghai Ninth People's Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200011, P.R. China.,Department of Ophthalmology, Children's Hospital of Fudan University, Shanghai Fudan University, School of Medicine, Shanghai 201102, P.R. China
| | - Yu Yue
- Department of Ophthalmology, Shanghai Ninth People's Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200011, P.R. China
| | - Dan-Ning Hu
- Departments of Ophthalmology and Pathology, New York Eye and Ear Infirmary of Mount Sinai, New York, NY 10003, USA
| | - Ji-Li Chen
- Department of Ophthalmology, Shibei Hospital, Shanghai 200435, P.R. China
| | - Ji-Bo Zhou
- Department of Ophthalmology, Shanghai Ninth People's Hospital, Shanghai Jiaotong University, School of Medicine, Shanghai 200011, P.R. China
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21
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Chen JL, Zou C, Chen Y, Zhu W, Liu W, Huang J, Liu Q, Wang D, Duan L, Xiong J, Cui J, Jia Z, Wang D. TGFβ1 induces hypertrophic change and expression of angiogenic factors in human chondrocytes. Oncotarget 2017; 8:91316-91327. [PMID: 29207646 PMCID: PMC5710926 DOI: 10.18632/oncotarget.20509] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2017] [Accepted: 08/04/2017] [Indexed: 11/25/2022] Open
Abstract
The transforming growth factor β1 (TGFβ1) plays an important role in cartilage development. However, whether TGFβ1 stimulates chondrocyte proliferation and cartilage regeneration in osteoarthritis (OA) remains elusive, especially in the context of different treatment and tissue environments. In the present study, we investigated the role of TGFβ1 in human chondrocyte culture in vitro, focusing on the morphological change of chondrocytes and the expression of angiogenic factors upon TGFβ1 stimulation. We found increased expression of biomarkers indicating chondrocyte hypertrophy and the chondrocytes aggregated to form networks when they were treated with TGFβ1. DNA microarray analysis revealed significantly increased expression of genes related to blood vessel formation in TGFβ1 treatment group compared to control group. Matrigel assay further demonstrated that chondrocytes had the potential to form network-like structure. These results suggested that TGFβ1 induces the hypertrophic change of chondrocytes culture in vitro and induce expression of angiogenic biomarkers. Therefore, application of TGFβ1 for chondrocyte culture in practice should be considered prudentially and targeting TGFβ1 or relevant receptors to block the signaling pathway might be a strategy to prevent or alleviate progression of osteoarthritis.
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Affiliation(s)
- Jie-Lin Chen
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Chang Zou
- Shenzhen Public Service Platform for Cancer Precision Medicine and Molecular Diagnosis, Shenzhen 518020, China.,Clinical Medical Research Center, The Second Clinical Medical College, Shenzhen People's Hospital, Jinan University, Shenzhen, 518020 China
| | - Yunfang Chen
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,The Eighth Affiliated Hospital, Sun Yat-sen University, Shenzhen 518035, Guangdong Province, China
| | - Weimin Zhu
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Wei Liu
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Jianghong Huang
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Qisong Liu
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Daming Wang
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Li Duan
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Jianyi Xiong
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Jiaming Cui
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Zhaofeng Jia
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
| | - Daping Wang
- Shenzhen Key Laboratory of Tissue Engineering, Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen 518035, Guangdong Province, China.,Shenzhen Centre for Sports Medicine and Engineering Technology, Shenzhen 518035, Guangdong Province, China
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22
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Ayerst BI, Merry CLR, Day AJ. The Good the Bad and the Ugly of Glycosaminoglycans in Tissue Engineering Applications. Pharmaceuticals (Basel) 2017; 10:E54. [PMID: 28608822 PMCID: PMC5490411 DOI: 10.3390/ph10020054] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2017] [Revised: 06/05/2017] [Accepted: 06/05/2017] [Indexed: 12/14/2022] Open
Abstract
High sulfation, low cost, and the status of heparin as an already FDA- and EMA- approved product, mean that its inclusion in tissue engineering (TE) strategies is becoming increasingly popular. However, the use of heparin may represent a naïve approach. This is because tissue formation is a highly orchestrated process, involving the temporal expression of numerous growth factors and complex signaling networks. While heparin may enhance the retention and activity of certain growth factors under particular conditions, its binding 'promiscuity' means that it may also inhibit other factors that, for example, play an important role in tissue maintenance and repair. Within this review we focus on articular cartilage, highlighting the complexities and highly regulated processes that are involved in its formation, and the challenges that exist in trying to effectively engineer this tissue. Here we discuss the opportunities that glycosaminoglycans (GAGs) may provide in advancing this important area of regenerative medicine, placing emphasis on the need to move away from the common use of heparin, and instead focus research towards the utility of specific GAG preparations that are able to modulate the activity of growth factors in a more controlled and defined manner, with less off-target effects.
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Affiliation(s)
- Bethanie I Ayerst
- Wellcome Trust Centre for Cell-Matrix Research, Division of Cell-Matrix Biology & Regenerative Medicine, School of Biology, Faculty of Biology, Medicine & Health, The University of Manchester, Manchester Academic Health Science Centre, Manchester M13 9PL, UK.
| | - Catherine L R Merry
- Stem Cell Glycobiology Group, Wolfson Centre for Stem Cells, Tissue Engineering & Modelling (STEM), Centre for Biomolecular Sciences, University of Nottingham, University Park, Nottingham NG7 2RD, UK.
| | - Anthony J Day
- Wellcome Trust Centre for Cell-Matrix Research, Division of Cell-Matrix Biology & Regenerative Medicine, School of Biology, Faculty of Biology, Medicine & Health, The University of Manchester, Manchester Academic Health Science Centre, Manchester M13 9PL, UK.
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23
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Diaz-Romero J, Kürsener S, Kohl S, Nesic D. S100B + A1 CELISA: A Novel Potency Assay and Screening Tool for Redifferentiation Stimuli of Human Articular Chondrocytes. J Cell Physiol 2016; 232:1559-1570. [DOI: 10.1002/jcp.25682] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2016] [Accepted: 11/08/2016] [Indexed: 12/19/2022]
Affiliation(s)
- Jose Diaz-Romero
- Osteoarticular Research Group; Department of Clinical Research; University of Bern; Bern Switzerland
| | - Sibylle Kürsener
- Osteoarticular Research Group; Department of Clinical Research; University of Bern; Bern Switzerland
| | - Sandro Kohl
- Department of Orthopedics and Traumatology; Inselspital; University of Bern; Bern Switzerland
| | - Dobrila Nesic
- Osteoarticular Research Group; Department of Clinical Research; University of Bern; Bern Switzerland
- Department of Orthopedics and Traumatology; Inselspital; University of Bern; Bern Switzerland
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24
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Chameettachal S, Midha S, Ghosh S. Regulation of Chondrogenesis and Hypertrophy in Silk Fibroin-Gelatin-Based 3D Bioprinted Constructs. ACS Biomater Sci Eng 2016; 2:1450-1463. [DOI: 10.1021/acsbiomaterials.6b00152] [Citation(s) in RCA: 65] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Affiliation(s)
- Shibu Chameettachal
- Department of Textile Technology, Indian Institute of Technology, Delhi, India
| | - Swati Midha
- Department of Textile Technology, Indian Institute of Technology, Delhi, India
| | - Sourabh Ghosh
- Department of Textile Technology, Indian Institute of Technology, Delhi, India
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25
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Chondrogenesis by bone marrow‐derived mesenchymal stem cells grown in chondrocyte‐conditioned medium for auricular reconstruction. J Tissue Eng Regen Med 2016; 11:2763-2773. [DOI: 10.1002/term.2171] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2015] [Revised: 01/15/2016] [Accepted: 02/10/2016] [Indexed: 01/10/2023]
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26
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Lolli A, Narcisi R, Lambertini E, Penolazzi L, Angelozzi M, Kops N, Gasparini S, van Osch GJ, Piva R. Silencing of Antichondrogenic MicroRNA-221 in Human Mesenchymal Stem Cells Promotes Cartilage Repair In Vivo. Stem Cells 2016; 34:1801-11. [DOI: 10.1002/stem.2350] [Citation(s) in RCA: 50] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2015] [Revised: 01/21/2016] [Accepted: 02/01/2016] [Indexed: 02/06/2023]
Affiliation(s)
- Andrea Lolli
- Department of Biomedical and Specialty Surgical Sciences; University of Ferrara; Ferrara Italy
| | - Roberto Narcisi
- Department of Orthopaedics; Erasmus MC, University Medical Center; CN Rotterdam The Netherlands
| | - Elisabetta Lambertini
- Department of Biomedical and Specialty Surgical Sciences; University of Ferrara; Ferrara Italy
| | - Letizia Penolazzi
- Department of Biomedical and Specialty Surgical Sciences; University of Ferrara; Ferrara Italy
| | - Marco Angelozzi
- Department of Biomedical and Specialty Surgical Sciences; University of Ferrara; Ferrara Italy
| | - Nicole Kops
- Department of Orthopaedics; Erasmus MC, University Medical Center; CN Rotterdam The Netherlands
| | - Simona Gasparini
- Department of Orthopaedics; Erasmus MC, University Medical Center; CN Rotterdam The Netherlands
| | - Gerjo J.V.M. van Osch
- Department of Orthopaedics; Erasmus MC, University Medical Center; CN Rotterdam The Netherlands
- Department of Otorhinolaryngology; Erasmus MC, University Medical Center; CN Rotterdam The Netherlands
| | - Roberta Piva
- Department of Biomedical and Specialty Surgical Sciences; University of Ferrara; Ferrara Italy
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27
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Ahn J, Kumar H, Cha BH, Park S, Arai Y, Han I, Park SG, Lee SH. AIMP1 downregulation restores chondrogenic characteristics of dedifferentiated/degenerated chondrocytes by enhancing TGF-β signal. Cell Death Dis 2016; 7:e2099. [PMID: 26890138 PMCID: PMC5399188 DOI: 10.1038/cddis.2016.17] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2015] [Revised: 12/10/2015] [Accepted: 12/11/2015] [Indexed: 12/11/2022]
Abstract
Dedifferentiation and degeneration of chondrocytes critically influences the efficiency of cartilage repair. One of the causes is the defect of transforming growth factor (TGF)-β signaling that promotes chondrogenic differentiation and degeneration. In the present study, we found that aminoacyl-tRNA synthetase-interacting multifunctional protein 1 (AIMP1) negatively regulates TGF-β signaling via interactions with Smad2 and Smad3 in immunoprecipitation assay and luciferase assay. In addition, we observed that the AIMP1 expression level was significantly increased in osteoarthritis (OA) patient-derived degenerated chondrocytes compared with healthy control. So, we hypothesized that downregulation of AIMP1 using small-interfering RNA (siRNA) technology in dedifferentiated (collected at passage #6) and degenerated (obtained from OA-affected areas) chondrocytes could lead to recover TGF-β signaling in both chondrocytes. Indeed, AIMP1 downregulation restored TGF-β signaling by promoting phosphorylation of Smad2 and Smad3, which shows redifferentiated characteristics in both dedifferentiated and degenerated chondrocytes. Additionally, implantation analyses using in vivo mouse model clearly showed that AIMP1 downregulation resulted in the increased chondrogenic potential as well as the enhanced cartilage tissue formation in both dedifferentiated and degenerated chondrocytes. Histological analyses clarified that AIMP1 downregulation increased expression levels of collagen type II (Col II) and aggrecan, but not Col I expression. Taken together, these data indicate that AIMP1 downregulation using siRNA is a novel tool to restore TGF-β signaling and thereby increases the chondrogenic potential of dedifferentiated/degenerated chondrocytes, which could be further developed as a therapeutic siRNA to treat OA.
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Affiliation(s)
- J Ahn
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - H Kumar
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea.,Department of Neurosurgery, Bundang Medical Center, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - B-H Cha
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - S Park
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - Y Arai
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - I Han
- Department of Neurosurgery, Bundang Medical Center, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
| | - S G Park
- Department of Pharmacy, College of Pharmacy, Ajou University, Suwon, Gyeonggi-do, Republic of Korea
| | - S-H Lee
- Department of Biomedical Science, CHA University, Seongnam-si, Gyeonggi-do, Republic of Korea
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28
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Giannoni P, Villa F, Cordazzo C, Zardi L, Fattori P, Quarto R, Fiorini M. Rheological properties, biocompatibility and in vivo performance of new hydrogel-based bone fillers. Biomater Sci 2016; 4:1691-1703. [DOI: 10.1039/c6bm00478d] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Three different heterologous substitutes for bone regeneration, manufactured with equine-derived cortical powder, cancellous chips and demineralized bone matrix granules, were compared in vitro and in vivo.
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Affiliation(s)
- Paolo Giannoni
- Stem Cell Laboratory
- Dept. of Experimental Medicine
- University of Genova
- c/o Advanced Biotechnology Centre
- 16132–Genova
| | - Federico Villa
- Stem Cell Laboratory
- Dept. of Experimental Medicine
- University of Genova
- c/o Advanced Biotechnology Centre
- 16132–Genova
| | - Cinzia Cordazzo
- Sirius-Biotech S.r.l
- c/o
- Advanced Biotechnology Centre
- 16132 Genova
- Italy
| | - Luciano Zardi
- Sirius-Biotech S.r.l
- c/o
- Advanced Biotechnology Centre
- 16132 Genova
- Italy
| | | | - Rodolfo Quarto
- Stem Cell Laboratory
- Dept. of Experimental Medicine
- University of Genova
- c/o Advanced Biotechnology Centre
- 16132–Genova
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29
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Zhong L, Huang X, Karperien M, Post JN. The Regulatory Role of Signaling Crosstalk in Hypertrophy of MSCs and Human Articular Chondrocytes. Int J Mol Sci 2015; 16:19225-47. [PMID: 26287176 PMCID: PMC4581295 DOI: 10.3390/ijms160819225] [Citation(s) in RCA: 101] [Impact Index Per Article: 10.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/07/2015] [Accepted: 08/07/2015] [Indexed: 12/26/2022] Open
Abstract
Hypertrophic differentiation of chondrocytes is a main barrier in application of mesenchymal stem cells (MSCs) for cartilage repair. In addition, hypertrophy occurs occasionally in osteoarthritis (OA). Here we provide a comprehensive review on recent literature describing signal pathways in the hypertrophy of MSCs-derived in vitro differentiated chondrocytes and chondrocytes, with an emphasis on the crosstalk between these pathways. Insight into the exact regulation of hypertrophy by the signaling network is necessary for the efficient application of MSCs for articular cartilage repair and for developing novel strategies for curing OA. We focus on articles describing the role of the main signaling pathways in regulating chondrocyte hypertrophy-like changes. Most studies report hypertrophic differentiation in chondrogenesis of MSCs, in both human OA and experimental OA. Chondrocyte hypertrophy is not under the strict control of a single pathway but appears to be regulated by an intricately regulated network of multiple signaling pathways, such as WNT, Bone morphogenetic protein (BMP)/Transforming growth factor-β (TGFβ), Parathyroid hormone-related peptide (PTHrP), Indian hedgehog (IHH), Fibroblast growth factor (FGF), Insulin like growth factor (IGF) and Hypoxia-inducible factor (HIF). This comprehensive review describes how this intricate signaling network influences tissue-engineering applications of MSCs in articular cartilage (AC) repair, and improves understanding of the disease stages and cellular responses within an OA articular joint.
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Affiliation(s)
- Leilei Zhong
- Developmental BioEngineering, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede 7500 AE, The Netherlands.
| | - Xiaobin Huang
- Developmental BioEngineering, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede 7500 AE, The Netherlands.
- School of Life Sciences, Chongqing University, Chongqing 400030, China.
| | - Marcel Karperien
- Developmental BioEngineering, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede 7500 AE, The Netherlands.
| | - Janine N Post
- Developmental BioEngineering, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede 7500 AE, The Netherlands.
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30
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Tekari A, Luginbuehl R, Hofstetter W, Egli RJ. Transforming growth factor beta signaling is essential for the autonomous formation of cartilage-like tissue by expanded chondrocytes. PLoS One 2015; 10:e0120857. [PMID: 25775021 PMCID: PMC4361600 DOI: 10.1371/journal.pone.0120857] [Citation(s) in RCA: 57] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2014] [Accepted: 01/27/2015] [Indexed: 02/07/2023] Open
Abstract
Cartilage is a tissue with limited self-healing potential. Hence, cartilage defects require surgical attention to prevent or postpone the development of osteoarthritis. For cell-based cartilage repair strategies, in particular autologous chondrocyte implantation, articular chondrocytes are isolated from cartilage and expanded in vitro to increase the number of cells required for therapy. During expansion, the cells lose the competence to autonomously form a cartilage-like tissue, that is in the absence of exogenously added chondrogenic growth factors, such as TGF-βs. We hypothesized that signaling elicited by autocrine and/or paracrine TGF-β is essential for the formation of cartilage-like tissue and that alterations within the TGF-β signaling pathway during expansion interfere with this process. Primary bovine articular chondrocytes were harvested and expanded in monolayer culture up to passage six and the formation of cartilage tissue was investigated in high density pellet cultures grown for three weeks. Chondrocytes expanded for up to three passages maintained the potential for autonomous cartilage-like tissue formation. After three passages, however, exogenous TGF-β1 was required to induce the formation of cartilage-like tissue. When TGF-β signaling was blocked by inhibiting the TGF-β receptor 1 kinase, the autonomous formation of cartilage-like tissue was abrogated. At the initiation of pellet culture, chondrocytes from passage three and later showed levels of transcripts coding for TGF-β receptors 1 and 2 and TGF-β2 to be three-, five- and five-fold decreased, respectively, as compared to primary chondrocytes. In conclusion, the autonomous formation of cartilage-like tissue by expanded chondrocytes is dependent on signaling induced by autocrine and/or paracrine TGF-β. We propose that a decrease in the expression of the chondrogenic growth factor TGF-β2 and of the TGF-β receptors in expanded chondrocytes accounts for a decrease in the activity of the TGF-β signaling pathway and hence for the loss of the potential for autonomous cartilage-like tissue formation.
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Affiliation(s)
- Adel Tekari
- Group for Bone Biology & Orthopaedic Research, Department Clinical Research, University of Bern, Bern, Switzerland
- Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland
| | | | - Willy Hofstetter
- Group for Bone Biology & Orthopaedic Research, Department Clinical Research, University of Bern, Bern, Switzerland
| | - Rainer J. Egli
- Group for Bone Biology & Orthopaedic Research, Department Clinical Research, University of Bern, Bern, Switzerland
- RMS Foundation, Bettlach, Switzerland
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31
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Abstract
Due to a blood supply shortage, articular cartilage has a limited capacity for self-healing once damaged. Articular chondrocytes, cartilage progenitor cells, embryonic stem cells, and mesenchymal stem cells are candidate cells for cartilage regeneration. Significant current attention is paid to improving chondrogenic differentiation capacity; unfortunately, the potential chondrogenic hypertrophy of differentiated cells is largely overlooked. Consequently, the engineered tissue is actually a transient cartilage rather than a permanent one. The development of hypertrophic cartilage ends with the onset of endochondral bone formation which has inferior mechanical properties. In this review, current strategies for inhibition of chondrogenic hypertrophy are comprehensively summarized; the impact of cell source options is discussed; and potential mechanisms underlying these strategies are also categorized. This paper aims to provide guidelines for the prevention of hypertrophy in the regeneration of cartilage tissue. This knowledge may also facilitate the retardation of osteophytes in the treatment of osteoarthritis.
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Affiliation(s)
- Song Chen
- Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA
- Department of Joint Surgery, Shanghai Changzheng Hospital, The Second Military Medical University, Shanghai 200003, China
| | - Peiliang Fu
- Department of Joint Surgery, Shanghai Changzheng Hospital, The Second Military Medical University, Shanghai 200003, China
| | - Ruijun Cong
- Department of Orthopaedics, The 10th People's Hospital of Shanghai, Affiliated with Tongji University, Shanghai 200072, China
| | - HaiShan Wu
- Department of Joint Surgery, Shanghai Changzheng Hospital, The Second Military Medical University, Shanghai 200003, China
| | - Ming Pei
- Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, WV 26506, USA
- Exercise Physiology, West Virginia University, Morgantown, WV 26506, USA
- Mechanical and Aerospace Engineering, West Virginia University, Morgantown, WV 26506, USA
- Corresponding author. Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, PO Box 9196, One Medical Center Drive, Morgantown, WV 26506-9196, USA. Tel.: +1 304 293 1072; fax: +1 304 293 7070.
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32
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Thompson EM, Matsiko A, Farrell E, Kelly DJ, O'Brien FJ. Recapitulating endochondral ossification: a promising route toin vivobone regeneration. J Tissue Eng Regen Med 2014; 9:889-902. [DOI: 10.1002/term.1918] [Citation(s) in RCA: 94] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2013] [Revised: 02/14/2014] [Accepted: 04/24/2014] [Indexed: 12/22/2022]
Affiliation(s)
- Emmet M. Thompson
- Tissue Engineering Research Group, Department of Anatomy; Royal College of Surgeons in Ireland; Dublin Ireland
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute; Trinity College Dublin; Ireland
- Advanced Materials and Bioengineering Research (AMBER) Centre; Dublin Ireland
| | - Amos Matsiko
- Tissue Engineering Research Group, Department of Anatomy; Royal College of Surgeons in Ireland; Dublin Ireland
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute; Trinity College Dublin; Ireland
- Advanced Materials and Bioengineering Research (AMBER) Centre; Dublin Ireland
| | - Eric Farrell
- Department of Oral and Maxillofacial Surgery, Erasmus MC; University Medical Centre Rotterdam; The Netherlands
| | - Daniel J. Kelly
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute; Trinity College Dublin; Ireland
- Advanced Materials and Bioengineering Research (AMBER) Centre; Dublin Ireland
- Department of Mechanical and Manufacturing Engineering, School of Engineering; Trinity College Dublin; Ireland
| | - Fergal J. O'Brien
- Tissue Engineering Research Group, Department of Anatomy; Royal College of Surgeons in Ireland; Dublin Ireland
- Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute; Trinity College Dublin; Ireland
- Advanced Materials and Bioengineering Research (AMBER) Centre; Dublin Ireland
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33
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Ahmed N, Iu J, Brown CE, Taylor DW, Kandel RA. Serum- and growth-factor-free three-dimensional culture system supports cartilage tissue formation by promoting collagen synthesis via Sox9-Col2a1 interaction. Tissue Eng Part A 2014; 20:2224-33. [PMID: 24606204 DOI: 10.1089/ten.tea.2013.0559] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
OBJECTIVE One of the factors preventing clinical application of regenerative medicine to degenerative cartilage diseases is a suitable source of cells. Chondrocytes, the only cell type of cartilage, grown in vitro under culture conditions to expand cell numbers lose their phenotype along with the ability to generate hyaline cartilaginous tissue. In this study we determine that a serum- and growth-factor-free three-dimensional (3D) culture system restores the ability of the passaged chondrocytes to form cartilage tissue in vitro, a process that involves sox9. METHODS Bovine articular chondrocytes were passaged twice to allow for cell number expansion (P2) and cultured at high density on 3D collagen-type-II-coated membranes in high glucose content media supplemented with insulin and dexamethasone (SF3D). The cells were characterized after monolayer expansion and following 3D culture by flow cytometry, gene expression, and histology. The early changes in signaling transduction pathways during redifferentiation were characterized. RESULTS The P2 cells showed a progenitor-like antigen profile of 99% CD44(+) and 40% CD105(+) and a gene expression profile suggestive of interzone cells. P2 in SF3D expressed chondrogenic genes and accumulated extracellular matrix. Downregulating insulin receptor (IR) with HNMPA-(AM3) or the PI-3/AKT kinase pathway (activated by insulin treatment) with Wortmannin inhibited collagen synthesis. HNMPA-(AM3) reduced expression of Col2, Col11, and IR genes as well as Sox6 and -9. Co-immunoprecipitation and chromatin immunoprecipitation analyses of HNMPA-(AM3)-treated cells showed binding of the coactivators Sox6 and Med12 with Sox9 but reduced Sox9-Col2a1 binding. CONCLUSIONS We describe a novel culture method that allows for increase in the number of chondrocytes and promotes hyaline-like cartilage tissue formation in part by insulin-mediated Sox9-Col2a1 binding. The suitability of the tissue generated via this approach for use in joint repair needs to be examined in vivo.
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Affiliation(s)
- Nazish Ahmed
- 1 CIHR-BioEngineering of Skeletal Tissues Team, Mount Sinai Hospital, University of Toronto , Toronto, Canada
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34
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Krase A, Abedian R, Steck E, Hurschler C, Richter W. BMP activation and Wnt-signalling affect biochemistry and functional biomechanical properties of cartilage tissue engineering constructs. Osteoarthritis Cartilage 2014; 22:284-92. [PMID: 24280245 DOI: 10.1016/j.joca.2013.11.011] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/28/2013] [Revised: 10/26/2013] [Accepted: 11/15/2013] [Indexed: 02/02/2023]
Abstract
OBJECTIVES Bone morphogenetic protein (BMP-) and Wnt-signalling play crucial roles in cartilage homeostasis. Our objective was to investigate whether activation of the BMP-pathway or stimulation of Wnt-signalling cascades effectively enhances cartilage-specific extracellular matrix (ECM) accumulation and functional biomechanical parameters of chondrocyte-seeded tissue engineering (TE)-constructs. DESIGN Articular chondrocytes were cultured in collagen-type-I/III-matrices over 6 weeks to create a biomechanical standard curve. Effects of stimulation with 100 ng/mL BMP-4/-7 heterodimer or 10 mM lithium chloride (LiCl) on ECM-deposition was quantified and characterized histologically. Biomechanical parameters were determined by the Very Low Rubber Hardness (VLRH) method and under confined compression stress relaxation. RESULTS BMP-4/-7 treatment resulted in stronger collagen type-II staining and significantly enhanced glycosaminoglycan (GAG) deposition (3.2-fold; *P < 0.01) correlating with improved hardness (∼1.7-fold; *P = 0.001) reaching 83% of native cartilage values after 28 days, a value not reached before 9 weeks without stimulation. LiCl treatment enhanced VLRH slightly, but significantly (∼1.3-fold; *P = 0.016) with a trend to more ECM-deposition. BMP-4/-7 treatment significantly enhanced the E Modulus (105.7 ± 34.1 kPa; *P = 0.000001) compared to controls (8.0 ± 4.2 kPa). Poisson's ratio was significantly improved by BMP-4/-7 treatment (0.0703 ± 0.0409; *P = 0.013) vs controls (0.0432 ± 0.0284) and a significantly lower permeability (5.8 ± 2.1056 × 10(-14) m4/N.s; *P = 0.00001) was detected compared to untreated scaffolds (4.4 ± 3.1289 × 10(-13) m4/N.s). CONCLUSIONS While Wnt-activation is less effective, BMP-4/-7 heterodimer stimulation approximated native cartilage features in less than 50% of standard culture time representing a promising strategy for functional cartilage TE to improve biomechanical parameters of engineered cartilage.
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Affiliation(s)
- A Krase
- Research Center for Experimental Orthopaedics, Department of Orthopaedics, Trauma Surgery and Paraplegiology, Heidelberg University Hospital, Heidelberg, Germany.
| | - R Abedian
- Laboratory for Biomechanics and Biomaterials, Orthopaedic Department, Hannover Medical School of Hannover, Hannover, Germany.
| | - E Steck
- Research Center for Experimental Orthopaedics, Department of Orthopaedics, Trauma Surgery and Paraplegiology, Heidelberg University Hospital, Heidelberg, Germany.
| | - C Hurschler
- Laboratory for Biomechanics and Biomaterials, Orthopaedic Department, Hannover Medical School of Hannover, Hannover, Germany.
| | - W Richter
- Research Center for Experimental Orthopaedics, Department of Orthopaedics, Trauma Surgery and Paraplegiology, Heidelberg University Hospital, Heidelberg, Germany.
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Zhu Y, Yuan M, Meng HY, Wang AY, Guo QY, Wang Y, Peng J. Basic science and clinical application of platelet-rich plasma for cartilage defects and osteoarthritis: a review. Osteoarthritis Cartilage 2013; 21:1627-37. [PMID: 23933379 DOI: 10.1016/j.joca.2013.07.017] [Citation(s) in RCA: 229] [Impact Index Per Article: 19.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/12/2013] [Revised: 07/26/2013] [Accepted: 07/30/2013] [Indexed: 02/02/2023]
Abstract
Cartilage defects (CDs) and the most common joint disease, osteoarthritis (OA), are characterized by degeneration of the articular cartilage that ultimately leads to joint destruction. Current treatment strategies are inadequate: none results in restoration of fully functional hyaline cartilage, for uncertain long-term prognosis. Tissue engineering of cartilage with auto-cartilage cells or appropriate mesenchymal stem cell (MSC)-derived cartilage cells is currently being investigated to search for new therapies. Platelet-rich plasma (PRP), an autologous source of factors obtained by centrifugation, possesses various functions. For culture of MSCs and cartilage cells, it might be substituted for fetal bovine serum (FBS) with high efficiency and safety. It enhances the regeneration of cartilage cells when added to cartilage tissue engineering constructs for repairing CDs and as regenerative injection therapy for OA. But challenges also remain. Some of the growth factors (GFs) present in PRP have negative effects on the OA joint. It is therefore unlikely that a mix of GFs some of which have negative effects in the OA joint, as present in PRP, will be of benefit in OA. Future directions of PRP application may concentrate on seeking an appropriate and innocuous agent like anti-VEGF antibody that can modulate and control the effect of PRP.
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Affiliation(s)
- Y Zhu
- Institute of Orthopedics, Chinese PLA General Hospital, Fuxing 28# Road, Beijing 100853, China
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Saito T, Yano F, Mori D, Ohba S, Hojo H, Otsu M, Eto K, Nakauchi H, Tanaka S, Chung UI, Kawaguchi H. Generation of Col2a1-EGFP iPS cells for monitoring chondrogenic differentiation. PLoS One 2013; 8:e74137. [PMID: 24066106 PMCID: PMC3774617 DOI: 10.1371/journal.pone.0074137] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2013] [Accepted: 07/27/2013] [Indexed: 01/04/2023] Open
Abstract
Induced pluripotent stem cells (iPSC) are a promising cell source for cartilage regenerative medicine; however, the methods for chondrocyte induction from iPSC are currently developing and not yet sufficient for clinical application. Here, we report the establishment of a fluorescent indicator system for monitoring chondrogenic differentiation from iPSC to simplify screening for effective factors that induce chondrocytes from iPSC. We generated iPSC from embryonic fibroblasts of Col2a1-EGFP transgenic mice by retroviral transduction of Oct4, Sox2, Klf4, and c-Myc. Among the 30 clones of Col2a1-EGFP iPSC we established, two clones showed high expression levels of embryonic stem cell (ESC) marker genes, similar to control ESC. A teratoma formation assay showed that the two clones were pluripotent and differentiated into cell types from all three germ layers. The fluorescent signal was observed during chondrogenic differentiation of the two clones concomitant with the increase in chondrocyte marker expression. In conclusion, Col2a1-EGFP iPSC are useful for monitoring chondrogenic differentiation and will contribute to research in cartilage regenerative medicine.
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Affiliation(s)
- Taku Saito
- Bone and Cartilage Regenerative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
- Sensory & Motor System Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
- * E-mail:
| | - Fumiko Yano
- Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Daisuke Mori
- Bone and Cartilage Regenerative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Shinsuke Ohba
- Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Hironori Hojo
- Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Makoto Otsu
- Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo, Japan
| | - Koji Eto
- Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo, Japan
- Clinical Application Department, Center for iPS Research and Application (CiRA), Kyoto University, Shogoin, Sakyo-ku, Kyoto, Japan
| | - Hiromitsu Nakauchi
- Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo, Japan
| | - Sakae Tanaka
- Sensory & Motor System Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Ung-il Chung
- Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
| | - Hiroshi Kawaguchi
- Sensory & Motor System Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, Japan
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Preitschopf A, Zwickl H, Li K, Lubec G, Joo G, Rosner M, Hengstschläger M, Mikula M. Chondrogenic differentiation of amniotic fluid stem cells and their potential for regenerative therapy. Stem Cell Rev Rep 2013; 8:1267-74. [PMID: 22869300 DOI: 10.1007/s12015-012-9405-4] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Abstract
Chronic articular cartilage defects are the most common disabling conditions of humans in the western world. The incidence for cartilage defects is increasing with age and the most prominent risk factors are overweight and sports associated overloading. Damage of articular cartilage frequently leads to osteoarthritis due to the aneural and avascular nature of articular cartilage, which impairs regeneration and repair. Hence, patients affected by cartilage defects will benefit from a cell-based transplantation strategy. Autologous chondrocytes, mesenchymal stem cells and embryonic stem cells are suitable donor cells for regeneration approaches and most recently the discovery of amniotic fluid stem cells has opened a plethora of new therapeutic options. It is the aim of this review to summarize recent advances in the use of amniotic fluid stem cells as novel cell sources for the treatment of articular cartilage defects. Molecular aspects of articular cartilage formation as well as degeneration are summarized and the role of growth factor triggered signaling pathways, scaffolds, hypoxia and autophagy during the process of chondrogenic differentiation are discussed.
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Affiliation(s)
- Andrea Preitschopf
- Institute of Medical Genetics, Medical University of Vienna, Währinger Strasse 10, 1090 Vienna, Austria
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Perrier-Groult E, Pasdeloup M, Malbouyres M, Galéra P, Mallein-Gerin F. Control of collagen production in mouse chondrocytes by using a combination of bone morphogenetic protein-2 and small interfering RNA targeting Col1a1 for hydrogel-based tissue-engineered cartilage. Tissue Eng Part C Methods 2013; 19:652-64. [PMID: 23311625 DOI: 10.1089/ten.tec.2012.0396] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Because articular cartilage does not self-repair, tissue-engineering strategies should be considered to regenerate this tissue. Autologous chondrocyte implantation is already used for treatment of focal damage of articular cartilage. Unfortunately, this technique includes a step of cell amplification, which results in dedifferentiation of chondrocytes, with expression of type I collagen, a protein characteristic of fibrotic tissues. Therefore, the risk of producing a fibrocartilage exists. The aim of this study was to propose a new strategy for authorizing the recovery of the differentiated status of the chondrocytes after their amplification on plastic. Because the bone morphogenetic protein (BMP)-2 and the transforming growth factor (TGF)-β1 are cytokines both proposed as stimulants for cartilage repair, we undertook a detailed comparative analysis of their biological effects on chondrocytes. As a cellular model, we used mouse chondrocytes after their expansion on plastic and we tested the capability of BMP-2 or TGF-β1 to drive their redifferentiation, with special attention given to the nature of the proteins synthesized by the cells. To prevent any fibrotic character of the newly synthesized extracellular matrix, we silenced type I collagen by transfecting small interfering RNA (siRNA) into the chondrocytes, before their exposure to BMP-2 or TGF-β1. Our results showed that addition of siRNA targeting the mRNA encoded by the Col1a1 gene (Col1a1 siRNA) and BMP-2 represents the most efficient combination to control the production of cartilage-characteristic collagen proteins. To go one step further toward scaffold-based cartilage engineering, Col1a1 siRNA-transfected chondrocytes were encapsulated in agarose hydrogel and cultured in vitro for 1 week. The analysis of the chondrocyte-agarose constructs by using real-time polymerase chain reaction, Western-blotting, immunohistochemistry, and electron microscopy techniques demonstrated that the BMP-2/Col1a1 siRNA combination is effective in reinitializing correct production and assembly of the cartilage-characteristic matrix in agarose hydrogel, without production of type I collagen. Because agarose is known to favor long-term expression of the chondrocyte phenotype and agarose-based hydrogels are approved for clinical trials, this strategy appears very promising to repair hyaline cartilage.
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No effect of subperiosteal growth factor application on periosteal neo-chondrogenesis in osteoperiosteal bone grafts for osteochondral defect repair. INTERNATIONAL ORTHOPAEDICS 2013; 37:1171-8. [PMID: 23503670 DOI: 10.1007/s00264-013-1827-3] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/05/2012] [Accepted: 02/03/2013] [Indexed: 02/06/2023]
Abstract
PURPOSE The purpose of this study was to examine the effect of subperiosteal injection of chondroinductive growth factors on the histological and biomechanical outcome of autologous osteoperiosteal grafts. METHODS Thirty six standardised osteochondral defects were created in the trochlear groove of 18 Göttinger Minipigs and evaluated after six, 12 and 52 weeks. Defects were treated with press-fit implantation of autologous osteoperiosteal cylindrical block-grafts with or without subperiosteal injection of a chondroinductive growth factor mixture (GFM). RESULTS Histomorphological analysis showed complete osseointegration of all grafts from six weeks. The periosteum remained in place in 35 of 36 cases. Fibrocartilagineous repair tissue formation occurred at the cambium layer with a maximum at 12 weeks in both groups. Histomorphological grading and biomechanical testing showed highest values at 12 weeks, with signs of tissue degradation at one year. There was no significant difference between both groups. CONCLUSION Transplantation of autologous osteoperiosteal grafts is an effective method to restore subchondral bone defects, but not the overlying cartilage as the repair tissue deteriorates in the long term. Subperiosteal growth factors injection did not stimulate tissue differentiation on a biomechanical and histomorphological level.
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Matmati M, Ng TF, Rosenzweig DH, Quinn TM. Protection of Bovine Chondrocyte Phenotype by Heat Inactivation of Allogeneic Serum in Monolayer Expansion Cultures. Ann Biomed Eng 2013; 41:894-903. [DOI: 10.1007/s10439-012-0732-z] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2012] [Accepted: 12/20/2012] [Indexed: 10/27/2022]
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TGFβ inhibition during expansion phase increases the chondrogenic re-differentiation capacity of human articular chondrocytes. Osteoarthritis Cartilage 2012; 20:1152-60. [PMID: 22772045 DOI: 10.1016/j.joca.2012.06.010] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/18/2012] [Revised: 05/30/2012] [Accepted: 06/21/2012] [Indexed: 02/02/2023]
Abstract
OBJECTIVE Autologous chondrocyte implantation is a cell-based treatment to repair articular cartilage defects, relying on the availability of expanded (de-differentiated) chondrocytes. Unfortunately, the expansion process causes several phenotypical changes, requiring re-establishment of the native chondrogenic phenotype to sustain proper repair. Among other proteins, transforming growth factor-β (TGFβ) is known to influence the chondrogenic re-differentiation of human articular chondrocytes (HACs) and their matrix deposition. Thus we investigated the effects of TGFβ-depletion during the expansion phase. DESIGN HACs were isolated from articular cartilage and expanded in the canonical serum-supplemented medium [fetal calf serum (FCS)] or in a chemically-defined (CD) medium, with or without anti-TGFβ antibody administration. The re-differentiation potential of the cells was assessed by pellet cultures, gene expression analysis and histology. RESULTS Cell proliferation proceeded more rapidly in CD-medium than in FCS-medium; it was not affected by the use of anti-TGFβ antibody but was further increased by addition of exogenous TGFβ1, via increased p-Smad1/5/8. Conversely, in FCS-medium, addition of anti-TGFβ antibody decreased both proliferation and p-Smad1/5/8 level. Challenging either FCS- or CD-medium with anti-TGFβ antibody during expansion enhanced chondrogenesis in the subsequent pellet cultures. Moreover, TGFβ-depletion during expansion in CD-medium inhibited mRNA expression of hypertrophic markers, collagen type-X (COL10) and matrix metalloproteinase-13 (MMP-13). Interestingly, the TGFβ1 level detected by enzyme-linked immunosorbent sandwich assay (ELISA) during cell expansion was correlated with COL10 mRNA expression after re-differentiation. CONCLUSION TGFβ-depletion during expansion improves the re-differentiation capacity of chondrocytes and inhibits hypertrophy. These results indicate the importance of the expansion medium composition to improve chondrogenic re-differentiation and to inhibit hypertrophy.
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