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Piccolo S, Grieco G, Visconte C, De Luca P, Taiana M, Zagra L, Ragni E, de Girolamo L. Starvation and Inflammation Modulate Adipose Mesenchymal Stromal Cells' Molecular Signature. J Pers Med 2024; 14:847. [PMID: 39202038 PMCID: PMC11355917 DOI: 10.3390/jpm14080847] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2024] [Revised: 07/23/2024] [Accepted: 08/08/2024] [Indexed: 09/03/2024] Open
Abstract
Mesenchymal stromal cells (MSCs) and their released factors (secretome) are intriguing options for regenerative medicine approaches based on the management of inflammation and tissue restoration, as in joint disorders like osteoarthritis (OA). Production strategy may modulate cells and secretome fingerprints, and for the latter, the effect of serum removal by starvation used in clinical-grade protocols has been underestimated. In this work, the effect of starvation on the molecular profile of interleukin 1 beta (IL1β)-primed adipose-derived MSCs (ASCs) was tested by assessing the expression level of 84 genes related to secreted factors and 84 genes involved in defining stemness potential. After validation at the protein level, the effect of starvation modulation in the secretomes was tested in a model of OA chondrocytes. IL1β priming in vitro led to an increase in inflammatory mediators' release and reduced anti-inflammatory potential on chondrocytes, features reversed by subsequent starvation. Therefore, when applying serum removal-based clinical-grade protocols for ASCs' secretome production, the effects of starvation must be carefully considered and investigated.
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Affiliation(s)
- Simona Piccolo
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Giulio Grieco
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Caterina Visconte
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Paola De Luca
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Michela Taiana
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Luigi Zagra
- Hip Department, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy;
| | - Enrico Ragni
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
| | - Laura de Girolamo
- Laboratorio di Biotecnologie Applicate all’Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via Cristina Belgioioso 173, 20157 Milano, Italy; (S.P.); (G.G.); (C.V.); (P.D.L.); (M.T.); (L.d.G.)
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Hou XY, Danzeng LM, Wu YL, Ma QH, Yu Z, Li MY, Li LS. Mesenchymal stem cells and their derived exosomes for the treatment of COVID-19. World J Stem Cells 2024; 16:353-374. [PMID: 38690515 PMCID: PMC11056634 DOI: 10.4252/wjsc.v16.i4.353] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/21/2023] [Revised: 02/17/2024] [Accepted: 03/15/2024] [Indexed: 04/25/2024] Open
Abstract
Coronavirus disease 2019 (COVID-19) is an acute respiratory infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 infection typically presents with fever and respiratory symptoms, which can progress to severe respiratory distress syndrome and multiple organ failure. In severe cases, these complications may even lead to death. One of the causes of COVID-19 deaths is the cytokine storm caused by an overactive immune response. Therefore, suppressing the overactive immune response may be an effective strategy for treating COVID-19. Mesenchymal stem cells (MSCs) and their derived exosomes (MSCs-Exo) have potent homing abilities, immunomodulatory functions, regenerative repair, and antifibrotic effects, promising an effective tool in treating COVID-19. In this paper, we review the main mechanisms and potential roles of MSCs and MSCs-Exo in treating COVID-19. We also summarize relevant recent clinical trials, including the source of cells, the dosage and the efficacy, and the clinical value and problems in this field, providing more theoretical references for the clinical use of MSCs and MSCs-Exo in the treatment of COVID-19.
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Affiliation(s)
- Xiang-Yi Hou
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun 130021, Jilin Province, China
| | - La-Mu Danzeng
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun 130021, Jilin Province, China
| | - Yi-Lin Wu
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun 130021, Jilin Province, China
| | - Qian-Hui Ma
- Department of Pharmacy, Jilin University, Changchun 130021, Jilin Province, China
| | - Zheng Yu
- The First Hospital of Jilin University, Jilin University, Changchun 130021, Jilin Province, China
| | - Mei-Ying Li
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun 130021, Jilin Province, China
| | - Li-Sha Li
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun 130021, Jilin Province, China.
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Yin H, Mao K, Huang Y, Guo A, Shi L. Tendon stem/progenitor cells are promising reparative cell sources for multiple musculoskeletal injuries of concomitant articular cartilage lesions associated with ligament injuries. J Orthop Surg Res 2023; 18:869. [PMID: 37968672 PMCID: PMC10647040 DOI: 10.1186/s13018-023-04313-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/16/2023] [Accepted: 10/23/2023] [Indexed: 11/17/2023] Open
Abstract
BACKGROUND Trauma-related articular cartilage lesions usually occur in conjunction with ligament injuries. Torn ligaments are frequently reconstructed with tendon autograft and has been proven to achieve satisfactory clinical outcomes. However, treatments for the concomitant articular cartilage lesions are still very insufficient. The current study was aimed to evaluate whether stem cells derived from tendon tissue can be considered as an alternative reparative cell source for cartilage repair. METHODS Primary human tendon stem/progenitor cells (hTSPCs) were isolated from 4 male patients (32 ± 8 years) who underwent ACL reconstruction surgery with autologous semitendinosus and gracilis tendons. The excessive tendon tissue after graft preparation was processed for primary cell isolation with an enzyme digestion protocol. Decellularization cartilage matrix (DCM) was used to provide a chondrogenic microenvironment for hTSPCs. Cell viability, cell morphology on the DCM, as well as their chondrogenic differentiation were evaluated. RESULTS DAPI staining and DNA quantitative analysis (61.47 μg per mg dry weight before and 2.64 μg/mg after decellularization) showed that most of the cells in the cartilage lacuna were removed after decellularization process. Whilst, the basic structure of the cartilage tissue was preserved and the main ECM components, collagen type II and sGAG were retained after decellularization, which were revealed by DMMB assay and histology. Live/dead staining and proliferative assay demonstrated that DCM supported attachment, survival and proliferation of hTSPCs with an excellent biocompatibility. Furthermore, gene expression analysis indicated that chondrogenic differentiation of hTSPC was induced by the DCM microenvironment, with upregulation of chondrogenesis-related marker genes, COL 2 and SOX9, without the use of exogenous growth factors. CONCLUSION DCM supported hTSPCs attachment and proliferation with high biocompatibility. Moreover, TSPCs underwent a distinct chondrogenesis after the induction of a chondrogenic microenvironment provided by DCM. These results indicated that TSPCs are promising reparative cell sources for promoting cartilage repair. Particularly, in the cohort that articular cartilage lesions occur in conjunction with ligament injuries, autologous TSPCs can be isolated from a portion of the tendon autograph harvested for ligaments reconstruction. In future clinical practice, combined ligament reconstruction with TSPCs- based therapy for articular cartilage repair can to be considered to achieve superior repair of these associated injuries, in which autologous TSPCs can be isolated from a portion of the tendon autograph harvested for ligaments reconstruction.
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Affiliation(s)
- Heyong Yin
- Department of Orthopaedics, Beijing Friendship Hospital, Capital Medical University, Beijing, 100053, China
| | - Kelei Mao
- Department of Orthopaedics, Beijing Friendship Hospital, Capital Medical University, Beijing, 100053, China
| | - Yufu Huang
- Department of Orthopaedics, Beijing Friendship Hospital, Capital Medical University, Beijing, 100053, China
| | - Ai Guo
- Department of Orthopaedics, Beijing Friendship Hospital, Capital Medical University, Beijing, 100053, China.
| | - Lin Shi
- Department of Orthopaedics, Beijing Friendship Hospital, Capital Medical University, Beijing, 100053, China.
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Different Sources of Mesenchymal Stem Cells for Tissue Regeneration: A Guide to Identifying the Most Favorable One in Orthopedics and Dentistry Applications. Int J Mol Sci 2022; 23:ijms23116356. [PMID: 35683035 PMCID: PMC9181542 DOI: 10.3390/ijms23116356] [Citation(s) in RCA: 59] [Impact Index Per Article: 19.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2022] [Revised: 05/31/2022] [Accepted: 06/03/2022] [Indexed: 12/04/2022] Open
Abstract
The success of regenerative medicine in various clinical applications depends on the appropriate selection of the source of mesenchymal stem cells (MSCs). Indeed, the source conditions, the quality and quantity of MSCs, have an influence on the growth factors, cytokines, extracellular vesicles, and secrete bioactive factors of the regenerative milieu, thus influencing the clinical result. Thus, optimal source selection should harmonize this complex setting and ensure a well-personalized and effective treatment. Mesenchymal stem cells (MSCs) can be obtained from several sources, including bone marrow and adipose tissue, already used in orthopedic regenerative applications. In this sense, for bone, dental, and oral injuries, MSCs could provide an innovative and effective therapy. The present review aims to compare the properties (proliferation, migration, clonogenicity, angiogenic capacity, differentiation potential, and secretome) of MSCs derived from bone marrow, adipose tissue, and dental tissue to enable clinicians to select the best source of MSCs for their clinical application in bone and oral tissue regeneration to delineate new translational perspectives. A review of the literature was conducted using the search engines Web of Science, Pubmed, Scopus, and Google Scholar. An analysis of different publications showed that all sources compared (bone marrow mesenchymal stem cells (BM-MSCs), adipose tissue mesenchymal stem cells (AT-MSCs), and dental tissue mesenchymal stem cells (DT-MSCs)) are good options to promote proper migration and angiogenesis, and they turn out to be useful for gingival, dental pulp, bone, and periodontal regeneration. In particular, DT-MSCs have better proliferation rates and AT and G-MSC sources showed higher clonogenicity. MSCs from bone marrow, widely used in orthopedic regenerative medicine, are preferable for their differentiation ability. Considering all the properties among sources, BM-MSCs, AT-MSCs, and DT-MSCs present as potential candidates for oral and dental regeneration.
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Ragni E, Viganò M, Torretta E, Perucca Orfei C, Colombini A, Tremolada C, Gelfi C, de Girolamo L. Characterization of Microfragmented Adipose Tissue Architecture, Mesenchymal Stromal Cell Content and Release of Paracrine Mediators. J Clin Med 2022; 11:2231. [PMID: 35456324 PMCID: PMC9026471 DOI: 10.3390/jcm11082231] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2022] [Revised: 04/01/2022] [Accepted: 04/13/2022] [Indexed: 12/15/2022] Open
Abstract
The use of microfragmented adipose tissue (µFAT) for the treatment of musculoskeletal disorders, especially osteoarthritis (OA), is gaining popularity, following positive results reported in recent case series and clinical trials. Although these outcomes were postulated to rely on paracrine signals, to date, a thorough fingerprint of released molecules is largely missing. The purpose of this study was to first characterize both structure and cell content of unprocessed lipoaspirate (LA) and µFAT, and further identify and frame the array of signaling factors in the context of OA disease, by means of high throughput qRT-PCR for extracellular-vesicle (EV) embedded miRNAs and proteomics for tissue and secreted factors. Cell count showed reduction of blood cells in µFAT, confirmed by histological and flow cytometry analyses, that also showed a conserved presence of structural, endothelial and stromal components and pericytes. In the secretome, 376 and 381 EV-miRNAs in LA and µFAT, respectively, were identified. In particular, most abundant and µFAT upregulated EV-miRNAs were mainly recapitulating those already reported as ASC-EVs-specific, with crucial roles in cartilage protection and M2 macrophage polarization, while only a scarce presence of those related to blood cells emerged. Furthermore, secretome proteomic analysis revealed reduction in µFAT of acute phase factors driving OA progression. Taken together, these results suggest that processing of LA into µFAT allows for removal of blood elements and maintenance of tissue structure and stromal cell populations, and possibly the increase of OA-protective molecular features. Thus, microfragmentation represents a safe and efficient method for the application of adipose tissue properties in the frame of musculoskeletal disorders.
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Affiliation(s)
- Enrico Ragni
- Laboratorio di Biotecnologie Applicate all'Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milano, Italy
| | - Marco Viganò
- Laboratorio di Biotecnologie Applicate all'Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milano, Italy
| | - Enrica Torretta
- Laboratorio di Proteomica e Scienze Separative, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milan, Italy
| | - Carlotta Perucca Orfei
- Laboratorio di Biotecnologie Applicate all'Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milano, Italy
| | - Alessandra Colombini
- Laboratorio di Biotecnologie Applicate all'Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milano, Italy
| | - Carlo Tremolada
- Image Regenerative Clinic, Via Mascagni 14, I-20122 Milan, Italy
| | - Cecilia Gelfi
- Laboratorio di Proteomica e Scienze Separative, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milan, Italy
- Department of Biomedical Sciences for Health, University of Milan, Via Fratelli Cervi 93, I-20054 Segrate, Italy
| | - Laura de Girolamo
- Laboratorio di Biotecnologie Applicate all'Ortopedia, IRCCS Istituto Ortopedico Galeazzi, Via R. Galeazzi 4, I-20161 Milano, Italy
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Zamudio-Cuevas Y, Plata-Rodríguez R, Fernández-Torres J, Flores KM, Cárdenas-Soria VH, Olivos-Meza A, Hernández-Rangel A, Landa-Solís C. Synovial membrane mesenchymal stem cells for cartilaginous tissues repair. Mol Biol Rep 2022; 49:2503-2517. [PMID: 35013859 DOI: 10.1007/s11033-021-07051-z] [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: 09/29/2021] [Accepted: 12/02/2021] [Indexed: 10/19/2022]
Abstract
BACKGROUND The present review is focused on general aspects of the synovial membrane as well as specialized aspects of its cellular constituents, particularly the composition and location of synovial membrane mesenchymal stem cells (S-MSCs). S-MSC multipotency properties are currently at the center of translational medicine for the repair of multiple joint tissues, such as articular cartilage and meniscus lesions. METHODS AND RESULTS We reviewed the results of in vitro and in vivo research on the current clinical applications of S-MSCs, surface markers, cell culture techniques, regenerative properties, and immunomodulatory mechanisms of S-MSCs as well as the practical limitations of the last twenty-five years (1996 to 2021). CONCLUSIONS Despite the poor interest in the development of new clinical trials for the application of S-MSCs in joint tissue repair, we found evidence to support the clinical use of S-MSCs for cartilage repair. S-MSCs can be considered a valuable therapy for the treatment of repairing joint lesions.
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Affiliation(s)
- Yessica Zamudio-Cuevas
- Laboratorio de Líquido Sinovial, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289 Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Ricardo Plata-Rodríguez
- Laboratorio de Líquido Sinovial, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289 Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Javier Fernández-Torres
- Laboratorio de Líquido Sinovial, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289 Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Karina Martínez Flores
- Laboratorio de Líquido Sinovial, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289 Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Víctor Hugo Cárdenas-Soria
- Unidad de Ingeniería de Tejidos, Terapia Celular y Medicina Regenerativa, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289. Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Anell Olivos-Meza
- Ortopedia del Deporte y Artroscopía, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289 Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico
| | - Adriana Hernández-Rangel
- Instituto Politécnico Nacional-ESIQIE, Av. Luis Enrique Erro S/N, Nueva Industrial Vallejo, Gustavo A. Madero, 07738, Mexico City, CDMX, Mexico
| | - Carlos Landa-Solís
- Unidad de Ingeniería de Tejidos, Terapia Celular y Medicina Regenerativa, Instituto Nacional de Rehabilitación Luis Guillermo Ibarra Ibarra, Calzada México-Xochimilco #289. Col. Arenal de Guadalupe, Delegación Tlalpan, 14389, Mexico City, Mexico.
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Leonardi EA, Xiao M, Murray IR, Robinson WH, Abrams GD. Tendon-Derived Progenitor Cells With Multilineage Potential Are Present Within Human Patellar Tendon. Orthop J Sports Med 2021; 9:23259671211023452. [PMID: 34435068 PMCID: PMC8381435 DOI: 10.1177/23259671211023452] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/21/2021] [Accepted: 02/24/2021] [Indexed: 01/13/2023] Open
Abstract
Background: Progenitor cells serve as a promising source of regenerative potential in a
variety of tissue types yet remain underutilized in tendinopathy.
Tendon-derived progenitor cells (TDPCs) have previously been isolated from
hamstring tendon but only as part of a concomitant medical procedure.
Determining the presence of TDPCs in patellar tendon may facilitate clinical
utilization of these cells because of the relative accessibility of this
location for tissue harvest. Purpose: To characterize TDPCs in human patellar tendon samples. Study Design: Descriptive laboratory study. Methods: Human patellar tendon samples were obtained during elective knee surgery.
TDPCs were isolated and seeded at an optimal low cell density and
subcultured to confluence for up to 2 passages. Flow cytometry was used to
analyze for the expression of CD90+, CD105+, CD44+, and CD31–, CD34–, and
CD45– markers. The multilineage differentiation potential of TDPCs was
tested in vitro via adipogenic, osteogenic, and chondrogenic culture with
subsequent cytochemical staining for Oil Red O, Alizarin Red, and Alcian
Blue, respectively. Enzyme-linked immunosorbent assay was used to quantify
the amount of adiponectin, alkaline phosphatase, and SRY-box transcription
factor 9 secreted into cell culture supernatant for further confirmation of
lineage differentiation. Results were analyzed statistically using the
2-tailed Student t test. Results: TDPCs demonstrated near-uniform expression of CD90, CD105, and CD44 with
minimal expression of CD34, CD31, and CD45. Adipogenic, osteogenic, and
chondrogenic differentiation of TDPCs was confirmed using qualitative
analysis. The expression of adiponectin, alkaline phosphatase, and SRY-box
transcription factor 9 were significantly increased in differentiated cells
versus undifferentiated TDPCs (P < .05). Conclusion: TDPCs can be successfully isolated from human patellar tendon samples, and
they exhibit characteristics of multipotent progenitor cells. Clinical Relevance: These data demonstrate the promise of patellar tendon tissue as a source of
progenitor cells for use in biologic therapies for the treatment of
tendinopathy.
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Affiliation(s)
- Erika A Leonardi
- Department of Orthopedic Surgery, Stanford University School of Medicine, Stanford, California, USA
| | - Michelle Xiao
- Department of Orthopedic Surgery, Stanford University School of Medicine, Stanford, California, USA
| | - Iain R Murray
- Department of Orthopedic Surgery, Stanford University School of Medicine, Stanford, California, USA
| | - William H Robinson
- Division of Rheumatology and Immunology, Department of Medicine, Stanford University School of Medicine, Stanford, California, USA.,Palo Alto Division, VA Palo Alto Health Care System, Palo Alto, California, USA
| | - Geoffrey D Abrams
- Department of Orthopedic Surgery, Stanford University School of Medicine, Stanford, California, USA
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Application of Stem Cell Therapy for ACL Graft Regeneration. Stem Cells Int 2021; 2021:6641818. [PMID: 34381504 PMCID: PMC8352687 DOI: 10.1155/2021/6641818] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2020] [Revised: 02/19/2021] [Accepted: 06/30/2021] [Indexed: 02/07/2023] Open
Abstract
Graft regeneration after anterior cruciate ligament (ACL) reconstruction surgery is a complex three-stage process, which usually takes a long duration and often results in fibrous scar tissue formation that exerts a detrimental impact on the patients' prognosis. Hence, as a regeneration technique, stem cell transplantation has attracted increasing attention. Several different stem cell types have been utilized in animal experiments, and almost all of these have shown good capacity in improving tendon-bone regeneration. Various differentiation inducers have been widely applied together with stem cells to enhance specific lineage differentiation, such as recombinant gene transfection, growth factors, and biomaterials. Among the various different types of stem cells, bone marrow-derived mesenchymal stem cells (BMSCs) have been investigated the most, while ligament stem progenitor cells (LDSCs) have demonstrated the best potential in generating tendon/ligament lineage cells. In the clinic, 4 relevant completed trials have been reported, but only one trial with BMSCs showed improved outcomes, while 5 relevant trials are still in progress. This review describes the process of ACL graft regeneration after implantation and summarizes the current application of stem cells from bench to bedside, as well as discusses future perspectives in this field.
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Bochon K, Zielniok K, Gawlak M, Zawada K, Zarychta-Wiśniewska W, Siennicka K, Struzik S, Pączek L, Burdzińska A. The Effect of L-Ascorbic Acid and Serum Reduction on Tenogenic Differentiation of Human Mesenchymal Stromal Cells. Int J Stem Cells 2021; 14:33-46. [PMID: 33122467 PMCID: PMC7904532 DOI: 10.15283/ijsc20023] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2020] [Revised: 08/02/2020] [Accepted: 08/17/2020] [Indexed: 12/19/2022] Open
Abstract
Background and Objectives Despite significant improvement in the treatment of tendon injuries, the full tissue recovery is often not possible because of its limited ability to auto-repair. The transplantation of mesenchymal stromal cells (MSCs) is considered as a novel approach in the treatment of tendinopathies. The question about the optimal culture conditions remains open. In this study we aimed to investigate if serum reduction, L-ascorbic acid supplementation or a combination of both factors can induce tenogenic differentiation of human adipose-derived MSCs (ASCs). Methods and Results Human ASCs from 3 healthy donors were used in the study. The tested conditions were: 0.5 mM of ascorbic acid 2-phosphate (AA-2P), reduced serum content (2% FBS) or combination of these two factors. The combination of AA-2P and 2% FBS was the only experimental condition that caused a significant increase of the expression of all analyzed genes related to tenogenesis (SCLERAXIS, MOHAWK, COLLAGEN_1, COLLAGEN_3, DECORIN) in comparison to the untreated control (evaluated by RT-PCR, 5th day of experiment). Moreover, this treatment significantly increased the synthesis of SCLERAXIS, MOHAWK, COLLAGEN_1, COLLAGEN_3 proteins at the same time point (evaluated by Western blot method). Double immunocytochemical staining revealed that AA-2P significantly increased the extracellular deposition of both types of collagens. Semi-quantitative Electron Spin Resonance analysis of ascorbyl free radical revealed that AA-2P do not induce harmful transition metals-driven redox reactions in cell culture media. Conclusions Obtained results justify the use of reduced content of serum with the addition of 0.5 mM of AA-2P in tenogenic inducing media.
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Affiliation(s)
- Karolina Bochon
- Department of Immunology, Transplantology and Internal Diseases, Medical University of Warsaw, Warsaw, Poland
| | - Katarzyna Zielniok
- Department of Immunology, Transplantology and Internal Diseases, Medical University of Warsaw, Warsaw, Poland
| | - Maciej Gawlak
- Department of Pharmacodynamics and Pathophysiology, Centre for Preclinical Research and Technology, Medical University of Warsaw, Warsaw, Poland
| | - Katarzyna Zawada
- Department of Physical Chemistry, Faculty of Pharmacy with the Laboratory Medicine Division, Medical University of Warsaw, Warsaw, Poland
| | | | - Katarzyna Siennicka
- Department of Regenerative Medicine, Maria Sklodowska-Curie National Research Institute of Oncology, Warsaw, Poland
| | - Sławomir Struzik
- Department of Orthopedics and Traumatology, Medical University of Warsaw, Warsaw, Poland
| | - Leszek Pączek
- Department of Immunology, Transplantology and Internal Diseases, Medical University of Warsaw, Warsaw, Poland.,Department of Bioinformatics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland
| | - Anna Burdzińska
- Department of Immunology, Transplantology and Internal Diseases, Medical University of Warsaw, Warsaw, Poland
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Alishahi M, Anbiyaiee A, Farzaneh M, Khoshnam SE. Human Mesenchymal Stem Cells for Spinal Cord Injury. Curr Stem Cell Res Ther 2021; 15:340-348. [PMID: 32178619 DOI: 10.2174/1574888x15666200316164051] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2019] [Revised: 08/03/2019] [Accepted: 09/17/2019] [Indexed: 12/13/2022]
Abstract
Spinal Cord Injury (SCI), as a devastating and life-altering neurological disorder, is one of the most serious health issues. Currently, the management of acute SCI includes pharmacotherapy and surgical decompression. Both the approaches have been observed to have adverse physiological effects on SCI patients. Therefore, novel therapeutic targets for the management of SCI are urgently required for developing cell-based therapies. Multipotent stem cells, as a novel strategy for the treatment of tissue injury, may provide an effective therapeutic option against many neurological disorders. Mesenchymal stem cells (MSCs) or multipotent stromal cells can typically self-renew and generate various cell types. These cells are often isolated from bone marrow (BM-MSCs), adipose tissues (AD-MSCs), umbilical cord blood (UCB-MSCs), and placenta (PMSCs). MSCs have remarkable potential for the development of regenerative therapies in animal models and humans with SCI. Herein, we summarize the therapeutic potential of human MSCs in the treatment of SCI.
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Affiliation(s)
- Masoumeh Alishahi
- Department of Biology, Tehran North Branch, Islamic Azad University, Tehran, Iran
| | - Amir Anbiyaiee
- Department of Surgery, School of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz 61357-15794, Iran
| | - Maryam Farzaneh
- Physiology Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
| | - Seyed E Khoshnam
- Physiology Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
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11
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3D Bioprinting of Human Adipose-Derived Stem Cells and Their Tenogenic Differentiation in Clinical-Grade Medium. Int J Mol Sci 2020; 21:ijms21228694. [PMID: 33218011 PMCID: PMC7698777 DOI: 10.3390/ijms21228694] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2020] [Revised: 11/11/2020] [Accepted: 11/13/2020] [Indexed: 12/20/2022] Open
Abstract
Defining the best combination of cells and biomaterials is a key challenge for the development of tendon tissue engineering (TE) strategies. Adipose-derived stem cells (ASCs) are ideal candidates for this purpose. In addition, controlled cell-based products adherent to good manufacturing practice (GMP) are required for their clinical scale-up. With this aim, in this study, ASC 3D bioprinting and GMP-compliant tenogenic differentiation were investigated. In detail, primary human ASCs were embedded within a nanofibrillar-cellulose/alginate bioink and 3D-bioprinted into multi-layered square-grid matrices. Bioink viscoelastic properties and scaffold ultrastructural morphology were analyzed by rheology and scanning electron microscopy (SEM). The optimal cell concentration for printing among 3, 6 and 9 × 106 ASC/mL was evaluated in terms of cell viability. ASC morphology was characterized by SEM and F-actin immunostaining. Tenogenic differentiation ability was then evaluated in terms of cell viability, morphology and expression of scleraxis and collagen type III by biochemical induction using BMP-12, TGF-β3, CTGF and ascorbic acid supplementation (TENO). Pro-inflammatory cytokine release was also assessed. Bioprinted ASCs showed high viability and survival and exhibited a tenocyte-like phenotype after biochemical induction, with no inflammatory response to the bioink. In conclusion, we report a first proof of concept for the clinical scale-up of ASC 3D bioprinting for tendon TE.
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12
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Colombini A, Perucca Orfei C, Vincenzi F, De Luca P, Ragni E, Viganò M, Setti S, Varani K, de Girolamo L. A2A adenosine receptors are involved in the reparative response of tendon cells to pulsed electromagnetic fields. PLoS One 2020; 15:e0239807. [PMID: 32998161 PMCID: PMC7527253 DOI: 10.1371/journal.pone.0239807] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2019] [Accepted: 09/14/2020] [Indexed: 11/19/2022] Open
Abstract
Tendinopathy is a degenerative disease in which inflammatory mediators have been found to be sometimes present. The interaction between inflammation and matrix remodeling in human tendon cells (TCs) is supported by the secretion of cytokines such as IL-1β, IL-6 and IL-33. In this context, it has been demonstrated that pulsed electromagnetic fields (PEMFs) were able to reduce inflammation and promote tendon marker synthesis. The aim of this study was to evaluate the anabolic and anti-inflammatory PEMF-mediated response on TCs in an in vitro model of inflammation. Moreover, since PEMFs enhance the anti-inflammatory efficacy of adenosine through the adenosine receptors (ARs), the study also focused on the role of A2AARs. Human TCs were exposed to PEMFs for 48 hours. After stimulation, A2AAR saturation binding experiments were performed. Along with 48 hours PEMF stimulation, TCs were treated with IL-1β and A2AAR agonist CGS-21680. IL-1Ra, IL-6, IL-8, IL-10, IL-33, VEGF, TGF-β1, PGE2 release and SCX, COL1A1, COL3A1, ADORA2A expression were quantified. PEMFs exerted A2AAR modulation on TCs and promoted COL3A1 upregulation and IL-33 secretion. In presence of IL-1β, TCs showed an upregulation of ADORA2A, SCX and COL3A1 expression and an increase of IL-6, IL-8, PGE2 and VEGF secretion. After PEMF and IL-1β exposure, IL-33 was upregulated, whereas IL-6, PGE2 and ADORA2A were downregulated. These findings demonstrated that A2AARs have a role in the promotion of the TC anabolic/reparative response to PEMFs and to IL-1β.
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Affiliation(s)
- Alessandra Colombini
- Orthopaedic Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, Milan, Italy
- * E-mail:
| | | | - Fabrizio Vincenzi
- Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy
| | - Paola De Luca
- Orthopaedic Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, Milan, Italy
| | - Enrico Ragni
- Orthopaedic Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, Milan, Italy
| | - Marco Viganò
- Orthopaedic Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, Milan, Italy
| | | | - Katia Varani
- Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy
| | - Laura de Girolamo
- Orthopaedic Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, Milan, Italy
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13
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Wang HN, Huang YC, Ni GX. Mechanotransduction of stem cells for tendon repair. World J Stem Cells 2020; 12:952-965. [PMID: 33033557 PMCID: PMC7524696 DOI: 10.4252/wjsc.v12.i9.952] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/03/2020] [Revised: 05/06/2020] [Accepted: 07/19/2020] [Indexed: 02/06/2023] Open
Abstract
Tendon is a mechanosensitive tissue that transmits force from muscle to bone. Physiological loading contributes to maintaining the homeostasis and adaptation of tendon, but aberrant loading may lead to injury or failed repair. It is shown that stem cells respond to mechanical loading and play an essential role in both acute and chronic injuries, as well as in tendon repair. In the process of mechanotransduction, mechanical loading is detected by mechanosensors that regulate cell differentiation and proliferation via several signaling pathways. In order to better understand the stem-cell response to mechanical stimulation and the potential mechanism of the tendon repair process, in this review, we summarize the source and role of endogenous and exogenous stem cells active in tendon repair, describe the mechanical response of stem cells, and finally, highlight the mechanotransduction process and underlying signaling pathways.
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Affiliation(s)
- Hao-Nan Wang
- School of Sport Medicine and Rehabilitation, Beijing Sport University, Beijing 100084, China
| | - Yong-Can Huang
- Shenzhen Engineering Laboratory of Orthopaedic Regenerative Technologies, Department of Spine Surgery, Peking University Shenzhen Hospital, Shenzhen 518036, Guangdong Province, China
| | - Guo-Xin Ni
- School of Sport Medicine and Rehabilitation, Beijing Sport University, Beijing 100084, China.
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14
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Viganò M, Lugano G, Perucca Orfei C, Menon A, Ragni E, Colombini A, De Luca P, Randelli P, de Girolamo L. Autologous microfragmented adipose tissue reduces inflammatory and catabolic markers in supraspinatus tendon cells derived from patients affected by rotator cuff tears. INTERNATIONAL ORTHOPAEDICS 2020; 45:419-426. [PMID: 32642826 DOI: 10.1007/s00264-020-04693-9] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/23/2020] [Accepted: 06/29/2020] [Indexed: 01/08/2023]
Abstract
PURPOSE Rotator cuff tears are common musculoskeletal disorders, and surgical repair is characterized by a high rate of re-tear. Regenerative medicine strategies, in particular mesenchymal stem cell-based therapies, have been proposed to enhance tendon healing and reduce the re-tear rate. Autologous microfragmented adipose tissue (μFAT) allows for the clinical application of cell therapies and showed the ability to improve tenocyte proliferation and viability in previous in vitro assessments. The hypothesis of this study is that μFAT paracrine action would reduce the catabolic and inflammatory marker expression in tendon cells (TCs) derived from injured supraspinatus tendon (SST). METHODS TCs derived from injured SST were co-cultured with autologous μFAT in transwell for 48 h. Metabolic activity, DNA content, the content of soluble mediators in the media, and the gene expression of tendon-specific, inflammatory, and catabolic markers were analyzed. RESULTS μFAT-treated TCs showed a reduced expression of PTGS2 and MMP-3 with respect to untreated controls. Increased IL-1Ra, VEGF, and IL-6 content were observed in the media of μFAT-treated samples, in comparison with untreated TCs. CONCLUSION μFAT exerted an anti-inflammatory action on supraspinatus tendon cells in vitro through paracrine action, resulting in the reduction of catabolic and inflammatory marker expression. These observations potentially support the use of μFAT as adjuvant therapy in the treatment of rotator cuff disease.
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Affiliation(s)
- Marco Viganò
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
| | - Gaia Lugano
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
| | - Carlotta Perucca Orfei
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy.
| | - Alessandra Menon
- Laboratory of Applied Biomechanics, Department of Biomedical Sciences for Health, Università degli Studi di Milano, Via Mangiagalli 31, 20133, Milan, Italy.,1° Clinica Ortopedica, ASST Centro Specialistico Ortopedico Traumatologico Gaetano Pini-CTO, Piazza Cardinal Ferrari 1, 20122, Milan, Italy
| | - Enrico Ragni
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
| | - Alessandra Colombini
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
| | - Paola De Luca
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
| | - Pietro Randelli
- Laboratory of Applied Biomechanics, Department of Biomedical Sciences for Health, Università degli Studi di Milano, Via Mangiagalli 31, 20133, Milan, Italy.,1° Clinica Ortopedica, ASST Centro Specialistico Ortopedico Traumatologico Gaetano Pini-CTO, Piazza Cardinal Ferrari 1, 20122, Milan, Italy
| | - Laura de Girolamo
- Orthopedics Biotechnology Lab, IRCCS Istituto Ortopedico Galeazzi, via Riccardo Galeazzi 4, 20161, Milan, Italy
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15
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Autologous Microfragmented Adipose Tissue Reduces the Catabolic and Fibrosis Response in an In Vitro Model of Tendon Cell Inflammation. Stem Cells Int 2019; 2019:5620286. [PMID: 31885616 PMCID: PMC6915130 DOI: 10.1155/2019/5620286] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2019] [Revised: 10/17/2019] [Accepted: 11/20/2019] [Indexed: 12/16/2022] Open
Abstract
Background Mesenchymal stem cells (MSCs) emerged as a promising therapy for tendon pathologies. Microfragmented adipose tissue (μFAT) represents a convenient autologous product for the application of MSC-based therapies in the clinical setting. In the present study, the ability of μFAT to counteract inflammatory processes induced by IL-1β on human tendon cells (TCs) was evaluated. Methods Cell viability and proliferation were evaluated after 48 hours of transwell coculture of TCs and autologous μFAT in the presence or absence of IL-1β. Gene expression of scleraxis, collagen type I and type III, metalloproteinases-1 and -3, and cyclooxygenase-2 was evaluated by real-time RT-PCR. The content of VEGF, IL-1Ra, TNFα, and IL-6 was evaluated by ELISA. Results IL-1β-treated TCs showed augmented collagen type III, metalloproteases, and cyclooxygenase-2 expression. μFAT was able to reduce the expression of collagen type III and metalloproteases-1 in a significant manner, and at the same time, it enhanced the production of VEGF, IL-1Ra, and IL-6. Conclusions In this in vitro model of tendon cell inflammation, the paracrine action of μFAT, exerted by anti-inflammatory molecules and growth factors, was able to inhibit the expression of fibrosis and catabolic markers. Then, these results suggest that the application of μFAT may represent an effective conservative or adjuvant therapy for the treatment of tendon disorders.
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16
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Rajpar I, Barrett JG. Optimizing growth factor induction of tenogenesis in three-dimensional culture of mesenchymal stem cells. J Tissue Eng 2019; 10:2041731419848776. [PMID: 31205672 PMCID: PMC6535701 DOI: 10.1177/2041731419848776] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/06/2019] [Accepted: 04/16/2019] [Indexed: 12/17/2022] Open
Abstract
Adult tissue stem cells have shown promise for the treatment of debilitating tendon injuries. However, few comparisons of stem cells from different tissue sources have been made to determine the optimum stem cell source for treating tendon. Moreover, it is likely that the application of tenogenic growth factors will improve tendon stem cell treatments further, and a comprehensive comparison of a number of growth factors is needed. Thus far, different types of stem cells cannot be evaluated in a high-throughput manner. To this end, we have developed an approach to culture mesenchymal stem cells isolated from bone marrow in collagen type I hydrogels with tenogenic growth factors using economical, commercially available supplies. To optimize growth factors for this assay, FGF-2, TGF-β1, IGF-1, and/or BMP-12 were tested singly and in novel combinations of (1) BMP-12 and IGF-1, (2) TGF-β1 and IGF-1, and/or (3) BMP-12 and FGF-2 over 10 days. Our data suggest that BMP-12 supplementation alone results in the strongest expression of tendon marker genes, controlled contractility of constructs, a higher degree of cell alignment, and tendon-like tissue morphology. This easy-to-use benchtop assay can be used to screen novel sources of stem cells and cell lines for tissue engineering and tendon healing applications.
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Affiliation(s)
- Ibtesam Rajpar
- Department of Large Animal Clinical Sciences, Marion duPont Scott Equine Medical Center, Virginia-Maryland College of Veterinary Medicine, Virginia Tech, Leesburg, VA, USA
| | - Jennifer G Barrett
- Department of Large Animal Clinical Sciences, Marion duPont Scott Equine Medical Center, Virginia-Maryland College of Veterinary Medicine, Virginia Tech, Leesburg, VA, USA
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17
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Stanco D, Caprara C, Ciardelli G, Mariotta L, Gola M, Minonzio G, Soldati G. Tenogenic differentiation protocol in xenogenic-free media enhances tendon-related marker expression in ASCs. PLoS One 2019; 14:e0212192. [PMID: 30753235 PMCID: PMC6372228 DOI: 10.1371/journal.pone.0212192] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2018] [Accepted: 01/29/2019] [Indexed: 12/03/2022] Open
Abstract
Adipose-derived stem cells (ASCs) are multipotent and immune-privileged mesenchymal cells, making them ideal candidates for therapeutic purposes to manage tendon disorders. Providing safe and regulated cell therapy products to patients requires adherence to good manufacturing practices. To this aim we investigated the in vitro tenogenic differentiation potential of ASCs using a chemically defined serum-free medium (SF) or a xenogenic-free human pooled platelet lysate medium (hPL) suitable for cell therapy and both supplemented with CTGF, TGFβ-3, BMP-12 and ascorbic acid (AA) soluble factors. Human ASCs were isolated from 4 healthy donors and they were inducted to differentiate until 14 days in both hPL and SF tenogenic media (hPL-TENO and SF-TENO). Cell viability and immunophenotype profile were analysed to evaluate mesenchymal stem cell (MSC) characteristics in both xenogenic-free media. Moreover, the expression of stemness and tendon-related markers upon cell differentiation by RT-PCR, protein staining and cytofluorimetric analysis were also performed. Our results showed the two xenogenic-free media well support cell viability of ASCs and maintain their MSC nature as demonstrated by their typical immunophenototype profile and by the expression of NANOG, OCT4 and Ki67 genes. Moreover, both hPL-TENO and SF-TENO expressed significant high levels of the tendon-related genes SCX, COL1A1, COL3A1, COMP, MMP3 and MMP13 already at early time points in comparison to the respective controls. Significant up-regulations in scleraxis, collagen and tenomodulin proteins were also demonstrated at in both differentiated SF and hPL ASCs. In conclusion, we demonstrated firstly the feasibility of both serum and xenogenic-free media tested to culture ASCs moving forward the GMP-compliant approaches for clinical scale expansion of human MSCs needed for therapeutical application of stem cells. Moreover, a combination of CTGF, BMP-12, TGFβ3 and AA factors strongly and rapidly induce human ASCs to differentiate into tenocyte-like cells.
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Affiliation(s)
- Deborah Stanco
- Swiss Stem Cell Foundation, Gentilino, Switzerland
- Department of Mechanical and Aerospace Engineering, Politecnico di Torino, Turin, Italy
| | | | - Gianluca Ciardelli
- Department of Mechanical and Aerospace Engineering, Politecnico di Torino, Turin, Italy
| | | | - Mauro Gola
- Swiss Stem Cell Foundation, Gentilino, Switzerland
| | | | - Gianni Soldati
- Swiss Stem Cell Foundation, Gentilino, Switzerland
- * E-mail:
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18
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Perucca Orfei C, Viganò M, Pearson JR, Colombini A, De Luca P, Ragni E, Santos-Ruiz L, de Girolamo L. In Vitro Induction of Tendon-Specific Markers in Tendon Cells, Adipose- and Bone Marrow-Derived Stem Cells is Dependent on TGFβ3, BMP-12 and Ascorbic Acid Stimulation. Int J Mol Sci 2019; 20:ijms20010149. [PMID: 30609804 PMCID: PMC6337430 DOI: 10.3390/ijms20010149] [Citation(s) in RCA: 34] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2018] [Accepted: 12/27/2018] [Indexed: 12/17/2022] Open
Abstract
Mesenchymal Stem Cells (MSCs) and tissue-specific progenitors have been proposed as useful tools for regenerative medicine approaches in bone, cartilage and tendon-related pathologies. The differentiation of cells towards the desired, target tissue-specific lineage has demonstrated advantages in the application of cell therapies and tissue engineering. Unlike osteogenic and chondrogenic differentiation, there is no consensus on the best tenogenic induction protocol. Many growth factors have been proposed for this purpose, including BMP-12, b-FGF, TGF-β3, CTGF, IGF-1 and ascorbic acid (AA). In this study, different combinations of these growth factors have been tested in the context of a two-step differentiation protocol, in order to define their contribution to the induction and maintenance of tendon marker expression in adipose tissue and bone marrow derived MSCs and tendon cells (TCs), respectively. Our results demonstrate that TGF-β3 is the main inducer of scleraxis, an early expressed tendon marker, while at the same time inhibiting tendon markers normally expressed later, such as decorin. In contrast, we find that decorin is induced by BMP-12, b-FGF and AA. Our results provide new insights into the effect of different factors on the tenogenic induction of MSCs and TCs, highlighting the importance of differential timing in TGF-β3 stimulation.
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Affiliation(s)
| | - Marco Viganò
- IRCCS Istituto Ortopedico Galeazzi, Orthopaedic Biotechnology Lab, 20161 Milan, Italy.
| | - John R Pearson
- Andalusian Centre for Nanomedicine and Biotechnology, BIONAND, 29590 Málaga, Spain.
| | - Alessandra Colombini
- IRCCS Istituto Ortopedico Galeazzi, Orthopaedic Biotechnology Lab, 20161 Milan, Italy.
| | - Paola De Luca
- IRCCS Istituto Ortopedico Galeazzi, Orthopaedic Biotechnology Lab, 20161 Milan, Italy.
| | - Enrico Ragni
- IRCCS Istituto Ortopedico Galeazzi, Orthopaedic Biotechnology Lab, 20161 Milan, Italy.
| | - Leonor Santos-Ruiz
- Andalusian Centre for Nanomedicine and Biotechnology, BIONAND, 29590 Málaga, Spain.
- Network Centre for Biomedical Research ⁻ Biotechnology, Biomaterials and Nanomedicine, CIBER-BBN, 50018 Zaragoza, Spain.
- Department of Cell Biology, Genetics and Physiology, Instituto de Investigación University of Málaga, 29016 Malaga, Spain.
| | - Laura de Girolamo
- IRCCS Istituto Ortopedico Galeazzi, Orthopaedic Biotechnology Lab, 20161 Milan, Italy.
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Zhang H, Liu MF, Liu RC, Shen WL, Yin Z, Chen X. Physical Microenvironment-Based Inducible Scaffold for Stem Cell Differentiation and Tendon Regeneration. TISSUE ENGINEERING PART B-REVIEWS 2018; 24:443-453. [PMID: 29724151 DOI: 10.1089/ten.teb.2018.0018] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Tendon injuries are common musculoskeletal system disorders, but the tendons have poor regeneration ability. To address this issue, tendon tissue engineering provides potential strategies for future therapeutic treatment. Elements of the physical microenvironment, such as the mechanical force and surface topography, play a vital role in regulating stem cell fate, enhancing the differentiation efficiency of seed cells in tendon tissue engineering. Various inducible scaffolds have been widely explored for tendon regeneration, and scaffold-enhancing modifications have been extensively studied. In this review, we systematically summarize the effects of the physical microenvironment on stem cell differentiation and tendon regeneration; we also provide an overview of the inducible scaffolds for stem cell tenogenic differentiation. Finally, we suggest some potential scaffold-based therapies for tendon injuries, presenting an interesting perspective on tendon regenerative medicine.
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Affiliation(s)
- Hong Zhang
- 1 School of Basic Medical Sciences, and Department of Orthopedic Surgery of The Second Affiliated Hospital, School of Medicine, Zhejiang University , Hangzhou, China .,2 Dr. Li Dak Sum & Yip Yio Chin Center for Stem Cell and Regenerative Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,3 Key Laboratory of Tissue Engineering and Regenerative Medicine of Zhejiang Province, School of Medicine, Zhejiang University , Hangzhou, China
| | - Meng-Fei Liu
- 1 School of Basic Medical Sciences, and Department of Orthopedic Surgery of The Second Affiliated Hospital, School of Medicine, Zhejiang University , Hangzhou, China .,2 Dr. Li Dak Sum & Yip Yio Chin Center for Stem Cell and Regenerative Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,3 Key Laboratory of Tissue Engineering and Regenerative Medicine of Zhejiang Province, School of Medicine, Zhejiang University , Hangzhou, China
| | - Ri-Chun Liu
- 4 Guangxi Collaborative Innovation Center for Biomedicine, Guangxi Medical University , Nanning, China
| | - Wei-Liang Shen
- 2 Dr. Li Dak Sum & Yip Yio Chin Center for Stem Cell and Regenerative Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,5 Department of Sports Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,6 China Orthopedic Regenerative Medicine Group (CORMed) , Hangzhou, China .,7 State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, School of Medicine, Zhejiang University , Hangzhou, China
| | - Zi Yin
- 1 School of Basic Medical Sciences, and Department of Orthopedic Surgery of The Second Affiliated Hospital, School of Medicine, Zhejiang University , Hangzhou, China .,2 Dr. Li Dak Sum & Yip Yio Chin Center for Stem Cell and Regenerative Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,3 Key Laboratory of Tissue Engineering and Regenerative Medicine of Zhejiang Province, School of Medicine, Zhejiang University , Hangzhou, China .,6 China Orthopedic Regenerative Medicine Group (CORMed) , Hangzhou, China
| | - Xiao Chen
- 1 School of Basic Medical Sciences, and Department of Orthopedic Surgery of The Second Affiliated Hospital, School of Medicine, Zhejiang University , Hangzhou, China .,2 Dr. Li Dak Sum & Yip Yio Chin Center for Stem Cell and Regenerative Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,3 Key Laboratory of Tissue Engineering and Regenerative Medicine of Zhejiang Province, School of Medicine, Zhejiang University , Hangzhou, China .,4 Guangxi Collaborative Innovation Center for Biomedicine, Guangxi Medical University , Nanning, China .,5 Department of Sports Medicine, School of Medicine, Zhejiang University , Hangzhou, China .,6 China Orthopedic Regenerative Medicine Group (CORMed) , Hangzhou, China
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Zhou K, Feng B, Wang W, Jiang Y, Zhang W, Zhou G, Jiang T, Cao Y, Liu W. Nanoscaled and microscaled parallel topography promotes tenogenic differentiation of ASC and neotendon formation in vitro. Int J Nanomedicine 2018; 13:3867-3881. [PMID: 30013341 PMCID: PMC6038871 DOI: 10.2147/ijn.s161423] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Background Topography at different scales plays an important role in directing mesenchymal stem cell differentiation including adipose-derived stem cells (ASCs) and the differential effect remains to be investigated. Purpose This study aimed to investigate the similarity and difference between micro- and nanoscaled aligned topography for inducing tenogenic differentiation of human ASCs (hASCs). Methods Parallel microgrooved PDMS membrane and a parallel aligned electrospun nanofibers of gelatin/poly-ε-caprolactone mixture were employed as the models for the study. Results Aligned topographies of both microscales and nanoscales could induce an elongated cell shape with parallel alignment, as supported by quantitative cell morphology analysis (cell area, cell body aspect, and cell body major axis angle). qPCR analysis also demonstrated that the aligned topography at both scales could induce the gene expressions of various tenogenic markers at the 7th day of in vitro culture including tenomodulin, collagen I and collagen VI, decorin, tenascin-C and biglycan, but with upregulated expression of scleraxis and tenascin-C only in microscaled topography. Additionally, tenogenic differentiation at the 3rd day was confirmed only at microscale. Furthermore, microscaled topography was confirmed for its tenogenic induction at tissue level as neotendon tissue was formed with the evidence of mature type I collagen fibers only in parallel aligned polyglycolic acid (PGA) microfibers after in vitro culture with mouse ASCs. Instead, only fat tissue was formed in random patterned PGA microfibers. Conclusion Both microscaled and nanoscaled aligned topographies could induce tenogenic differentiation of hASCs and micro-scaled topography seemed better able to induce elongated cell shape and stable tenogenic marker expression when compared to nanoscaled topography. The microscaled inductive effect was also confirmed at tissue level by neotendon formation in vitro.
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Affiliation(s)
- Kaili Zhou
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ;
| | - Bei Feng
- Shanghai Children's Medical Center, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China
| | - Wenbo Wang
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ;
| | - Yongkang Jiang
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ;
| | - Wenjie Zhang
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ; .,National Tissue Engineering Center of China, Shanghai, People's Republic of China, ;
| | - Guangdong Zhou
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ; .,National Tissue Engineering Center of China, Shanghai, People's Republic of China, ;
| | - Ting Jiang
- Department of Burn and Plastic Surgery, Nanchong Central Hospital, the Second Clinical College of North Sichuan Medical College, Nanchong, Sichuan, People's Republic of China
| | - Yilin Cao
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ; .,National Tissue Engineering Center of China, Shanghai, People's Republic of China, ;
| | - Wei Liu
- Department of Plastic and Reconstructive Surgery, Shanghai Key Laboratory of Tissue Engineering, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University - School of Medicine, Shanghai, People's Republic of China, ; .,National Tissue Engineering Center of China, Shanghai, People's Republic of China, ;
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Viganò M, Perucca Orfei C, de Girolamo L, Pearson JR, Ragni E, De Luca P, Colombini A. Housekeeping Gene Stability in Human Mesenchymal Stem and Tendon Cells Exposed to Tenogenic Factors. Tissue Eng Part C Methods 2018; 24:360-367. [PMID: 29676207 DOI: 10.1089/ten.tec.2017.0518] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
The use of biochemical inducers of mesenchymal stem cell (MSC) differentiation into tenogenic lineage represents an investigated aspect of tendon disorder treatment. Bone morphogenetic protein 12 (BMP-12) is a widely studied factor, representing along with ascorbic acid (AA) and basic fibroblast growth factor (bFGF) one of the most promising stimulus in this context so far. Quantitative gene expression of specific tenogenic marker is commonly used to assess the efficacy of these supplements. Nevertheless, the reliability of these data is strongly associated with the choice of stable housekeeping genes. To date, no published studies have evaluated the stability of housekeeping genes in MSCs during tenogenic induction. Three candidate housekeeping genes (YWHAZ, RPL13A, and GAPDH) in human MSCs from bone marrow (BMSCs), adipose tissue (ASCs), and tendon cells (TCs) supplemented with BMP-12 or AA and bFGF in comparison with control untreated cells for 3 and 10 days were evaluated. GeNorm, NormFinder, and BestKeeper tools and the comparative ΔCt method were used to evaluate housekeeping gene stability and the overall ranking was determined by using by the RefFinder algorithm. In all culture conditions, YWHAZ was the most stable gene and RPL13A was the second choice. YWHAZ and RPL13A were the two most stable genes also for ASCs and BMSCs, regardless of the time point analyzed, and for TCs at 10 days of tenogenic induction. Only for TCs at 3 days of tenogenic induction were GAPDH and YWHAZ the best performers. In conclusion, our findings will be useful for the proper selection of housekeeping genes in studies involving MSCs cultured in the presence of tenogenic factors, to obtain accurate and high-quality data from quantitative gene expression analysis.
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Affiliation(s)
- Marco Viganò
- 1 Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute , Milan, Italy
| | | | - Laura de Girolamo
- 1 Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute , Milan, Italy
| | - John R Pearson
- 2 Nano-imaging Unit, Andalusian Centre for Nanomedicine and Biotechnology, BIONAND , Málaga, Spain
| | - Enrico Ragni
- 1 Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute , Milan, Italy
| | - Paola De Luca
- 1 Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute , Milan, Italy
| | - Alessandra Colombini
- 1 Orthopaedic Biotechnology Lab, IRCCS Galeazzi Orthopaedic Institute , Milan, Italy
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22
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Yang Y, Yang JT, Chen XH, Qin BG, Li FG, Chen YX, Gu LQ, Zhu JK, Li P. Construction of tissue-engineered lymphatic vessel using human adipose derived stem cells differentiated lymphatic endothelial like cells and decellularized arterial scaffold: A preliminary study. Biotechnol Appl Biochem 2017; 65:428-434. [PMID: 28981171 DOI: 10.1002/bab.1618] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2017] [Accepted: 09/29/2017] [Indexed: 12/19/2022]
Abstract
We have previously demonstrated that human adipose-derived stem cells (hADSCs) can be differentiated into lymphatic endothelial like cells. The purpose of this study was to investigate the feasibility of utilizing the induced lymphatic endothelial like cells and decellularized arterial scaffold to construct the tissue-engineered lymphatic vessel. The hADSCs were isolated from adipose tissue in healthy adults and were characterized the multilineage differentiation potential. Decellularized arterial scaffold was prepared using the Triton x-100 method. ADSCs were differentiated into lymphatic-like endothelial cells, and the induced cells were then seeded onto the decellularized arterial scaffold to engineer the lymphatic vessel. The histological analyses were performed to examine the endothelialized construct. The decellularized arterial scaffold was successfully obtained and was able to maintain its vessel morphology. The isolated ADSCs can be differentiated into osteocytes and adipocytes. After seeding onto the scaffold, the seeded cells attached and grew well on the decellularized arterial scaffold. Our preliminary results demonstrated that the induced lymphatic endothelial like cells combined with decellularized arterial scaffold could be utilized to successfully engineer the lymphatic vessel. Our findings may be helpful for the development of tissue-engineering of the lymphatic graft.
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Affiliation(s)
- Yi Yang
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Jian-Tao Yang
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Xiao-Hu Chen
- Department of Orthopedic Trauma, The Hui Ya Hospital of Sun Yat-sen University, Huizhou, People's Republic of China
| | - Ben-Gang Qin
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Fu-Gui Li
- Department of Cancer Institute, The Zhong Shan Hospital of Sun Yat-sen University, Zhongshan, People's Republic of China
| | - Yun-Xian Chen
- Department of Hematology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Li-Qiang Gu
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Jia-Kai Zhu
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
| | - Ping Li
- Department of Microsurgery and Orthopedic Trauma, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, People's Republic of China
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23
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Different culture conditions affect the growth of human tendon stem/progenitor cells (TSPCs) within a mixed tendon cells (TCs) population. J Exp Orthop 2017; 4:8. [PMID: 28244027 PMCID: PMC5328904 DOI: 10.1186/s40634-017-0082-8] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/14/2016] [Accepted: 02/10/2017] [Indexed: 12/16/2022] Open
Abstract
Background Tendon resident cells (TCs) are a mixed population made of terminally differentiated tenocytes and tendon stem/progenitor cells (TSPCs). Since the enrichment of progenitors proportion could enhance the effectiveness of treatments based on these cell populations, the interest on the effect of culture conditions on the TSPCs is growing. In this study the clonal selection and the culture in presence or absence of basic fibroblast growth factor (bFGF) were used to assess their influences on the stemness properties and phenotype specific features of tendon cells. Methods Cells cultured with the different methods were analyzed in terms of clonogenic and differentiation abilities, stem and tendon specific genes expression and immunophenotype at passage 2 and passage 4. Results The clonal selection allowed to isolate cells with a higher multi-differentiation potential, but at the same time a lower proliferation rate in comparison to the whole population. Moreover, the clones express a higher amounts of stemness marker OCT4 and tendon specific transcription factor Scleraxis (SCX) mRNA, but a lower level of decorin (DCN). On the other hand, the number of cells obtained by clonal selection was extremely low and most of the clones were unable to reach a high number of passages in cultures. The presence of bFGF influences TCs morphology, enhance their proliferation rate and reduce their clonogenic ability. Interestingly, the expression of CD54, a known mesenchymal stem cell marker, is reduced in presence of bFGF at early passages. Nevertheless, bFGF does not affect the chondrogenic and osteogenic potential of TCs and the expression of tendon specific markers, while it was able to downregulate the OCT4 expression. Conclusion This study showed that clonal selection enhance progenitors content in TCs populations, but the extremely low number of cells produced with this method could represent an insurmountable obstacle to its application in clinical approaches. We observed that the addition of bFGF to the culture medium promotes the maintenance of a higher number of differentiated cells, reducing the proportion of progenitors within the whole population. Overall our findings demonstrated the importance of the use of specific culture protocols to obtain tendon cells for possible clinical applications.
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24
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Zarychta-Wiśniewska W, Burdzinska A, Kulesza A, Gala K, Kaleta B, Zielniok K, Siennicka K, Sabat M, Paczek L. Bmp-12 activates tenogenic pathway in human adipose stem cells and affects their immunomodulatory and secretory properties. BMC Cell Biol 2017; 18:13. [PMID: 28214472 PMCID: PMC5316159 DOI: 10.1186/s12860-017-0129-9] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2016] [Accepted: 02/08/2017] [Indexed: 02/07/2023] Open
Abstract
BACKGROUND Cell-based therapy is a treatment method in tendon injuries. Bone morphogenic protein 12 (BMP-12) possesses tenogenic activity and was proposed as a differentiating factor for stem cells directed to transplantation. However, BMPs belong to pleiotropic TGF-β superfamily and have diverse effect on cells. Therefore, the aim of this study was to determine if BMP-12 induces tenogenic differentiation of human adipose stem cells (hASCs) and how it affects other features of this population. RESULTS Human ASCs from 6 healthy donors were treated or not with BMP-12 (50 or 100 ng/ml, 7 days) and tested for gene expression (COLL1, SCX, MKH, DCN, TNC, RUNX2), protein expression (COLL1, COLL3, MKH), proliferation, migration, secretory activity, immunomodulatory properties and susceptibility to oxidative stress. RT-PCR revealed up-regulation of SCX, MKH and RUNX2 genes in BMP-12 treated cells (2.05, 2.65 and 1.87 fold in comparison to control, respectively, p < 0.05) and Western Blot revealed significant increase of COLL1 and MHK expression after BMP-12 treatment. Addition of BMP-12 significantly enhanced secretion of VEGF, IL-6, MMP-1 and MPP-8 by hASCs while had no effect on TGF-β, IL-10, EGF and MMP-13. Moreover, BMP-12 presence in medium attenuated inhibitory effect of hASCs on allo-activated lymphocytes proliferation. At the same time BMP-12 displayed no influence on hASCs proliferation, migration and susceptibility to oxidative stress. CONCLUSION BMP-12 activates tenogenic pathway in hASCs but also affects secretory activity and impairs immunomodulatory potential of this population that can influence the clinical outcome after cell transplantation.
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Affiliation(s)
- Weronika Zarychta-Wiśniewska
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland
| | - Anna Burdzinska
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland.
| | - Agnieszka Kulesza
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland
| | - Kamila Gala
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland
| | - Beata Kaleta
- Department of Clinical Immunology, Transplantation Institute, Medical University of Warsaw, Warsaw, Poland
| | - Katarzyna Zielniok
- Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences, Warsaw, Poland
| | - Katarzyna Siennicka
- Department of Regenerative Medicine, Maria Sklodowska-Curie Memorial Cancer Center, Warsaw, Poland
| | - Marek Sabat
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland
| | - Leszek Paczek
- Department of Immunology, Transplantology and Internal Medicine, Transplantation Institute, Medical University of Warsaw, Nowogrodzka str. 59, 02-006, Warsaw, Poland.,Department of Bioinformatics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland
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25
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Yang G, Rothrauff BB, Lin H, Yu S, Tuan RS. Tendon-Derived Extracellular Matrix Enhances Transforming Growth Factor-β3-Induced Tenogenic Differentiation of Human Adipose-Derived Stem Cells. Tissue Eng Part A 2017; 23:166-176. [PMID: 27809678 DOI: 10.1089/ten.tea.2015.0498] [Citation(s) in RCA: 40] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Because of the limited and unsatisfactory outcomes of clinical tendon repair, tissue engineering approaches using adult mesenchymal stem cells are being considered a promising alternative strategy to heal tendon injuries. Successful and functional tendon tissue engineering depends on harnessing the biochemical cues presented by the native tendon extracellular matrix (ECM) and the embedded tissue-specific biofactors. In this study, we have prepared and characterized the biological activities of a soluble extract of decellularized tendon ECM (tECM) on adult adipose-derived stem cells (ASCs), on the basis of histological, biochemical, and gene expression analyses. The results showed that tECM enhances the proliferation and transforming growth factor (TGF)-β3-induced tenogenesis of ASCs in both plate and scaffold cultures in vitro, and modulates matrix deposition of ASCs seeded in scaffolds. These findings suggest that combining tendon ECM extract with TGF-β3 treatment is a possible alternative approach to induce tenogenesis for ASCs.
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Affiliation(s)
- Guang Yang
- 1 Center for Cellular and Molecular Engineering, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,3 Department of Bioengineering, University of Pittsburgh Swanson School of Engineering , Pittsburgh, Pennsylvania
| | - Benjamin B Rothrauff
- 1 Center for Cellular and Molecular Engineering, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,4 Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania
| | - Hang Lin
- 1 Center for Cellular and Molecular Engineering, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,4 Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania
| | - Shuting Yu
- 1 Center for Cellular and Molecular Engineering, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,5 School of Medicine, Tsinghua University , Beijing, China
| | - Rocky S Tuan
- 1 Center for Cellular and Molecular Engineering, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.,3 Department of Bioengineering, University of Pittsburgh Swanson School of Engineering , Pittsburgh, Pennsylvania.,4 Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania
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26
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Viganò M, Sansone V, d'Agostino MC, Romeo P, Perucca Orfei C, de Girolamo L. Mesenchymal stem cells as therapeutic target of biophysical stimulation for the treatment of musculoskeletal disorders. J Orthop Surg Res 2016; 11:163. [PMID: 27986082 PMCID: PMC5162101 DOI: 10.1186/s13018-016-0496-5] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/17/2016] [Accepted: 11/28/2016] [Indexed: 12/20/2022] Open
Abstract
BACKGROUND Musculoskeletal disorders are regarded as a major cause of worldwide morbidity and disability, and they result in huge costs for national health care systems. Traditional therapies frequently turned out to be poorly effective in treating bone, cartilage, and tendon disorders or joint degeneration. As a consequence, the development of novel biological therapies that can treat more effectively these conditions should be the highest priority in regenerative medicine. Mesenchymal stem cells (MSCs) represent one of the most promising tools in musculoskeletal tissue regenerative medicine, thanks to their proliferation and differentiation potential and their immunomodulatory and trophic ability. Indeed, MSC-based approaches have been proposed for the treatment of almost all orthopedic conditions, starting from different cell sources, alone or in combination with scaffolds and growth factors, and in one-step or two-step procedures. While all these approaches would require cell harvesting and transplantation, the possibility to stimulate the endogenous MSCs to enhance their tissue homeostasis activity represents a less-invasive and cost-effective therapeutic strategy. Nowadays, the role of tissue-specific resident stem cells as possible therapeutic target in degenerative pathologies is underinvestigated. Biophysical stimulations, and in particular extracorporeal shock waves treatment and pulsed electromagnetic fields, are able to induce proliferation and support differentiation of MSCs from different origins and affect their paracrine production of growth factors and cytokines. SHORT CONCLUSIONS The present review reports the attempts to exploit the resident stem cell potential in musculoskeletal pathologies, highlighting the role of MSCs as therapeutic target of currently applied biophysical treatments.
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Affiliation(s)
- Marco Viganò
- IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.,Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milan, Italy
| | - Valerio Sansone
- IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.,Department of Biomedical Science for Health, University of Milan, Milan, Italy
| | | | - Pietro Romeo
- IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy
| | - Carlotta Perucca Orfei
- IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.,Department of Drug Sciences, University of Pavia, Pavia, Italy
| | - Laura de Girolamo
- IRCCS Galeazzi Orthopaedic Institute, Via R. Galeazzi 4, 20161, Milan, Italy.
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27
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Lui PPY, Wong OT, Lee YW. Transplantation of tendon-derived stem cells pre-treated with connective tissue growth factor and ascorbic acid in vitro promoted better tendon repair in a patellar tendon window injury rat model. Cytotherapy 2016; 18:99-112. [PMID: 26719200 DOI: 10.1016/j.jcyt.2015.10.005] [Citation(s) in RCA: 52] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2015] [Revised: 09/26/2015] [Accepted: 10/05/2015] [Indexed: 12/16/2022]
Abstract
BACKGROUND AIMS Treatment of tendon-derived stem cells (TDSCs) with connective tissue growth factor (CTGF) and ascorbic acid promoted their tenogenic differentiation. We investigated the effects of TDSCs pre-treated with CTGF and ascorbic acid on tendon repair in a patellar tendon window injury rat model. METHODS Green fluorescent protein-TDSCs (GFP-TDSCs) were pre-treated with or without CTGF and ascorbic acid for 2 weeks before transplantation. The patellar tendons of rats were injured and divided into three groups: fibrin glue-only group (control group), untreated and treated TDSC group. The rats were followed up until week 16. RESULTS The treated TDSCs accelerated and enhanced the quality of tendon repair compared with untreated TDSCs up to week 8, which was better than that in the controls up to week 16 as shown by histology, ultrasound imaging and biomechanical test. The fibrils in the treated TDSC group showed better alignment and larger size compared with those in the control group at week 8 (P = 0.004). There was lower risk of ectopic mineralization after transplantation of treated or untreated TDSCs (all P ≤ 0.050). The transplanted cells proliferated and could be detected in the window wound up to weeks 2 to 4 and week 8 for the untreated and treated TDSC groups, respectively. CONCLUSIONS The transplantation of TDSCs promoted tendon repair up to week 16, with CTGF and ascorbic acid pre-treatment showing the best results up to week 8. Pre-treatment of TDSCs with CTGF and ascorbic acid may be used to further enhance the rate and quality of tendon repair after injury.
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Affiliation(s)
| | - On Tik Wong
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, China
| | - Yuk Wa Lee
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, China
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28
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Perucca Orfei C, Lovati AB, Viganò M, Stanco D, Bottagisio M, Di Giancamillo A, Setti S, de Girolamo L. Dose-Related and Time-Dependent Development of Collagenase-Induced Tendinopathy in Rats. PLoS One 2016; 11:e0161590. [PMID: 27548063 PMCID: PMC4993508 DOI: 10.1371/journal.pone.0161590] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2016] [Accepted: 08/08/2016] [Indexed: 12/20/2022] Open
Abstract
Tendinopathy is a big burden in clinics and it represents 45% of musculoskeletal lesions. Despite the relevant social impact, both pathogenesis and development of the tendinopathy are still under-investigated, thus limiting the therapeutic advancement in this field. The purpose of this study was to evaluate the dose-dependent and time-related tissue-level changes occurring in a collagenase-induced tendinopathy in rat Achilles tendons, in order to establish a standardized model for future pre-clinical studies. With this purpose, 40 Sprague Dawley rats were randomly divided into two groups, treated by injecting collagenase type I within the Achilles tendon at 1 mg/mL (low dose) or 3 mg/mL (high dose). Tendon explants were histologically evaluated at 3, 7, 15, 30 and 45 days. Our results revealed that both the collagenase doses induced a disorganization of collagen fibers and increased the number of rounded resident cells. In particular, the high dose treatment determined a greater neovascularization and fatty degeneration with respect to the lower dose. These changes were found to be time-dependent and to resemble the features of human tendinopathy. Indeed, in our series, the acute phase occurred from day 3 to day 15, and then progressed towards the proliferative phase from day 30 to day 45 displaying a degenerative appearance associated with a very precocious and mild remodeling process. The model represents a good balance between similarity with histological features of human tendinopathy and feasibility, in terms of tendon size to create lesions and costs when compared to other animal models. Moreover, this model could contribute to improve the knowledge in this field, and it could be useful to properly design further pre-clinical studies to test innovative treatments for tendinopathy.
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Affiliation(s)
- Carlotta Perucca Orfei
- Orthopaedic Biotechnology Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
- Department of Drug Sciences, University of Pavia, Pavia, Italy
| | - Arianna B. Lovati
- Cell and Tissue Engineering Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
| | - Marco Viganò
- Orthopaedic Biotechnology Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
- Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milan, Italy
| | - Deborah Stanco
- Orthopaedic Biotechnology Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
| | - Marta Bottagisio
- Cell and Tissue Engineering Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
- Department of Veterinary Medicine (DiMeVet), University of Milan, Milan, Italy
| | | | | | - Laura de Girolamo
- Orthopaedic Biotechnology Laboratory, IRCCS Galeazzi Orthopaedic Institute, Milan, Italy
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29
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Adipose-Derived Regenerative Cells Promote Tendon-Bone Healing in a Rabbit Model. Arthroscopy 2016; 32:851-9. [PMID: 26790583 DOI: 10.1016/j.arthro.2015.10.012] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/21/2015] [Revised: 08/15/2015] [Accepted: 10/15/2015] [Indexed: 02/08/2023]
Abstract
PURPOSE To evaluate the therapeutic effect of adipose-derived regenerative cell (ADRC) administration on tendon-bone healing in a rabbit anterior cruciate ligament (ACL) reconstruction model. METHODS ACL reconstruction with semitendinosus tendon autograft was performed in the right knees of adult white rabbits. Eighty rabbits were divided into 2 groups: the treatment group, in which the graft was coated with ADRCs mixed in a fibrin glue carrier during surgery, and the control group, in which the graft was coated with fibrin glue only. At 2, 4, 6, 8, and 12 weeks postoperatively, 8 rabbits were killed in each group. Three were used for histologic evaluation at the tendon-bone interface and 5 for biomechanical examination. RESULTS On histologic analysis, chondroid cells appeared more orderly and more regular in size and shape and Sharpey-like fibers, which connected the tendon graft and bone tissue, appeared earlier in ADRC-treated tissues than in control tissues. On biomechanical analysis, the ultimate failure load in the ADRC-treated group was significantly greater than that in the control group at 2 weeks (29.5 ± 7.2 N v 20.9 ± 2.7 N, P = .016) and 4 weeks (32.3 ± 3.9 N v 22.8 ± 5.4 N, P = .016). Stiffness was significantly higher in the ADRC-treated group than in the control group at 6 weeks (21.7 ± 5.9 N/mm v 12.6 ± 4.9 N/mm, P = .037). Although the ultimate failure load and stiffness of the ADRC-treated limbs were higher than those of the limbs in the control group at 8 and 12 weeks, these differences were not significant. CONCLUSIONS Local administration of ADRCs promoted the early healing process at the tendon-bone junction, both histologically and mechanically, in a rabbit ACL reconstruction model. CLINICAL RELEVANCE ADRCs could be used to enhance graft healing in ACL reconstruction.
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30
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Markers for the identification of tendon-derived stem cells in vitro and tendon stem cells in situ - update and future development. Stem Cell Res Ther 2015; 6:106. [PMID: 26031740 PMCID: PMC4451873 DOI: 10.1186/s13287-015-0097-y] [Citation(s) in RCA: 50] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
The efficacy of tendon-derived stem cells (TDSCs) for the promotion of tendon and tendon-bone junction repair has been reported in animal studies. Modulation of the tendon stem cell niche in vivo has also been reported to influence tendon structure. There is a need to have specific and reliable markers that can define TDSCs in vitro and tendon stem cells in situ for several reasons: to understand the basic biology of TDSCs and their subpopulations in vitro; to understand the identity, niches and functions of tendon/progenitor stem cells in vivo; to meet the governmental regulatory requirements for quality of TDSCs when translating the exciting preclinical findings into clinical trial/practice; and to develop new treatment strategies for mobilizing endogenous stem/progenitor cells in tendon. TDSCs were reported to express the common mesenchymal stem cell (MSC) markers and some embryonic stem cell (ESC) markers, and there were attempts to use these markers to label tendon stem cells in situ. Are these stem cell markers useful for the identification of TDSCs in vitro and tracking of tendon stem cells in situ? This review aims to discuss the values of the panel of MSC, ESC and tendon-related markers for the identification of TDSCs in vitro. Important factors influencing marker expression by TDSCs are discussed. The usefulness and limitations of the panel of MSC, ESC and tendon-related markers for tracking stem cells in tendon, especially tendon stem cells, in situ are then reviewed. Future research directions are proposed.
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