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Wada S, Fujii T, Carballo CB, Suzuki Y, Song Z, Liu Y, Zhang X, Croen BJ, Bhandari R, Deng XH, Rodeo SA. Remodeling Process of the Tendon Graft After Anterior Cruciate Ligament Reconstruction: Comprehensive Analysis With RNA Sequencing in a Murine Model. J Orthop Res 2025; 43:1122-1131. [PMID: 40064586 DOI: 10.1002/jor.26065] [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: 07/21/2024] [Revised: 01/06/2025] [Accepted: 02/18/2025] [Indexed: 05/13/2025]
Abstract
The tendon graft is known to undergo a remodeling process after anterior cruciate ligament (ACL) reconstruction. However, little is known about the transcriptional profile of this process. The aim of the present study is to identify differentially expressed genes inside the remodeling ACL graft in the early phase after ACL reconstruction in our murine model using RNA sequencing (RNAseq). Fifty four male C57BL/6 mice were used in this study. The mice were euthanized at 1, 2, and 4 weeks after surgery and used for histological evaluations and RNAseq of the tendon graft. Histologically, there was a progressive decrease in the tendon-bone interface gap space and increased tissue continuity between the grafted tendon and the bone tunnel over time. At 1 and 2 weeks after surgery, cell increase and loss of collagen fiber organization inside the tendon graft were observed. RNAseq showed that genes related to inflammation, matrix metalloproteinases, bone metabolism, chemokines and signaling pathways were upregulated at 1 and 2 weeks after surgery compared to the control group (p < 0.0001). Our transcriptional profiling data suggests that expression of inflammatory mediators and bone remodeling genes may play an important role in the early events in graft-to-bone healing. Further validation at the protein level is necessary to draw firm conclusions about the role of these mediators in graft remodeling and healing. Understanding the remodeling process of the grafted tendons may lead to the identification of new approaches to improve clinical outcomes after ACL reconstruction.
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Affiliation(s)
- Susumu Wada
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
- Department of Orthopaedic Surgery, St. Luke's International Hospital, Tokyo, Japan
| | - Takayuki Fujii
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
- Department of Orthopaedic Surgery, Kyoto University Graduate School of Medicine, Kyoto, Japan
| | - Camila B Carballo
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Yuki Suzuki
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
- Department of Orthopaedic Surgery, Faculty of Medicine and Graduate School of Medicine, Hokkaido University, Sapporo, Japan
| | - Zhe Song
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Yake Liu
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Xueying Zhang
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Brett J Croen
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Reyna Bhandari
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Xiang-Hua Deng
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
| | - Scott A Rodeo
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York, USA
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Fang F, Casserly M, Robbins J, Thomopoulos S. Hedgehog signaling directs cell differentiation and plays a critical role in tendon enthesis healing. NPJ Regen Med 2025; 10:3. [PMID: 39833191 PMCID: PMC11747568 DOI: 10.1038/s41536-025-00392-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2024] [Accepted: 01/10/2025] [Indexed: 01/22/2025] Open
Abstract
A high prevalence of rotator cuff tears presents a major clinical challenge. A better understanding of the molecular mechanisms underlying enthesis development and healing is needed for developing treatments. We recently identified hedgehog (Hh)-lineage cells critical for enthesis development and repair. This study revealed cell-cell communication within the Hh-lineage cell population. To further characterize the role of Hh signaling, we used mouse models to activate and inactivate the Hh pathway in enthesis progenitors. Activation of Hh target genes during enthesis development increased its mineralization and mechanical properties. Activation of Hh signaling at the injured mature enthesis promoted fibrocartilage formation, enhanced mineralization, and increased expression of chondrogenic and osteogenic markers, which implies that Hh signaling drives cell differentiation to regenerate the damaged enthesis. Conversely, deletion of Hh target genes impaired enthesis healing. In summary, this study revealed a new strategy for enthesis repair via activation of Hh signaling in endogenous cells.
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Affiliation(s)
- Fei Fang
- Leni and Peter W. May Department of Orthopaedics, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
| | - Matthew Casserly
- Leni and Peter W. May Department of Orthopaedics, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Julia Robbins
- Leni and Peter W. May Department of Orthopaedics, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Stavros Thomopoulos
- Department of Orthopedic Surgery, Columbia University, New York, NY, USA
- Department of Biomedical Engineering, Columbia University, New York, NY, USA
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Dyment NA, Kamalitdinov TB, Kuntz AF. The 2024 Kappa Delta Young Investigator Award: Leveraging Insights From Development to Improve Adult Repair: Hedgehog Signaling as a Master Regulator of Enthesis Fibrocartilage Formation. J Am Acad Orthop Surg 2024; 32:1074-1086. [PMID: 39589737 PMCID: PMC11753257 DOI: 10.5435/jaaos-d-24-00996] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/30/2024] [Accepted: 08/31/2024] [Indexed: 11/27/2024] Open
Abstract
The work in this article summarizes findings from our group on key biochemical cues that govern the formation and repair of tendons and ligaments. Specifically, we summarize the journey that started with a serendipitous discovery that is now being translated into novel therapies to improve tendon-to-bone repair outcomes. This journey began with the discovery that the Hedgehog (Hh) signaling pathway was expressed within the enthesis during development and that its primary role was to promote fibrocartilage production and maturation. Next, we developed an anterior cruciate ligament reconstruction model in novel transgenic mice that allowed us to discover that the Hh pathway promotes fibrocartilaginous tendon-to-bone attachments during the integration process. In addition, we established that the coordinated stages of zonal tendon-to-bone integration after anterior cruciate ligament reconstruction were comparable with the stages required for enthesis formation during development. Now that we have demonstrated that the Hh pathway is a potent therapeutic target, we are currently advancing these findings to develop drug delivery systems to improve tendon-to-bone repair. Ultimately, our group aims to establish key mechanisms that govern tendon and ligament formation that can be leveraged for novel regenerative therapies to improve clinical care.
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Affiliation(s)
- Nathaniel A Dyment
- From the Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA (Dr. Dyment, Dr. Kamalitdinov, and Dr. Kuntz), and the Department of Bioengineering, University of Pennsylvania, Philadelphia, PA (Dr. Dyment and Dr. Kamalitdinov)
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Little D, Amadio PC, Awad HA, Cone SG, Dyment NA, Fisher MB, Huang AH, Koch DW, Kuntz AF, Madi R, McGilvray K, Schnabel LV, Shetye SS, Thomopoulos S, Zhao C, Soslowsky LJ. Preclinical tendon and ligament models: Beyond the 3Rs (replacement, reduction, and refinement) to 5W1H (why, who, what, where, when, how). J Orthop Res 2023; 41:2133-2162. [PMID: 37573480 PMCID: PMC10561191 DOI: 10.1002/jor.25678] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/08/2023] [Revised: 07/21/2023] [Accepted: 08/02/2023] [Indexed: 08/14/2023]
Abstract
Several tendon and ligament animal models were presented at the 2022 Orthopaedic Research Society Tendon Section Conference held at the University of Pennsylvania, May 5 to 7, 2022. A key objective of the breakout sessions at this meeting was to develop guidelines for the field, including for preclinical tendon and ligament animal models. This review summarizes the perspectives of experts for eight surgical small and large animal models of rotator cuff tear, flexor tendon transection, anterior cruciate ligament tear, and Achilles tendon injury using the framework: "Why, Who, What, Where, When, and How" (5W1H). A notable conclusion is that the perfect tendon model does not exist; there is no single gold standard animal model that represents the totality of tendon and ligament disease. Each model has advantages and disadvantages and should be carefully considered in light of the specific research question. There are also circumstances when an animal model is not the best approach. The wide variety of tendon and ligament pathologies necessitates choices between small and large animal models, different anatomic sites, and a range of factors associated with each model during the planning phase. Attendees agreed on some guiding principles including: providing clear justification for the model selected, providing animal model details at publication, encouraging sharing of protocols and expertise, improving training of research personnel, and considering greater collaboration with veterinarians. A clear path for translating from animal models to clinical practice was also considered as a critical next step for accelerating progress in the tendon and ligament field.
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Affiliation(s)
- Dianne Little
- Department of Basic Medical Sciences, The Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, USA
| | - Peter C Amadio
- Department of Orthopaedic Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Hani A Awad
- Department of Orthopaedics, Department of Biomedical Engineering, The Center for Musculoskeletal Research, University of Rochester, Rochester, New York, USA
| | - Stephanie G Cone
- Department of Biomedical Engineering, University of Delaware, Newark, Delaware, USA
| | - Nathaniel A Dyment
- McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Matthew B Fisher
- Joint Department of Biomedical Engineering, College of Engineering, North Carolina State University-University of North Carolina at Chapel Hill, Raleigh, North Carolina, USA
| | - Alice H Huang
- Department of Orthopedic Surgery, Department of Biomedical Engineering, Columbia University, New York, New York, USA
| | - Drew W Koch
- Department of Clinical Sciences, College of Veterinary Medicine, and Comparative Medicine Institute, North Carolina State University, Raleigh, North Carolina, USA
| | - Andrew F Kuntz
- McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Rashad Madi
- McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Kirk McGilvray
- Department of Mechanical Engineering, School of Biomedical Engineering, Colorado State University, Fort Collins, Colorado, USA
| | - Lauren V Schnabel
- Department of Clinical Sciences, College of Veterinary Medicine, and Comparative Medicine Institute, North Carolina State University, Raleigh, North Carolina, USA
| | - Snehal S Shetye
- McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, Pennsylvania, USA
| | - Stavros Thomopoulos
- Department of Orthopedic Surgery, Department of Biomedical Engineering, Columbia University, New York, New York, USA
| | - Chunfeng Zhao
- Department of Orthopaedic Surgery, Mayo Clinic, Rochester, Minnesota, USA
| | - Louis J Soslowsky
- McKay Orthopaedic Research Laboratory, University of Pennsylvania, Philadelphia, Pennsylvania, USA
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Kamalitdinov TB, Fujino K, Keith Lang S, Jiang X, Madi R, Evans MK, Zgonis MH, Kuntz AF, Dyment NA. Targeting the hedgehog signaling pathway to improve tendon-to-bone integration. Osteoarthritis Cartilage 2023; 31:1202-1213. [PMID: 37146960 PMCID: PMC10524548 DOI: 10.1016/j.joca.2023.04.013] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/21/2022] [Revised: 04/24/2023] [Accepted: 04/29/2023] [Indexed: 05/07/2023]
Abstract
OBJECTIVE While the role of hedgehog (Hh) signaling in promoting zonal fibrocartilage production during development is well-established, whether this pathway can be leveraged to improve tendon-to-bone repair in adults is unknown. Our objective was to genetically and pharmacologically stimulate the Hh pathway in cells that give rise to zonal fibrocartilaginous attachments to promote tendon-to-bone integration. DESIGN Hh signaling was stimulated genetically via constitutive Smo (SmoM2 construct) activation of bone marrow stromal cells or pharmacologically via systemic agonist delivery to mice following anterior cruciate ligament reconstruction (ACLR). To assess tunnel integration, we measured mineralized fibrocartilage (MFC) formation in these mice 28 days post-surgery and performed tunnel pullout testing. RESULTS Hh pathway-related genes increased in cells forming the zonal attachments in wild-type mice. Both genetic and pharmacologic stimulation of the Hh pathway increased MFC formation and integration strength 28 days post-surgery. We next conducted studies to define the role of Hh in specific stages of the tunnel integration process. We found Hh agonist treatment increased the proliferation of the progenitor pool in the first week post-surgery. Additionally, genetic stimulation led to continued MFC production in the later stages of the integration process. These results indicate that Hh signaling plays an important biphasic role in cell proliferation and differentiation towards fibrochondrocytes following ACLR. CONCLUSION This study reveals a biphasic role for Hh signaling during the tendon-to-bone integration process after ACLR. In addition, the Hh pathway is a promising therapeutic target to improve tendon-to-bone repair outcomes.
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Affiliation(s)
- Timur B Kamalitdinov
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA; Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA
| | - Keitaro Fujino
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA; Osaka Medical and Pharmaceutical University, Takatsuki, Osaka Prefecture, Japan
| | - Sinaia Keith Lang
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA; Department of Biology, University of Pennsylvania, Philadelphia, PA, USA
| | - Xi Jiang
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Rashad Madi
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Mary Kate Evans
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA; Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA
| | - Miltiadis H Zgonis
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Andrew F Kuntz
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Nathaniel A Dyment
- Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA; Department of Bioengineering, University of Pennsylvania, Philadelphia, PA, USA.
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Li FQ, Chen WB, Luo ZW, Chen YS, Sun YY, Su XP, Sun JM, Chen SY. Bone marrow mesenchymal stem cell-derived exosomal microRNAs target PI3K/Akt signaling pathway to promote the activation of fibroblasts. World J Stem Cells 2023; 15:248-267. [PMID: 37181002 PMCID: PMC10173806 DOI: 10.4252/wjsc.v15.i4.248] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Revised: 01/19/2023] [Accepted: 03/23/2023] [Indexed: 04/26/2023] Open
Abstract
BACKGROUND Fibroblast plays a major role in tendon-bone healing. Exosomes derived from bone marrow mesenchymal stem cells (BMSCs) can activate fibroblasts and promote tendon-bone healing via the contained microRNAs (miRNAs). However, the underlying mechanism is not comprehensively understood. Herein, this study aimed to identify overlapped BMSC-derived exosomal miRNAs in three GSE datasets, and to verify their effects as well as mechanisms on fibroblasts. AIM To identify overlapped BMSC-derived exosomal miRNAs in three GSE datasets and verify their effects as well as mechanisms on fibroblasts. METHODS BMSC-derived exosomal miRNAs data (GSE71241, GSE153752, and GSE85341) were downloaded from the Gene Expression Omnibus (GEO) database. The candidate miRNAs were obtained by the intersection of three data sets. TargetScan was used to predict potential target genes for the candidate miRNAs. Functional and pathway analyses were conducted using the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, respectively, by processing data with the Metascape. Highly interconnected genes in the protein-protein interaction (PPI) network were analyzed using Cytoscape software. Bromodeoxyuridine, wound healing assay, collagen contraction assay and the expression of COL I and α-smooth muscle actin positive were applied to investigate the cell proliferation, migration and collagen synthesis. Quantitative real-time reverse transcription polymerase chain reaction was applied to determine the cell fibroblastic, tenogenic, and chondrogenic potential. RESULTS Bioinformatics analyses found two BMSC-derived exosomal miRNAs, has-miR-144-3p and has-miR-23b-3p, were overlapped in three GSE datasets. PPI network analysis and functional enrichment analyses in the GO and KEGG databases indicated that both miRNAs regulated the PI3K/Akt signaling pathway by targeting phosphatase and tensin homolog (PTEN). In vitro experiments confirmed that miR-144-3p and miR-23b-3p stimulated proliferation, migration and collagen synthesis of NIH3T3 fibroblasts. Interfering with PTEN affected the phosphorylation of Akt and thus activated fibroblasts. Inhibition of PTEN also promoted the fibroblastic, tenogenic, and chondrogenic potential of NIH3T3 fibroblasts. CONCLUSION BMSC-derived exosomes promote fibroblast activation possibly through the PTEN and PI3K/Akt signaling pathways, which may serve as potential targets to further promote tendon-bone healing.
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Affiliation(s)
- Fang-Qi Li
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China
| | - Wen-Bo Chen
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China
| | - Zhi-Wen Luo
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China
| | - Yi-Sheng Chen
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China
| | - Ya-Ying Sun
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China
| | - Xiao-Ping Su
- Guangxi Key Laboratory of Oral and Maxillofacial Rehabilitation and Reconstruction, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
| | - Jun-Ming Sun
- Laboratory Animal Center, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
| | - Shi-Yi Chen
- Department of Sports Medicine, Huashan Hospital, Fudan University, Shanghai 200040, China.
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Fu S, Lan Y, Wang G, Bao D, Qin B, Zheng Q, Liu H, Wong VKW. External stimulation: A potential therapeutic strategy for tendon-bone healing. Front Bioeng Biotechnol 2023; 11:1150290. [PMID: 37064229 PMCID: PMC10102526 DOI: 10.3389/fbioe.2023.1150290] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2023] [Accepted: 03/23/2023] [Indexed: 04/03/2023] Open
Abstract
Injuries at the tendon-bone interface are very common in the field of sports medicine, and healing at the tendon-bone interface is complex. Injuries to the tendon-bone interface can seriously affect a patient’s quality of life, so it is essential to restore stability and promote healing of the tendon-bone interface. In addition to surgical treatment, the healing of tendons and bones can also be properly combined with extracorporeal stimulation therapy during the recovery process. In this review, we discuss the effects of extracorporeal shock waves (ESWs), low-intensity pulsed ultrasound (LIPUS), and mechanical stress on tendon-bone healing, focusing on the possible mechanisms of action of mechanical stress on tendon-bone healing in terms of transcription factors and biomolecules. The aim is to provide possible therapeutic approaches for subsequent clinical treatment.
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Affiliation(s)
- Shijie Fu
- Faculty of Chinese Medicine, Macau University of Science and Technology, Taipa, Macao SAR, China
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
- Dr. Neher’s Biophysics Laboratory for Innovative Drug Discovery, State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Taipa, Macao SAR, China
| | - Yujian Lan
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
| | - Guoyou Wang
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
| | - Dingsu Bao
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
| | - Bo Qin
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
| | - Qiu Zheng
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
| | - Huan Liu
- Department of Orthopedics, Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China
- *Correspondence: Huan Liu, ; Vincent Kam Wai Wong,
| | - Vincent Kam Wai Wong
- Dr. Neher’s Biophysics Laboratory for Innovative Drug Discovery, State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Taipa, Macao SAR, China
- *Correspondence: Huan Liu, ; Vincent Kam Wai Wong,
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Tian B, Zhang M, Kang X. Strategies to promote tendon-bone healing after anterior cruciate ligament reconstruction: Present and future. Front Bioeng Biotechnol 2023; 11:1104214. [PMID: 36994361 PMCID: PMC10040767 DOI: 10.3389/fbioe.2023.1104214] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2022] [Accepted: 03/02/2023] [Indexed: 03/16/2023] Open
Abstract
At present, anterior cruciate ligament (ACL) reconstruction still has a high failure rate. Tendon graft and bone tunnel surface angiogenesis and bony ingrowth are the main physiological processes of tendon-bone healing, and also the main reasons for the postoperative efficacy of ACL reconstruction. Poor tendon-bone healing has been also identified as one of the main causes of unsatisfactory treatment outcomes. The physiological process of tendon-bone healing is complicated because the tendon-bone junction requires the organic fusion of the tendon graft with the bone tissue. The failure of the operation is often caused by tendon dislocation or scar healing. Therefore, it is important to study the possible risk factors for tendon-bone healing and strategies to promote it. This review comprehensively analyzed the risk factors contributing to tendon-bone healing failure after ACL reconstruction. Additionally, we discuss the current strategies used to promote tendon-bone healing following ACL reconstruction.
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Liu Y, Liu S, Song Z, Chen D, Album Z, Green S, Deng X, Rodeo SA. GLI1 Deficiency Impairs the Tendon-Bone Healing after Anterior Cruciate Ligament Reconstruction: In Vivo Study Using Gli1-Transgenic Mice. J Clin Med 2023; 12:jcm12030999. [PMID: 36769647 PMCID: PMC9917856 DOI: 10.3390/jcm12030999] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2022] [Revised: 12/31/2022] [Accepted: 01/05/2023] [Indexed: 01/31/2023] Open
Abstract
Hedgehog (Hh) signaling plays a fundamental role in the enthesis formation process and GLI-Kruppel family member GLI1 (Gli1) is a key downstream mediator. However, the role of Gli1 in tendon-bone healing after anterior cruciate ligament reconstruction (ACLR) is unknown. To evaluate the tendon-bone healing after ACLR in Gli1LacZ/LacZ (GLI1-NULL) mice, and compare Gli1LacZ/WT (GLI1-HET) and Gli1WT/WT wild type (WT) mice, a total of 45 mice, 15 mice each of GLI1-NULL, GLI1-HET and WT were used in this study. All mice underwent microsurgical ACLR at 12 weeks of age. Mice were euthanized at 4 weeks after surgery and were used for biomechanical testing, histological evaluation, and micro-CT analysis. The GLI1-NULL group had significantly lower biomechanical failure force, poorer histological healing, and lower BV/TV when compared with the WT and GLI1-HET groups. These significant differences were only observed at the femoral tunnel. Immunohistology staining showed positive expression of Indian hedgehog (IHH) and Patched 1(PTCH1) in all three groups, which indicated the activation of the Hh signal pathway. The GLI1 was negative in the GLI1-NULL group, validating the absence of GLI1 protein in these mice. These results proved that activation of the Hh signaling pathway occurs during ACL graft healing, and the function of Gli1 was necessary for tendon-bone healing. Healing in the femoral tunnel is more obviously impaired by Gli1 deficiency. Our findings provide further insight into the molecular mechanism of tendon-bone healing and suggest that Gli1 might represent a novel therapeutic target to improve tendon-bone healing after ACLR.
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Affiliation(s)
- Yake Liu
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
- Department of Orthopedic, Affiliated Hospital of Nantong University, Nantong 226001, China
| | - Shaohua Liu
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Zhe Song
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Daoyun Chen
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Zoe Album
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Samuel Green
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Xianghua Deng
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
| | - Scott A. Rodeo
- Laboratory for Joint Tissue Repair and Regeneration, Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY 10021, USA
- Correspondence:
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Cardona-Ramirez S, Cook JL, Stoker AM, Ma R. Small laboratory animal models of anterior cruciate ligament reconstruction. J Orthop Res 2022; 40:1967-1980. [PMID: 35689508 DOI: 10.1002/jor.25395] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/09/2021] [Revised: 04/19/2022] [Accepted: 06/08/2022] [Indexed: 02/04/2023]
Abstract
Anterior cruciate ligament (ACL) injuries are common knee ligament injuries. While generally successful, ACL reconstruction that uses a tendon graft to stabilize the knee is still associated with a notable percentage of failures and long-term morbidities. Preclinical research that uses small laboratory species (i.e., mice, rats, and rabbits) to model ACL reconstruction are important to evaluate factors that can impact graft incorporation or posttraumatic osteoarthritis after ACL reconstruction. Small animal ACL reconstruction models are also used for proof-of-concept studies for the development of emerging biological strategies aimed at improving ACL reconstruction healing. The objective of this review is to provide an overview on the use of common small animal laboratory species to model ACL reconstruction. The review includes a discussion on comparative knee anatomy, technical considerations including types of tendon grafts employed amongst the small laboratory species (i.e., mice, rats, and rabbits), and common laboratory evaluative methods used to study healing and outcomes after ACL reconstruction in small laboratory animals. The review will also highlight common research questions addressed with small animal models of ACL reconstruction.
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Affiliation(s)
- Sebastian Cardona-Ramirez
- Thompson Laboratory for Regenerative Orthopaedics, Missouri Orthopaedic Institute, University of Missouri, Columbia, Missouri, USA
| | - James L Cook
- Thompson Laboratory for Regenerative Orthopaedics, Missouri Orthopaedic Institute, University of Missouri, Columbia, Missouri, USA
| | - Aaron M Stoker
- Thompson Laboratory for Regenerative Orthopaedics, Missouri Orthopaedic Institute, University of Missouri, Columbia, Missouri, USA
| | - Richard Ma
- Thompson Laboratory for Regenerative Orthopaedics, Missouri Orthopaedic Institute, University of Missouri, Columbia, Missouri, USA
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Geng R, Lin Y, Ji M, Chang Q, Li Z, Xu L, Zhang W, Lu J. MFG-E8 promotes tendon-bone healing by regualting macrophage efferocytosis and M2 polarization after anterior cruciate ligament reconstruction. J Orthop Translat 2022; 34:11-21. [PMID: 35615640 PMCID: PMC9109120 DOI: 10.1016/j.jot.2022.04.002] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/12/2022] [Revised: 03/18/2022] [Accepted: 04/15/2022] [Indexed: 11/24/2022] Open
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12
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Liu Y, Deng XH, Zhang X, Cong T, Chen D, Hall AJ, Ying L, Rodeo SA. The Role of Indian Hedgehog Signaling in Tendon Response to Subacromial Impingement: Evaluation Using a Mouse Model. Am J Sports Med 2022; 50:362-370. [PMID: 34904906 DOI: 10.1177/03635465211062244] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
BACKGROUND The underlying cellular and molecular mechanisms involved in the development of tendinopathy due to subacromial supraspinatus tendon (SST) impingement and the response to subsequent removal of impingement remain unknown. PURPOSE To investigate the involvement of Indian hedgehog (IHH) signaling in the development of SST tendinopathy and the subsequent healing process after the relief of subacromial impingement in a novel mouse shoulder impingement model. STUDY DESIGN Controlled laboratory study. METHODS A total of 48 male wild-type C57BL/6 mice were used in this study. Supraspinatus tendinopathy was induced by inserting a microsurgical clip into the subacromial space bilaterally. Eleven mice were sacrificed at 4 weeks after surgery to establish impingement baseline; 24 mice underwent clip removal at 4 weeks after surgery and then were euthanized at 2 or 4 weeks after clip removal. Thirteen mice without surgical intervention were utilized as the control group. All SSTs were evaluated with biomechanical testing; quantitative histomorphometry after staining with hematoxylin and eosin, Alcian blue, and picrosirius red; and immunohistochemical staining (factor VIII, IHH, Patched1 [PTCH1], and glioma-associated oncogene homolog 1 [GLI1]). RESULTS The mean failure force and stiffness in the 4-week impingement group decreased significantly compared with the control group (P < .001) and gradually increased at 2 and 4 weeks after clip removal. Histological analysis demonstrated increased cellularity and disorganized collagen fibers in the SST, with higher modified Bonar scores at 4 weeks, followed by gradual improvement after clip removal. The IHH-positive area and PTCH1- and GLI1-positive cell percentages significantly increased after 4 weeks of clip impingement (20.64% vs 2.06%, P < .001; 53.9% vs 28.03%, P = .016; and 30% vs 12.19%, P = .036, respectively) and continuously increased after clip removal. CONCLUSION The authors' findings suggest that the hedgehog signaling pathway and its downstream signaling mediator and target GLI1 may play a role in the development and healing process of rotator cuff tendinopathy due to extrinsic rotator cuff impingement. CLINICAL RELEVANCE This study suggests the potential for the hedgehog pathway, together with its downstream targets, as candidates for further study as potential therapeutic targets in the treatment of supraspinatus tendinopathy.
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Affiliation(s)
- Yulei Liu
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA.,Institute of Sports Medicine, Peking University Third Hospital, Beijing, China
| | - Xiang-Hua Deng
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Xueying Zhang
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Ting Cong
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Daoyun Chen
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Arielle Jordan Hall
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Liang Ying
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
| | - Scott A Rodeo
- Orthopedic Soft Tissue Research Program, Hospital for Special Surgery, New York, New York, USA
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Remnants-preserving ACL reconstruction using direct tendinous graft fixation: a new rat model. J Orthop Surg Res 2022; 17:7. [PMID: 34986843 PMCID: PMC8729105 DOI: 10.1186/s13018-021-02890-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/06/2021] [Accepted: 12/15/2021] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Anterior cruciate ligament (ACL) repair techniques are new emerging strategies prevailing, in selected cases, over standard reconstruction of the ACL with excision of its remnants. Mid-substance ACL tears represent a challenge for ACL repair techniques, and remnants-preserving ACL reconstruction (rp-ACLR) using an autograft remains the recommended treatment in this situation. However, morbidity associated with the autograft harvesting prompts the need for alternative surgical strategies based on the use of synthetic scaffolds. Relevant small animal models of mid-substance tears with ACL remnants preservation and reconstruction are necessary to establish the preliminary proof of concept of these new strategies. METHODS A rat model of rp-ACLR using a tendinous autograft after complete mid-substance ACL transection was established. Twelve weeks following surgery, clinical outcomes and knee joints were assessed through visual gait analysis, Lachman tests, thigh perimeter measurements, magnetic resonance imaging, micro-computed tomography, and histology, to evaluate the morbidity of the procedure, accuracy of bone tunnel positioning, ACL remnants fate, osteoarthritis, and autograft bony integration. Results were compared with those obtained with isolated ACL transection without reconstruction and to right non-operated knees. RESULTS AND DISCUSSION Most operated animals were weight-bearing the day following surgery, and no adverse inflammatory reaction has been observed for the whole duration of the study. Autograft fixation with cortical screws provided effective graft anchorage until sacrifice. Healing of the transected ACL was not observed in the animals in which no graft reconstruction was performed. rp-ACLR was associated with a reduced degeneration of the ACL remnants (p = 0.004) and cartilages (p = 0.0437). Joint effusion and synovitis were significantly lower in the reconstructed group compared to the transected ACL group (p = 0.004). Most of the bone tunnel apertures were anatomically positioned in the coronal and/or sagittal plane. The most deviated bone tunnel apertures were the tibial ones, located in median less than 1 mm posteriorly to anatomical ACL footprint center. CONCLUSION This study presents a cost-effective, new relevant and objective rat model associated with low morbidity for the preliminary study of bio-implantable materials designed for remnants-preserving ACL surgery after mid-substance ACL tear.
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Fang F, Sup M, Luzzi A, Ferrer X, Thomopoulos S. Hedgehog signaling underlying tendon and enthesis development and pathology. Matrix Biol 2022; 105:87-103. [PMID: 34954379 PMCID: PMC8821161 DOI: 10.1016/j.matbio.2021.12.001] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Revised: 12/18/2021] [Accepted: 12/19/2021] [Indexed: 02/08/2023]
Abstract
Hedgehog (Hh) signaling has been widely acknowledged to play essential roles in many developmental processes, including endochondral ossification and growth plate maintenance. Furthermore, a rising number of studies have shown that Hh signaling is necessary for tendon enthesis development. Specifically, the well-tuned regulation of Hh signaling during development drives the formation of a mineral gradient across the tendon enthesis fibrocartilage. However, aberrant Hh signaling can also lead to pathologic heterotopic ossification in tendon or osteophyte formation at the enthesis. Therefore, the therapeutic potential of Hh signaling modulation for treating tendon and enthesis diseases remains uncertain. For example, increased Hh signaling may enhance tendon-to-bone healing by promoting the formation of mineralized fibrocartilage at the healing interface, but pathologic heterotopic ossification may also be triggered in the adjacent tendon. Further work is needed to elucidate the distinct functions of Hh signaling in the tendon and enthesis to support the development of therapies that target the pathway.
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Affiliation(s)
- Fei Fang
- Department of Orthopedic Surgery, Columbia University, Black Building, Room 1408, 650W 168 ST, New York, NY 10032-3702, United States
| | - McKenzie Sup
- Department of Biomedical Engineering, Columbia University, New York, NY, United States
| | - Andrew Luzzi
- Department of Orthopedic Surgery, Columbia University, Black Building, Room 1408, 650W 168 ST, New York, NY 10032-3702, United States
| | - Xavier Ferrer
- Department of Orthopedic Surgery, Columbia University, Black Building, Room 1408, 650W 168 ST, New York, NY 10032-3702, United States
| | - Stavros Thomopoulos
- Department of Orthopedic Surgery, Columbia University, Black Building, Room 1408, 650W 168 ST, New York, NY 10032-3702, United States; Department of Biomedical Engineering, Columbia University, New York, NY, United States.
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15
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Tendon and multiomics: advantages, advances, and opportunities. NPJ Regen Med 2021; 6:61. [PMID: 34599188 PMCID: PMC8486786 DOI: 10.1038/s41536-021-00168-6] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2020] [Accepted: 09/01/2021] [Indexed: 02/08/2023] Open
Abstract
Tendons heal by fibrosis, which hinders function and increases re-injury risk. Yet the biology that leads to degeneration and regeneration of tendons is not completely understood. Improved understanding of the metabolic nuances that cause diverse outcomes in tendinopathies is required to solve these problems. 'Omics methods are increasingly used to characterize phenotypes in tissues. Multiomics integrates 'omic datasets to identify coherent relationships and provide insight into differences in molecular and metabolic pathways between anatomic locations, and disease stages. This work reviews the current literature pertaining to multiomics in tendon and the potential of these platforms to improve tendon regeneration. We assessed the literature and identified areas where 'omics platforms contribute to the field: (1) Tendon biology where their hierarchical complexity and demographic factors are studied. (2) Tendon degeneration and healing, where comparisons across tendon pathologies are analyzed. (3) The in vitro engineered tendon phenotype, where we compare the engineered phenotype to relevant native tissues. (4) Finally, we review regenerative and therapeutic approaches. We identified gaps in current knowledge and opportunities for future study: (1) The need to increase the diversity of human subjects and cell sources. (2) Opportunities to improve understanding of tendon heterogeneity. (3) The need to use these improvements to inform new engineered and regenerative therapeutic approaches. (4) The need to increase understanding of the development of tendon pathology. Together, the expanding use of various 'omics platforms and data analysis resulting from these platforms could substantially contribute to major advances in the tendon tissue engineering and regenerative medicine field.
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Cardona-Ramirez S, Stoker AM, Cook JL, Ma R. Fibroblasts From Common Anterior Cruciate Ligament Tendon Grafts Exhibit Different Biologic Responses to Mechanical Strain. Am J Sports Med 2021; 49:215-225. [PMID: 33259232 DOI: 10.1177/0363546520971852] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
BACKGROUND Different tendons are chosen for anterior cruciate ligament (ACL) reconstruction based on perceived advantages and disadvantages, yet there is a relative paucity of information regarding biologic responsiveness of commonly used tendon grafts to mechanical strain. PURPOSE To evaluate the in vitro responses of graft fibroblasts derived from tendons used for ACL reconstruction to clinically relevant strain levels. STUDY DESIGN Controlled laboratory study. METHODS Twelve quadriceps tendons (QTs), 12 patellar tendons (PTs), and 9 hamstring tendons (HTs) were harvested from skeletally mature dogs (n = 16). Tendon fibroblasts were isolated and seeded onto BioFlex plates (1 × 105 cells/well). Cells were subjected to 3 strain conditions (stress deprivation, 0%; physiologic, 4%; high, 10%) for 5 days. Media were collected for proinflammatory and metabolic assays. RNA was extracted for gene expression analysis using real-time reverse transcription polymerase chain reaction. RESULTS Stress deprivation elicited significantly higher metabolic activity from HT and PT cells than from QT cells (P < .001 and P = .001, respectively). There were no differences in metabolic activity among all 3 graft fibroblasts at physiologic and high strain. COL-1 expression was significantly higher in PT versus HT during physiologic strain (P = .007). No significant differences with COL-3 expression were seen. TIMP-1 (P = .01) expression was higher in PT versus HT under physiologic strain. Scleraxis expression was higher in PT versus HT (P = .007) under physiologic strain. A strain-dependent increase in PGE2 levels occurred for all grafts. At physiologic strain conditions, HT produced significantly higher levels of PGE2 versus QT (P < .001) and PT (P = .005). CONCLUSION Fibroblasts from common ACL graft tissues exhibited different metabolic responses to mechanical strain. On the basis of these data, we conclude that early production of extracellular matrix and proinflammatory responses from ACL grafts are dependent on mechanical loading and graft source. CLINICAL RELEVANCE Graft-specific differences in ACL reconstruction outcomes are known to exist. Our results suggest that there are differences in the biologic responsiveness of cells from the tendon grafts used in ACL reconstruction, which are dependent on strain levels and graft source. The biologic properties of the tissue used for ACL reconstruction should be considered when selecting graft source.
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Affiliation(s)
- Sebastian Cardona-Ramirez
- Department of Orthopaedic Surgery, Thompson Laboratory for Regenerative Orthopaedics, University of Missouri, Columbia, Missouri, USA
| | - Aaron M Stoker
- Department of Orthopaedic Surgery, Thompson Laboratory for Regenerative Orthopaedics, University of Missouri, Columbia, Missouri, USA
| | - James L Cook
- Department of Orthopaedic Surgery, Thompson Laboratory for Regenerative Orthopaedics, University of Missouri, Columbia, Missouri, USA
| | - Richard Ma
- Department of Orthopaedic Surgery, Thompson Laboratory for Regenerative Orthopaedics, University of Missouri, Columbia, Missouri, USA
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He X, Li Y, Guo J, Xu J, Zu H, Huang L, Tim-Yun Ong M, Shu-Hang Yung P, Qin L. Biomaterials developed for facilitating healing outcome after anterior cruciate ligament reconstruction: Efficacy, surgical protocols, and assessments using preclinical animal models. Biomaterials 2020; 269:120625. [PMID: 33395579 DOI: 10.1016/j.biomaterials.2020.120625] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2020] [Revised: 12/16/2020] [Accepted: 12/18/2020] [Indexed: 12/17/2022]
Abstract
Anterior cruciate ligament (ACL) reconstruction is the recommended treatment for ACL tear in the American Academy of Orthopaedic Surgeons (AAOS) guideline. However, not a small number of cases failed because of the tunnel bone resorption, unsatisfactory bone-tendon integration, and graft degeneration. The biomaterials developed and designed for improving ACL reconstruction have been investigated for decades. According to the Food and Drug Administration (FDA) and the International Organization for Standardization (ISO) regulations, animal studies should be performed to prove the safety and bioeffect of materials before clinical trials. In this review, we first evaluated available biomaterials that can enhance the healing outcome after ACL reconstruction in animals and then discussed the animal models and assessments for testing applied materials. Furthermore, we identified the relevance and knowledge gaps between animal experimental studies and clinical expectations. Critical analyses and suggestions for future research were also provided to design the animal study connecting basic research and requirements for future clinical translation.
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Affiliation(s)
- Xuan He
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Ye Li
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Jiaxin Guo
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Jiankun Xu
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Haiyue Zu
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Le Huang
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Michael Tim-Yun Ong
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Patrick Shu-Hang Yung
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region
| | - Ling Qin
- Musculoskeletal Research Laboratory of Department of Orthopaedics & Traumatology and Innovative Orthopaedic Biomaterial and Drug Translational Research Laboratory of Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong Special Administrative Region.
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Biomechanical, histologic, and molecular characteristics of graft-tunnel healing in a murine modified ACL reconstruction model. J Orthop Translat 2020; 24:103-111. [PMID: 32775202 PMCID: PMC7390781 DOI: 10.1016/j.jot.2020.05.004] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/05/2019] [Revised: 05/15/2020] [Accepted: 05/20/2020] [Indexed: 02/07/2023] Open
Abstract
Purpose The purpose of our study was to introduce and validate a metal-free, reproducible and reliable mouse model of anterior cruciate ligament (ACL) reconstruction (ACLR) surgery as an effective tool for a better understanding of molecular mechanisms of graft-tunnel healing after ACLR. Methods A total of 150 C57BL/6 mice were randomly allocated into five Groups: Group 1 (mice with intact ACL), Group 2–4 (mice underwent modified ACLR surgery and sacrificed 1-, 2-, and 4-weeks after surgery), and Group 5 (mice underwent unmodified ACLR surgery and sacrificed 4 weeks after surgery). Micro-computed tomography (CT), biomechanical histological as well as immunohistochemical (IHC) analyses were performed to characterize the modified ACLR. Results Micro-CT analysis demonstrated there is a non-significant increase in BV/TV and BMD of the bone tunnel during the tendon-to-bone healing following ACLR. Biomechanical tests showed that the mean load-to-failure forces of Group 3 and 4 are equal to 31.7% and 46.0% of that in Group 1, while the stiffness was 33.1% and 57.2% of that of Group 1, respectively. And no obvious difference in biomechanical parameters was found between Group 4 and 5. Histological analysis demonstrated that formation of fibrovascular tissue in the tibial tunnel and aperture in Groups 4 and 5 and direct junction appeared between tendon graft and tunnel both in Groups 4 and 5. IHC results showed that there are gradually enhanced expression of Patched1, Smoothened and Gli2 concomitant with decreased Gli3 protein in the tendon-bone interface during the tendon-bone healing process. Conclusion We introduced a metal-free, reproducible and reliable mouse model of ACLR compared to the unmodified ACLR procedure, and characterized the expression pattern of key molecules in Ihh signaling during the graft healing process. The translational potential of this article In the present study we introduced and validated, for the first time, a metal-free, reproducible and reliable ACLR mouse model, which could be used to investigate the detailed molecular mechanisms of graft-tunnel healing after ACLR. We also explored new strategies to promote the healing of tendon-to-bone integration.
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Key Words
- ACL, Anterior cruciate ligament
- ACLR, ACL reconstruction
- Anterior cruciate ligament
- BMD, Bone mineral density
- BV/TV, Bone volume/total volume
- CI, Confidence interval
- CT, Computed tomography
- Gli1, Glioma-associated oncogene homologue 1
- Gli2, Glioma-associated oncogene homologue 2
- Gli3, Glioma-associated oncogene homologue 3
- H&E, Haematoxylin-eosin
- Hedgehog signaling
- Ihh, Indian hedgehog
- Mouse model
- NS, Non-significant
- Ptch1, Patched1
- Smo, Smoothened
- Tendon-bone healing
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Kamalitdinov TB, Fujino K, Shetye SS, Jiang X, Ye Y, Rodriguez AB, Kuntz AF, Zgonis MH, Dyment NA. Amplifying Bone Marrow Progenitors Expressing α-Smooth Muscle Actin Produce Zonal Insertion Sites During Tendon-to-Bone Repair. J Orthop Res 2020; 38:105-116. [PMID: 31228280 PMCID: PMC6917878 DOI: 10.1002/jor.24395] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/31/2019] [Accepted: 06/06/2019] [Indexed: 02/04/2023]
Abstract
Traditional tendon-to-bone repair where the tendon is reattached to bone via suture anchors often results in disorganized scar production rather than the formation of a zonal insertion. In contrast, ligament reconstructions where tendon grafts are passed through bone tunnels can yield zonal tendon-to-bone attachments between the graft and adjacent bone. Therefore, ligament reconstructions can be used to study mechanisms that regulate zonal tendon-to-bone repair in the adult. Anterior cruciate ligament (ACL) reconstructions are one of the most common reconstruction procedures and while we know that cells from outside the graft produce the attachments, we have not yet established specific cell populations that give rise to this tissue. To address this knowledge gap, we performed ACL reconstructions in lineage tracing mice where α-smooth muscle actin (αSMACreERT2) was used to label αSMA-expressing progenitors within the bone marrow that produced zonal attachments. Expression of αSMA was increased during early stages of the repair process such that the contribution of SMA-labeled cells to the tunnel integration was highest when tamoxifen was delivered in the first week post-surgery. The zonal attachments shared features with normal entheses, including tidemarks oriented perpendicularly to collagen fibers, Col1a1-expressing cells, alkaline phosphatase activity, and proteoglycan-rich staining. Finally, the integration strength increased with time, requiring 112% greater force to remove the graft from the tunnel at 28 days compared with 14 days post-surgery. Future studies will target these progenitor cells to define the pathways that regulate zonal tendon-to-bone repair in the adult. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:105-116, 2020.
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Affiliation(s)
- Timur B. Kamalitdinov
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Keitaro Fujino
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA,Department of Orthopedic Surgery, Osaka Medical College, Osaka, Japan
| | - Snehal S. Shetye
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Xi Jiang
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Yaping Ye
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Ashley B. Rodriguez
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Andrew F. Kuntz
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Miltiadis H. Zgonis
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
| | - Nathaniel A. Dyment
- McKay Orthopaedic Research Laboratory, Department of Orthopaedic Surgery, University of Pennsylvania, Philadelphia, PA, USA
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Yu S, Tang Q, Xie M, Zhou X, Long Y, Xie Y, Guo F, Chen L. Circadian BMAL1 regulates mandibular condyle development by hedgehog pathway. Cell Prolif 2020; 53:e12727. [PMID: 31747713 PMCID: PMC6985652 DOI: 10.1111/cpr.12727] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2019] [Revised: 10/17/2019] [Accepted: 10/31/2019] [Indexed: 12/13/2022] Open
Abstract
OBJECTIVE Chondrogenesis and endochondral ossification in mandibular condyle play crucial roles in maxillofacial morphogenesis and function. Circadian regulator brain and muscle arnt-like 1 (BMAL1) is proven to be essential for embryonic and postnatal development. The goal of this study was to define the functions of BMAL1 in the embryonic and postnatal growth of mandibular condylar cartilages (MCC). MATERIALS AND METHODS Micro-CT, TUNEL staining and EdU assay were performed using BMAL1-deficient mice model, and in vitro experiments were performed using rat chondrocytes isolated from MCC. RNA sequencing in mandibular condyle tissues from Bmal1-/- mice and the age-matched wild-type mice was used for transcriptional profiling at different postnatal stages. RESULTS The expression levels of BMAL1 decrease gradually in MCC. BMAL1 is proved to regulate sequential chondrocyte differentiation, and its deficiency can result in the impairment of endochondral ossification of MCC. RNA sequencing reveals hedgehog signalling pathway is the potential target of BMAL1. BMAL1 regulates hedgehog signalling and affects its downstream cascades through directly binding to the promoters of Ptch1 and Ihh, modulating targets of hedgehog signalling which is indispensable for endochondral ossification. Importantly, the short stature phenotypes caused by BMAL1 deficiency can be rescued by hedgehog signalling activator. CONCLUSIONS Collectively, these results indicate that BMAL1 plays critical roles on chondrogenesis and endochondral ossification of MCC, giving a new insight on potential therapeutic strategies for facial dysmorphism.
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Affiliation(s)
- Shaoling Yu
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Qingming Tang
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Mengru Xie
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Xin Zhou
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Yanlin Long
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Yanling Xie
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Fengyuan Guo
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
| | - Lili Chen
- Department of StomatologyUnion HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina
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A multi-chamber tissue culture device for load-dependent parallel evaluation of tendon explants. BMC Musculoskelet Disord 2019; 20:549. [PMID: 31739778 PMCID: PMC6862789 DOI: 10.1186/s12891-019-2896-2] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/06/2018] [Accepted: 10/14/2019] [Indexed: 11/29/2022] Open
Abstract
Background Injuries in the musculoskeletal system, such as tendon and ligament ruptures, are challenging to manage and often require surgical reconstructions with limited long-term success. Thus, characterizations of these tissues are urgently needed to better understand cellular mechanisms that regulate tissue homeostasis and healing. Explant culturing systems allow for ex vivo analysis of tissues in an environment that mimics the native microenvironment in vivo. Methods Collaborative efforts within our institution facilitated the establishment of a novel explant culturing system. Tissue specimens cultured in single wells, with individual applied loading and/or biological environment, allowed characterization of tissue cultured under a variety of biological loading conditions. Quantitative PCR analysis for selected gene markers was our primary outcome. Results Data were stratified for analysis by either culture environment or loading condition. Our gene expression results show that specimens clustered by culture condition may differ in molecular markers related to ECM production (e.g., Col1a1, Adamts4) and/or organization (e.g., Tnc, Dnc). In contrast, loading condition did significantly alter the median gene expression levels of tissues in comparison to unloaded control samples, although gene expression values related to ECM degradation (e.g., Mmp1, Mmp10) were altered in tendons cultured under tension in the device. Conclusion Our study demonstrates promising utility of a novel explant culturing system for further characterization of musculoskeletal tissues such as native tendons and ligaments, as well as pathologic fibrotic tissues resulting from arthrofibrosis or Dupuytren’s disease.
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Walters M, Crew M, Fyfe G. Bone Surface Micro‐Topography at Craniofacial Entheses: Insights on Osteogenic Adaptation at Muscle Insertions. Anat Rec (Hoboken) 2019; 302:2140-2155. [DOI: 10.1002/ar.24215] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2018] [Revised: 02/04/2019] [Accepted: 03/06/2019] [Indexed: 12/19/2022]
Affiliation(s)
- Mark Walters
- School of Human SciencesThe University of Western Australia Crawley Perth Western Australia
- Department of Plastic and Reconstructive SurgeryPerth Children's Hospital Nedlands Perth Western Australia
| | - Michael Crew
- Health Department of Western Australia and Faculty of Health SciencesCurtin University Western Australia
| | - Georgina Fyfe
- Faculty of Health SciencesCurtin University Perth Western Australia
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23
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Wada S, Lebaschi AH, Nakagawa Y, Carballo CB, Uppstrom TJ, Cong GT, Album ZM, Hall AJ, Ying L, Deng XH, Rodeo SA. Postoperative Tendon Loading With Treadmill Running Delays Tendon-to-Bone Healing: Immunohistochemical Evaluation in a Murine Rotator Cuff Repair Model. J Orthop Res 2019; 37:1628-1637. [PMID: 30977544 DOI: 10.1002/jor.24300] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/27/2018] [Accepted: 03/08/2019] [Indexed: 02/04/2023]
Abstract
Mechanical stress has an important effect on tendon-to-bone healing. The purpose of the present study was to compare tendon-to-bone healing in animals exposed to either tendon unloading (botulinum toxin injection) or excessive loading (treadmill running) in a murine rotator cuff repair model. Forty-eight C57BL/6 mice underwent unilateral supraspinatus tendon detachment and repair. Mice in the unloaded group were injected with botulinum toxin to the supraspinatus muscle. The contralateral shoulder of the unloaded group was used as a control. Mice were euthanized at 1, 2, and 4 weeks after surgery and evaluated with hematoxylin-eosin and immunohistochemical (IHC) staining for Ihh, Gli1, Wnt3a, and β-catenin. The positive staining area on IHC and the Modified Tendon Maturing Score were measured. The score of the unloaded group was significantly higher (better healing) than that of the treadmill group at 4 weeks. Ihh and the glioma-associated oncogene homolog 1 (Gli1) positive area in the unloaded group were significantly higher than those of the control group at 1 week. The peak time-points of the Ihh and Gli1 positive area was 1 week for the unloaded group and 2 weeks for the treadmill group. The Wnt3a positive area in the unloaded group was significantly higher than that of the control group at 2 weeks. The β-catenin positive area in the unloaded group was significantly higher than that of the treadmill group and the control group at 1 week. Our data indicated that the unloaded group has superior tendon maturation compared to the treadmill running group. Excessive tendon loading may delay the tendon healing process by affecting the activity of Ihh and Wnt/β-Catenin pathways. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1628-1637, 2019.
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Affiliation(s)
- Susumu Wada
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Amir H Lebaschi
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Yusuke Nakagawa
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Camila B Carballo
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Tyler J Uppstrom
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Guang-Ting Cong
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Zoe M Album
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Arielle J Hall
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Liang Ying
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Xiang-Hua Deng
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
| | - Scott A Rodeo
- Laboratory for Joint Tissue Repair and Regeneration, Orthopedic Soft Tissue Research Program, The Hospital for Special Surgery, New York, New York
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24
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Rothrauff BB, Smith CA, Ferrer GA, Novaretti JV, Pauyo T, Chao T, Hirsch D, Beaudry MF, Herbst E, Tuan RS, Debski RE, Musahl V. The effect of adipose-derived stem cells on enthesis healing after repair of acute and chronic massive rotator cuff tears in rats. J Shoulder Elbow Surg 2019; 28:654-664. [PMID: 30527883 DOI: 10.1016/j.jse.2018.08.044] [Citation(s) in RCA: 40] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/25/2018] [Revised: 08/26/2018] [Accepted: 08/29/2018] [Indexed: 02/01/2023]
Abstract
BACKGROUND Chronic massive rotator cuff tears heal poorly and often retear. This study investigated the effect of adipose-derived stem cells (ADSCs) and transforming growth factor-β3 (TGF-β3) delivered in 1 of 2 hydrogels (fibrin or gelatin methacrylate [GelMA]) on enthesis healing after repair of acute or chronic massive rotator cuff tears in rats. METHODS Adult male Lewis rats underwent bilateral transection of the supraspinatus and infraspinatus tendons with intramuscular injection of botulinum toxin A (n = 48 rats). After 8 weeks, animals received 1 of 8 interventions (n = 12 shoulders/group): (1) no repair, (2) repair only, or repair augmented with (3) fibrin, (4) GelMA, (5) fibrin + ADSCs, (6) GelMA + ADSCs, (7) fibrin + ADSCs + TGF-β3, or (8) GelMA + ADSCs + TGF-β3. An equal number of animals underwent acute tendon transection and immediate application of 1 of 8 interventions. Enthesis healing was evaluated 4 weeks after the repair by microcomputed tomography, histology, and mechanical testing. RESULTS Increased bone loss and reduced structural properties were seen in chronic compared with acute tears. Bone mineral density of the proximal humerus was higher in repairs of chronic tears augmented with fibrin + ADSCs and GelMA + ADSCs than in unrepaired chronic tears. Similar improvement was not seen in acute tears. No intervention enhanced histologic appearance or structural properties in acute or chronic tears. CONCLUSIONS Surgical repair augmented with ADSCs may provide more benefit in chronic tears compared with acute tears, although there was no added benefit to supplementing ADSCs with TGF-β3.
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Affiliation(s)
- Benjamin B Rothrauff
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Catherine A Smith
- Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA
| | - Gerald A Ferrer
- Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA
| | - João V Novaretti
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Thierry Pauyo
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Tom Chao
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - David Hirsch
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Mason F Beaudry
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Elmar Herbst
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA
| | - Rocky S Tuan
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA; Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA
| | - Richard E Debski
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA; Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA
| | - Volker Musahl
- Department of Orthopaedic Surgery, University of Pittsburgh, Pittsburgh, PA, USA; Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA, USA.
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25
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Deng XH, Lebaschi A, Camp CL, Carballo CB, Coleman NW, Zong J, Grawe BM, Rodeo SA. Expression of Signaling Molecules Involved in Embryonic Development of the Insertion Site Is Inadequate for Reformation of the Native Enthesis: Evaluation in a Novel Murine ACL Reconstruction Model. J Bone Joint Surg Am 2018; 100:e102. [PMID: 30063598 PMCID: PMC6661256 DOI: 10.2106/jbjs.16.01066] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/01/2023]
Abstract
BACKGROUND Since healing of anterior cruciate ligament (ACL) grafts occurs by formation of a fibrovascular scar-tissue interface rather than by reformation of the native fibrocartilage transition zone, the purpose of our study was to examine expression of various signaling molecules and transcription factors that are known to be involved in embryologic insertion-site development following ACL reconstruction. We also aimed to characterize a murine model of ACL reconstruction to allow future study of the molecular mechanisms of healing. METHODS Seventy-nine mice underwent reconstruction of the ACL with autograft. Healing was assessed using histology in 12 mice and quantitative real-time polymerase chain reaction (qRT-PCR) gene-expression analysis in 3 mice at 1 week postoperatively (Group-1 mice) and by biomechanical analysis in 7, histological analysis in 7, immunohistochemical analysis in 5, microcomputed tomography analysis in 5, and qRT-PCR analyses in 8 at 2 weeks (Group-2 mice) and 4 weeks (Group-3 mice) postoperatively. Fifteen additional mice did not undergo surgery and were used for biomechanical (7 mice), qRT-PCR (3 mice), and immunohistochemical (5 mice) analyses to obtain baseline data for the native ACL. RESULTS Histological analysis demonstrated healing by formation of fibrovascular tissue at the tendon-bone interface. Immunohistochemical analysis showed a positive expression of proteins in the Indian hedgehog, Wnt, and parathyroid hormone-related protein (PTHrP) pathways. There was minimal Sox-9 expression. Gene-expression analysis showed an initial increase in markers of tissue repair and turnover, followed by a subsequent decline. Mean failure force and stiffness of the native ACL were 5.60 N and 3.44 N/mm, respectively. Mean failure force and stiffness were 1.29 N and 2.28 N/mm, respectively, in Group 2 and were 1.79 N and 2.59 N/mm, respectively, in Group 3, with 12 of 14 failures in these study groups occurring by tunnel pull-out. CONCLUSIONS The spatial and temporal pattern of expression of signaling molecules that direct embryologic insertion-site formation was not adequate to restore the structure and composition of the native insertion site. CLINICAL RELEVANCE Development of a murine model to study ACL reconstruction will allow the use of transgenic animals to investigate the cellular, molecular, and biomechanical aspects of tendon-to-bone healing following ACL reconstruction, ultimately suggesting methods to improve healing in patients.
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Affiliation(s)
- Xiang-Hua Deng
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY
| | - Amir Lebaschi
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY
| | - Christopher L. Camp
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY,Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota
| | - Camila B. Carballo
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY
| | - Nathan W. Coleman
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY
| | - Jianchun Zong
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY
| | - Brian M. Grawe
- Sports Medicine and Shoulder Reconstruction, Department of Orthopaedics, University of Cincinnati Academic Health Center, Cincinnati, Ohio
| | - Scott A. Rodeo
- Orthopaedic Soft Tissue Research Program, Hospital for Special Surgery, New York, NY,E-mail address for S.A. Rodeo:
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26
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Jensen PT, Lambertsen KL, Frich LH. Assembly, maturation, and degradation of the supraspinatus enthesis. J Shoulder Elbow Surg 2018; 27:739-750. [PMID: 29329904 DOI: 10.1016/j.jse.2017.10.030] [Citation(s) in RCA: 41] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/02/2017] [Revised: 10/22/2017] [Accepted: 10/27/2017] [Indexed: 02/01/2023]
Abstract
The development of the rotator cuff enthesis is still poorly understood. The processes in the early and late developmental steps are gradually elucidated, but it is still unclear how cell activities are coordinated during development and maturation of the structured enthesis. This review summarizes current knowledge about development and age-related degradation of the supraspinatus enthesis. Healing and repair of an injured and degenerated supraspinatus enthesis also remain a challenge, as the original graded transitional tissue of the fibrocartilaginous insertion is not re-created after the tendon is surgically reattached to bone. Instead, mechanically inferior and disorganized tissue forms at the healing site because of scar tissue formation. Consequently, the enthesis never reaches mechanical properties comparable to those of the native enthesis. So far, no novel biologic healing approach has been successful in enhancing healing of the injured enthesis. The results revealed in this review imply the need for further research to pave the way for better treatment of patients with rotator cuff disorder.
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Affiliation(s)
- Peter T Jensen
- Department of Clinical Research, University of Southern Denmark, Odense, Denmark
| | - Kate L Lambertsen
- Department of Clinical Research, University of Southern Denmark, Odense, Denmark; Department of Neurobiology Research, Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark
| | - Lars H Frich
- Department of Clinical Research, University of Southern Denmark, Odense, Denmark; Department of Orthopaedics and Traumatology, Odense University Hospital, Odense, Denmark.
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27
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Growth factor delivery strategies for rotator cuff repair and regeneration. Int J Pharm 2018; 544:358-371. [PMID: 29317260 DOI: 10.1016/j.ijpharm.2018.01.006] [Citation(s) in RCA: 61] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2017] [Revised: 12/21/2017] [Accepted: 01/01/2018] [Indexed: 12/21/2022]
Abstract
The high incidence of degenerative tears and prevalence of retears (20-95%) after surgical repair makes rotator cuff injuries a significant health problem. This high retear rate is attributed to the failure of the repaired tissue to regenerate the native tendon-to-bone insertion (enthesis). Biological augmentation of surgical repair such as autografts, allografts, and xenografts are confounded by donor site morbidity, immunogenicity, and disease transmission, respectively. In contrast, these risks may be alleviated via growth factor therapy, which can actively influence the healing environment to promote functional repair. Several challenges have to be overcome before growth factor delivery can translate into clinical practice such as the selection of optimal growth factor(s) or combination, identification of the most efficient stage and duration of delivery, and the design considerations for the delivery device. Emerging insight into the injury-repair microenvironment and our understanding of growth factor mechanisms in healing are informing the design of advanced delivery scaffolds to effectively treat rotator cuff tears. Here, we review potential growth factor candidates, design parameters and material selection for growth factor delivery, innovative and dynamic delivery scaffolds, and novel therapeutic targets from tendon and developmental biology for the structural and functional healing of rotator cuff repair.
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28
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Lim J, Munivez E, Jiang MM, Song IW, Gannon F, Keene DR, Schweitzer R, Lee BH, Joeng KS. mTORC1 Signaling is a Critical Regulator of Postnatal Tendon Development. Sci Rep 2017; 7:17175. [PMID: 29215029 PMCID: PMC5719403 DOI: 10.1038/s41598-017-17384-0] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2017] [Accepted: 11/24/2017] [Indexed: 12/11/2022] Open
Abstract
Tendons transmit contractile forces between musculoskeletal tissues. Whereas the biomechanical properties of tendons have been studied extensively, the molecular mechanisms regulating postnatal tendon development are not well understood. Here we examine the role of mTORC1 signaling in postnatal tendon development using mouse genetic approaches. Loss of mTORC1 signaling by removal of Raptor in tendons caused severe tendon defects postnatally, including decreased tendon thickness, indicating that mTORC1 is necessary for postnatal tendon development. By contrast, activation of mTORC1 signaling in tendons increased tendon cell numbers and proliferation. In addition, Tsc1 conditional knockout mice presented severely disorganized collagen fibers and neovascularization in the tendon midsubstance. Interestingly, collagen fibril diameter was significantly reduced in both Raptor and Tsc1 conditional knockout mice, albeit with variations in severity. We performed RNA-seq analysis using Achilles tendons to investigate the molecular changes underlying these tendon phenotypes. Raptor conditional knockout mice showed decreased extracellular matrix (ECM) structure-related gene expression, whereas Tsc1 conditional knockout mice exhibited changes in genes regulating TGF-β/BMP/FGF signaling, as well as in genes controlling ECM structure and disassembly. Collectively, our studies suggest that maintaining physiological levels of mTORC1 signaling is essential for postnatal tendon development and maturation.
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Affiliation(s)
- Joohyun Lim
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
| | - Elda Munivez
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
| | - Ming-Ming Jiang
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
| | - I-Wen Song
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
| | - Francis Gannon
- Department of Pathology, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
| | - Douglas R Keene
- Research Center, Shriners Hospital for Children, Portland, OR, 97239, USA
| | - Ronen Schweitzer
- Research Center, Shriners Hospital for Children, Portland, OR, 97239, USA
| | - Brendan H Lee
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA.
| | - Kyu Sang Joeng
- Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA
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29
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Li G, Hosseini A, Gadikota H, Gill T. A Novel Graft Fixation Technique for Anterior Cruciate Ligament Reconstruction Using Hamstring Tendon Grafts. J Med Device 2017. [DOI: 10.1115/1.4038307] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
This study evaluated the biomechanical efficacy of single-tunnel double-bundle anterior cruciate ligament (ACL) reconstruction technique. The graft construct is achieved using a novel fixation device that splits an ACL (SPACL) graft into two bundles, recreating the anteromedial (AM) and posterolateral (PL) bundles for ACL reconstruction. A pullout strength test of the SPACL was performed using a 7-mm bovine digital extensor tendon graft. The capability in restoration of knee kinematics after SPACL reconstruction was investigated using cadaveric human knees on a robotic testing system under an anterior tibial load of 134 N and a simulated quadriceps load of 400 N. The data indicated that the SPACL graft has a pullout strength of 823.7±172.3 N. Under the 134 N anterior tibial load, the anteroposterior joint laxity had increased constraint using the SPACL reconstruction but not significantly (p > 0.05) at all selected flexion angles. Under the 400 N quadriceps load, no significant differences were observed between the anterior tibial translation of intact knee and SPACL conditions at all selected flexion angles, but the SPACL graft induced a significant increase in external tibial rotation compared to the intact knee condition at all selected flexion angles with a maximal external rotation of −3.20 deg ±3.6 deg at 90 deg flexion. These data showed that the SPACL technique is equivalent or superior to existing ACL reconstruction techniques in restoration of knee laxity and kinematics. The new SPACL reconstruction technique could provide a valuable alternation to contemporary ACL reconstruction surgery by more closely recreating native ACL kinematics.
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Affiliation(s)
- Guoan Li
- Orthopaedic Biomechanics Lab, Department of Orthopaedic Surgery, Newton-Wellesley Hospital/Harvard Medical School, Newton, MA 02462 e-mail:
| | - Ali Hosseini
- Orthopaedic Biomechanics Lab, Department of Orthopaedic Surgery, Newton-Wellesley Hospital/Harvard Medical School, Newton, MA 02462
| | - Hemanth Gadikota
- Orthopaedic Biomechanics Lab, Department of Orthopaedic Surgery, Newton-Wellesley Hospital/Harvard Medical School, Newton, MA 02462
| | - Thomas Gill
- Orthopaedic Biomechanics Lab, Department of Orthopaedic Surgery, Newton-Wellesley Hospital/Harvard Medical School, Newton, MA 02462
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30
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Liu Y, Suen CW, Zhang JF, Li G. Current concepts on tenogenic differentiation and clinical applications. J Orthop Translat 2017; 9:28-42. [PMID: 29662797 PMCID: PMC5822963 DOI: 10.1016/j.jot.2017.02.005] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/15/2016] [Revised: 02/21/2017] [Accepted: 02/23/2017] [Indexed: 12/16/2022] Open
Abstract
Tendon is a tissue that transmits force from muscle to bone. Chronic or acute tendon injuries are very common, and are always accompanied by pain and a limited range of motion in patients. In clinical settings, management of tendon injuries still remains a big challenge. Cell therapies, such as the application of stem cells for tenogenic differentiation, were suggested to be an ideal strategy for clinical translation. However, there is still a lack of specific methods for tenogenic differentiation due to the limited understanding of tendon biology currently. This review focuses on the summary of current published strategies for tenogenic differentiation, such as the application of growth factors, mechanical stimulation, biomaterials, coculture, or induced pluripotent stem cells. Current clinical applications of stem cells for treatment of tendon injuries and their limitations have also been discussed in this review.
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Affiliation(s)
- Yang Liu
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China
| | - Chun-Wai Suen
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China
| | - Jin-fang Zhang
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China
| | - Gang Li
- Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China
- Stem Cells and Regenerative Medicine Laboratory, Lui Che Woo Institute of Innovative Medicine, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China
- Key Laboratory for Regenerative Medicine, Ministry of Education, School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, China
- The CUHK-ACC Space Medicine Centre on Health Maintenance of Musculoskeletal System, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China
- Corresponding author. Department of Orthopaedics and Traumatology and Li Ka Shing Institute of Health Sciences, Prince of Wales Hospital, The Chinese University of Hong Kong, 30-32 Ngan Shing Street, Shatin, New Territories, Hong Kong, China.Department of Orthopaedics and Traumatology and Li Ka Shing Institute of Health SciencesPrince of Wales HospitalThe Chinese University of Hong Kong30-32 Ngan Shing StreetShatinNew TerritoriesHong Kong, China
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31
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Schwartz AG, Galatz LM, Thomopoulos S. Enthesis regeneration: a role for Gli1+ progenitor cells. Development 2017; 144:1159-1164. [PMID: 28219952 DOI: 10.1242/dev.139303] [Citation(s) in RCA: 43] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2016] [Accepted: 02/13/2017] [Indexed: 12/14/2022]
Abstract
The tendon enthesis originates from a specific pool of hedgehog-active Gli1+ progenitor cells that differentiate and produce mineralized fibrocartilage. The current study investigated the regenerative capacity of this cell population by comparing the responses of early postnatal and mature entheses to injury. Lineage tracing studies demonstrated that the original Gli1+ cell population had the capacity to heal immature entheses after injury, but this capacity was lost after the cells differentiated into mature fibrochondrocytes. To further examine the involvement of Gli1+ cells and hedgehog signaling in enthesis healing, Gli1 expression was examined via lineage tracing approaches and the effect of Smo deletion was examined in the injured entheses. Immature injured entheses retained high levels of Gli1 expression, a marker of hedgehog activation, consistent with non-injured controls. In contrast, injured mature entheses had few Gli1+ cells early in the healing process, with limited recovery of the cell population later in the healing process. These results suggest that the presence of activated hedgehog signaling in enthesis cells early in the healing process may enhance healing of enthesis injuries by mimicking developmental processes.
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Affiliation(s)
- Andrea G Schwartz
- Department of Orthopedic Surgery, Washington University, St. Louis, MO 63110, USA
| | - Leesa M Galatz
- Department of Orthopedic Surgery, Icahn School of Medicine at Mount Sinai Hospital, Mount Sinai Health System, New York, NY 10029, USA
| | - Stavros Thomopoulos
- Department of Orthopedic Surgery, Columbia University, New York, NY 10032, USA .,Department of Biomedical Engineering, Columbia University, New York, NY 10032, USA
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32
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Song F, Jiang D, Wang T, Wang Y, Chen F, Xu G, Kang Y, Zhang Y. Mechanical Loading Improves Tendon-Bone Healing in a Rabbit Anterior Cruciate Ligament Reconstruction Model by Promoting Proliferation and Matrix Formation of Mesenchymal Stem Cells and Tendon Cells. Cell Physiol Biochem 2017; 41:875-889. [DOI: 10.1159/000460005] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2016] [Accepted: 12/21/2016] [Indexed: 12/17/2022] Open
Abstract
Background/Aims: This study investigated the effect of mechanical stress on tendon-bone healing in a rabbit anterior cruciate ligament (ACL) reconstruction model as well as cell proliferation and matrix formation in co-culture of bone-marrow mesenchymal stem cells (BMSCs) and tendon cells (TCs). Methods: The effect of continuous passive motion (CPM) therapy on tendon-bone healing in a rabbit ACL reconstruction model was evaluated by histological analysis, biomechanical testing and gene expressions at the tendon-bone interface. Furthermore, the effect of mechanical stretch on cell proliferation and matrix synthesis in BMSC/TC co-culture was also examined. Results: Postoperative CPM therapy significantly enhanced tendon-bone healing, as evidenced by increased amount of fibrocartilage, elevated ultimate load to failure levels, and up-regulated gene expressions of Collagen I, alkaline phosphatase, osteopontin, Tenascin C and tenomodulin at the tendon-bone junction. In addition, BMSC/TC co-culture treated with mechanical stretch showed a higher rate of cell proliferation and enhanced expressions of Collagen I, Collagen III, alkaline phosphatase, osteopontin, Tenascin C and tenomodulin than that of controls. Conclusion: These results demonstrated that proliferation and differentiation of local precursor cells could be enhanced by mechanical stimulation, which results in enhanced regenerative potential of BMSCs and TCs in tendon-bone healing.
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33
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Rothrauff BB, Pauyo T, Debski RE, Rodosky MW, Tuan RS, Musahl V. The Rotator Cuff Organ: Integrating Developmental Biology, Tissue Engineering, and Surgical Considerations to Treat Chronic Massive Rotator Cuff Tears. TISSUE ENGINEERING PART B-REVIEWS 2017; 23:318-335. [PMID: 28084902 DOI: 10.1089/ten.teb.2016.0446] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
The torn rotator cuff remains a persistent orthopedic challenge, with poor outcomes disproportionately associated with chronic, massive tears. Degenerative changes in the tissues that comprise the rotator cuff organ, including muscle, tendon, and bone, contribute to the poor healing capacity of chronic tears, resulting in poor function and an increased risk for repair failure. Tissue engineering strategies to augment rotator cuff repair have been developed in an effort to improve rotator cuff healing and have focused on three principal aims: (1) immediate mechanical augmentation of the surgical repair, (2) restoration of muscle quality and contractility, and (3) regeneration of native enthesis structure. Work in these areas will be reviewed in sequence, highlighting the relevant pathophysiology, developmental biology, and biomechanics, which must be considered when designing therapeutic applications. While the independent use of these strategies has shown promise, synergistic benefits may emerge from their combined application given the interdependence of the tissues that constitute the rotator cuff organ. Furthermore, controlled mobilization of augmented rotator cuff repairs during postoperative rehabilitation may provide mechanotransductive cues capable of guiding tissue regeneration and restoration of rotator cuff function. Present challenges and future possibilities will be identified, which if realized, may provide solutions to the vexing condition of chronic massive rotator cuff tears.
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Affiliation(s)
- Benjamin B Rothrauff
- 1 Department of Orthopaedic Surgery, Center for Cellular and Molecular Engineering, University of Pittsburgh , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh , Pittsburgh, Pennsylvania
| | - Thierry Pauyo
- 3 Division of Sports Medicine, Department of Orthopaedic Surgery, University of Pittsburgh , Pittsburgh, Pennsylvania
| | - Richard E Debski
- 2 McGowan Institute for Regenerative Medicine, University of Pittsburgh , Pittsburgh, Pennsylvania
| | - Mark W Rodosky
- 3 Division of Sports Medicine, Department of Orthopaedic Surgery, University of Pittsburgh , Pittsburgh, Pennsylvania
| | - Rocky S Tuan
- 1 Department of Orthopaedic Surgery, Center for Cellular and Molecular Engineering, University of Pittsburgh , Pittsburgh, Pennsylvania.,2 McGowan Institute for Regenerative Medicine, University of Pittsburgh , Pittsburgh, Pennsylvania
| | - Volker Musahl
- 2 McGowan Institute for Regenerative Medicine, University of Pittsburgh , Pittsburgh, Pennsylvania.,3 Division of Sports Medicine, Department of Orthopaedic Surgery, University of Pittsburgh , Pittsburgh, Pennsylvania.,4 Orthopaedic Robotics Laboratory, Department of Orthopaedic Surgery, University of Pittsburgh , Pittsburgh, Pennsylvania
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Lebaschi A, Deng XH, Zong J, Cong GT, Carballo CB, Album ZM, Camp C, Rodeo SA. Animal models for rotator cuff repair. Ann N Y Acad Sci 2016; 1383:43-57. [DOI: 10.1111/nyas.13203] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2016] [Revised: 07/12/2016] [Accepted: 07/18/2016] [Indexed: 01/08/2023]
Affiliation(s)
- Amir Lebaschi
- Tissue Engineering; Repair, and Regeneration Program
| | | | - Jianchun Zong
- Tissue Engineering; Repair, and Regeneration Program
| | | | | | - Zoe M. Album
- Tissue Engineering; Repair, and Regeneration Program
| | - Christopher Camp
- Tissue Engineering; Repair, and Regeneration Program
- Sports Medicine and Shoulder Service; Hospital for Special Surgery; New York New York
| | - Scott A. Rodeo
- Tissue Engineering; Repair, and Regeneration Program
- Sports Medicine and Shoulder Service; Hospital for Special Surgery; New York New York
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