Publications by authors named "Farshid Guilak"

304 Publications

Fresh Osteochondral and Chondral Allograft Preservation and Storage Media: A Systematic Review of the Literature.

Am J Sports Med 2021 Jul 26:3635465211016832. Epub 2021 Jul 26.

University of California, San Diego, San Diego, California, USA.

Background: Storage procedures and parameters have a significant influence on the health of fresh osteochondral allograft (OCA) cartilage. To date, there is a lack of agreement on the optimal storage conditions for OCAs.

Purpose: To systematically review the literature on (1) experimental designs and reporting of key variables of ex vivo (laboratory) studies, (2) the effects of various storage solutions and conditions on cartilage health ex vivo, and (3) in vivo animal studies and human clinical studies evaluating the effect of fresh OCA storage on osteochondral repair and outcomes.

Study Design: Systematic review; Level of evidence, 5.

Methods: A systematic review was performed using the PubMed, Embase, and Cochrane databases. The inclusion criteria were laboratory studies (ex vivo) reporting cartilage health outcomes after prolonged storage (>3 days) of fresh osteochondral or chondral tissue explants and animal studies (in vivo) reporting outcomes of fresh OCA. The inclusion criteria for clinical studies were studies (>5 patients) that analyzed the relationship of storage time or chondrocyte viability at time of implantation to patient outcomes. Frozen, cryopreserved, decellularized, synthetic, or tissue-engineered grafts were excluded.

Results: A total of 55 peer-reviewed articles met the inclusion criteria. Ex vivo studies reported a spectrum of tissue sources and storage solutions and conditions, although the majority of studies lacked complete reporting of key variables, including storage solution formula and environmental conditions. The effect of various conditions (eg, temperature) and storage solutions on cartilage health were inconsistent. Although 60% of animal models suggest that storage time may influence outcomes and 80% indicate inferior outcomes with frozen OCA as compared with fresh OCA, 75% of clinical studies report no correlation between storage time and outcomes.

Conclusion: Given the variability in experimental designs and lack of reporting across studies, it is still not possible to determine optimal storage conditions, although animal studies suggest that storage time and chondrocyte viability influence osteochondral repair outcomes. A list of recommendations was developed to encourage reporting of key variables, such as media formulation, environmental factors, and methodologies used. High-quality clinical data are needed to investigate the effects of storage and graft health on outcomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/03635465211016832DOI Listing
July 2021

Taxonomic changes in the gut microbiota are associated with cartilage damage independent of adiposity, high fat diet, and joint injury.

Sci Rep 2021 Jul 15;11(1):14560. Epub 2021 Jul 15.

Department of Orthopaedic Surgery, Washington University, Couch Building Room 3213, 4523 Clayton Avenue, St Louis, MO, 63110, USA.

Lipodystrophic mice are protected from cartilage damage following joint injury. This protection can be reversed by the implantation of a small adipose tissue graft. The purpose of this study was to evaluate the relationship between the gut microbiota and knee cartilage damage while controlling for adiposity, high fat diet, and joint injury using lipodystrophic (LD) mice. LD and littermate control (WT) mice were fed a high fat diet, chow diet, or were rescued with fat implantation, then challenged with destabilization of the medial meniscus surgery to induce osteoarthritis (OA). 16S rRNA sequencing was conducted on feces. MaAslin2 was used to determine associations between taxonomic relative abundance and OA severity. While serum LPS levels between groups were similar, synovial fluid LPS levels were increased in both limbs of HFD WT mice compared to all groups, except for fat transplanted animals. The Bacteroidetes:Firmicutes ratio of the gut microbiota was significantly reduced in HFD and OA-rescued animals when compared to chow. Nine novel significant associations were found between gut microbiota taxa and OA severity. These findings suggest the presence of causal relationships the gut microbiome and cartilage health, independent of diet or adiposity, providing potential therapeutic targets through manipulation of the microbiome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-021-94125-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8282619PMC
July 2021

Cartilage from human-induced pluripotent stem cells: comparison with neo-cartilage from chondrocytes and bone marrow mesenchymal stromal cells.

Cell Tissue Res 2021 Jul 9. Epub 2021 Jul 9.

Dept. of Biomedical Data Sciences, Section Molecular Epidemiology, Leiden University Medical Center, LUMC, Leiden, The Netherlands.

Cartilage has little intrinsic capacity for repair, so transplantation of exogenous cartilage cells is considered a realistic option for cartilage regeneration. We explored whether human-induced pluripotent stem cells (hiPSCs) could represent such unlimited cell sources for neo-cartilage comparable to human primary articular chondrocytes (hPACs) or human bone marrow-derived mesenchymal stromal cells (hBMSCs). For this, chondroprogenitor cells (hiCPCs) and hiPSC-derived mesenchymal stromal cells (hiMSCs) were generated from two independent hiPSC lines and characterized by morphology, flow cytometry, and differentiation potential. Chondrogenesis was compared to hBMSCs and hPACs by histology, immunohistochemistry, and RT-qPCR, while similarities were estimated based on Pearson correlations using a panel of 20 relevant genes. Our data show successful differentiations of hiPSC into hiMSCs and hiCPCs. Characteristic hBMSC markers were shared between hBMSCs and hiMSCs, with the exception of CD146 and CD45. However, neo-cartilage generated from hiMSCs showed low resemblances when compared to hBMSCs (53%) and hPACs (39%) characterized by lower collagen type 2 and higher collagen type 1 expression. Contrarily, hiCPC neo-cartilage generated neo-cartilage more similar to hPACs (65%), with stronger expression of matrix deposition markers. Our study shows that taking a stepwise approach to generate neo-cartilage from hiPSCs via chondroprogenitor cells results in strong similarities to neo-cartilage of hPACs within 3 weeks following chondrogenesis, making them a potential candidate for regenerative therapies. Contrarily, neo-cartilage deposited by hiMSCs seems more prone to hypertrophic characteristics compared to hPACs. We therefore compared chondrocytes derived from hiMSCs and hiCPCs with hPACs and hBMSCs to outline similarities and differences between their neo-cartilage and establish their potential suitability for regenerative medicine and disease modelling.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00441-021-03498-5DOI Listing
July 2021

Immunoengineering the next generation of arthritis therapies.

Acta Biomater 2021 Apr 3. Epub 2021 Apr 3.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO 63110, USA; Shriners Hospitals for Children - St. Louis, St. Louis, MO 63110, USA; Department of Biomedical Engineering, Washington University, St. Louis, MO 63110, USA; Center of Regenerative Medicine, Washington University, St. Louis, MO 63110, USA. Electronic address:

Immunoengineering continues to revolutionize healthcare, generating new approaches for treating previously intractable diseases, particularly in regard to cancer immunotherapy. In joint diseases, such as osteoarthritis (OA) and rheumatoid arthritis (RA), biomaterials and anti-cytokine treatments have previously been at that forefront of therapeutic innovation. However, while many of the existing anti-cytokine treatments are successful for a subset of patients, these treatments can also pose severe risks, adverse events and off-target effects due to continuous delivery at high dosages or a lack of disease-specific targets. The inadequacy of these current treatments has motivated the development of new immunoengineering strategies that offer safer and more efficacious alternative therapies through the precise and controlled targeting of specific upstream immune responses, including direct and mechanistically-driven immunoengineering approaches. Advances in the understanding of the immunomodulatory pathways involved in musculoskeletal disease, in combination with the growing emphasis on personalized medicine, stress the need for carefully considering the delivery strategies and therapeutic targets when designing therapeutics to better treat RA and OA. Here, we focus on recent advances in biomaterial and cell-based immunomodulation, in combination with genetic engineering, for therapeutic applications in joint diseases. The application of immunoengineering principles to the study of joint disease will not only help to elucidate the mechanisms of disease pathogenesis but will also generate novel disease-specific therapeutics by harnessing cellular and biomaterial responses. STATEMENT OF SIGNIFICANCE: It is now apparent that joint diseases such as osteoarthritis and rheumatoid arthritis involve the immune system at both local (i.e., within the joint) and systemic levels. In this regard, targeting the immune system using both biomaterial-based or cellular approaches may generate new joint-specific treatment strategies that are well-controlled, safe, and efficacious. In this review, we focus on recent advances in immunoengineering that leverage biomaterials and/or genetically engineered cells for therapeutic applications in joint diseases. The application of such approaches, especially synergistic strategies that target multiple immunoregulatory pathways, has the potential to revolutionize our understanding, treatment, and prevention of joint diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.actbio.2021.03.062DOI Listing
April 2021

Perlecan in Pericellular Mechanosensory Cell-Matrix Communication, Extracellular Matrix Stabilisation and Mechanoregulation of Load-Bearing Connective Tissues.

Int J Mol Sci 2021 Mar 8;22(5). Epub 2021 Mar 8.

Graduate School of Biomedical Engineering, University of New South Wales, Sydney, NSW 2052, Australia.

In this study, we review mechanoregulatory roles for perlecan in load-bearing connective tissues. Perlecan facilitates the co-acervation of tropoelastin and assembly of elastic microfibrils in translamellar cross-bridges which, together with fibrillin and elastin stabilise the extracellular matrix of the intervertebral disc annulus fibrosus. Pericellular perlecan interacts with collagen VI and XI to define and stabilize this matrix compartment which has a strategic position facilitating two-way cell-matrix communication between the cell and its wider extracellular matrix. Cues from the extracellular matrix are fed through this pericellular matrix back to the chondrocyte, allowing it to perceive and respond to subtle microenvironmental changes to regulate tissue homeostasis. Thus perlecan plays a key regulatory role in chondrocyte metabolism, and in chondrocyte differentiation. Perlecan acts as a transport proteoglycan carrying poorly soluble, lipid-modified proteins such as the Wnt or Hedgehog families facilitating the establishment of morphogen gradients that drive tissue morphogenesis. Cell surface perlecan on endothelial cells or osteocytes acts as a flow sensor in blood and the lacunar canalicular fluid providing feedback cues to smooth muscle cells regulating vascular tone and blood pressure, and the regulation of bone metabolism by osteocytes highlighting perlecan's multifaceted roles in load-bearing connective tissues.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms22052716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7962540PMC
March 2021

Inflammatory signaling sensitizes Piezo1 mechanotransduction in articular chondrocytes as a pathogenic feed-forward mechanism in osteoarthritis.

Proc Natl Acad Sci U S A 2021 Mar;118(13)

Department of Neurology, Duke University, Durham, NC 27710;

Osteoarthritis (OA) is a painful and debilitating condition of synovial joints without any disease-modifying therapies [A. M. Valdes, T. D. Spector, 7, 23-32 (2011)]. We previously identified mechanosensitive PIEZO channels, PIEZO1 and PIEZO2, both expressed in articular cartilage, to function in chondrocyte mechanotransduction in response to injury [W. Lee , 111, E5114-E5122 (2014); W. Lee, F. Guilak, W. Liedtke, 79, 263-273 (2017)]. We therefore asked whether interleukin-1-mediated inflammatory signaling, as occurs in OA, influences iezo gene expression and channel function, thus indicative of maladaptive reprogramming that can be rationally targeted. Primary porcine chondrocyte culture and human osteoarthritic cartilage tissue were studied. We found that interleukin-1α (IL-1α) up-regulated Piezo1 in porcine chondrocytes. Piezo1 expression was significantly increased in human osteoarthritic cartilage. Increased Piezo1 expression in chondrocytes resulted in a feed-forward pathomechanism whereby increased function of Piezo1 induced excess intracellular Ca at baseline and in response to mechanical deformation. Elevated resting state Ca in turn rarefied the F-actin cytoskeleton and amplified mechanically induced deformation microtrauma. As intracellular substrates of this OA-related inflammatory pathomechanism, in porcine articular chondrocytes exposed to IL-1α, we discovered that enhanced Piezo1 expression depended on p38 MAP-kinase and transcription factors HNF4 and ATF2/CREBP1. CREBP1 directly bound to the proximal gene promoter. Taken together, these signaling and genetic reprogramming events represent a detrimental Ca-driven feed-forward mechanism that can be rationally targeted to stem the progression of OA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2001611118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8020656PMC
March 2021

Mapping the musculoskeletal system one cell at a time.

Nat Rev Rheumatol 2021 05;17(5):247-248

The Botnar Research Centre, Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Science, University of Oxford, Oxford, UK.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41584-021-00600-7DOI Listing
May 2021

A synthetic mechanogenetic gene circuit for autonomous drug delivery in engineered tissues.

Sci Adv 2021 Jan 27;7(5). Epub 2021 Jan 27.

Department of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USA.

Mechanobiologic signals regulate cellular responses under physiologic and pathologic conditions. Using synthetic biology and tissue engineering, we developed a mechanically responsive bioartificial tissue that responds to mechanical loading to produce a preprogrammed therapeutic biologic drug. By deconstructing the signaling networks induced by activation of the mechanically sensitive ion channel transient receptor potential vanilloid 4 (TRPV4), we created synthetic TRPV4-responsive genetic circuits in chondrocytes. We engineered these cells into living tissues that respond to mechanical loading by producing the anti-inflammatory biologic drug interleukin-1 receptor antagonist. Chondrocyte TRPV4 is activated by osmotic loading and not by direct cellular deformation, suggesting that tissue loading is transduced into an osmotic signal that activates TRPV4. Either osmotic or mechanical loading of tissues transduced with TRPV4-responsive circuits protected constructs from inflammatory degradation by interleukin-1α. This synthetic mechanobiology approach was used to develop a mechanogenetic system to enable long-term, autonomously regulated drug delivery driven by physiologically relevant loading.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/sciadv.abd9858DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7840132PMC
January 2021

Single-cell RNA sequencing reveals the induction of novel myeloid and myeloid-associated cell populations in visceral fat with long-term obesity.

FASEB J 2021 03;35(3):e21417

Department of Orthopaedic Surgery, Washington University in St. Louis, St. Louis, MO, USA.

Macrophages and other immune cells are important contributors to obesity-associated inflammation; however, the cellular identities of these specific populations remain unknown. In this study, we identified individual populations of myeloid cells found in mouse epididymal/visceral adipose tissue by single-cell RNA sequencing, immunofluorescence, and flow cytometry. Multiple canonical correlation analysis identified 11 unique myeloid and myeloid-associate cell populations. In obese mice, we detected an increased percentage of monocyte-derived pro-inflammatory cells expressing Cd9 and Trem2, as well as significantly decreased percentages of multiple cell populations, including tissue-resident cells expressing Lyve1, Mafb, and Mrc1. We have identified and validated a novel myeloid/macrophage population defined by Ly6a expression, exhibiting both myeloid and mesenchymal characteristics, which increased with obesity and showed high pro-fibrotic characteristics in vitro. Our mouse adipose tissue myeloid cell atlas provides an important resource to investigate obesity-associated inflammation and fibrosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.202001970RDOI Listing
March 2021

Single Cell Omics for Musculoskeletal Research.

Curr Osteoporos Rep 2021 Apr 9;19(2):131-140. Epub 2021 Feb 9.

Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN, USA.

Purpose Of Review: The ability to analyze the molecular events occurring within individual cells as opposed to populations of cells is revolutionizing our understanding of musculoskeletal tissue development and disease. Single cell studies have the great potential of identifying cellular subpopulations that work in a synchronized fashion to regenerate and repair damaged tissues during normal homeostasis. In addition, such studies can elucidate how these processes break down in disease as well as identify cellular subpopulations that drive the disease. This review highlights three emerging technologies: single cell RNA sequencing (scRNA-seq), Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq), and Cytometry by Time-Of-Flight (CyTOF) mass cytometry.

Recent Findings: Technological and bioinformatic tools to analyze the transcriptome, epigenome, and proteome at the individual cell level have advanced rapidly making data collection relatively easy; however, understanding how to access and interpret the data remains a challenge for many scientists. It is, therefore, of paramount significance to educate the musculoskeletal community on how single cell technologies can be used to answer research questions and advance translation. This article summarizes talks given during a workshop on "Single Cell Omics" at the 2020 annual meeting of the Orthopedic Research Society. Studies that applied scRNA-seq, ATAC-seq, and CyTOF mass cytometry to cartilage development and osteoarthritis are reviewed. This body of work shows how these cutting-edge tools can advance our understanding of the cellular heterogeneity and trajectories of lineage specification during development and disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s11914-021-00662-2DOI Listing
April 2021

Regulation of chondrocyte biosynthetic activity by dynamic hydrostatic pressure: the role of TRP channels.

Connect Tissue Res 2021 Jan 25:1-13. Epub 2021 Jan 25.

Department of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO, USA.

Introduction: Chondrocytes perceive and respond to mechanical loading as signals that regulate their metabolism. Joint loading exposes chondrocytes to multiple modes of mechanical stress, including hydrostatic pressure; however, the mechanisms by which chondrocytes sense physiologically relevant levels of hydrostatic pressure are not well understood. We hypothesized that hydrostatic pressure is transduced to an intracellular signal through mechanosensitive membrane ion channels of chondrocytes. The goals of this study were to examine the effect of hydrostatic loading on the development of engineered cartilage tissue and the contribution of mechanosensitive ion channels on these hydrostatic loading effects.

Methods: Using a 3D model of porcine chondrocytes in agarose, we applied specific chemical inhibitors to determine the role of transient receptor potential (TRP) ion channels TRPV1, TRPV4, TRPC3, and TRPC1 in transducing hydrostatic pressure.

Results: Hydrostatic loading caused a frequency and magnitude-dependent decrease in sulfated glycosaminoglycans (S-GAG), without changes in DNA content. Inhibiting TRPC3 and TRPV4 decreased S-GAG content; however, only the inhibition of TRPV1 partially attenuated the hydrostatic loading-induced reduction in S-GAG content.

Conclusions: Our findings indicate that TRPV1 may serve as a transducer of hydrostatic pressure in chondrocytes, and provide further support for the role of TRPV4 in regulating chondrocyte anabolism, as well as initial evidence implicating TRPC3 in chondrogenesis. These findings add to our further understanding of the chondrocyte "channelome" and suggest that a range of ion channels mediate the transduction of different biophysical stimuli such as hydrostatic pressure, membrane stretch, or osmotic stress.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/03008207.2020.1871475DOI Listing
January 2021

Adipose tissue is a critical regulator of osteoarthritis.

Proc Natl Acad Sci U S A 2021 01;118(1)

Department of Orthopaedic Surgery, Washington University, St. Louis, MO 63110;

Osteoarthritis (OA), the leading cause of pain and disability worldwide, disproportionally affects individuals with obesity. The mechanisms by which obesity leads to the onset and progression of OA are unclear due to the complex interactions among the metabolic, biomechanical, and inflammatory factors that accompany increased adiposity. We used a murine preclinical model of lipodystrophy (LD) to examine the direct contribution of adipose tissue to OA. Knee joints of LD mice were protected from spontaneous or posttraumatic OA, on either a chow or high-fat diet, despite similar body weight and the presence of systemic inflammation. These findings indicate that adipose tissue itself plays a critical role in the pathophysiology of OA. Susceptibility to posttraumatic OA was reintroduced into LD mice using implantation of a small adipose tissue depot derived from wild-type animals or mouse embryonic fibroblasts that undergo spontaneous adipogenesis, implicating paracrine signaling from fat, rather than body weight, as a mediator of joint degeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2021096118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7817130PMC
January 2021

Single cell transcriptomic analysis of human pluripotent stem cell chondrogenesis.

Nat Commun 2021 01 13;12(1):362. Epub 2021 Jan 13.

Dept. of Orthopaedic Surgery, Washington University in Saint Louis, St. Louis, MO, 63110, USA.

The therapeutic application of human induced pluripotent stem cells (hiPSCs) for cartilage regeneration is largely hindered by the low yield of chondrocytes accompanied by unpredictable and heterogeneous off-target differentiation of cells during chondrogenesis. Here, we combine bulk RNA sequencing, single cell RNA sequencing, and bioinformatic analyses, including weighted gene co-expression analysis (WGCNA), to investigate the gene regulatory networks regulating hiPSC differentiation under chondrogenic conditions. We identify specific WNTs and MITF as hub genes governing the generation of off-target differentiation into neural cells and melanocytes during hiPSC chondrogenesis. With heterocellular signaling models, we further show that WNT signaling produced by off-target cells is responsible for inducing chondrocyte hypertrophy. By targeting WNTs and MITF, we eliminate these cell lineages, significantly enhancing the yield and homogeneity of hiPSC-derived chondrocytes. Collectively, our findings identify the trajectories and molecular mechanisms governing cell fate decision in hiPSC chondrogenesis, as well as dynamic transcriptome profiles orchestrating chondrocyte proliferation and differentiation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-20598-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806634PMC
January 2021

Formation of Osteochondral Organoids from Murine Induced Pluripotent Stem Cells.

Tissue Eng Part A 2020 Dec 22. Epub 2020 Dec 22.

Department of Orthopaedic Surgery, Washington University in St. Louis, St. Louis, Missouri, USA.

Osteoarthritis is a debilitating joint disease that is characterized by pathologic changes in both cartilage and bone, potentially involving cross talk between these tissues that is complicated by extraneous factors that are difficult to study . To create a model system of these cartilage-bone interactions, we developed an osteochondral organoid from murine induced pluripotent stem cells (iPSCs). Using this approach, we grew organoids from a single cell type through time-dependent sequential exposure of growth factors, namely transforming growth factor β-3 and bone morphogenic protein 2, to mirror bone development through endochondral ossification. The result is a cartilaginous region and a calcified bony region comprising an organoid with the potential for joint disease drug screening and investigation of genetic risk in a patient or disease-specific manner. Furthermore, we also investigated the possibility of the differentiated cells within the organoid to revert to a pluripotent state. It was found that while the cells themselves maintain the capacity for reinduction of pluripotency, encapsulation in the newly formed 3D matrix prevents this process from occurring, which could have implications for future clinical use of iPSCs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/ten.TEA.2020.0273DOI Listing
December 2020

Initial displacement of the intra-articular surface after articular fracture correlates with PTA in C57BL/6 mice but not "superhealer" MRL/MpJ mice.

J Orthop Res 2020 Nov 12. Epub 2020 Nov 12.

Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina, USA.

Posttraumatic arthritis (PTA) occurs commonly after articular fracture and may arise, in part, from joint surface incongruity after injury. MRL/MpJ (MRL) "super-healer" mice are protected from PTA compared to C57BL/6 (B6) mice following articular fracture. However, the relationship between the initial displacement of the articular surface, biologic response, and susceptibility to PTA after fracture remains unclear. The objective of this study was to assess whether joint incongruity after articular fracture, as measured by in vivo micro-computed tomography (microCT), could predict pathomechanisms of PTA in mice. B6 and MRL mice (n = 12/strain) received a closed articular fracture (fx) of the left tibial plateau. Articular incongruity was quantified as bone surface deviations (BSD) for each in vivo microCT scan obtained from pre-fx to 8 weeks post-fx, followed by histologic assessment of arthritis. Serum concentrations of bone formation (PINP) and bone resorption (CTX-I) biomarkers were quantified longitudinally. Both strains showed increases in surface incongruity over time, as measured by increases in BSD. In B6 mice, acute surface incongruity was significantly correlated to the severity of PTA (R  = 0.988; p = .0006), but not in MRL mice (R  = 0.224; p = .220). PINP concentrations significantly decreased immediately post-fx in B6 mice (p = .023) but not in MRL mice, indicating higher bone synthesis in MRL mice. MRL/MpJ mice demonstrate a unique biologic response to articular fracture such that the observed articular bone surface displacement does not correlate with the severity of subsequent PTA. Clinical Relevance: Identifying therapies to enhance acute biologic repair following articular fracture may mitigate the risk of articular surface displacement for PTA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jor.24912DOI Listing
November 2020

An immortalized human adipose-derived stem cell line with highly enhanced chondrogenic properties.

Biochem Biophys Res Commun 2020 09 6;530(1):252-258. Epub 2020 Aug 6.

Departments of Orthopaedic Surgery, Washington University, St. Louis, MO, USA; Department of Biomedical Engineering, Washington University, St. Louis, MO, USA; Shriners Hospitals for Children, St. Louis, MO, USA; Center of Regenerative Medicine, Washington University, St. Louis, MO, USA; Department of Mechanical Engineering and Materials Science, Washington University, St. Louis, MO, USA. Electronic address:

Human adipose-derived stem cells (ASCs) are a commonly used cell type for cartilage tissue engineering. However, donor-to-donor variability, cell heterogeneity, inconsistent chondrogenic potential, and limited expansion potential can hinder the use of these cells for modeling chondrogenesis, in vitro screening of drugs and treatments for joint diseases, or translational applications for tissue engineered cartilage repair. The goal of this study was to create an immortalized ASC line that showed enhanced and consistent chondrogenic potential for applications in cartilage tissue engineering as well as to provide a platform for investigation of biological and mechanobiological pathways involved in cartilage homeostasis and disease. Starting with the ASC52telo cell line, a hTERT-immortalized ASC line, we used lentivirus to overexpress SOX9, a master regulator of chondrogenesis, and screened several clonal populations of SOX9 overexpressing cells to form a new stable cell line with high chondrogenic potential. One clonal line, named ASC52telo-SOX9, displayed increased GAG and type II collagen synthesis and was found to be responsive to both mechanical and inflammatory stimuli in a manner similar to native chondrocytes. The development of a clonal line such as ASC52telo-SOX9 has the potential to be a powerful tool for studying cartilage homeostasis and disease mechanisms in vitro, and potentially as a platform for in vitro drug screening for diseases that affect articular cartilage. Our findings provide an approach for the development of other immortalized cell lines with improved chondrogenic capabilities in ASCs or other adult stem cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbrc.2020.07.016DOI Listing
September 2020

The miRNA-mRNA interactome of murine induced pluripotent stem cell-derived chondrocytes in response to inflammatory cytokines.

FASEB J 2020 09 7;34(9):11546-11561. Epub 2020 Aug 7.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA.

Osteoarthritis (OA) is a degenerative joint disease, and inflammation within an arthritic joint plays a critical role in disease progression. Pro-inflammatory cytokines, specifically IL-1 and TNF-α, induce aberrant expression of catabolic and degradative enzymes and inflammatory cytokines in OA and result in a challenging environment for cartilage repair and regeneration. MicroRNAs (miRNAS) are small noncoding RNAs and are important regulatory molecules that act by binding to target messenger RNAs (mRNAs) to reduce protein synthesis and have been implicated in many diseases, including OA. The goal of this study was to understand the mechanisms of miRNA regulation of the transcriptome of tissue-engineered cartilage in response to IL-1β and TNF-α using an in vitro murine induced pluripotent stem cell (miPSC) model system. We performed miRNA and mRNA sequencing to determine the temporal and dynamic responses of genes to specific inflammatory cytokines as well as miRNAs that are differentially expressed (DE) in response to both cytokines or exclusively to IL-1β or TNF-α. Through integration of mRNA and miRNA sequencing data, we created networks of miRNA-mRNA interactions which may be controlling the response to inflammatory cytokines. Within the networks, hub miRNAs, miR-29b-3p, miR-17-5p, and miR-20a-5p, were identified. As validation of these findings, we found that delivery of miR-17-5p and miR-20a-5p mimics significantly decreased degradative enzyme activity levels while also decreasing expression of inflammation-related genes in cytokine-treated cells. This study utilized an integrative approach to determine the miRNA interactome controlling the response to inflammatory cytokines and novel mediators of inflammation-driven degradation in tissue-engineered cartilage.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.202000889RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7725885PMC
September 2020

Is Obesity a Disease of Stem Cells?

Cell Stem Cell 2020 07;27(1):15-18

Department of Orthopaedic Surgery, Washington University in St. Louis, St. Louis, MO, USA; Shriners Hospitals for Children, St. Louis, MO, USA; Center of Regenerative Medicine, Washington University in St. Louis, St. Louis, MO, USA. Electronic address:

Obesity disrupts physiological homeostasis and alters both systemic and local microenvironments that impact stem cell plasticity and impair regenerative capacity. We present growing evidence that reveals the bidirectionality of obesity-induced stem cell dysfunction and how the molecular changes in stem cells residing in obese environments may accelerate disease severity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.stem.2020.04.019DOI Listing
July 2020

Transcriptomic analysis of bone and fibrous tissue morphogenesis during digit tip regeneration in the adult mouse.

FASEB J 2020 07 7;34(7):9740-9754. Epub 2020 Jun 7.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA.

Humans have limited regenerative potential of musculoskeletal tissues following limb or digit loss. The murine digit has been used to study mammalian regeneration, where stem/progenitor cells (the "blastema") completely regenerate the digit tip after distal, but not proximal, amputation. However, the molecular mechanisms responsible for this response remain to be determined. Here, we evaluated the spatiotemporal formation of bone and fibrous tissues after level-dependent amputation of the murine terminal phalanx and quantified the transcriptome of the repair tissue. Distal (regenerative) and proximal (non-regenerative) amputations showed significant differences in temporal gene expression and tissue regrowth over time. Genes that direct skeletal system development and limb morphogenesis are transiently upregulated during blastema formation and differentiation, including distal Hox genes. Overall, our results suggest that digit tip regeneration is controlled by a gene regulatory network that recapitulates aspects of limb development, and that failure to activate this developmental program results in fibrotic wound healing.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.202000330RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501216PMC
July 2020

Gene therapy for follistatin mitigates systemic metabolic inflammation and post-traumatic arthritis in high-fat diet-induced obesity.

Sci Adv 2020 May 8;6(19):eaaz7492. Epub 2020 May 8.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA.

Obesity-associated inflammation and loss of muscle function play critical roles in the development of osteoarthritis (OA); thus, therapies that target muscle tissue may provide novel approaches to restoring metabolic and biomechanical dysfunction associated with obesity. Follistatin (FST), a protein that binds myostatin and activin, may have the potential to enhance muscle formation while inhibiting inflammation. Here, we hypothesized that adeno-associated virus 9 (AAV9) delivery of FST enhances muscle formation and mitigates metabolic inflammation and knee OA caused by a high-fat diet in mice. AAV-mediated FST delivery exhibited decreased obesity-induced inflammatory adipokines and cytokines systemically and in the joint synovial fluid. Regardless of diet, mice receiving FST gene therapy were protected from post-traumatic OA and bone remodeling induced by joint injury. Together, these findings suggest that FST gene therapy may provide a multifactorial therapeutic approach for injury-induced OA and metabolic inflammation in obesity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/sciadv.aaz7492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7209997PMC
May 2020

Transgenic conversion of ω-6 to ω-3 polyunsaturated fatty acids via fat-1 reduces the severity of post-traumatic osteoarthritis.

Arthritis Res Ther 2020 04 15;22(1):83. Epub 2020 Apr 15.

Department of Orthopaedic Surgery, Washington University in St. Louis, Campus Box 8233, Couch Biomedical Research Bldg, Room 3121, St. Louis, MO, 63110, USA.

Background: Dietary fatty acid (FA) content has been shown to influence the development of post-traumatic osteoarthritis (PTOA) in obesity. We used the fat-1 transgenic mouse to examine the hypothesis that endogenous reduction of ω-6 to ω-3 FA ratio, under the same dietary conditions, would mitigate metabolic inflammation and the pathogenesis of PTOA in obese male and female mice.

Methods: Male and female fat-1 and wild-type littermates were fed either a control diet or an ω-6 FA-rich high-fat diet and underwent destabilization of the medial meniscus (DMM) surgery to induce PTOA. OA severity, synovitis, and osteophyte formation were determined histologically, while biomarker and lipidomic analyses were performed to evaluate levels of adipokines, insulin, pro-/anti-inflammatory cytokines, and FAs in serum and joint synovial fluid. Multivariable models were performed to elucidate the associations of dietary, metabolic, and mechanical factors with PTOA.

Results: We found that elevated serum levels of ω-3 FAs in fat-1 mice as compared to wild-type controls fed the same diet resulted in reduced OA and synovitis in a sex- and diet-dependent manner, despite comparable body weights. The fat-1 mice showed trends toward decreased serum pro-inflammatory cytokines and increased anti-inflammatory cytokines. Multivariable analysis for variables predicting OA severity in mice resulted in correlations with serum FA levels, but not with body weight.

Conclusions: This study provides further evidence that circulating FA composition and systemic metabolic inflammation, rather than body weight, may be the major risk factor for obesity-associated OA. We also demonstrate the potential genetic use of ω-3 FA desaturase in mitigating PTOA in obese patients following injury.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13075-020-02170-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7160898PMC
April 2020

Long non-coding RNA enhances chondrogenesis via suppression of the interferon type II signaling pathway.

Elife 2020 03 23;9. Epub 2020 Mar 23.

Department of Orthopaedic Surgery, Washington University, St Louis, United States.

The roles of long noncoding RNAs (lncRNAs) in musculoskeletal development, disease, and regeneration remain poorly understood. Here, we identified the novel lncRNA (originally named ) as a regulator of mesenchymal stem cell (MSC) chondrogenesis. , a primate-specific lncRNA, is upregulated during MSC chondrogenesis and appears to act directly downstream of SOX9, but not TGF-β3. We showed that the silencing of resulted in lower accumulation of cartilage-like extracellular matrix in a pellet assay, while overexpression - either via transgene ectopic expression or by endogenous activation via CRISPR-dCas9-VP64 - significantly enhanced cartilage matrix production. acts to inhibit IFN-γ by binding to EIF2AK2, and we further demonstrated that exhibits a protective effect in engineered cartilage against interferon type II. Our results indicate an important role of in regulating stem cell chondrogenesis, as well as its therapeutic potential in the treatment of cartilage-related diseases, such as osteoarthritis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7554/eLife.49558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7202894PMC
March 2020

Prospective isolation of chondroprogenitors from human iPSCs based on cell surface markers identified using a CRISPR-Cas9-generated reporter.

Stem Cell Res Ther 2020 02 18;11(1):66. Epub 2020 Feb 18.

Department of Orthopaedic Surgery, Washington University, St. Louis, MO, 63110, USA.

Background: Articular cartilage shows little or no capacity for intrinsic repair, generating a critical need of regenerative therapies for joint injuries and diseases such as osteoarthritis. Human-induced pluripotent stem cells (hiPSCs) offer a promising cell source for cartilage tissue engineering and in vitro human disease modeling; however, off-target differentiation remains a challenge during hiPSC chondrogenesis. Therefore, the objective of this study was to identify cell surface markers that define the true chondroprogenitor population and use these markers to purify iPSCs as a means of improving the homogeneity and efficiency of hiPSC chondrogenic differentiation.

Methods: We used a CRISPR-Cas9-edited COL2A1-GFP knock-in reporter hiPSC line, coupled with a surface marker screen, to identify a novel chondroprogenitor population. Single-cell RNA sequencing was then used to analyze the distinct clusters within the population. An unpaired t test with Welch's correction or an unpaired Kolmogorov-Smirnov test was performed with significance reported at a 95% confidence interval.

Results: Chondroprogenitors expressing CD146, CD166, and PDGFRβ, but not CD45, made up an average of 16.8% of the total population. Under chondrogenic culture conditions, these triple-positive chondroprogenitor cells demonstrated decreased heterogeneity as measured by single-cell RNA sequencing with fewer clusters (9 clusters in unsorted vs. 6 in sorted populations) closer together. Additionally, there was more robust and homogenous matrix production (unsorted: 1.5 ng/ng vs. sorted: 19.9 ng/ng sGAG/DNA; p < 0.001) with significantly higher chondrogenic gene expression (i.e., SOX9, COL2A1, ACAN; p < 0.05).

Conclusions: Overall, this study has identified a unique hiPSC-derived subpopulation of chondroprogenitors that are CD146/CD166/PDGFRβ/CD45 and exhibit high chondrogenic potential, providing a purified cell source for cartilage tissue engineering or disease modeling studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13287-020-01597-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7026983PMC
February 2020

Single cell RNA-sequencing reveals cellular heterogeneity and trajectories of lineage specification during murine embryonic limb development.

Matrix Biol 2020 07 24;89:1-10. Epub 2019 Dec 24.

Department of Orthopaedic Surgery, Washington University, 4515 McKinley Ave, St. Louis, MO, 63110, USA; Shriners Hospital for Children - St. Louis, 4400 Clayton Ave, St. Louis, MO, 63110, USA; Center of Regenerative Medicine, Washington University, St. Louis, MO, 63110, USA. Electronic address:

The coordinated spatial and temporal regulation of gene expression in the murine hindlimb determines the identity of mesenchymal progenitors and the development of diversity of musculoskeletal tissues they form. Hindlimb development has historically been studied with lineage tracing of individual genes selected a priori, or at the bulk tissue level, which does not allow for the determination of single cell transcriptional programs yielding mature cell types and tissues. To identify the cellular trajectories of lineage specification during limb bud development, we used single cell mRNA sequencing (scRNA-seq) to profile the developing murine hindlimb between embryonic days (E)11.5-E18.5. We found cell type heterogeneity at all time points, and the expected cell types that form the mouse hindlimb. In addition, we used RNA fluorescence in situ hybridization (FISH) to examine the spatial locations of cell types and cell trajectories to understand the ancestral continuum of cell maturation. This data provides a resource for the transcriptional program of hindlimb development that will support future studies of musculoskeletal development and generate hypotheses for tissue regeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.matbio.2019.12.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7282974PMC
July 2020

Nanotherapy Targeting NF-κB Attenuates Acute Pain After Joint Injury.

Precis Nanomed 2019 12;2(1):245-248. Epub 2019 Jan 12.

Department of Medicine, Washington University School of Medicine, St. Louis, MO.

Inflammation after joint injury leads to joint responses that result in eventual osteoarthritis development. Blockade of inflammation, by suppressing NF-κB expression, has been shown to reduce joint injury-induced chondrocyte apoptosis and reactive synovitis . Herein, we demonstrate that the suppression of NF-κB p65 expression also significantly mitigates the acute pain sensitivity induced by mechanical injury to the joint. These results suggest that early intervention with anti-NF-κB nanotherapy mitigates both structural and pain-related outcomes, which in turn may impact the progression of post-traumatic osteoarthritis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.33218/prnano2(1).181129.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6824724PMC
January 2019

Intergenerational Transmission of Diet-Induced Obesity, Metabolic Imbalance, and Osteoarthritis in Mice.

Arthritis Rheumatol 2020 04 5;72(4):632-644. Epub 2020 Mar 5.

Washington University in St. Louis and Shriners Hospitals for Children, St. Louis, Missouri.

Objective: Obesity and osteoarthritis (OA) are 2 major public health issues affecting millions of people worldwide. Whereas parental obesity affects the predisposition to diseases such as cancer or diabetes in children, transgenerational influences on musculoskeletal conditions such as OA are poorly understood. This study was undertaken to assess the intergenerational effects of a parental/grandparental high-fat diet on the metabolic and skeletal phenotype, systemic inflammation, and predisposition to OA in 2 generations of offspring in mice.

Methods: Metabolic phenotype and predisposition to OA were investigated in the first and second (F1 and F2) generations of offspring (n = 10-16 mice per sex per diet) bred from mice fed a high-fat diet (HFD) or a low-fat control diet. OA was induced by destabilizing the medial meniscus. OA, synovitis, and adipose tissue inflammation were determined histologically, while bone changes were measured using micro-computed tomography. Serum and synovial cytokines were measured by multiplex assay.

Results: Parental high-fat feeding showed an intergenerational effect, with inheritance of increased weight gain (up to 19% in the F1 generation and 9% in F2), metabolic imbalance, and injury-induced OA in at least 2 generations of mice, despite the fact that the offspring were fed the low-fat diet. Strikingly, both F1 and F2 female mice showed an increased predisposition to injury-induced OA (48% higher predisposition in F1 and 19% in F2 female mice fed the HFD) and developed bone microarchitectural changes that were attributable to parental and grandparental high-fat feeding.

Conclusion: The results of this study reveal a detrimental effect of parental HFD and obesity on the musculoskeletal integrity of 2 generations of offspring, indicating the importance of further investigation of these effects. An improved understanding of the mechanisms involved in the transmissibility of diet-induced changes through multiple generations may help in the development of future therapies that would target the effects of obesity on OA and related conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/art.41147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7113102PMC
April 2020

Optimization of Meniscus Cell Transduction Using Lentivirus and Adeno-Associated Virus for Gene Editing and Tissue Engineering Applications.

Cartilage 2019 Oct 14:1947603519880321. Epub 2019 Oct 14.

Department of Orthopaedic Surgery, Duke University School of Medicine, Durham, NC, USA.

Objectives: The utilization of viral vectors to deliver genes of interest directly to meniscus cells and promote long-term modulation of gene expression may prove useful to enhance meniscus repair and regeneration. The objective of this study was to optimize and compare the potential of lentivirus (LV) and adeno-associated virus (AAV) to deliver transgenes to meniscus cells in both intact meniscus tissue and isolated primary cells in monolayer.

Design: Porcine meniscus tissue explants and primary meniscus cells in monolayer were transduced with LV or self-complementary AAV2 (scAAV2) encoding green fluorescent protein (GFP). Following transduction, explants were enzymatically digested to isolate meniscus cells, and monolayer cells were trypsinized. Isolated cells were analyzed by flow cytometry to determine percent transduction.

Results: LV and scAAV2 showed a high transduction efficiency in monolayer meniscus cells. scAAV2 was most effective at transducing cells within intact meniscus tissue but the efficiency was less than 20%. Outer zone meniscus cells were more readily transduced by both LV and scAAV2 than the inner zone cells. Higher virus titers and higher cell density resulted in improved transduction efficiency. Polybrene was necessary for the highest transduction efficiency with LV, but it reduced scAAV2 transduction.

Conclusions: Both LV and scAAV2 efficiently transduce primary meniscus cells but only scAAV2 can modestly transduce cells embedded in meniscus tissue. This work lays the foundation for viral gene transfer to be utilized to deliver bioactive transgenes or gene editing machinery, which can induce long-term and tunable expression of therapeutic proteins from tissue-engineered constructs for meniscus repair and regeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1177/1947603519880321DOI Listing
October 2019

Combined Experimental Approach and Finite Element Modeling of Small Molecule Transport Through Joint Synovium to Measure Effective Diffusivity.

J Biomech Eng 2020 04;142(4)

Department of Biomedical Engineering, Washington University in St. Louis, Whitaker Hall, 1 Brookings Dr., St. Louis, MO 63130.

Trans-synovial solute transport plays a critical role in the clearance of intra-articularly (IA) delivered drugs. In this study, we present a computational finite element model (FEM) of solute transport through the synovium validated by experiments on synovial explants. Unsteady diffusion of urea, a small uncharged molecule, was measured through devitalized porcine and human synovium using custom-built diffusion chambers. A multiphasic computational model was constructed and optimized with the experimental data to extract effective diffusivity for urea within the synovium. A monotonic decrease in urea concentration was observed in the donor bath over time, with an effective diffusivity found to be an order of magnitude lower in synovium versus that measured in free solution. Parametric studies incorporating an intimal cell layer with varying thickness and varying effective diffusivities were performed, revealing a dependence of drug clearance kinetics on both parameters. The findings of this study indicate that the synovial matrix impedes urea solute transport out of the joint with little retention of the solute in the matrix.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1115/1.4044892DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7104772PMC
April 2020

Genome Engineering for Osteoarthritis: From Designer Cells to Disease-Modifying Drugs.

Tissue Eng Regen Med 2019 08 5;16(4):335-343. Epub 2019 Jan 5.

1Department of Orthopaedic Surgery, Washington University in St. Louis, 1 Brookings Dr, St. Louis, MO 63130 USA.

Background: Osteoarthritis (OA) is a highly prevalent degenerative joint disease involving joint cartilage and its surrounding tissues. OA is the leading cause of pain and disability worldwide. At present, there are no disease-modifying OA drugs, and the primary therapies include exercise and nonsteroidal anti-inflammatory drugs until total joint replacement at the end-stage of the disease.

Methods: In this review, we summarized the current state of knowledge in genetic and epigenetic associations and risk factors for OA and their potential diagnostic and therapeutic applications.

Results: Genome-wide association studies and analysis of epigenetic modifications (such as miRNA expression, DNA methylation and histone modifications) conducted across various populations support the notion that there is a genetic basis for certain subsets of OA pathogenesis.

Conclusion: With recent advances in the development of genome editing technologies such as the CRISPR-Cas9 system, these genetic and epigenetic alternations in OA can be used as platforms from which potential biomarkers for the diagnosis, prognosis, drug response, and development of potential personalized therapeutic targets for OA can be approached. Furthermore, genome editing has allowed the development of "designer" cells, whereby the receptors, gene regulatory networks, or transgenes can be modified as a basis for new cell-based therapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s13770-018-0172-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6675820PMC
August 2019

Stem cell-derived extracellular vesicles attenuate the early inflammatory response after tendon injury and repair.

J Orthop Res 2020 01 26;38(1):117-127. Epub 2019 Jul 26.

Department of Orthopaedic Surgery, Washington University School of Medicine, 660 South Euclid Avenue, Campus Box 8233, St. Louis, Missouri.

Adipose-derived stem cells (ASCs) have the potential to enhance tendon repair via paracrine regulation of the inflammatory response to injury. Extracellular vesicles (EVs), which are secreted by ASCs, have shown promise in mediating this process. This study was designed to evaluate the effect of ASC EVs on early tendon healing using a mouse Achilles tendon injury and repair model. EVs were isolated from the conditioned medium of naïve and interferonγ-primed ASCs and applied to the repair site via a collagen sheet. Tendon healing was assessed in nuclear factor-κB (NF-κB)-luciferase reporter mice up to 7 days after suture repair. As anticipated, repair site NF-κB activity increased greater than twofold following tendon repair. Treatment with EVs from primed but not naïve ASCs effectively suppressed the response. Accordingly, the pro-inflammatory genes Il1b and Ifng were both dramatically increased in repaired tendons, while primed, but not naïve ASC EVs attenuated the response. Compared with control repairs, primed ASC EVs further reduced the rate of post-repair tendon gap formation and rupture and facilitated collagen formation at the injury site. Additional experiments demonstrated that EVs target macrophages and that primed ASC EVs were most effective in blocking macrophage NF-κB activity. Collectively, the findings of this study demonstrate that primed ASC EVs, similar to ASCs, attenuate the early tendon inflammatory response after injury via modulation of the macrophage inflammatory response. Statement of clinical significance: These findings introduce a new cell-free therapy, derived from stem cells, for tendon repair with the potential for improved therapeutic efficacy and safety. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:117-127, 2020.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jor.24406DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6917960PMC
January 2020
-->