The Experts below are selected from a list of 1878 Experts worldwide ranked by ideXlab platform

R K W Smith - One of the best experts on this subject based on the ideXlab platform.

  • macrophage sub populations and the lipoxin a4 receptor implicate active inflammation during Equine Tendon repair
    PLOS ONE, 2012
    Co-Authors: Stephanie G Dakin, R K W Smith, Dirk Werling, Andrew P Hibbert, D R E Abayasekara, Natalie Jayne Young, Jayesh Dudhia
    Abstract:

    Macrophages (Mϕ) orchestrate inflammatory and reparatory processes in injured connective tissues but their role during different phases of Tendon healing is not known. We investigated the contribution of different Mϕ subsets in an Equine model of naturally occurring Tendon injury. Post mortem tissues were harvested from normal (uninjured), sub-acute (3–6 weeks post injury) and chronically injured (>3 months post injury) superficial digital flexor Tendons. To determine if inflammation was present in injured Tendons, Mϕ sub-populations were quantified based on surface antigen expression of CD172a (pan Mϕ), CD14highCD206low (pro-inflammatory M1Mϕ), and CD206high (anti-inflammatory M2Mϕ) to assess potential polarised phenotypes. In addition, the Lipoxin A4 receptor (FPR2/ALX) was used as marker for resolving inflammation. Normal Tendons were negative for both Mϕ and FPR2/ALX. In contrast, M1Mϕ predominated in sub-acute injury, whereas a potential phenotype-switch to M2Mϕ polarity was seen in chronic injury. Furthermore, FPR2/ALX expression by tenocytes was significantly upregulated in sub-acute but not chronic injury. Expression of the FPR2/ALX ligand Annexin A1 was also significantly increased in sub-acute and chronic injuries in contrast to low level expression in normal Tendons. The combination of reduced FPR2/ALX expression and persistence of the M2Mϕ phenotype in chronic injury suggests a potential mechanism for incomplete resolution of inflammation after Tendon injury. To investigate the effect of pro-inflammatory mediators on lipoxin A4 (LXA4) production and FPR2/ALX expression in vitro, normal Tendon explants were stimulated with interleukin-1 beta and prostaglandin E2. Stimulation with either mediator induced LXA4 release and maximal upregulation of FPR2/ALX expression after 72 hours. Taken together, our data suggests that although tenocytes are capable of mounting a protective mechanism to counteract inflammatory stimuli, this appears to be of insufficient duration and magnitude in natural Tendon injury, which may potentiate chronic inflammation and fibrotic repair, as indicated by the presence of M2Mϕ.

  • implantation of bone marrow derived mesenchymal stem cells demonstrates improved outcome in horses with overstrain injury of the superficial digital flexor Tendon
    Equine Veterinary Journal, 2012
    Co-Authors: E E Godwin, N J Young, Jayesh Dudhia, I C Beamish, R K W Smith
    Abstract:

    Summary Reasons for performing study: Mesenchymal stem (progenitor; stromal) cell (MSC) therapy has gained popularity for the treatment of Equine Tendon injuries but without reports of long-term follow-up. Objectives: To evaluate the safety and reinjury rate of racehorses after intralesional MSC injection in a large study of naturally occurring superficial digital flexor tendinopathy and to compare these data with those published for other treatments. Methods: Safety was assessed clinically, ultrasonographically, scintigraphically and histologically in a cohort of treated cases: 141 client-owned treated racehorses followed-up for a minimum of 2 years after return to full work. Reinjury percentages were compared to 2 published studies of other treatments with similar selection criteria and follow-up. The number of race starts, discipline, age, number of MSCs injected and interval between injury and treatment were analysed. Results: There were no adverse effects of the treatment with no aberrant tissue on histological examination. The reinjury percentage of all racehorses with follow-up (n = 113) undergoing MSC treatment was 27.4%, with the rate for flat (n = 8) and National Hunt (n = 105) racehorses being 50 and 25.7%, respectively. This was significantly less than published for National Hunt racehorses treated in other ways. No relationship between outcome and age, discipline, number of MSCs injected or injury to implantation interval was found. Conclusions: Whilst recognising the limitations of historical controls, this study has shown that MPC implantation is safe and appears to reduce the reinjury rate after superficial digital flexor tendinopathy, especially in National Hunt racehorses. Potential relevance: This study has provided evidence for the long-term efficacy of MSC treatment for tendinopathy in racehorses and provides support for translation to human Tendon injuries.

  • correlation of cartilage oligomeric matrix protein comp levels in Equine Tendon with mechanical properties a proposed role for comp in determining function specific mechanical characteristics of locomotor Tendons
    Equine Veterinary Journal, 2010
    Co-Authors: R K W Smith, Helen L Birch, M Gerard, B A Dowling, A J Dart, A E Goodship
    Abstract:

    Summary Over-strain injury of the superficial digital flexor Tendon (SDFT) is a common injury in the horse. Tendon appears to adapt to loads placed on it during development, but fatigue damage accumulates after skeletal maturity, which is inadequately repaired and predisposes to clinical tendinitis. In any population of horses, there is a wide variation in SDFT mechanical properties. A noncollagenous protein, cartilage oligomeric matrix protein (COMP), is particularly abundant during growth in the Equine SDFT and has been proposed to have an organisational role in the formation of collagenous matrices. This study aimed to determine whether COMP levels were correlated to mechanical properties at skeletal maturity. Tendons from 2 groups of 12 horses were analysed: Group 1 horses with restricted age, 2 years ± 2 months, showed a significant correlation between both ultimate tensile stress modulus of elasticity and stiffness and COMP, while Group 2 mature horses with varying age did not, because of age- and exercise-induced loss of COMP. These data supports the hypothesis that COMP is an important mediator in the growth of Tendon. This data would suggest that the identification of low COMP levels in Tendon during growth would indicate horses prone to Tendon injury and methods of promoting COMP synthesis during growth would potentially improve Tendon quality and reduce the risk of subsequent tendinitis.

  • aging enhances a mechanically induced reduction in Tendon strength by an active process involving matrix metalloproteinase activity
    Aging Cell, 2007
    Co-Authors: Jayesh Dudhia, Dick Heinegård, Charlotte M Scott, Edward R C Draper, A A Pitsillides, R K W Smith
    Abstract:

    Age-associated and degenerative loss of functional integrity in soft tissues develops from effects of cumulative and subtle changes in their extracellular matrix (ECM). The highly ordered Tendon ECM provides the tissue with its tensile strength during loading. As age and exercise collude in the high incidence of tendinopathies, we hypothesized that aged Tendons fail due to cumulative damage resulting from a combination of diminished matrix repair and fragmentation of ECM proteins induced by prolonged cyclical loading, and that this is an active cell-mediated process. We developed an Equine Tendon explant model to examine the effect of age on the influence of prolonged cyclical loading at physiologically relevant strain rates (5% strain, 1 Hz for 24 h) on tissue mechanical properties, loss of ECM protein and matrix metalloproteinase (MMP) expression. We show significantly diminished mechanical strength of cyclically loaded tissue compared to controls (39.7 +/- 12%, P <= 0.05) this reduction was dependent on the presence of both viable cells and metalloproteinase activity. Furthermore, Tendon from older specimens was more susceptible to weakening (11-30 years, 50% P <= 0.05) compared to immature and young mature tissue (1-3 years, 34%; 4-10 years, 35%, respectively). Cyclical load also induced release of degraded cartilage oligomeric matrix protein, an integral ECM protein, an effect that could be mimicked by culture with fibronectin fragments. These findings indicate prolonged cyclical loading of physiological magnitude decreases Tendon tensile strength by an active process, and that MMPs may contribute to loss of functional competence, exaggerated by age, via load-induced proteolytic disruption of the ECM.

  • the distribution of cartilage oligomeric matrix protein comp in Tendon and its variation with Tendon site age and load
    Matrix Biology, 1997
    Co-Authors: R K W Smith, L Zunino, P M Webbon, Dick Heinegård
    Abstract:

    A protein prominent in guanidine hydrochloride extracts of adult bovine and Equine digital flexor Tendons was confirmed to be Cartilage Oligomeric Matrix Protein (COMP) by non-reducing and reducing SDS-PAGE, reaction with rabbit anti-COMP polyclonal antiserum on Western blots, trypsin digestion followed by HPLC on a C2/C18 column, and identification of COMP mRNA from Tendon on Northern blots. Immunohistochemistry and Western blots of extracts showed COMP to be present in all regions of digital flexor Tendons. Equine Tendon COMP was purified by ion exchange chromatography and gel filtration and used in a heterologous inhibition ELISA to quantify COMP in Equine digital flexor Tendons at different ages, and in other Tendons and ligaments. Mean COMP levels in digital flexor Tendon were approximately 2-5mg/g wet weight, but they showed a large variation. Levels were low in neonatal Tendon but rose rapidly during growth, with the metacarpal (tensional) superficial digital flexor Tendon having the highest levels (approximately 10mg/g wet weight). Levels subsequently declined in this region, while in areas which experience a variable amount of compression, levels increased less but then remained constant. Extensor Tendons and collateral ligaments, which experience less loading in vivo, had levels similar to those in neonatal Tendon. COMP was identified in scarred skin and granulation tissue but not in normal skin, chronic fibrosis, or a fibrosarcomatous skin growth. A unilateral non-weight-bearing growing animal contained three to six times more COMP in the weight-bearing digital flexor Tendons compared to the paralyzed limb, while the extensor Tendons had similar amounts in both limbs. With the recent discovery of a COMP gene mutation causing pseudoachondroplasia (Hecht et al., 1995), in which lax Tendons and ligaments are a feature, the present data suggest that COMP is synthesized in response to, and is necessary for Tendon to resist, load.

Peter M. Stadler - One of the best experts on this subject based on the ideXlab platform.

  • Tracking of autologous adipose tissue-derived mesenchymal stromal cells with in vivo magnetic resonance imaging and histology after intralesional treatment of artificial Equine Tendon lesions--a pilot study.
    Stem Cell Research & Therapy, 2016
    Co-Authors: Florian Geburek, Kathrin Mundle, Sabine Conrad, Maren Hellige, Ulrich Walliser, Hans T. M. Van Schie, René Van Weeren, Thomas Skutella, Peter M. Stadler
    Abstract:

    Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat Equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into Equine superficial digital flexor Tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology. Four adult warmblood horses received a unilateral injection of 10 × 106 autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment. AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated Tendons contained high numbers of SPIO- and GFP-labelled cells. Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created Equine SDFT lesions. Intralesional injection of 10 × 106 AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created Tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing Tendon tissue is an essential pathway after intralesional administration. Injection techniques have to be chosen deliberately to avoid reflux of the cell substrate injected. In vivo low-field MRI may be used as a non-invasive tool to monitor homing and engraftment of AT-MSCs in horses with tendinopathy of the SDFT.

  • Tracking of autologous adipose tissue-derived mesenchymal stromal cells with in vivo magnetic resonance imaging and histology after intralesional treatment of artificial Equine Tendon lesions - a pilot study
    Stem Cell Research & Therapy, 2016
    Co-Authors: Florian Geburek, Kathrin Mundle, Sabine Conrad, Maren Hellige, Ulrich Walliser, Thomas Skutella, Hans T. M. Van Schie, René Van Weeren, Peter M. Stadler
    Abstract:

    Background Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat Equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into Equine superficial digital flexor Tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology. Methods Four adult warmblood horses received a unilateral injection of 10 × 10^6 autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment. Results AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated Tendons contained high numbers of SPIO- and GFP-labelled cells. Conclusions Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created Equine SDFT lesions. Intralesional injection of 10 × 10^6 AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created Tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing Tendon tissue is an essential pathway after intralesional administration. Injection techniques have to be chosen deliberately to avoid reflux of the cell substrate injected. In vivo low-field MRI may be used as a non-invasive tool to monitor homing and engraftment of AT-MSCs in horses with tendinopathy of the SDFT.

P D Clegg - One of the best experts on this subject based on the ideXlab platform.

  • Insulin-like growth factor binding protein (IGFBP6) is a cross-species Tendon marker.
    'European Cells and Materials', 2019
    Co-Authors: Aj Turlo, Aj Mueller-breckenridge, De Zamboulis, Eg Canty-laird, P D Clegg
    Abstract:

    The main challenge in Tendon injury management is suboptimal tissue healing that fails to re-establish original Tendon function. Tissue bioengineering is a promising approach for Tendon therapy, with potential to improve its functional outcomes. However, evaluation criteria for tissue-engineered Tendon are unclear due to the lack of specific markers of differentiated Tendon. The study aim was to identify a panel of genes that characterised Tendons in comparison to cartilage or muscles and validate those genes, both in human and key species used as models for Tendon diseases. Gene expression profiling of rat Tendon and cartilage in whole-tissue samples and primary tenocytes and chondrocytes was undertaken using two independent microarray platforms. Genes that demonstrated high expression correlation across two assays were validated by qRT-PCR in rat Tendon relative to cartilage and muscle. Five genes demonstrating the highest Tendon-related expression in the validation experiment (ASPN, ECM1, IGFBP6, TNMD, THBS4) were further evaluated by qRT-PCR in ovine, Equine and human tissue. The group of Tendon markers, identified by unbiased transcriptomic analysis of rat musculoskeletal tissues, demonstrated species-dependent profiles of expression. Insulin-like growth factor binding protein 6 (IGFBP6) was identified as the only universal Tendon marker. Further investigation in Equine Tendon showed that IGFBP6 expression was not affected by ageing or Tendon function but decreased in anatomical regions subjected to elevated compressive force. IGFBP6 is a robust cross-species marker of Tendon phenotype and may find application in evaluation of Tendon physiology and guided differentiation of permissive cells towards functional tenocytes

  • the stem cell niche in Tendon and ligament investigating alterations with ageing and disease
    Orthopaedic Proceedings, 2018
    Co-Authors: Katie Joanna Lee, P D Clegg, Comerford Ejv, E G Cantylaird
    Abstract:

    IntroductionTendon is prone to degeneration through ageing and injury and current therapies are largely ineffective. The recent identification of a cell population within Tendon with stem cell-like characteristics holds potential for regeneration of Tendon. The local stem cell environment (niche) is important for stem cell maintenance and function. This study aims to characterize extracellular matrix (ECM) components of the stem cell niche in Equine Tendon, which is prone to age-related degeneration and rupture.Materials and MethodsPutative Tendon stem cells (TSCs) were isolated from Equine superficial digital flexor Tendon by low-density plating and differential adhesion to fibronectin. Cells were analysed by flow cytometry using antibodies to mesenchymal stem cell markers, as well as qRT-PCR for stem cell and tenogenic markers. The multipotency of cells was assessed using tri-lineage differentiation assays. ECM components of the tenocyte and TSC niche were analysed using radio-isotope labelling, immunoh...

  • identification of equid herpesvirus 2 in tissue engineered Equine Tendon
    Wellcome Open Research, 2017
    Co-Authors: Roisin Wardle, P D Clegg, Jane A Pullman, Sam Haldenby, Lorenzo Ressel, Marion Pope, Alan D Radford, James P Stewart, Mohammed Alsaadi, Philip Dyer
    Abstract:

    Background: Incidental findings of virus-like particles were identified following electron microscopy of tissue-engineered Tendon constructs (TETC) derived from Equine tenocytes. We set out to determine the nature of these particles, as there are few studies which identify virus in Tendons per se, and their presence could have implications for tissue-engineering using allogenic grafts. Methods: Virus particles were identified in electron microscopy of TETCs. Virion morphology was used to initially hypothesise the virus identity.  Next generation sequencing was implemented to identify the virus. A pan herpesvirus PCR was used to validate the RNASeq findings using an independent platform. Histological analysis and biochemical analysis was undertaken on the TETCs. Results: Morphological features suggested the virus to be either a retrovirus or herpesvirus. Subsequent next generation sequencing mapped reads to Equid herpesvirus 2 (EHV2). Histological examination and biochemical testing for collagen content revealed no significant differences between virally affected TETCs and non-affected TETCs. An independent set of Equine superficial digital flexor Tendon tissue (n=10) examined using designed primers for specific EHV2 contigs identified at sequencing were negative. These data suggest that EHV is resident in some Equine Tendon. Conclusions: EHV2 was demonstrated in Equine tenocytes for the first time; likely from in vivo infection. The presence of EHV2 could have implications to both tissue-engineering and tendinopathy.

  • the stem cell niche in Tendon and ligament investigating alterations with ageing and disease
    Journal of Bone and Joint Surgery-british Volume, 2015
    Co-Authors: Katie Joanna Lee, P D Clegg, Ejv Comerford, E G Cantylaird
    Abstract:

    Introduction Tendon is prone to degeneration through ageing and injury and current therapies are largely ineffective. The recent identification of a cell population within Tendon with stem cell-like characteristics holds potential for regeneration of Tendon. The local stem cell environment (niche) is important for stem cell maintenance and function. This study aims to characterize extracellular matrix (ECM) components of the stem cell niche in Equine Tendon, which is prone to age-related degeneration and rupture. Materials and Methods Putative Tendon stem cells (TSCs) were isolated from Equine superficial digital flexor Tendon by low-density plating and differential adhesion to fibronectin. Cells were analysed by flow cytometry using antibodies to mesenchymal stem cell markers, as well as qRT-PCR for stem cell and tenogenic markers. The multipotency of cells was assessed using tri-lineage differentiation assays. ECM components of the tenocyte and TSC niche were analysed using radio-isotope labelling, immunohistochemistry and histology. Results Putative TSCs were able to form colonies, and both tenocytes and TSCs expressed CD90, CD105 and CD73 as determined by flow cytometry. However, TSCs did not exhibit increased expression of stem cell marker genes when compared with tenocytes. TSCs and tenocytes both displayed osteogenic and chondrogenic differentiation, however not adipogenic differentiation. Tenocytes and TSCs labelled with 14C-labelled amino acids both displayed similar labelling profiles. Histological analysis of Tendon tissue highlighted the varied structure and composition of Tendon, with tenascin C expression confined to the interfascicular matrix. Discussion TSCs do not highly express stem cell markers when compared with tenocytes, indicating that these cells may not be true stem cells. In addition the similar labelling profiles of the two cell types indicates that a stem cell population has not been differentially isolated, however the tri-lineage differentiation assays suggest the cells may possess some stem cell-like properties. It is possible that the Equine Tendon cell population consists of a heterogeneous mixture of cells at different stages of differentiation.

  • 54 the stem cell niche in Tendon and ligament investigating alterations with ageing and disease
    British Journal of Sports Medicine, 2014
    Co-Authors: Katie Lee, P D Clegg, Ejv Comerford, Kate Williamson, E G Cantylaird
    Abstract:

    Introduction Ligament and Tendon are prone to degeneration through ageing and injury and current therapies are largely ineffective. The identification of a cell population within Tendon with stem cell-like characteristics (Bi, 2007) holds potential for regeneration of Tendon and ligament. Tendon stem cells differentiate into tenocytes (Zhang, 2010); the predominant cell type within Tendon, responsible for producing extracellular matrix (ECM). The local stem cell environment (niche) is vital for stem cell maintenance and function in many tissues, and tenascin C in particular has been shown to play an important role within stem cell niches (Garcion, 2004). Tendon and ligament are composed of fascicles and interfascicular matrix (IFM) which vary considerably in composition providing definitive niches within the tissue. This study aims to characterise ECM components of the stem cell niche in Equine Tendon and canine ligament, which are prone to age-related degeneration. The goal of this research is to produce an in vitro environment for stem cells which mimics the stem cell niche, for treatment of Tendon and ligament disease. Methods Putative stem cells were isolated from Equine superficial digital flexor Tendon (SDFT) and canine anterior cruciate ligament (ACL) by low-density plating and differential adhesion to plastic and fibronectin substrates. Cells were analysed by flow cytometry using antibodies to mesenchymal stem cell markers CD90, CD73 and CD105, as well as qRT-PCR for stem cell and tenogenic markers. ECM components of the fibroblast and stem cell niche were analysed using radioisotope labelling. Cells were labelled with 14 C-labelled amino acids to specifically label newly synthesised collagenous (proline) and non-collagenous (lysine/arginine) ECM, prior to extraction of ECM. Immuno-histochemistry and histology were conducted to analyse the structure and composition of SDFT. Results Tendon and ligament cells formed colonies after low-density plating, however only ligament cells formed colonies after differential adhesion to fibronectin. A subpopulation of Tendon cells expressed CD90 in both freshly isolated cells and putative stem cells, but were CD105 and CD73 negative. Putative Tendon stem cells, isolated by differential fibronectin adhesion did not exhibit increased expression of stem cell markers when compared with tenocytes. However there was a significant increase in expression of stem cell markers in putative ligament stem cells compared with ligamentocytes. Tenocytes and putative Tendon stem cells (isolated by low-density plating) labelled with 14 C-labelled amino acids both displayed similar labelling profiles. Histological analysis of SDFT tissue highlighted the varied structure and composition of Tendon, with tenascin C expression confined to IFM (see Figure.1). Conclusion The absence of stem cell marker expression in putative stem cell populations indicates that further testing of stem cell isolation procedures is required. Published techniques for Tendon stem cell isolation in humans and other mammals do not appear to be effective for isolation of Equine Tendon stem cells. Alternatively it is possible that the Equine Tendon cell population consists of a heterogenous mixture of cells at different stages of differentiation. We are currently optimising stem cell isolation techniques and conducting tri-lineage differentiation assays. Tendon stem cells in other species show tri-lineage differentiation, however it is possible that Equine Tendon stem cells may be restricted to tenogenic differentiation. Future experiments aim to identify ECM components of the stem cell niche by mass spectrometry and microarray comparison of Tendon and ligament tissue, stem cells and fibroblasts. References Bi et al . Nat Med. 2007;13: 1219–1227 Garcion et al . Development. 131:3423–3432 Zhang et al . Am J Sport Med. 38:2477–2486

Deborah J. Guest - One of the best experts on this subject based on the ideXlab platform.

  • Equine mesenchymal stromal cells and embryo derived stem cells are immune privileged in vitro
    Stem Cell Research & Therapy, 2014
    Co-Authors: Yasmin Z Paterson, N Rash, Elaine R Garvican, R Paillot, Deborah J. Guest
    Abstract:

    Autologous mesenchymal stem cells (MSCs) are an attractive concept in regenerative medicine, but their mechanism of action remains poorly defined. No immune response is reported after in vivo injection of allogeneic Equine MSCs or embryo-derived stem cells (ESCs) into the Equine Tendon, which may be due to the cells’ immune-privileged properties. This study further investigates these properties to determine their potential for clinical application in other tissues. Mitomycin C-treated MSCs, ESCs, or differentiated ESCs (dESCs) were cultured with allogeneic Equine peripheral blood mononuclear cells (PBMCs), and their effect on PBMC proliferation, in the presence or absence of interferon-gamma (IFN-γ) was determined. MSCs and super-antigen (sAg)-stimulated PBMCs were co-cultured directly or indirectly in transwells, and PBMC proliferation examined. Media from MSC culture were harvested and used for PBMC culture; subsequent PBMC proliferation and gene expression were evaluated and media assayed for IFN-γ, tumor necrosis factor alpha (TNF-α), and interleukin (IL)-10 and IL-6 proteins with enzyme-linked immunosorbent assay (ELISA). Co-culture of PBMCs with ESCs or dESCs did not affect baseline proliferation, whereas co-culture with MSCs significantly suppressed baseline proliferation. Stimulation of PBMC proliferation by using super-antigens (sAgs) was also suppressed by co-culture with MSCs. Inhibition was greatest with direct contact, but significant inhibition was produced in transwell culture and by using MSC-conditioned media, suggesting that soluble factors play a role in MSC-mediated immune suppression. The MSCs constitutively secrete IL-6, even in the absence of co-culture with PBMCs. MSC-conditioned media also brought about a change in the cytokine-expression profile of sAg-stimulated PBMCs, significantly reducing PBMC expression of IL-6, IFN-γ, and TNF-α. Equine MSCs and ESCs possess a degree of innate immune privilege, and MSCs secrete soluble factors that suppress PBMC proliferation and alter cytokine expression. These properties may make possible the future clinical use of allogeneic stem cells to help standardize and broaden the scope of treatment of tissue injuries.

  • Equine mesenchymal stromal cells and embryo derived stem cells are immune privileged in vitro
    Stem Cell Research & Therapy, 2014
    Co-Authors: Yasmin Z Paterson, N Rash, Elaine R Garvican, R Paillot, Deborah J. Guest
    Abstract:

    Introduction Autologous mesenchymal stem cells (MSCs) are an attractive concept in regenerative medicine, but their mechanism of action remains poorly defined. No immune response is reported after in vivo injection of allogeneic Equine MSCs or embryo-derived stem cells (ESCs) into the Equine Tendon, which may be due to the cells’ immune-privileged properties. This study further investigates these properties to determine their potential for clinical application in other tissues.

  • transforming growth factor beta3 promotes Tendon differentiation of Equine embryo derived stem cells
    Tissue Engineering Part A, 2013
    Co-Authors: Tom Barsby, Deborah J. Guest
    Abstract:

    Tendon injuries occur frequently in horses and have a poor capacity to regenerate, which leads to high re-injury rates. Equine embryo-derived stem cells (ESCs) survive in high numbers in the injured horse Tendon and we hypothesized that they differentiate into tenocytes in vivo. Immunocytochemistry revealed that in the injured horse Tendon ESCs express the Tendon progenitor marker scleraxis and that there is a local upregulation of the transforming growth factor-β (TGF-β) at the injury site. The aim of this study was to determine if TGF-β signaling was able to drive tenocyte differentiation by ESCs. Exposure of differentiating ESCs to TGF-β in vitro produced an upregulation of scleraxis at the gene and protein level with the greatest effect being produced in the presence of TGF-β3. TGF-β3 treatment of differentiating ESCs also promotes a significant upregulation of other Tendon-associated genes and proteins suggesting it can promote ESC differentiation into tenocytes. Our results demonstrate that Equine ESCs can differentiate into a therapeutically relevant cell type and that TGF-β driven differentiation of ESCs may provide a model to study Tendon development and better understand the transcriptional networks that are involved in Equine Tendon cell differentiation from the early embryonic stages.

Stephen J. Matcher - One of the best experts on this subject based on the ideXlab platform.

  • Absolute fast axis determination using non-polarization-maintaining fiber-based polarization-sensitive optical coherence tomography
    Optics letters, 2012
    Co-Authors: Stephen J. Matcher
    Abstract:

    We report on a new calibration technique that permits the accurate extraction of sample Jones matrix and hence fast-axis orientation by using fiber-based polarization-sensitive optical coherence tomography (PS-OCT) that is completely based on non-polarization-maintaining fiber such as SMF-28. In this technique, two quarter-wave plates (QWPs) are used to completely specify the parameters of the system fibers in the sample arm so that the Jones matrix of the sample can be determined directly. The device was validated on measurements of a QWP and an Equine Tendon sample by a single-mode fiber-based swept-source PS-OCT system.

  • optic axis determination by fibre based polarization sensitive swept source optical coherence tomography
    Physics in Medicine and Biology, 2011
    Co-Authors: Deepa K Kasaragod, Stephen J. Matcher
    Abstract:

    We describe a fibre-based variable-incidence angle (VIA) polarization-sensitive swept-source optical coherence tomography (PS-SS-OCT) system to determine the 3D optical axis of birefringent biological tissues. Single-plane VIA-PS-OCT is also explored which requires measurement of the absolute fast-axis orientation. A state-of-the-art PS-SS-OCT system with some improvements both in hardware and software was used to determine the apparent optical birefringence of Equine Tendon for a number of different illumination directions. Polar and azimuthal angles of cut Equine Tendon were produced by the VIA method and compared with the nominal values. A quarter waveplate (QWP) and Equine Tendon were used as test targets to validate the fast-axis measurements using the system. Polar and azimuthal angles of cut Equine Tendon broadly agreed with the expected values within about 8% of the nominal values. A theoretical and experimental analysis of the effect of the sample arm fibre on determination of optical axis orientation using a proposed definition based on the orientation of the eigenpolarization ellipse experimentally confirms that this algorithm only works correctly for special settings of the sample arm fibre. A proposed algorithm based on the angle between Stokes vectors on the Poincare sphere is confirmed to work for all settings of the sample arm fibre. A calibration procedure is proposed to remove the sign ambiguity of the measured orientation and was confirmed experimentally by using the QWP.

  • the collagen structure of bovine intervertebral disc studied using polarization sensitive optical coherence tomography
    Physics in Medicine and Biology, 2004
    Co-Authors: Stephen J. Matcher, Peter C Winlove, Sergei V Gangnus
    Abstract:

    Polarization-sensitive optical coherence tomography (PS-OCT) is used to measure the birefringence properties of bovine intervertebral disc and Equine flexor Tendon. For Equine Tendon the birefringence ?n is (6.0 ? 0.2) ? 10?3 at a wavelength of 1.3 ?m. This is somewhat larger than the values reported for bovine Tendon. The surface region of the annulus fibrosus of a freshly excised intact bovine intervertebral disc displays an identical value of birefringence, ?n = (6.0 ? 0.6) ? 10?3 at 1.3 ?m. The nucleus pulposus does not display birefringence, the measured apparent value of ?n = (0.39 ? 0.01) ? 10?3 being indistinguishable from the effects of depolarization due to multiple scattering. A clear difference is found between the depth-resolved retardance of Equine Tendon and that of bovine intervertebral disc. This apparently relates to the lamellar structure of the latter tissue, in which the collagen fibre orientation alternates between successive lamellae. A semi-empirical model based on Jones calculus shows that the measurements are in reasonable agreement with previous optical and x-ray data. These results imply that PS-OCT could be a useful tool to study collagen organization within the intervertebral disc in vitro and possibly in vivo and its variation with applied load and disease.