Publications by authors named "Moeed Akbar"

19 Publications

  • Page 1 of 1

Translational targeting of inflammation and fibrosis in frozen shoulder: Molecular dissection of the T cell/IL-17A axis.

Proc Natl Acad Sci U S A 2021 09;118(39)

Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences,University of Glasgow, Glasgow G12 8QQ, United Kingdom;

Frozen shoulder is a common fibroproliferative disease characterized by the insidious onset of pain and restricted range of shoulder movement with a significant socioeconomic impact. The pathophysiological mechanisms responsible for chronic inflammation and matrix remodeling in this prevalent fibrotic disorder remain unclear; however, increasing evidence implicates dysregulated immunobiology. IL-17A is a key cytokine associated with inflammation and tissue remodeling in numerous musculoskeletal diseases, and thus, we sought to determine the role of IL-17A in the immunopathogenesis of frozen shoulder. We demonstrate an immune cell landscape that switches from a predominantly macrophage population in nondiseased tissue to a T cell-rich environment in disease. Furthermore, we observed a subpopulation of IL-17A-producing T cells capable of inducing profibrotic and inflammatory responses in diseased fibroblasts through enhanced expression of the signaling receptor IL-17RA, rendering diseased cells more sensitive to IL-17A. We further established that the effects of IL-17A on diseased fibroblasts was TRAF-6/NF-κB dependent and could be inhibited by treatment with an IKKβ inhibitor or anti-IL-17A antibody. Accordingly, targeting of the IL-17A pathway may provide future therapeutic approaches to the management of this common, debilitating disease.
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http://dx.doi.org/10.1073/pnas.2102715118DOI Listing
September 2021

Targeting the CCR6/CCL20 axis in entheseal and cutaneous inflammation.

Arthritis Rheumatol 2021 Jun 3. Epub 2021 Jun 3.

Department of Dermatology, University of California, Davis, Sacramento, CA, USA.

Objectives: To assess the involvement of the CCR6/CCL20 axis in psoriatic arthritis (PsA) and psoriasis (PsO) and to evaluate its potential as a therapeutic target.

Methods: First, we quantified CCL20 levels in peripheral blood and synovial fluid of PsA patients and the presence of CCR6 cells in synovial and tendon tissue. Utilizing an IL-23 minicircle DNA (MC) mouse model exhibiting key features of both PsO and PsA, we investigated CCR6 and CCL20 expression and the preventive and therapeutical effect of CCL20 blockade. Healthy tendon stromal cells were stimulated in vitro with IL-1β to assess the production of CCL20 by qPCR and ELISA. The effect of conditioned media from stimulated tenocytes in inducing T cell migration was interrogated with a transwell system.

Results: We observed an upregulation of both CCR6 and CCL20 in the enthesis of IL-23 MC-treated mice, which was confirmed in human biopsies. Specific targeting of the CCR6/CCL20 axis with a CCL20 locked dimer (CCL20LD) blocked entheseal inflammation, leading to profound reductions in clinical and proinflammatory markers in the joints and skin of IL-23 MC-treated mice. The stromal compartment in the tendon was the main source of CCL20 in this model and accordingly, in vitro activated human tendon cells were able to produce this chemokine and to induce CCR6+ T cell migration, the latter of which could be blocked by CCL20LD.

Conclusions: Our studies highlight the pathogenic role of CCR6-CCL20 axis in enthesitis and raise the prospect of a novel therapeutic approach for treating patients with PsO and PsA.
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http://dx.doi.org/10.1002/art.41882DOI Listing
June 2021

Single cell and spatial transcriptomics in human tendon disease indicate dysregulated immune homeostasis.

Ann Rheum Dis 2021 May 17. Epub 2021 May 17.

Institute of Infection, Immunity and Inflammation, College of Medical Veterinary and Life Sciences, University of Glasgow, Glasgow, UK

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http://dx.doi.org/10.1136/annrheumdis-2021-220256DOI Listing
May 2021

Novel self-amplificatory loop between T cells and tenocytes as a driver of chronicity in tendon disease.

Ann Rheum Dis 2021 Mar 10. Epub 2021 Mar 10.

Institute of Infection, Immunity and Inflammation, University of Glasgow College of Medical Veterinary and Life Sciences, Glasgow, UK

Objectives: Increasing evidence suggests that inflammatory mechanisms play a key role in chronic tendon disease. After observing T cell signatures in human tendinopathy, we explored the interaction between T cells and tendon stromal cells or tenocytes to define their functional contribution to tissue remodelling and inflammation amplification and hence disease perpetuation.

Methods: T cells were quantified and characterised in healthy and tendinopathic tissues by flow cytometry (FACS), imaging mass cytometry (IMC) and single cell RNA-seq. Tenocyte activation induced by conditioned media from primary damaged tendon or interleukin-1β was evaluated by qPCR. The role of tenocytes in regulating T cell migration was interrogated in a standard transwell membrane system. T cell activation (cell surface markers by FACS and cytokine release by ELISA) and changes in gene expression in tenocytes (qPCR) were assessed in cocultures of T cells and explanted tenocytes.

Results: Significant quantitative differences were observed in healthy compared with tendinopathic tissues. IMC showed T cells in close proximity to tenocytes, suggesting tenocyte-T cell interactions. On activation, tenocytes upregulated inflammatory cytokines, chemokines and adhesion molecules implicated in T cell recruitment and activation. Conditioned media from activated tenocytes induced T cell migration and coculture of tenocytes with T cells resulted in reciprocal activation of T cells. In turn, these activated T cells upregulated production of inflammatory mediators in tenocytes, while increasing the pathogenic collagen 3/collagen 1 ratio.

Conclusions: Interaction between T cells and tenocytes induces the expression of inflammatory cytokines/chemokines in tenocytes, alters collagen composition favouring collagen 3 and self-amplifies T cell activation via an auto-regulatory feedback loop. Selectively targeting this adaptive/stromal interface may provide novel translational strategies in the management of human tendon disorders.
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http://dx.doi.org/10.1136/annrheumdis-2020-219335DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8292554PMC
March 2021

Basic Science of Tendinopathy and Novel Treatments: An Update.

Instr Course Lect 2021 ;70:551-562

Tendinopathy describes a spectrum of degenerative and inflammatory changes occurring in tendons, usually caused by mechanical loading. It is associated with pain and reduced function and can often lead to tendon rupture. Although advances have been made in recent years, it remains challenging to manage tendinopathy because its definition and the understanding of its risk factors and pathophysiology are still evolving. The objective of this chapter is to present current ideas on the basic science of tendinopathy and in particular new findings in regard to pathophysiology. Current therapies and insights into potential novel therapies for tendinopathy are also discussed.
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January 2021

Vancomycin Wrap for Anterior Cruciate Ligament Surgery: Molecular Insights.

Am J Sports Med 2021 02 6;49(2):426-434. Epub 2021 Jan 6.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow, UK.

Background: The use of the vancomycin wrap to pretreat the hamstring graft in anterior cruciate ligament reconstruction (ACLR) has grown in popularity since it was first described in 2012 and has significantly reduced rates of postoperative infection. However, it remains unknown if this antibiotic treatment affects the molecular composition of the graft.

Purpose: To establish whether treatment with vancomycin at 5 mg/mL, the most commonly used concentration, alters the molecular function of the hamstring graft in ACLR.

Study Design: Controlled laboratory study.

Methods: Surplus hamstring tendon collected after routine ACLR surgery was used for in vitro cell culture and ex vivo tissue experiments. Vancomycin was used at 5 mg/mL in RPMI or saline diluent to treat cells and tendon tissue, respectively, with diluent control conditions. Cell viability at 30, 60, and 120 minutes was assessed via colorimetric viability assay. Tendon cells treated with control and experimental conditions for 1 hour was evaluated using semiquantitative reverse transcription analysis, immunohistochemistry staining, and protein quantitation via enzyme-linked immunosorbent assay for changes in apoptotic, matrix, and inflammatory gene and protein expression.

Results: Vancomycin treatment at 5 mg/mL significantly reduced tenocyte viability in vitro after 60 minutes of treatment ( < .05); however, this was not sustained at 120 minutes. Vancomycin-treated tendon tissue showed no significant increase in apoptotic gene expression, or apoptotic protein levels in tissue or supernatant, ex vivo. Vancomycin was associated with a reduction in inflammatory proteins from treated tendon supernatants (IL-6; < .05).

Conclusion: Vancomycin did not significantly alter the molecular structure of the hamstring graft. Reductions in matrix protein and inflammatory cytokine release point to a potential beneficial effect of vancomycin in generating a homeostatic environment.

Clinical Relevance: Vancomycin ACL wrap does not alter the molecular structure of the ACL hamstring graft and may improve graft integrity.
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http://dx.doi.org/10.1177/0363546520981570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7859666PMC
February 2021

Attenuation of Dupuytren's fibrosis via targeting of the STAT1 modulated IL-13Rα1 response.

Sci Adv 2020 Jul 10;6(28):eaaz8272. Epub 2020 Jul 10.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow, Scotland, UK.

Fibrotic disorders represent common complex disease pathologies that are therapeutically challenging. Inflammation is associated with numerous fibrotic pathogeneses; however, its role in the multifaceted mechanisms of fibrosis remains unclear. IL-13 is implicated in aberrant responses involved in fibrotic disease, and we aimed to understand its role in the inflammatory processes of a common fibrotic disorder, Dupuytren's disease. We demonstrated T-cells produced IFN-g, which induced IL-13 secretion from mast cells and up-regulated IL-13Ra1 on fibroblasts, rendering them more reactive to IL-13. Consequently, diseased myofibroblasts demonstrated enhanced fibroproliferative effects upon IL-13 stimulation. We established IFN-g and IL-13 responses involved STAT dependent pathways, and STAT targeting (tofacitinib) could inhibit IL-13 production from mast cells, IL-13Ra1 up-regulation in fibroblasts and fibroproliferative effects of IL-13 on diseased myofibroblasts. Accordingly, utilizing Dupuytren's as an accessible human model of fibrosis, we propose targeting STAT pathways may offer previously unidentified therapeutic approaches in the management of fibrotic disease.
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http://dx.doi.org/10.1126/sciadv.aaz8272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7351483PMC
July 2020

Fibroblast activation and inflammation in frozen shoulder.

PLoS One 2019 23;14(4):e0215301. Epub 2019 Apr 23.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow, Scotland, United Kingdom.

Introduction: Frozen shoulder is a common, fibro-proliferative disease characterised by the insidious onset of pain and progressively restricted range of shoulder movement. Despite the prevalence of this disease, there is limited understanding of the molecular mechanisms underpinning the pathogenesis of this debilitating disease. Previous studies have identified increased myofibroblast differentiation and proliferation, immune cell influx and dysregulated cytokine production. We hypothesised that subpopulations within the fibroblast compartment may take on an activated phenotype, thus initiating the inflammatory processes observed in frozen shoulder. Therefore, we sought to evaluate the presence and possible pathogenic role of known stromal activation proteins in Frozen shoulder.

Methods: Shoulder capsule samples were collected from 10 patients with idiopathic frozen shoulder and 10 patients undergoing shoulder stabilisation surgery. Fibroblast activation marker expression (CD248, CD146, VCAM and PDPN, FAP) was quantified using immunohistochemistry. Control and diseased fibroblasts were cultured for in vitro studies from capsule biopsies from instability and frozen shoulder surgeries, respectively. The inflammatory profile and effects of IL-1β upon diseased and control fibroblasts was assessed using ELISA, immunohistochemistry and qPCR.

Results: Immunohistochemistry demonstrated increased expression of fibroblast activation markers CD248, CD146, VCAM and PDPN in the frozen shoulder group compared with control (p < 0.05). Fibroblasts cultured from diseased capsule produced elevated levels of inflammatory protein (IL-6, IL-8 & CCL-20) in comparison to control fibroblasts. Exposing control fibroblasts to an inflammatory stimuli, (IL-1ß) significantly increased stromal activation marker transcript and protein expression (CD248, PDPN and VCAM).

Conclusions: These results show that fibroblasts have an activated phenotype in frozen shoulder and this is associated with inflammatory cytokine dysregulation. Furthermore, it supports the hypothesis that activated fibroblasts may be involved in regulating the inflammatory and fibrotic processes involved in this disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0215301PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6478286PMC
February 2020

Targeting the NF-κB signaling pathway in chronic tendon disease.

Sci Transl Med 2019 02;11(481)

Department of Orthopedic Surgery, Columbia University, 650 W 168th St, New York, NY 10032, USA.

Tendon disorders represent the most common musculoskeletal complaint for which patients seek medical attention; inflammation drives tendon degeneration before tearing and impairs healing after repair. Clinical evidence has implicated the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway as a correlate of pain-free return to function after surgical repair. However, it is currently unknown whether this response is a reaction to or a driver of pathology. Therefore, we aimed to understand the clinically relevant involvement of the NF-κB pathway in tendinopathy, to determine its potential causative roles in tendon degeneration, and to test its potential as a therapeutic candidate. Transcriptional profiling of early rotator cuff tendinopathy identified increases in NF-κB signaling, including increased expression of the regulatory serine kinase subunit IKKβ, which plays an essential role in inflammation. Using cre-mediated overexpression of IKKβ in tendon fibroblasts, we observed degeneration of mouse rotator cuff tendons and the adjacent humeral head. These changes were associated with increases in proinflammatory cytokines and innate immune cells within the joint. Conversely, genetic deletion of IKKβ in tendon fibroblasts partially protected mice from chronic overuse-induced tendinopathy. Furthermore, conditional knockout of IKKβ improved outcomes after surgical repair, whereas overexpression impaired tendon healing. Accordingly, targeting of the IKKβ/NF-κB pathway in tendon stromal cells may offer previously unidentified therapeutic approaches in the management of human tendon disorders.
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http://dx.doi.org/10.1126/scitranslmed.aav4319DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6534967PMC
February 2019

S100A8 & S100A9: Alarmin mediated inflammation in tendinopathy.

Sci Rep 2019 02 6;9(1):1463. Epub 2019 Feb 6.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow, Scotland, UK.

Alarmins S100A8 and S100A9 are endogenous molecules released in response to environmental triggers and cellular damage. They are constitutively expressed in immune cells such as monocytes and neutrophils and their expression is upregulated under inflammatory conditions. The molecular mechanisms that regulate inflammatory pathways in tendinopathy are largely unknown therefore identifying early immune effectors is essential to understanding the pathology. Based on our previous investigations highlighting tendinopathy as an alarmin mediated pathology we sought evidence of S100A8 & A9 expression in a human model of tendinopathy and thereafter, to explore mechanisms whereby S100 proteins may regulate release of inflammatory mediators and matrix synthesis in human tenocytes. Immunohistochemistry and quantitative RT-PCR showed S100A8 & A9 expression was significantly upregulated in tendinopathic tissue compared with control. Furthermore, treating primary human tenocytes with exogenous S100A8 & A9 significantly increased protein release of IL-6, IL-8, CCL2, CCL20 and CXCL10; however, no alterations in genes associated with matrix remodelling were observed at a transcript level. We propose S100A8 & A9 participate in early pathology by modulating the stromal microenvironment and influencing the inflammatory profile observed in tendinopathy. S100A8 and S100A9 may participate in a positive feedback mechanism involving enhanced leukocyte recruitment and release of pro-inflammatory cytokines from tenocytes that perpetuates the inflammatory response within the tendon in the early stages of disease.
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http://dx.doi.org/10.1038/s41598-018-37684-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6365574PMC
February 2019

Alarmins in Frozen Shoulder: A Molecular Association Between Inflammation and Pain.

Am J Sports Med 2018 03 30;46(3):671-678. Epub 2017 Nov 30.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences, University of Glasgow, Glasgow, Scotland, UK.

Background: The pathophysiological mechanisms behind proliferation of fibroblasts and deposition of dense collagen matrix in idiopathic frozen shoulder remain unclear. Alarmins (also known as danger signals) are endogenous molecules that are released into the extracellular milieu after infection or tissue injury and that signal cell and tissue damage.

Purpose: To investigate whether the presence of alarmins is higher in patients with idiopathic frozen shoulder than in control subjects.

Study Design: Controlled laboratory study.

Methods: Shoulder capsule samples were collected from 10 patients with idiopathic frozen shoulder and 10 patients with unstable shoulders (control). The samples were stained with hematoxylin and eosin (H&E) and analyzed by immunohistochemistry using antibodies against alarmin molecules including high-mobility group protein B1 (HMGB1), interleukin 33, S100A8, S100A9, and the peripheral nerve marker PGP9.5. Immunoreactivities were rated in a blinded fashion from "none" to "strong." Immunohistochemical distribution within the capsule was noted. Before surgery, patient-ranked pain frequency, severity, stiffness, and the range of passive shoulder motion were recorded and statistically analyzed.

Results: Compared with control patients, patients with frozen shoulder had greater frequency and severity of self-reported pain ( P = .02) and more restricted range of motion in all planes ( P < .05). H&E-stained capsular tissue from frozen shoulder showed fibroblastic hypercellularity and increased subsynovial vascularity. Immunoreactivity of alarmins was significantly stronger in frozen shoulder capsules compared with control capsules ( P < .05). Furthermore, the expression of the alarmin molecule HMGB1 significantly correlated ( r > 0.9, P < .05) with the severity of patient-reported pain.

Conclusion: This study demonstrates a potential role for key molecular danger signals in frozen shoulder and suggests an association between the expression of danger molecules and the pain experienced by patients.
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http://dx.doi.org/10.1177/0363546517741127DOI Listing
March 2018

Targeting danger molecules in tendinopathy: the HMGB1/TLR4 axis.

RMD Open 2017 28;3(2):e000456. Epub 2017 Jul 28.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences, University of Glasgow, Glasgow, Scotland, UK.

Objectives: To seek evidence of the danger molecule, high-mobility group protein B1 (HMGB1) expression in human tendinopathy and thereafter, to explore mechanisms where HMGB1 may regulate inflammatory mediators and matrix regulation in human tendinopathy.

Methods: Torn supraspinatus tendon (established pathology) and matched intact subscapularis tendon (representing 'early pathology') biopsies were collected from patients undergoing arthroscopic shoulder surgery. Control samples of subscapularis tendon were collected from patients undergoing arthroscopic stabilisation surgery. Markers of inflammation and HMGB1 were quantified by reverse transcriptase PCR (RT-PCR) and immunohistochemistry. Human tendon-derived primary cells were derived from hamstring tendon tissue obtained during hamstring tendon anterior cruciate ligament reconstruction and used through passage 3. In vitro effects of recombinant HMGB1 on tenocyte matrix and inflammatory potential were measured using quantitative RT-PCR, ELISA and immunohistochemistry staining.

Results: Tendinopathic tissues demonstrated significantly increased levels of the danger molecule HMGB1 compared with control tissues with early tendinopathy tissue showing the greatest expression. The addition of recombinant human HMGB1 to tenocytes led to significant increase in expression of a number of inflammatory mediators, including interleukin 1 beta (IL-1β), IL-6, IL-33, CCL2 and CXCL12, in vitro. Further analysis demonstrated rhHMGB1 treatment resulted in increased expression of genes involved in matrix remodelling. Significant increases were observed in Col3, Tenascin-C and Decorin. Moreover, blocking HMGB1 signalling via toll-like receptor 4 (TLR4) silencing reversed these key inflammatory and matrix changes.

Conclusion: HMGB1 is present in human tendinopathy and can regulate inflammatory cytokines and matrix changes. We propose HMGB1 as a mediator driving the inflammatory/matrix crosstalk and manipulation of the HMGB1/TLR4 axis may offer novel therapeutic approaches targeting inflammatory mechanisms in the management of human tendon disorders.
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http://dx.doi.org/10.1136/rmdopen-2017-000456DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5574425PMC
July 2017

MicroRNA29a Treatment Improves Early Tendon Injury.

Mol Ther 2017 10 28;25(10):2415-2426. Epub 2017 Jul 28.

Institute of Infection, Immunity, and Inflammation, College of Medicine, Veterinary, and Life Sciences, University of Glasgow, Glasgow G12 8TA, UK. Electronic address:

Tendon injuries (tendinopathies) are common in human and equine athletes and characterized by dysregulated collagen matrix, resulting in tendon damage. We have previously demonstrated a functional role for microRNA29a (miR29a) as a post-transcriptional regulator of collagen 3 expression in murine and human tendon injury. Given the translational potential, we designed a randomized, blinded trial to evaluate the potential of a miR29a replacement therapy as a therapeutic option to treat tendinopathy in an equine model that closely mimics human disease. Tendon injury was induced in the superficial digital flexor tendon (SDFT) of 17 horses. Tendon lesions were treated 1 week later with an intralesional injection of miR29a or placebo. miR29a treatment reduced collagen 3 transcript levels at week 2, with no significant changes in collagen 1. The relative lesion cross-sectional area was significantly lower in miR29a tendons compared to control tendons. Histology scores were significantly better for miR29a-treated tendons compared to control tendons. These data support the mechanism of microRNA-mediated modulation of early pathophysiologic events that facilitate tissue remodeling in the tendon after injury and provides a strong proof of principle that a locally delivered miR29a therapy improves early tendon healing.
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http://dx.doi.org/10.1016/j.ymthe.2017.07.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5628866PMC
October 2017

IL-17A mediates inflammatory and tissue remodelling events in early human tendinopathy.

Sci Rep 2016 06 6;6:27149. Epub 2016 Jun 6.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow, Scotland UK.

Increasingly, inflammatory mediators are considered crucial to the onset and perpetuation of tendinopathy. We sought evidence of interleukin 17A (IL-17A) expression in early human tendinopathy and thereafter, explored mechanisms whereby IL-17A mediated inflammation and tissue remodeling in human tenocytes. Torn supraspinatus tendon (established pathology) and matched intact subscapularis tendon (representing 'early pathology') along with control biopsies were collected from patients undergoing shoulder surgery. Markers of inflammation and IL-17A were quantified by RT-PCR and immunohistochemistry. Human tendon cells were derived from hamstring tendon obtained during ACL reconstruction. In vitro effects of IL-17A upon tenocytes were measured using RT-PCR, multiplex cytokine assays, apoptotic proteomic profiling, immunohistochemistry and annexin V FACS staining. Increased expression of IL-17A was detected in 'early tendinopathy' compared to both matched samples and non-matched control samples (p < 0.01) by RT-PCR and immunostaining. Double immunofluoresence staining revealed IL-17A expression in leukocyte subsets including mast cells, macrophages and T cells. IL-17A treated tenocytes exhibited increased production of proinflammatory cytokines (p < 0.001), altered matrix regulation (p < 0.01) with increased Collagen type III and increased expression of several apoptosis related factors. We propose IL-17A as an inflammatory mediator within the early tendinopathy processes thus providing novel therapeutic approaches in the management of tendon disorders.
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http://dx.doi.org/10.1038/srep27149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4893609PMC
June 2016

MicroRNA29a regulates IL-33-mediated tissue remodelling in tendon disease.

Nat Commun 2015 Apr 10;6:6774. Epub 2015 Apr 10.

Institute of Infection, Immunity and Inflammation, College of Medicine, Veterinary and Life Sciences University of Glasgow, Glasgow G12 8QQ, UK.

MicroRNA (miRNA) has the potential for cross-regulation and functional integration of discrete biological processes during complex physiological events. Utilizing the common human condition tendinopathy as a model system to explore the cross-regulation of immediate inflammation and matrix synthesis by miRNA we observed that elevated IL-33 expression is a characteristic of early tendinopathy. Using in vitro tenocyte cultures and in vivo models of tendon damage, we demonstrate that such IL-33 expression plays a pivotal role in the transition from type 1 to type 3 collagen (Col3) synthesis and thus early tendon remodelling. Both IL-33 effector function, via its decoy receptor sST2, and Col3 synthesis are regulated by miRNA29a. Downregulation of miRNA29a in human tenocytes is sufficient to induce an increase in Col3 expression. These data provide a molecular mechanism of miRNA-mediated integration of the early pathophysiologic events that facilitate tissue remodelling in human tendon after injury.
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http://dx.doi.org/10.1038/ncomms7774DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4403384PMC
April 2015

Notch signaling drives multiple myeloma induced osteoclastogenesis.

Oncotarget 2014 Nov;5(21):10393-406

Department of Health Sciences, Università degli Studi di Milano, Milano, Italy.

Multiple myeloma (MM) is closely associated with bone destruction. Once migrated to the bone marrow, MM cells unbalance bone formation and resorption via the recruitment and maturation of osteoclast precursors. The Notch pathway plays a key role in different types of cancer and drives several biological processes relevant in MM, including cell localization within the bone marrow, proliferation, survival and pharmacological resistance. Here we present evidences that MM can efficiently drive osteoclastogenesis by contemporaneously activating Notch signaling on tumor cells and osteoclasts through the aberrant expression of Notch ligands belonging to the Jagged family. Active Notch signaling in MM cells induces the secretion of the key osteoclastogenic factor, RANKL, which can be boosted in the presence of stromal cells. In turn, MM cells-derived RANKL causes the upregulation of its receptor, RANK, and Notch2 in pre-osteoclasts. Notch2 stimulates osteoclast differentiation by promoting autocrine RANKL signaling. Finally, MM cells through Jagged ligands expression can also activate Notch signaling in pre-osteoclast by direct contact. Such synergism between tumor cells and pre-osteoclasts in MM-induced osteoclastogenesis can be disrupted by silencing tumor-derived Jagged1 and 2. These results make the Jagged ligands new promising therapeutic targets in MM to contrast bone disease and the associated co-morbidities.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4279381PMC
http://dx.doi.org/10.18632/oncotarget.2084DOI Listing
November 2014

Acute inflammatory response to cobalt chromium orthopaedic wear debris in a rodent air-pouch model.

J R Soc Interface 2012 Sep 18;9(74):2109-19. Epub 2012 Apr 18.

Bioengineering Unit, University of Strathclyde, Wolfson Centre, Glasgow, UK.

This study used a rodent air-pouch model to assess the acute inflammatory response to cobalt-chromium (CoCr) alloy wear debris from a metal-on-metal hip resurfacing implant that may contribute to joint failure. Air-pouches were injected with either sterile phosphate-buffered saline, 1 μg lipopolysaccharide (LPS) or 2.5 mg CoCr wear debris. The in situ inflammatory response was monitored 4, 24, 48 and 72 h and 7 days later. A flow cytometric analysis of the inflammatory exudates showed that CoCr wear debris induced a different inflammatory pattern compared with LPS. LPS induced a strong early (4 h) neutrophil influx, with monocyte/macrophage influx peaking at 24 h, whereas CoCr wear debris initiated almost equal numbers of early monocyte/macrophage and neutrophil recruitment. Histological analyses also showed CoCr debris accumulated in the pouch wall and this was accompanied by vast cellular infiltration and fibrosis around the debris throughout the duration of the experiment. Assessment of inflammatory gene transcripts from air-pouch tissue showed that CoCr wear debris increased the expression of cytokines involved in promoting inflammation and fibrosis (IL-1β, TGF-β) and chemokines that promote the recruitment of neutrophils and monocytes/macrophages (CXCL2 and CCL2). The data suggest that inflammatory responses to CoCr debris induce a specific acute process in which the recruitment of monocytes/macrophages is key.
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http://dx.doi.org/10.1098/rsif.2012.0006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3405742PMC
September 2012

Distribution of metal released from cobalt-chromium alloy orthopaedic wear particles implanted into air pouches in mice.

J Biomed Mater Res A 2012 Jun 15;100(6):1529-38. Epub 2012 Mar 15.

Bioengineering Unit, University of Strathclyde, Wolfson Center, Glasgow G4 0NW, United Kingdom.

Metal-on-metal hip replacement implants generate wear debris and release ions both locally and systemically in patients. To investigate dissemination of metal, we determined blood and organ levels of cobalt (Co), chromium (Cr), and molybdenum (Mo) following the implantation of Co-Cr alloy wear debris in mice using skin pouches as a model system. We observed increased metal levels in blood for up to 72 h; the levels of Co were highest and remained elevated for 7 days. Co levels were elevated in all organs studied (liver, kidney, spleen, lung, heart, brain, and testes), with the peak at 48 h; highest levels were measured in liver and kidney (838.9 ± 223.7 ng/g in liver, and 938.8 ± 131.6 ng/g in kidney). Organ Cr levels were considerably lower than Co levels, for example, Cr in kidney was 117.2 ± 12.6 ng/g tissue at 48 h. Co is more mobile than Cr, reaching higher levels at earlier time points. This could be due to local tissue binding of Cr. Exposure to Co-Cr particles in vivo altered antioxidant enzyme expression and activities. We observed induction of catalase protein in the liver and glutathione reductase (GR) and peroxidase (GPx) proteins in the spleen. Activities of catalase and GPx in the liver were significantly increased while that of GR was decreased in the kidney. Organs of mice with Co-Cr particle implantation were exposed to increased metal levels capable of inducing reactive oxygen species scavenging enzymes, suggesting the tissue may be subjected to oxidative stress; however, the overall antioxidant defence system was not markedly disturbed.
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http://dx.doi.org/10.1002/jbm.a.34091DOI Listing
June 2012

Effect of chromium and cobalt ions on primary human lymphocytes in vitro.

J Immunotoxicol 2011 Jun 29;8(2):140-9. Epub 2011 Mar 29.

Bioengineering Unit, University of Strathclyde, Wolfson Centre, Glasgow G4 0NW, UK.

Cobalt-chromium (Co-Cr) alloy metal-on-metal hip resurfacing is increasingly common among younger more active patients suffering from osteoarthritis. Recent reports have increased awareness of metal ions leaching from metallic articulations; this ion exposure may have adverse effects on the immune system. As previous studies reported alterations in lymphocyte number and function in patients with Co-Cr implants, we investigated effects of clinically relevant concentrations of Cr(6+) and Co(2+) on primary human lymphocytes in vitro. Here, both resting and activated (anti-CD3 ± anti-CD28 antibodies) primary human lymphocytes were exposed to Cr(6+) or Co(2+) (0.1-100 µM). Following 24 or 48 h of exposure, cell viability, proliferation, cytokine [interferon-γ (IFNγ and interleukin-2 (IL-2)] release, and apoptosis (with and without pre-treatment of cells with a caspase-3 inhibitor) were assessed. Exposure to 10 and 100 µM Cr(6+) significantly decreased cell viability and increased apoptosis in both resting and activated lymphocytes. Cell proliferation and cytokine release were also significantly reduced in activated lymphocytes following exposure. The exposure of resting lymphocytes to 100 µM Co(2+) resulted in significant decreases in cell viability accompanied by a significant increase in apoptosis. Activated lymphocytes also showed this response after exposure to 100 µM Co(2+); in fact, activated cells were significantly more sensitive to Co(2+) toxicity. Exposure to 10 µM Co(2+) led to significant decreases in cell proliferation and cytokine release, but no significant increase in apoptosis, in activated cells. The results indicate that exposure to high concentrations of metal ions initiate apoptosis that results in decreased lymphocyte proliferation. IL-2 release is inhibited by both metal ions at concentrations that are not overtly toxic. However, metal ion concentrations not directly cytotoxic to lymphocytes may affect events at a molecular level, thereby impeding lymphocyte proliferation. Hence, this may contribute to altered immune system function in patients with Co-Cr implants.
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http://dx.doi.org/10.3109/1547691X.2011.553845DOI Listing
June 2011
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