Publications by authors named "Tonia L Vincent"

61 Publications

Local depletion of proteoglycans mediates cartilage tissue repair in an ex vivo integration model.

Acta Biomater 2022 Sep 30;149:179-188. Epub 2022 Jun 30.

Centre for Craniofacial and Regenerative Biology, King's College London, London SE1 9RT, UK. Electronic address:

Successfully replacing damaged cartilage with tissue-engineered constructs requires integration with the host tissue and could benefit from leveraging the native tissue's intrinsic healing capacity; however, efforts are limited by a poor understanding of how cartilage repairs minor defects. Here, we investigated the conditions that foster natural cartilage tissue repair to identify strategies that might be exploited to enhance the integration of engineered/grafted cartilage with host tissue. We damaged porcine articular cartilage explants and using a combination of pulsed SILAC-based proteomics, ultrastructural imaging, and catabolic enzyme blocking strategies reveal that integration of damaged cartilage surfaces is not driven by neo-matrix synthesis, but rather local depletion of proteoglycans. ADAMTS4 expression and activity are upregulated in injured cartilage explants, but integration could be reduced by inhibiting metalloproteinase activity with TIMP3. These observations suggest that catabolic enzyme-mediated proteoglycan depletion likely allows existing collagen fibrils to undergo cross-linking, fibrillogenesis, or entanglement, driving integration. Catabolic enzymes are often considered pathophysiological markers of osteoarthritis. Our findings suggest that damage-induced upregulation of metalloproteinase activity may be a part of a healing response that tips towards tissue destruction under pathological conditions and in osteoarthritis, but could also be harnessed in tissue engineering strategies to mediate repair. STATEMENT OF SIGNIFICANCE: Cartilage tissue engineering strategies require graft integration with the surrounding tissue; however, how the native tissue repairs minor injuries is poorly understood. We applied pulsed SILAC-based proteomics, ultrastructural imaging, and catabolic enzyme blocking strategies to a porcine cartilage explant model and found that integration of damaged cartilage surfaces is driven by catabolic enzyme-mediated local depletion of proteoglycans. Although catabolic enzymes have been implicated in cartilage destruction in osteoarthritis, our findings suggest that damage-induced upregulation of metalloproteinase activity may be a part of a healing response that tips towards tissue destruction under pathological conditions. They also suggest that this natural cartilage tissue repair process could be harnessed in tissue engineering strategies to enhance the integration of engineered cartilage with host tissue.
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http://dx.doi.org/10.1016/j.actbio.2022.06.032DOI Listing
September 2022

The Extracellular Matrix of Articular Cartilage Controls the Bioavailability of Pericellular Matrix-Bound Growth Factors to Drive Tissue Homeostasis and Repair.

Int J Mol Sci 2022 May 26;23(11). Epub 2022 May 26.

Norwich Medical School, University of East Anglia, Norwich, Norwich NR4 7UQ, UK.

The extracellular matrix (ECM) has long been regarded as a packing material; supporting cells within the tissue and providing tensile strength and protection from mechanical stress. There is little surprise when one considers the dynamic nature of many of the individual proteins that contribute to the ECM, that we are beginning to appreciate a more nuanced role for the ECM in tissue homeostasis and disease. Articular cartilage is adapted to be able to perceive and respond to mechanical load. Indeed, physiological loads are essential to maintain cartilage thickness in a healthy joint and excessive mechanical stress is associated with the breakdown of the matrix that is seen in osteoarthritis (OA). Although the trigger by which increased mechanical stress drives catabolic pathways remains unknown, one mechanism by which cartilage responds to increased compressive load is by the release of growth factors that are sequestered in the pericellular matrix. These are heparan sulfate-bound growth factors that appear to be largely chondroprotective and displaced by an aggrecan-dependent sodium flux. Emerging evidence suggests that the released growth factors act in a coordinated fashion to drive cartilage repair. Thus, we are beginning to appreciate that the ECM is the key mechano-sensor and mechano-effector in cartilage, responsible for directing subsequent cellular events of relevance to joint health and disease.
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http://dx.doi.org/10.3390/ijms23116003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9181404PMC
May 2022

Mechanical forces couple bone matrix mineralization with inhibition of angiogenesis to limit adolescent bone growth.

Nat Commun 2022 06 1;13(1):3059. Epub 2022 Jun 1.

Pitzer Laboratory of Osteoarthritis Research, German Rheumatism Research Centre (DRFZ), a Leibniz Institute, Berlin, Germany.

Bone growth requires a specialised, highly angiogenic blood vessel subtype, so-called type H vessels, which pave the way for osteoblasts surrounding these vessels. At the end of adolescence, type H vessels differentiate into quiescent type L endothelium lacking the capacity to promote bone growth. Until now, the signals that switch off type H vessel identity and thus limit adolescent bone growth have remained ill defined. Here we show that mechanical forces, associated with increased body weight at the end of adolescence, trigger the mechanoreceptor PIEZO1 and thereby mediate enhanced production of the kinase FAM20C in osteoblasts. FAM20C, the major kinase of the secreted phosphoproteome, phosphorylates dentin matrix protein 1, previously identified as a key factor in bone mineralization. Thereupon, dentin matrix protein 1 is secreted from osteoblasts in a burst-like manner. Extracellular dentin matrix protein 1 inhibits vascular endothelial growth factor signalling by preventing phosphorylation of vascular endothelial growth factor receptor 2. Hence, secreted dentin matrix protein 1 transforms type H vessels into type L to limit bone growth activity and enhance bone mineralization. The discovered mechanism may suggest new options for the treatment of diseases characterised by aberrant activity of bone and vessels such as osteoarthritis, osteoporosis and osteosarcoma.
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http://dx.doi.org/10.1038/s41467-022-30618-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9160028PMC
June 2022

Imaging articular cartilage in osteoarthritis using targeted peptide radiocontrast agents.

PLoS One 2022 10;17(5):e0268223. Epub 2022 May 10.

Centre for OA Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, United Kingdom.

Background: Established MRI and emerging X-ray contrast agents for non-invasive imaging of articular cartilage rely on non-selective electrostatic interactions with negatively charged proteoglycans. These contrast agents have limited prognostic utility in diseases such as osteoarthritis (OA) due to the characteristic high turnover of proteoglycans. To overcome this limitation, we developed a radiocontrast agent that targets the type II collagen macromolecule in cartilage and used it to monitor disease progression in a murine model of OA.

Methods: To confer radiopacity to cartilage contrast agents, the naturally occurring tyrosine derivative 3,5-diiodo-L-tyrosine (DIT) was introduced into a selective peptide for type II collagen. Synthetic DIT peptide derivatives were synthesised by Fmoc-based solid-phase peptide synthesis and binding to ex vivo mouse tibial cartilage evaluated by high-resolution micro-CT. Di-Iodotyrosinated Peptide Imaging of Cartilage (DIPIC) was performed ex vivo and in vivo 4, 8 and 12 weeks in mice after induction of OA by destabilisation of the medial meniscus (DMM). Finally, human osteochondral plugs were imaged ex vivo using DIPIC.

Results: Fifteen DIT peptides were synthesised and tested, yielding seven leads with varying cartilage binding strengths. DIPIC visualised ex vivo murine articular cartilage comparably to the ex vivo contrast agent phosphotungstic acid. Intra-articular injection of contrast agent followed by in vivo DIPIC enabled delineation of damaged murine articular cartilage. Finally, the translational potential of the contrast agent was confirmed by visualisation of ex vivo human cartilage explants.

Conclusion: DIPIC has reduction and refinement implications in OA animal research and potential clinical translation to imaging human disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0268223PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9089912PMC
May 2022

Osteoarthritis Pathophysiology: Therapeutic Target Discovery may Require a Multifaceted Approach.

Clin Geriatr Med 2022 05;38(2):193-219

Raymond Purves Bone and Joint Research Laboratories, Kolling Institute University of Sydney Faculty of Medicine and Health at Royal North Shore Hospital, St. Leonards, New South Wales 2065, Australia. Electronic address:

Molecular understanding of osteoarthritis (OA) has greatly increased through careful analysis of tissue samples, preclinical models, and large-scale agnostic "-omic" studies. There is broad acceptance that systemic and biomechanical signals affect multiple tissues of the joint, each of which could potentially be targeted to improve patient outcomes. In this review six experts in different aspects of OA pathogenesis provide their independent view on what they believe to be good tractable approaches to OA target discovery. We conclude that molecular discovery has been high but future transformative studies require a multidisciplinary holistic approach to develop therapeutic strategies with high clinical efficacy.
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http://dx.doi.org/10.1016/j.cger.2021.11.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9107912PMC
May 2022

HAILO: A Sensorised Hand Splint for the Exploration of Interaction Forces.

IEEE Trans Biomed Eng 2022 Mar 1;PP. Epub 2022 Mar 1.

This study presents the design and development of an instrumented splint for measuring the biomechanical effects of hand splinting, and for assessing interface loading characteristics for people with arthritis. Sixteen multi-axial soft load-sensing nodes were mounted on the splint-skin interface of a custom 3D printed thumb splint. The splint was used to measure the interface forces between splint and hand in 12 healthy participants in 6 everyday tasks. Forces were compared between a baseline relaxed hand position and during states of active use. These data were used to generate a measure of sensor activity across the splint surface. Through direct comparison with a commercial splint, the 3D printed splint was deemed to provide similar levels of support. Observation of the activity across the 16 sensors showed that active areas of the splint surface varied between tasks but were commonly focused at the base of the thumb. Our findings show promise in the ability to detect the changing forces imparted on the hand by the splint surface, objectively characterising their behaviour. This opens the opportunity for future study into the biomechanical effects of splints on arthritic thumbs to improve this important intervention and improve quality of life.
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http://dx.doi.org/10.1109/TBME.2022.3155589DOI Listing
March 2022

Ciliary IFT88 Protects Coordinated Adolescent Growth Plate Ossification From Disruptive Physiological Mechanical Forces.

J Bone Miner Res 2022 06 20;37(6):1081-1096. Epub 2022 Feb 20.

Centre for OA Pathogenesis Versus Arthritis, The Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

Compared with our understanding of endochondral ossification, much less is known about the coordinated arrest of growth defined by the narrowing and fusion of the cartilaginous growth plate. Throughout the musculoskeletal system, appropriate cell and tissue responses to mechanical force delineate morphogenesis and ensure lifelong health. It remains unclear how mechanical cues are integrated into many biological programs, including those coordinating the ossification of the adolescent growth plate at the cessation of growth. Primary cilia are microtubule-based organelles tuning a range of cell activities, including signaling cascades activated or modulated by extracellular biophysical cues. Cilia have been proposed to directly facilitate cell mechanotransduction. To explore the influence of primary cilia in the mouse adolescent limb, we conditionally targeted the ciliary gene Intraflagellar transport protein 88 (Ift88 ) in the juvenile and adolescent skeleton using a cartilage-specific, inducible Cre (AggrecanCreER Ift88 ). Deletion of IFT88 in cartilage, which reduced ciliation in the growth plate, disrupted chondrocyte differentiation, cartilage resorption, and mineralization. These effects were largely restricted to peripheral tibial regions beneath the load-bearing compartments of the knee. These regions were typified by an enlarged population of hypertrophic chondrocytes. Although normal patterns of hedgehog signaling were maintained, targeting IFT88 inhibited hypertrophic chondrocyte VEGF expression and downstream vascular recruitment, osteoclastic activity, and the replacement of cartilage with bone. In control mice, increases to physiological loading also impair ossification in the peripheral growth plate, mimicking the effects of IFT88 deletion. Limb immobilization inhibited changes to VEGF expression and epiphyseal morphology in Ift88cKO mice, indicating the effects of depletion of IFT88 in the adolescent growth plate are mechano-dependent. We propose that during this pivotal phase in adolescent skeletal maturation, ciliary IFT88 protects uniform, coordinated ossification of the growth plate from an otherwise disruptive heterogeneity of physiological mechanical forces. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).
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http://dx.doi.org/10.1002/jbmr.4502DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9304194PMC
June 2022

The Musculoskeletal Manifestations of Marfan Syndrome: Diagnosis, Impact, and Management.

Curr Rheumatol Rep 2021 11 26;23(11):81. Epub 2021 Nov 26.

Kennedy Institute of Rheumatology, NDORMS, University of Oxford, Oxford, UK.

Purpose Of Review: Marfan syndrome (MFS) is an autosomal dominant heritable disorder of fibrillin-1 (FBN1) with predominantly ocular, cardiovascular, and musculoskeletal manifestations that has a population prevalence of approximately 1 in 5-10,000 (Chiu et al. Mayo Clin Proc. 89(1):34-42, 146, Dietz 3, Loeys et al. J Med Genet. 47(7):476-85, 4).

Recent Findings: The vascular complications of MFS still pose the greatest threat, but effective management options, such as regular cardiac monitoring and elective surgical intervention, have reduced the risk of life-threatening cardiovascular events, such as aortic dissection. Although cardiovascular morbidity and mortality remains high, these improvements in cardiovascular management have extended the life expectancy of those with MFS by perhaps 30-50 years from an estimated mean of 32 years in 1972 (Dietz 3, Gott et al. Eur J Cardio-thoracic Surg. 10(3):149-58, 147, Murdoch et al. N Engl J Med. 286(15):804-8, 148). The musculoskeletal manifestations of MFS, which to date have received less attention, can also have a significant impact on the quality of life and are likely to become more important as the age of the Marfan syndrome population increases (Hasan et al. Int J Clin Pract. 61(8):1308-1320, 127). In addition, musculoskeletal manifestations are often critically important in the diagnosis of MFS. Here, we review the main clinically relevant and diagnostically useful musculoskeletal features of MFS, which together contribute to the "systemic features score" (referred to hereafter as systemic score), part of the revised Ghent nosology for MFS. We discuss current treatment strategies and highlight the need for a multidisciplinary approach to diagnosis and management. Finally, we review new pharmacological approaches that may be disease modifying and could help to improve the outcome for individuals with this syndrome.
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http://dx.doi.org/10.1007/s11926-021-01045-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8626407PMC
November 2021

Age-dependent changes in protein incorporation into collagen-rich tissues of mice by in vivo pulsed SILAC labelling.

Elife 2021 09 28;10. Epub 2021 Sep 28.

Kennedy Institute of Rheumatology, Arthritis Research UK Centre for OA Pathogenesis, University of Oxford, Oxford, United Kingdom.

Collagen-rich tissues have poor reparative capacity that predisposes to common age-related disorders such as osteoporosis and osteoarthritis. We used in vivo pulsed SILAC labelling to quantify new protein incorporation into cartilage, bone, and skin of mice across the healthy life course. We report dynamic turnover of the matrisome, the proteins of the extracellular matrix, in bone and cartilage during skeletal maturation, which was markedly reduced after skeletal maturity. Comparing young adult with older adult mice, new protein incorporation was reduced in all tissues. STRING clustering revealed changes in epigenetic modulators across all tissues, a decline in chondroprotective growth factors such as FGF2 and TGFβ in cartilage, and clusters indicating mitochondrial dysregulation and reduced collagen synthesis in bone. Several pathways were implicated in age-related disease. Fewer changes were observed for skin. This methodology provides dynamic protein data at a tissue level, uncovering age-related molecular changes that may predispose to disease.
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http://dx.doi.org/10.7554/eLife.66635DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8478409PMC
September 2021

Clinical and molecular associations with outcomes at 2 years after acute knee injury: a longitudinal study in the Knee Injury Cohort at the Kennedy (KICK).

Lancet Rheumatol 2021 Sep 24;3(9):e648-e658. Epub 2021 Jun 24.

Centre for Osteoarthritis Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, NDORMS, University of Oxford, Oxford, UK.

Background: Joint injury is a major risk factor for osteoarthritis and provides an opportunity to prospectively examine early processes associated with osteoarthritis. We investigated whether predefined baseline demographic and clinical factors, and protein analytes in knee synovial fluid and in plasma or serum, were associated with clinically relevant outcomes at 2 years after knee injury.

Methods: This longitudinal cohort study recruited individuals aged 16-50 years between Nov 1, 2010, and Nov 28, 2014, across six hospitals and clinics in London, UK. Participants were recruited within 8 weeks of having a clinically significant acute knee injury (effusion and structural injury on MRI), which was typically treated surgically. We measured several predefined clinical variables at baseline (eg, time from injury to sampling, extent and type of joint injury, synovial fluid blood staining, presence of effusion, self-reported sex, age, and BMI), and measured 12 synovial fluid and four plasma or serum biomarkers by immunoassay at baseline and 3 months. The primary outcome was Knee Injury and Osteoarthritis Outcome Score (KOOS) at 2 years, adjusted for baseline score, assessed in all patients. Linear and logistic regression models adjusting for predefined covariates were used to assess associations between baseline variables and 2-year KOOS. This study is registered with ClinicalTrials.gov, number NCT02667756.

Findings: We enrolled 150 patients at a median of 17 days (range 1-59, IQR 9-26) after knee injury. 123 (82%) were male, with a median age of 25 years (range 16-50, IQR 21-30). 98 (65%) of 150 participants completed a KOOS at 2 (or 3) years after enrolment (50 participants were lost to follow-up and two were withdrawn due to adverse events unrelated to study participation); 77 (51%) participants had all necessary variables available and were included in the core variable adjusted analysis. In the 2-year dataset mean KOOS improved from 38 (SD 18) at baseline to 79 (18) at 2 years. Baseline KOOS medium-to-large knee effusion, and moderate-to-severe synovial blood staining and their interaction significantly predicted 2-year KOOS (n=77; coefficient -20·5, 95% CI -34·8 to -6·18; p=0·0060). The only predefined biomarkers that showed independent associations with 2-year KOOS were synovial fluid MCP-1 (n=77; -0·015, 0·027 to -0·004 per change in 1 pg/mL units; p=0·011) and IL-6 (n=77; -0·0005, -0·0009 to -0·0001 per change in 1 pg/mL units; p=0·017). These biomarkers, combined with the interaction of effusion and blood staining, accounted for 39% of outcome variability. Two adverse events occurred that were linked to study participation, both at the time of blood sampling (one presyncopal episode, one tenderness and pain at the site of venepuncture).

Interpretation: The combination of effusion and haemarthrosis was significantly associated with symptomatic outcomes after acute knee injury. The synovial fluid molecular protein response to acute knee injury (best represented by MCP-1 and IL-6) was independently associated with symptomatic outcomes but not with structural outcomes, with the biomarkers overall playing a minor role relative to clinical predictors. The relationship between symptoms and structure after acute knee injury and their apparent dissociation early in this process need to be better understood to make clinical progress.

Funding: Versus Arthritis, Kennedy Trust for Rheumatology Research, and NIHR Oxford Biomedical Research Centre.
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http://dx.doi.org/10.1016/S2665-9913(21)00116-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8390381PMC
September 2021

Matrix-Bound Growth Factors are Released upon Cartilage Compression by an Aggrecan-Dependent Sodium Flux that is Lost in Osteoarthritis.

Function (Oxf) 2021 2;2(5):zqab037. Epub 2021 Aug 2.

Kennedy Institute of Rheumatology, University of Oxford, Oxford, OX3 7FY, UK.

Articular cartilage is a dense extracellular matrix-rich tissue that degrades following chronic mechanical stress, resulting in osteoarthritis (OA). The tissue has low intrinsic repair especially in aged and osteoarthritic joints. Here, we describe three pro-regenerative factors; fibroblast growth factor 2 (FGF2), connective tissue growth factor, bound to transforming growth factor-beta (CTGF-TGFβ), and hepatoma-derived growth factor (HDGF), that are rapidly released from the pericellular matrix (PCM) of articular cartilage upon mechanical injury. All three growth factors bound heparan sulfate, and were displaced by exogenous NaCl. We hypothesised that sodium, sequestered within the aggrecan-rich matrix, was freed by injurious compression, thereby enhancing the bioavailability of pericellular growth factors. Indeed, growth factor release was abrogated when cartilage aggrecan was depleted by IL-1 treatment, and in severely damaged human osteoarthritic cartilage. A flux in free matrix sodium upon mechanical compression of cartilage was visualised by Na -MRI just below the articular surface. This corresponded to a region of reduced tissue stiffness, measured by scanning acoustic microscopy and second harmonic generation microscopy, and where Smad2/3 was phosphorylated upon cyclic compression. Our results describe a novel intrinsic repair mechanism, controlled by matrix stiffness and mediated by the free sodium concentration, in which heparan sulfate-bound growth factors are released from cartilage upon injurious load. They identify aggrecan as a depot for sequestered sodium, explaining why osteoarthritic tissue loses its ability to repair. Treatments that restore matrix sodium to allow appropriate release of growth factors upon load are predicted to enable intrinsic cartilage repair in OA.

Significance Statement: Osteoarthritis is the most prevalent musculoskeletal disease, affecting 250 million people worldwide. We identify a novel intrinsic repair response in cartilage, mediated by aggrecan-dependent sodium flux, and dependent upon matrix stiffness, which results in the release of a cocktail of pro-regenerative growth factors after injury. Loss of aggrecan in late-stage osteoarthritis prevents growth factor release and likely contributes to disease progression. Treatments that restore matrix sodium in osteoarthritis may recover the intrinsic repair response to improve disease outcome.
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http://dx.doi.org/10.1093/function/zqab037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8374957PMC
August 2021

2021: The Year We Rewrite the Osteoarthritis Textbooks?

Authors:
Tonia L Vincent

Function (Oxf) 2021 21;2(1):zqaa043. Epub 2020 Dec 21.

Centre for OA Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

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http://dx.doi.org/10.1093/function/zqaa043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8248874PMC
December 2020

Hand Osteoarthritis: investigating Pain Effects of estrogen-containing therapy (HOPE-e): a protocol for a feasibility randomised placebo-controlled trial.

Pilot Feasibility Stud 2021 Jun 24;7(1):133. Epub 2021 Jun 24.

Centre for Osteoarthritis Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, NDORMS, University of Oxford, Oxford, UK.

Background: Hand osteoarthritis (OA) is a common condition, causing pain, stiffness and reduced quality of life. Incidence is higher amongst women, particularly around the age of the menopause. Whilst the relationship between sex hormones and OA has been studied in vitro, in epidemiological studies and in clinical trials of hormone replacement therapy (HRT), this study is the first to investigate the effect of estrogen-containing therapy on hand pain in post-menopausal women with symptomatic hand OA in a randomised study design.

Methods: This is a feasibility study of a double-blinded placebo-controlled intervention with 1:1 randomisation to either a combination of conjugated estrogens 0.45 mg and bazedoxifene acetate 20 mg (Duavive) or placebo. The target population is post-menopausal women with symptomatic hand OA, aiming to recruit 60-90 study participants. The primary objective is to assess the feasibility of a future fully powered randomised controlled trial (RCT). Participants will take the study medication for 24 weeks and be followed up for 28 weeks after randomisation. The primary outcomes used to determine feasibility are eligible participant identification rates and routes; recruitment, randomisation and retention rates of eligible participants; study medication compliance; and the likelihood of unintentional unblinding. Secondary outcomes include measures of hand pain, function, appearance and menopausal symptoms. An end of study questionnaire and focus groups will help to refine the final protocol for a full study.

Discussion: Identifying new treatments for symptomatic hand OA is a recognised research priority. The study will help us to understand whether there are sufficient interested and eligible individuals in this target population who would consider HRT for their hand symptoms. It will provide proof-of-concept RCT data on the effects of HRT on hand pain and other clinically relevant outcomes in this population. The study will gain valuable information on the feasibility of a full RCT and how best to run this. The findings will be published in a peer-reviewed journal and presented at a relevant conference.

Trial Registration: ISRCTN12196200 registered on 15 January 2019.
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http://dx.doi.org/10.1186/s40814-021-00869-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8223359PMC
June 2021

Role of Ciliary Protein Intraflagellar Transport Protein 88 in the Regulation of Cartilage Thickness and Osteoarthritis Development in Mice.

Arthritis Rheumatol 2022 01 16;74(1):49-59. Epub 2021 Dec 16.

University of Oxford, Oxford, UK.

Objective: Mechanical and biologic cues drive cellular signaling in cartilage development, health, and disease. Primary cilia proteins, which are implicated in the transduction of biologic and physiochemical signals, control cartilage formation during skeletal development. This study was undertaken to assess the influence of the ciliary protein intraflagellar transport protein 88 (IFT88) on postnatal cartilage from mice with conditional knockout of the Ift88 gene (Ift88-KO).

Methods: Ift88 and aggrecanCre mice were crossed to create a strain of cartilage-specific Ift88-KO mice (aggrecanCre ;Ift88 ). In these Ift88-KO mice and Ift88 control mice, tibial articular cartilage thickness was assessed by histomorphometry, and the integrity of the cartilage was assessed using Osteoarthritis Research Society International (OARSI) damage scores, from adolescence through adulthood. In situ mechanisms of cartilage damage were investigated in the microdissected cartilage sections using immunohistochemistry, RNAScope analysis, and quantitative polymerase chain reaction. Osteoarthritis (OA) was induced in aggrecanCre ;Ift88 mice and Ift88 control mice using surgical destabilization of the medial meniscus (DMM). Following tamoxifen injection and DMM surgery, the mice were given free access to exercise on a wheel.

Results: Deletion of Ift88 resulted in progressive reduction in the thickness of the medial tibial cartilage in adolescent mice, as well as marked atrophy of the cartilage in mice during adulthood. In aggrecanCre ;Ift88 mice at age 34 weeks, the median thickness of the medial tibial cartilage was 89.42 μm (95% confidence interval [95% CI] 84.00-93.49), whereas in Ift88 controls at the same age, the median cartilage thickness was 104.00 μm (95% CI 100.30-110.50; P < 0.0001). At all time points, the median thickness of the calcified cartilage was reduced. In some mice, atrophy of the medial tibial cartilage was associated with complete, spontaneous degradation of the cartilage. Following DMM, aggrecanCre ;Ift88 mice were found to have increased OARSI scores of cartilage damage. In articular cartilage from maturing mice, atrophy was not associated with obvious increases in aggrecanase-mediated destruction or chondrocyte hypertrophy. Of the 44 candidate genes analyzed, only Tcf7l2 expression levels correlated with Ift88 expression levels in the microdissected cartilage. However, RNAScope analysis revealed that increased hedgehog (Hh) signaling (as indicated by increased expression of Gli1) was associated with the reductions in Ift88 expression in the tibial cartilage from Ift88-deficient mice. Wheel exercise restored both the articular cartilage thickness and levels of Hh signaling in these mice.

Conclusion: Our results in a mouse model of OA demonstrate that IFT88 performs a chondroprotective role in articular cartilage by controlling the calcification of cartilage via maintenance of a threshold of Hh signaling during physiologic loading.
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http://dx.doi.org/10.1002/art.41894DOI Listing
January 2022

Comparison of LABORAS with static incapacitance testing for assessing spontaneous pain behaviour in surgically-induced murine osteoarthritis.

Osteoarthr Cartil Open 2020 Dec;2(4):100101

Centre for Osteoarthritis Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, University of Oxford, Roosevelt Drive, Oxford, UK.

Objective: Evoked responses following mechanical or thermal stimulation are typically used to assess pain behaviour in murine osteoarthritis (OA). However, there is no consensus on how best to measure spontaneous pain behaviour.

Method: OA by partial meniscectomy (PMX), or sham surgery was performed in 10-week old C57BL/6 male mice. Collagen-induced arthritis (CIA) was induced in 10 week old DBA1 male mice. Spontaneous pain behaviour, either at the time of active inflammatory disease (CIA), or over the 12 weeks after induction of OA, was assessed by static incapacitance testing (measuring percentage of weight placed through each hindlimb), and Laboratory Animal Behaviour Observation Registration and Analysis System (LABORAS) (translating cage vibrations of singly house animals into specific activities). Data were analysed by repeated measures two way ANOVA with post hoc testing comparing experimental groups with either sham operated or naïve controls.

Results: By incapacitance testing, two phases of painful behaviour were evident after PMX: a transient, post-operative phase, which resolved within one week, and a late OA pain phase starting 8 weeks post surgery and reaching statistical significance at week 12 (95% CI: sham 89.51-98.19, PMX 76.18-98.16). LABORAS, was able to detect pain behaviour in mice with CIA, but no statistically significant pain behaviour was observed in OA mice either post operatively (once analgesia had been controlled for) or at any later time points for any activity compared with the sham group.

Conclusion: Static incapacitance testing is superior to LABORAS for measuring spontaneous pain behaviour in surgically induced murine OA.
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http://dx.doi.org/10.1016/j.ocarto.2020.100101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762826PMC
December 2020

Peripheral pain mechanisms in osteoarthritis.

Authors:
Tonia L Vincent

Pain 2020 09;161 Suppl 1:S138-S146

NDORMS, Centre for OA Pathogenesis Versus Arthritis, Kennedy Institute for Rheumatology, University of Oxford, Oxford, United Kingdom.

There is a well-established historical observation that structural joint damage by plain X-ray correlates poorly with symptomatic disease in osteoarthritis (OA). This is often attributed to the inability to visualise soft-tissue pathology within the joint and the recognition of heterogeneous patient factors that drive central pain sensitisation. A major issue is the relative paucity of mechanistic studies in which molecular pathogenesis of pain is interrogated in relation to tissue pathology. Nonetheless, in recent years, three broad approaches have been deployed to attempt to address this: correlative clinical studies of peripheral and central pain outcomes using magnetic resonance imaging, where soft-tissue processes can be visualised; molecular studies on tissue from patients with OA; and careful molecular interrogation of preclinical models of OA across the disease time course. Studies have taken advantage of established clinical molecular targets such as nerve growth factor. Not only is the regulation of nerve growth factor within the joint being used to explore the relationship between tissue pathology and the origins of pain in OA, but it also provides a core model on which other molecules present within the joint can modulate the pain response. In this narrative review, how molecular and pathological tissue change relates to joint pain in OA will be discussed. Finally, a model for how tissue damage may lead to pain over the disease course will be proposed.
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http://dx.doi.org/10.1097/j.pain.0000000000001923DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7434216PMC
September 2020

TSG-6 Is Weakly Chondroprotective in Murine OA but Does not Account for FGF2-Mediated Joint Protection.

ACR Open Rheumatol 2020 Oct 7;2(10):605-615. Epub 2020 Oct 7.

Kennedy Institute of Rheumatology, Arthritis Research UK Centre for OA Pathogenesis, University of Oxford, UK.

Objective: Tumor necrosis factor α-stimulated gene 6 (TSG-6) is an anti-inflammatory protein highly expressed in osteoarthritis (OA), but its influence on the course of OA is unknown.

Methods: Cartilage injury was assessed by murine hip avulsion or by recutting rested explants. Forty-two previously validated injury genes were quantified by real-time polymerase chain reaction in whole joints following destabilization of the medial meniscus (DMM) (6 hours and 7 days). Joint pathology was assessed at 8 and 12 weeks following DMM in 10-week-old male and female fibroblast growth factor 2 (FGF2) , TSG-6 , TSG-6 (overexpressing), FGF2 ;TSG-6 (8 weeks only) mice, as well as strain-matched, wild-type controls. In vivo cartilage repair was assessed 8 weeks following focal cartilage injury in TSG-6 and control mice. FGF2 release following cartilage injury was measured by enzyme-linked immunosorbent assay.

Results: TSG-6 messenger RNA upregulation was strongly FGF2-dependent upon injury in vitro and in vivo. Fifteeen inflammatory genes were significantly increased in TSG-6 joints, including IL1α, Ccl2, and Adamts5 compared with wild type. Six genes were significantly suppressed in TSG-6 joints including Timp1, Inhibin βA, and podoplanin (known FGF2 target genes). FGF2 release upon cartilage injury was not influenced by levels of TSG-6. Cartilage degradation was significantly increased at 12 weeks post-DMM in male TSG-6 mice, with a nonsignificant 30% reduction in disease seen in TSG-6 mice. No differences were observed in cartilage repair between genotypes. TSG-6 overexpression was unable to prevent accelerated OA in FGF2 mice.

Conclusion: TSG-6 influences early gene regulation in the destabilized joint and exerts a modest late chondroprotective effect. Although strongly FGF2 dependent, TSG-6 does not explain the strong chondroprotective effect of FGF2.
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http://dx.doi.org/10.1002/acr2.11176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7571392PMC
October 2020

Of mice and men: converging on a common molecular understanding of osteoarthritis.

Authors:
Tonia L Vincent

Lancet Rheumatol 2020 Oct 23;2(10):e633-e645. Epub 2020 Sep 23.

Centre for Osteoarthritis Pathogenesis, Versus Arthritis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

Despite an increasing burden of osteoarthritis in developed societies, target discovery has been slow and there are currently no approved disease-modifying osteoarthritis drugs. This lack of progress is due in part to a series of misconceptions over the years: that osteoarthritis is an inevitable consequence of ageing, that damaged articular cartilage cannot heal itself, and that osteoarthritis is driven by synovial inflammation similar to that seen in rheumatoid arthritis. Molecular interrogation of disease through ex-vivo tissue analysis, in-vitro studies, and preclinical models have radically reshaped the knowledge landscape. Inflammation in osteoarthritis appears to be distinct from that seen in rheumatoid arthritis. Recent randomised controlled trials, using treatments repurposed from rheumatoid arthritis, have largely been unsuccessful. Genome-wide studies point to defects in repair pathways, which accords well with recent promise using growth factor therapies or Wnt pathway antagonism. Nerve growth factor has emerged as a robust target in osteoarthritis pain in phase 2-3 trials. These studies, both positive and negative, align well with those in preclinical surgical models of osteoarthritis, indicating that pathogenic mechanisms identified in mice can lead researchers to valid human targets. Several novel candidate pathways are emerging from preclinical studies that offer hope of future translational impact. Enhancing trust between industry, basic, and clinical scientists will optimise our collective chance of success.
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http://dx.doi.org/10.1016/S2665-9913(20)30279-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7511206PMC
October 2020

Does Pain at an Earlier Stage of Chondropathy Protect Female Mice Against Structural Progression After Surgically Induced Osteoarthritis?

Arthritis Rheumatol 2020 12 4;72(12):2083-2093. Epub 2020 Nov 4.

Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

Objective: Female C57BL/6 mice exhibit less severe chondropathy than male mice. This study was undertaken to test the robustness of this observation and explore underlying mechanisms.

Methods: Osteoarthritis was induced in male and female C57BL/6 or DBA/1 mice (n = 6-15 per group) by destabilization of the medial meniscus (DMM) or partial meniscectomy (PMX). Some mice were ovariectomized (OVX) (n = 30). In vivo repair after focal cartilage defect or joint immobilization (sciatic neurectomy) following DMM was assessed. Histologic analysis, evaluation of gene expression in whole knees, and behavioral analysis using Laboratory Animal Behavior Observation Registration and Analysis System (LABORAS) and Linton incapacitance testing (n = 7-10 mice per group) were performed.

Results: Female mice displayed less severe chondropathy (20-75% reduction) across both strains and after both surgeries. Activity levels after PMX were similar for male and female mice. Some repair-associated genes were increased in female mouse joints after surgery, but no repair differences were evident in vivo. Despite reduced chondropathy, female mice developed pain-like behavior at the same time as male mice. At the time of established pain-like behavior (10 weeks after PMX), pain-associated genes were significantly up-regulated in female mice, including Gdnf (mean ± SEM fold change 2.54 ± 0.30), Nrtn (6.71 ± 1.24), Ntf3 (1.92 ± 0.27), and Ntf5 (2.89 ± 0.48) (P < 0.01, P < 0.01, P < 0.05, and P < 0.001, respectively, versus male mice). Inflammatory genes were not regulated in painful joints in mice of either sex.

Conclusion: We confirm strong structural joint protection in female mice that is not due to activity or intrinsic repair differences. Female mice develop pain at the same time as males, but induce a distinct set of neurotrophins. We speculate that heightened pain sensitivity in female mice protects the joint by preventing overuse.
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http://dx.doi.org/10.1002/art.41421DOI Listing
December 2020

Ciliary proteins specify the cell inflammatory response by tuning NFκB signalling, independently of primary cilia.

J Cell Sci 2020 07 8;133(13). Epub 2020 Jul 8.

Kennedy Institute of Rheumatology Research, Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Sciences, Medical Sciences Division, University of Oxford, Oxford OX3 7FY, UK

Complex inflammatory signalling cascades define the response to tissue injury but also control development and homeostasis, limiting the potential for these pathways to be targeted therapeutically. Primary cilia are subcellular regulators of cellular signalling, controlling how signalling is organized, encoded and, in some instances, driving or influencing pathogenesis. Our previous research revealed that disruption of ciliary intraflagellar transport (IFT), altered the cell response to IL-1β, supporting a putative link emerging between cilia and inflammation. Here, we show that IFT88 depletion affects specific cytokine-regulated behaviours, changing cytosolic NFκB translocation dynamics but leaving MAPK signalling unaffected. RNA-seq analysis indicates that IFT88 regulates one third of the genome-wide targets, including the pro-inflammatory genes , and Through microscopy, we find altered NFκB dynamics are independent of assembly of a ciliary axoneme. Indeed, depletion of IFT88 inhibits inflammatory responses in the non-ciliated macrophage. We propose that ciliary proteins, including IFT88, KIF3A, TTBK2 and NPHP4, act outside of the ciliary axoneme to tune cytoplasmic NFκB signalling and specify the downstream cell response. This is thus a non-canonical function for ciliary proteins in shaping cellular inflammation.This article has an associated First Person interview with the first author of the paper.
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http://dx.doi.org/10.1242/jcs.239871DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7358134PMC
July 2020

Design and Evaluation of Magnetic Hall Effect Tactile Sensors for Use in Sensorized Splints.

Sensors (Basel) 2020 Feb 19;20(4). Epub 2020 Feb 19.

School of Mechanical Engineering, University of Leeds, Leeds LS2 9JT, UK.

Splinting techniques are widely used in medicine to inhibit the movement of arthritic joints. Studies into the effectiveness of splinting as a method of pain reduction have generally yielded positive results, however, no significant difference has been found in clinical outcomes between splinting types. Tactile sensing has shown great promise for the integration into splinting devices and may offer further information into applied forces to find the most effective methods of splinting. Hall effect-based tactile sensors are of particular interest in this application owing to their low-cost, small size, and high robustness. One complexity of the sensors is the relationship between the elastomer geometry and the measurement range. This paper investigates the design parameters of Hall effect tactile sensors for use in hand splinting. Finite element simulations are used to locate the areas in which sensitivity is high in order to optimise the deflection range of the sensor. Further simulations then investigate the mechanical response and force ranges of the elastomer layer under loading which are validated with experimental data. A 4 mm radius, 3 mm-thick sensor is identified as meeting defined sensing requirements for range and sensitivity. A prototype sensor is produced which exhibits a pressure range of 45 kPa normal and 6 kPa shear. A proof of principle prototype demonstrates how this can be integrated to form an instrumented splint with multi-axis sensing capability and has the potential to inform clinical practice for improved splinting.
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http://dx.doi.org/10.3390/s20041123DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7070306PMC
February 2020

Mechanoflammation in osteoarthritis pathogenesis.

Authors:
Tonia L Vincent

Semin Arthritis Rheum 2019 12;49(3S):S36-S38

Kennedy Institute of Rheumatology, Centre for OA Pathogenesis Versus Arthritis, University of Oxford, Roosevelt Drive, OX3 7FY, United Kingdom. Electronic address:

Mechanical injury is the most important risk factor in osteoarthritis (OA) development. Although once considered a passive disease of mechanical attrition, injury drives active mechanosensitive intracellular signalling which affects the structural and symptomatic course of disease. Mechanosensitive signalling in cartilage has been elucidated over the years and two principal responses emerge: those that cause the release of growth factors from the matrix and which stimulate repair, and those that drive inflammatory signalling, a process that we have termed "mechanoflammation". The up-stream activator of mechanoflammation remains unknown, but it results in rapid activation of NFkB and the inflammatory mitogen activated protein (MAP) kinases and this controls the bioavailability of aggrecanase and regulation of nerve growth factor (NGF), causing pain. The precise relationship between mechanoflammation and cartilage repair is currently unclear but it is likely that chronic mechanoflammation will contribute to disease by also suppressing intrinisic tissue repair.
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http://dx.doi.org/10.1016/j.semarthrit.2019.09.018DOI Listing
December 2019

IL-1 in osteoarthritis: time for a critical review of the literature.

Authors:
Tonia L Vincent

F1000Res 2019 21;8. Epub 2019 Jun 21.

Centre for OA Pathogenesis Versus Arthritis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, OX3 7FY, UK.

The concept of interleukin-1 (IL-1) as a target in osteoarthritis (OA) has been an attractive one for many years. It is a highly potent inducer of cartilage degradation, causing the induction of mRNA and controlling the bioavailability of disease-relevant proteases such as ADAMTS5 and MMP13. It drives synovitis and can induce other disease-relevant genes such as nerve growth factor, a key pain sensitiser in OA. However, the quality of evidence for its involvement in disease is modest. Descriptive studies have demonstrated expression of IL-1α and β in OA cartilage and elevated levels in the synovial fluid of some patients. Agnostic transcriptomic and genomic analyses do not identify IL-1 as a key pathway. models show a conflicting role for this molecule; early studies using therapeutic approaches in large animal models show a benefit, but most murine studies fail to demonstrate protection where the ligands (IL-1α/β), the cytokine activator (IL-1-converting enzyme), or the receptor (IL-1R) have been knocked out. Recently, a number of large double-blind randomised controlled clinical studies targeting IL-1 have failed. Enthusiasm for IL-1 as a target in OA is rapidly dwindling.
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http://dx.doi.org/10.12688/f1000research.18831.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6589928PMC
June 2020

Active immunisation targeting nerve growth factor attenuates chronic pain behaviour in murine osteoarthritis.

Ann Rheum Dis 2019 05 12;78(5):672-675. Epub 2019 Mar 12.

Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK

Objectives: Nerve growth factor (NGF) has emerged as a key driver of pain in osteoarthritis (OA) and antibodies to NGF are potent analgesics in human disease. Here, we validate a novel vaccine strategy to generate anti-NGF antibodies for reversal of pain behaviour in a surgical model of OA.

Methods: Virus-like particles were derived from the cucumber mosaic virus (CuMV) and coupled to expressed recombinant NGF to create the vaccine. 10-week-old male mice underwent partial meniscectomy to induce OA or sham-surgery. Spontaneous pain behaviour was measured by Linton incapacitance and OA severity was quantified using OARSI histological scoring. Mice (experimental and a sentinel cohort) were inoculated with CuMVtt (Vax) or CuMVtt (Mock) either before surgery or once pain was established. Efficacy of anti-NGF from the plasma of sentinel vaccinated mice was measured in vitro using a neurite outgrowth assay in PC12 cells.

Results: Anti-NGF titres were readily detectable in the vaccinated but not mock vaccinated mice. Regular boosting with fresh vaccine was required to maintain anti-NGF titres as measured in the sentinel cohort. Both prophylactic and therapeutic vaccination demonstrated a reversal of pain behaviour by incapacitance testing, and a meta-analysis of the two studies showing analgesia at peak anti-NGF titres was highly statistically significant. Serum anti-NGF was able to inhibit neurite outgrowth equivalent to around 150 ug/mL of recombinant monoclonal antibody.

Conclusions: This study demonstrates therapeutic efficacy of a novel NGF vaccine strategy that reversibly alleviates spontaneous pain behaviour in surgically induced murine OA.
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http://dx.doi.org/10.1136/annrheumdis-2018-214489DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6517802PMC
May 2019

Heparan Sulfate Proteoglycan Synthesis Is Dysregulated in Human Osteoarthritic Cartilage.

Am J Pathol 2019 03 14;189(3):632-647. Epub 2018 Dec 14.

Arthritis Research UK Centre for Osteoarthritis Pathogenesis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, United Kingdom; Norwich Medical School, University of East Anglia, Norwich, United Kingdom. Electronic address:

Osteoarthritis (OA) is a common degenerative joint disease, characterized by cartilage loss and subchondral bone remodeling in response to abnormal mechanical load. Heparan sulfate (HS) proteoglycans bind to many proteins that regulate cartilage homeostasis, including growth factors, morphogens, proteases, and their inhibitors, and modulate their localization, retention, and biological activity. Changes in HS expression and structure may thus have important consequences for joint health. We analyzed normal and osteoarthritic human knee cartilage, and found HS biosynthesis was markedly disrupted in OA, with 45% of the 38 genes analyzed differentially regulated in diseased cartilage. The expression of several HS core proteins, biosynthesis, and modification enzymes was increased in OA cartilage, whereas the expression of the HS proteoglycans syndecan 4 and betaglycan was reduced. The structure of HS was also altered, with increased levels of 6-O-sulfation in osteoarthritic samples, which correlated with increased expression of HS6ST1, a 6-O-sulfotransferase, and GLCE, an epimerase that promotes 6-O-sulfation. siRNA silencing of HS6ST1 expression in primary OA chondrocytes inhibited extracellular signal-regulated kinase phosphorylation in response to fibroblast growth factor 2, showing that changes in 6-O-sulfation impact a key cartilage signaling pathway. Given the broad range of homeostatic and repair pathways that HS regulates, these changes in proteoglycan expression and HS structure are likely to have significant effects on joint health and progression of OA.
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http://dx.doi.org/10.1016/j.ajpath.2018.11.011DOI Listing
March 2019

Hand osteoarthritis: clinical phenotypes, molecular mechanisms and disease management.

Nat Rev Rheumatol 2018 11;14(11):641-656

Research Institute for Primary Care and Health Sciences, Arthritis Research UK Primary Care Centre, David Weatherall Building, Keele University, Keele, UK.

Osteoarthritis (OA) is a highly prevalent condition, and the hand is the most commonly affected site. Patients with hand OA frequently report symptoms of pain, functional limitations and frustration in undertaking everyday activities. The condition presents clinically with changes to the bone, ligaments, cartilage and synovial tissue, which can be observed using radiography, ultrasonography or MRI. Hand OA is a heterogeneous disorder and is considered to be multifactorial in aetiology. This Review provides an overview of the epidemiology, presentation and burden of hand OA, including an update on hand OA imaging (including the development of novel techniques), disease mechanisms and management. In particular, areas for which new evidence has substantially changed the way we understand, consider and treat hand OA are highlighted. For example, genetic studies, clinical trials and careful prospective imaging studies from the past 5 years are beginning to provide insights into the pathogenesis of hand OA that might uncover new therapeutic targets in the disease.
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http://dx.doi.org/10.1038/s41584-018-0095-4DOI Listing
November 2018

Studying Osteoarthritis Pathogenesis in Mice.

Curr Protoc Mouse Biol 2018 Dec 21;8(4):e50. Epub 2018 Sep 21.

Disease Model Discovery, Mammalian Genetics Unit, MRC Harwell Institute, Oxfordshire, United Kingdom.

With the increasing availability and complexity of mouse models of disease, either spontaneous or induced, there is a concomitant increase in their use in the analysis of pathogenesis. Among such diseases is osteoarthritis, a debilitating disease with few treatment options. While advances in our understanding of the pathogenesis of osteoarthritis has advanced through clinical investigations and genome-wide association studies, there is still a large gap in our knowledge, hindering advances in therapy. Patient samples are available ex vivo, but these are generally in the very late stages of disease. However, with mice, we are able to induce disease at a defined time and track the progression in vivo and ex vivo, from inception to end stage, to delineate the processes involved in disease development. © 2018 by John Wiley & Sons, Inc.
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http://dx.doi.org/10.1002/cpmo.50DOI Listing
December 2018

Connective tissue growth factor contributes to joint homeostasis and osteoarthritis severity by controlling the matrix sequestration and activation of latent TGFβ.

Ann Rheum Dis 2018 09 20;77(9):1372-1380. Epub 2018 Jun 20.

Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

Objectives: One mechanism by which cartilage responds to mechanical load is by releasing heparin-bound growth factors from the pericellular matrix (PCM). By proteomic analysis of the PCM, we identified connective tissue growth factor (CTGF) and here investigate its function and mechanism of action.

Methods: Recombinant CTGF (rCTGF) was used to stimulate human chondrocytes for microarray analysis. Endogenous CTGF was investigated by in vitro binding assays and confocal microscopy. Its release from cut cartilage (injury CM) was analysed by Western blot under reducing and non-reducing conditions. A postnatal, conditional Ctgf mouse was generated for cartilage injury experiments and to explore the course of osteoarthritis (OA) by destabilisation of the medial meniscus. siRNA knockdown was performed on isolated human chondrocytes.

Results: The biological responses of rCTGF were TGFβ dependent. CTGF displaced latent TGFβ from cartilage and both were released on cartilage injury. CTGF and latent TGFβ migrated as a single high molecular weight band under non-reducing conditions, suggesting that they were in a covalent (disulfide) complex. This was confirmed by immunoprecipitation. Using Ctgf mice, CTGF was required for sequestration of latent TGFβ in the matrix and activation of the latent complex at the cell surface through TGFβR3. In vivo deletion of CTGF increased the thickness of the articular cartilage and protected mice from OA.

Conclusions: CTGF is a latent TGFβ binding protein that controls the matrix sequestration and activation of TGFβ in cartilage. Deletion of CTGF in vivo caused a paradoxical increase in Smad2 phosphorylation resulting in thicker cartilage that was protected from OA.
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http://dx.doi.org/10.1136/annrheumdis-2018-212964DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6104679PMC
September 2018

Mechanoadaptation: articular cartilage through thick and thin.

J Physiol 2019 03 29;597(5):1271-1281. Epub 2018 Jul 29.

Arthritis Research UK Centre for OA Pathogenesis, Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK.

The articular cartilage is exquisitely sensitive to mechanical load. Its structure is largely defined by the mechanical environment and destruction in osteoarthritis is the pathophysiological consequence of abnormal mechanics. It is often overlooked that disuse of joints causes profound loss of volume in the articular cartilage, a clinical observation first described in polio patients and stroke victims. Through the 1980s, the results of studies exploiting experimental joint immobilisation supported this. Importantly, this substantial body of work was also the first to describe metabolic changes that resulted in decreased synthesis of matrix molecules, especially sulfated proteoglycans. The molecular mechanisms that underlie disuse atrophy are poorly understood despite the identification of multiple mechanosensing mechanisms in cartilage. Moreover, there has been a tendency to equate cartilage loss with osteoarthritic degeneration. Here, we review the historic literature and clarify the structural, metabolic and clinical features that clearly distinguish cartilage loss due to disuse atrophy and those due to osteoarthritis. We speculate on the molecular sensing pathways in cartilage that may be responsible for cartilage mechanoadaptation.
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http://dx.doi.org/10.1113/JP275451DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6395418PMC
March 2019

Functional Characterization of the Osteoarthritis Genetic Risk Residing at ALDH1A2 Identifies rs12915901 as a Key Target Variant.

Arthritis Rheumatol 2018 10 23;70(10):1577-1587. Epub 2018 Aug 23.

Newcastle University, Newcastle upon Tyne, UK.

Objective: To identify the functional single-nucleotide polymorphisms (SNPs) and mechanisms conferring increased risk of hand osteoarthritis (OA) at the ALDH1A2 locus, which is a retinoic acid regulatory gene.

Methods: Tissue samples from 247 patients with knee, hip, or hand OA who had undergone joint surgery were included. RNA-sequencing analysis was used to investigate differential expression of ALDH1A2 and other retinoic acid signaling pathway genes in cartilage. Expression of ALDH1A2 in joint tissues obtained from multiple sites was quantified using quantitative reverse transcription-polymerase chain reaction. Allelic expression imbalance (AEI) was measured by pyrosequencing. The consequences of ALDH1A2 depletion by RNA interference were assessed in primary human chondrocytes. In silico and in vitro analyses were used to pinpoint which, among 62 highly correlated SNPs, could account for the association at the locus.

Results: ALDH1A2 expression was observed across multiple joint tissue samples, including osteochondral tissue from the hand. The expression of ALDH1A2 and of several retinoic acid signaling genes was different in diseased cartilage compared to non-diseased cartilage, with ALDH1A2 showing lower levels in OA cartilage. Experimental depletion of ALDH1A2 resulted in changes in the expression levels of a number of chondrogenic markers, including SOX9. In addition, reduced expression of the OA risk-conferring allele was witnessed in a number of joint tissues, with the strongest effect in cartilage. The intronic SNP rs12915901 recapitulated the AEI observed in patient tissues, while the Ets transcription factors were identified as potential mediators of this effect.

Conclusion: The ALDH1A2 locus seems to increase the risk of hand OA through decreased expression of ALDH1A2 in joint tissues, with the effect dependent on rs12915901. These findings indicate a mechanism that may now be targeted to modulate OA risk.
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http://dx.doi.org/10.1002/art.40545DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6175168PMC
October 2018
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