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Objectives: \ud 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.\ud \ud Methods: \ud 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.\ud \ud Results: \ud 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.\ud \ud Conclusions: \ud 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.

Background:\ud 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.\ud \ud Purpose:\ud 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.\ud \ud Study Design:\ud Controlled laboratory study.\ud \ud Methods:\ud 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.\ud \ud Results:\ud Vancomycin treatment at 5 mg/mL significantly reduced tenocyte viability in vitro after 60 minutes of treatment (P < .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; P < .05).\ud \ud Conclusion:\ud 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.\ud \ud Clinical Relevance:\ud Vancomycin ACL wrap does not alter the molecular structure of the ACL hamstring graft and may improve graft integrity.

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.\ud \ud 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.\ud \ud 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.\ud \ud 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.

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

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.