The Role of Tumor Necrosis Factor-Stimulated Gene 6 Protein (TSG-6) in Osteoarthritis

Links

Abstract

In our studies, we developed a novel method for extracting high quality nucleic acid from joint tissue that relies on finely grinding the desired regions of the knee tibial plateau under liquid nitrogen. By controlling the depth of drill penetration we could reliably separate and isolate articular cartilage and site-matched subchondral bone with high fidelity. We also set up a model system based on specific regions of interest within a joint corresponding to a gradient of disease severities; this system can be used to represent early (relative intact), intermediate (mild to moderate damaged) and late (severe damaged) disease within the knee joints. We performed microarray profiling to identify disease relevant genes and pathways associated with knee OA progression in human articular cartilage. Tumor Necrosis Factor-Stimulated Gene 6 (TSG-6) was identified to be significantly associated with OA progression and validated in both medial and lateral compartment dominant OA samples.

Next, we explored mechanisms underlying the association of TSG-6 with OA progression. We characterized the effects of TSG-6, in the presence and absence of interalpha-inhibitor (IαI), on anti-plasmin activity, evaluated for the in vivo presence of heavy chains (HCs) in cartilage matrix (evidence of TSG-6 activity), evaluated the gene expression of IαI components and TSG-6 in cartilage, and explored effects of TSG-6 on matrix assembly in vitro.

TSG-6 synergized with IαI to inhibit plasmin activity by 39.3%; the presence of HA (full length or fragments) reduced the inhibitory effect. TSG-6 activity was highly correlated (R = 0.6392, P = 0.0006) with TSG-6 protein concentrations in synovial fluid (SF) from knee OA joints. TSG-6 protein and RNA were highly expressed in damaged cartilage from knee tibial and meniscal cartilage and chondrocytes treated with cytokines. The components of the IαI complex, ITIH1, ITIH2, and AMBP genes were either not expressed or expressed at a low level in intact or damaged cartilage of OA joints and chondrocytes. Rate limiting amounts of IαI were demonstrated by spiking-in exogenous IαI into cartilage extracts. This suggested that the source of IαI for TSG-6 mediated HC transfer onto damaged cartilage is likely the synovial fluid and not the cartilage itself. Interestingly, SF TSG-6 activity was significantly positively associated with the inflammatory reactants TIMP-1, A2M, VEGF, VCAM-1, ICAM, MMP-3, TNFR2, cytokines IL-6, IL-8, IL-18, quantity of activated macrophages in synovium and soluble macrophage-associated marker CD14 and number of low molecular weight (proinflammatory) hyaluronan (HA) molecules. Moreover, TSG-6 impaired HA-aggrecan assembly, but TSG-6 mediated HA-HC formation reduced this negative effect.

IαI enhances the anti-protease activity of TSG6, preserves matrix assembly capabilities by enabling TSG6 transfer of HC to HA and mitigates inhibition of matrix assembly by TSG-6. During OA progression, inflammatory mediators increase production of TSG-6, but IαI originating outside cartilage acting in a rate-limiting manner and may only moderately interact with TSG-6 released from damaged cartilage. Along with previous work, these findings suggest that the net beneficial effect of TSG-6 is therefore dependent upon the availability of IαI in knee OA.

Cited Works (11)

Year	Entry
2010	Goldring MB, Goldring SR. Articular cartilage and subchondral bone in the pathogenesis of osteoarthritis. Annals NY Acad Sci. March 2010;1192(1):230-237.
2002	Ding M, Odgaard A, Danielsen CC, Hvid I. Mutual associations among microstructural, physical and mechanical properties of human cancellous bone. J Bone Joint Surg. August 2002;84B(6):900-907.
2006	Pritzker KPH, Gay S, Jimenez SA, Ostergaard K, Pelletier J-P, Revell PA, Salter D, van den Berg WB. Osteoarthritis cartilage histopathology: grading and staging. Osteoarthritis Cartilage. January 2006;14(1):13-29.
2008	Lawrence RC, Felson DT, Helmick CG, Arnold LM, Choi H, Deyo RA, Gabriel S, Hirsch R, Hochberg MC, Hunder GG, Jordan JM, Katz JN, Kremers HM, Wolfe F; National Arthritis Data Workgroup. Estimates of the prevalence of arthritis and other rheumatic conditions in the United States: part II. Arthritis Rheum. January 2008;58(1):26-35.
2011	Guilak F. Biomechanical factors in osteoarthritis. Best Pract Res Clin Rheumatol. December 2011;25(6):815-823.
1991	Grynpas MD, Alpert B, Katz I, Lieberman I, Pritzker KPH. Subchondral bone in osteoarthritis. Calcif Tiss Int. January 1991;49(1):20-26.
2004	Carter DR, Beaupré GS, Wong M, Smith RL, Andriacchi TP, Schurman DJ. The mechanobiology of articular cartilage development and degeneration. Clin Orthop Relat Res. October 2004;427(suppl):S69-S77.
2005	Lee JH, Fitzgerald JB, DiMicco MA, Grodzinsky AJ. Mechanical injury of cartilage explants causes specific time-dependent changes in chondrocyte gene expression. Arthritis Rheum. August 2005;52(8):2386-2395.
2004	Burr DB. Anatomy and physiology of the mineralized tissues: role in the pathogenesis of osteoarthrosis. Osteoarthritis Cartilage. 2004;12(suppl):S20-S30.
2014	Evans CH, Kraus VB, Setton LA. Progress in intra-articular therapy. Nat Rev Rheumatol. January 2014;10(1):11-22.
2009	Fox AJS, Bedi A, Rodeo SA. The basic science of articular cartilage: structure, composition, and function. Sports Health. November–December 2009;1(6):461-468.