TSG-6, an Arthritis-Associated Hyaluronan Binding Protein, Forms a Stable Complex with the Serum Protein Inter-.alpha.-inhibitor

Abstract

TSG-6 is a secreted 35-kDa glycoprotein, inducible by TNF and IL-1. The N-terminal portion of TSG-6 shows sequence homology to members of the cartilage link protein family of hyaluronan binding proteins. The C-terminal half of TSG-6 contains a so-called CUB domain, characteristic for developmentally regulated proteins. High levels of TSG-6 protein are found in the synovial fluid of patients with rheumatoid arthritis and some other arthritic diseases. Here we show that TSG-6 readily formed a complex with a protein present in human, bovine, rabbit, and mouse serum. This complex was stable during SDS-PAGE under reducing conditions, and in the presence of 8 M urea. The protein that binds TSG-6 was purified from human serum and identified as inter-alpha-inhibitor (I alpha I) by N-terminal microsequencing. Microsequencing of the complex itself revealed the presence of TSG-6 and two of the three polypeptide chains of I alpha I (bikunin and HC2). Experiments with recombinant TSG-6 and I alpha I purified from human serum showed that the TSG-6/I alpha I complex is rapidly formed even in the apparent absence of other proteins at 37 degrees C, but not at 4 degrees C. The TSG-6/I alpha I complex was cleaved by chondroitin sulfate ABC lyase, suggesting that cross-linking by chondroitin sulfate is required for the stability of the complex.(ABSTRACT TRUNCATED AT 250 WORDS)

Enhanced neutrophil extravasation and rapid progression of proteoglycan‐induced arthritis in TSG‐6–knockout mice

“…The antiinflammatory properties of Tnfip6 have been mainly attributed to its ability to potentiate the antiplasmin activity of I␣I upon association with this protease inhibitor, leading to the subsequent downregulation of the activity of a number of matrixdegrading proteases (7)(8)(9)(10). However, a recent study (11) found that the isolated Link module domain of Tnfip6, which lacks the ability to associate with I␣I, could still inhibit the influx of neutrophils to the site of inflammation.…”

“…The inhibitory effect of the Tnfip6-I␣I complex appears to be specific for plasmin, since no increase was observed in the inhibitory activity against other serine proteases (10). Since plasmin is a key activator of matrix-degrading metalloproteases, it has been postulated that Tnfip6 exerts its antiinflammatory and chondroprotective effects through the inactivation of the metalloprotease network (7,9,10).…”

“…Tnfip6 binds I␣I, a powerful serine protease inhibitor in serum, and this interaction has been shown to potentiate the inhibitory effect of I␣I on plasmin (9). Thus, plasmin activity was expected to be elevated in the joints with inflammation.…”

Molecular Heterogeneity of the SHAP-Hyaluronan Complex

“…In fractions 5-15 of the third gradient, no band was detected at the positions corresponding to ITI (180 kDa), P␣I (120 kDa), and the covalent ITI/TSG-6 complex (120 kDa) by Coomassie Blue staining and immunostaining. TSG-6 (tumor necrosis factor-stimulated gene 6), an inflammatory HABP of 35 kDa, has been shown to be present in synovial fluid of patients with rheumatoid arthritis and some other forms of arthritis but not in normal human joints (25). Interestingly, this molecule in the patient synovial fluid has been found to form a covalent complex with ITI molecules via the chondroitin sulfate chains (25).…”

TNF?-stimulated gene product (TSG-6) and its binding protein, I?I, in the human intervertebral disc: new molecules for the disc

“…TSG-6 is known to form a stable, $120 kDa, complex with inter-a-inhibitor (IaI), a 180 kDa serine protease inhibitor found in serum [23,31]. IaI-TSG-6 complexes of this size have been detected in the synovial fluids of arthritis patients [30].…”