Glycosaminoglycans Interact Selectively with Chemokines and Modulate Receptor Binding and Cellular Responses

Abstract

Chemokines selectively recruit and activate a variety of cells during inflammation. Interactions between cell surface glycosaminoglycans (GAGs) and chemokines drive the formation of haptotactic or immobilized gradients of chemokines at the site of inflammation, directing this recruitment. Chemokines bind to glycosaminoglycans on human umbilical vein endothelial cells (HUVECs) with affinities in the micromolar range: RANTES > MCP-1 > IL-8 > MIP-1alpha. This binding can be competed with by soluble glycosaminoglycans: heparin, heparin sulfate, chondroitin sulfate, and dermatan sulfate. RANTES binding showed the widest discrimination between glycosaminoglycans (700-fold), whereas MIP-1alpha was the least selective. Almost identical results were obtained in an assay using heparin sulfate beads as the source of immobilized glycosaminoglycan. The binding of chemokines to glycosaminoglycan fragments has a strong length dependence, and optimally requires both N- and O-sulfation. Isothermal titration calorimetry data confirm these results; IL-8 binds heparin fragments with a K(d) of 0.39-2.63 microM, and requires five saccharide units to bind each monomer of chemokine. In membranes from cells expressing the G-protein-coupled chemokine receptors CXCR1, CXCR2, and CCR1, soluble GAGs inhibit the binding of chemokine ligands to their receptors. Consistent with this, heparin and heparin sulfate could inhibit IL-8-induced neutrophil calcium flux. Chemokines can therefore form complexes with both cell surface and soluble GAGs; these interactions have different functions. Soluble GAG chemokines complexes are unable to bind the receptor, resulting in a block of the biological activity. Previously, we have shown that cell surface GAGs present chemokines to the G-protein-coupled receptors, by increasing the local concentration of protein. A model is presented which brings together all of these data. The selectivity in the chemokine-GAG interaction suggests selective disruption of the haptotactic gradient may be an achievable therapeutic approach in inflammatory disease.

Chemokines: attractive mediators of the immune response

“…Chemokines probably do not act as soluble entities, but as solid phase bound aggregates. Chemokines can bind proteoglycans through interactions with the glysoaminoglycan (GAG) component of the proteins [16]. The association of chemokines with GAGs of the extracelluar matrix and on the cell surface has important implications in terms of presentation and increasing local chemokine levels.…”

“…Using GAG deficient Chinese hamster ovary cells transfected with CCR1 and CCR5, Ali et al [17] demonstrated that surface GAGs increase the concentration of chemokines presented to the CCRs at the molecular level. CCL5 has been shown to bind GAGs with distinct specificity [16] suggesting that CCL5 may be crucial in mediating inflammatory responses. Furthermore, release of a GAG, chondroitin sulfate A, from platelets in vivo blocked CCL5 binding and CCR5 signaling [18].…”

Induction of a CXCL8 binding site on endothelial syndecan‐3 in rheumatoid synovium

“…The binding affinities have been variously reported in the nanomolar or micromolar range for CXCL8, CCL2, CCL3, and CCL5 (14)(15)(16). Chemokines display differences in their strength of interactions with glycosaminoglycans.…”

“…Chemokines display differences in their strength of interactions with glycosaminoglycans. For example, CXCL8 shows the following range: heparin Ͼ heparan sulfate Ͼ chondroitin sulfate ϭ dermatan sulfate (14). Different chemokines show variation in their affinities for glycosaminoglycans that are not solely based on charge considerations, suggesting selectivity in these interactions (14,17,18).…”

Interfering with the CCL2–glycosaminoglycan axis as a potential approach to modulate neuroinflammation

“…In the CNS GAGs/ proteoglycans can be found as ECM components and in the basement membrane [25,26]. Although specificity/ selectivity in the chemokine/GAG interaction has not been fully elucidated so far, the great variety in GAG structures on the one side and the reported different affinities for GAGs displayed towards chemokines on the other side, suggest that chemokines interact with disease specific GAG structures in vivo [27,28].…”