Chondroitin sulfate (CS) is a glycosaminoglycan (GAG) side chain of proteoglycans (PGs) which are widely distributed in the extracellular matrix and at cell surface. CS shows a highly structural diversity in not only molecular weight (MW) but sulfonation pattern. CS has been reported to exert anti-inflammatory activity by having effects on cytokine production by helper T cells. In this study, we focused on the structures of CS chains, especially MW of CS, and investigated effect of the different MW of CS on binding affinity with L-selectin and cytokine production by murine splenocytes. Firstly, we fractionated CS by employing gel filtration chromatography and obtained several CS fractions with different MW. Then the interaction between fractionated CS and L-selectin was analyzed by surface plasmon resonance (SPR). Finally, the influence of MW of CS on cytokine production by murine splenocytes was investigated in vitro. The results showed that interferon-gamma production was significantly increased by mouse splenocytes cocultivated with CS. On the contrary, CS inhibited interleukin 5 production by murine splenocytes depending on MW of the cocultivated CS. These results strongly indicate the existence of the optimal molecular size for an anti-inflammatory effect of CS through cytokine production by murine splenocytes. 1. Introduction Chondroitin sulfate (CS) belongs to a family of glycosaminoglycans (GAG), which is a linear, sulfonated polysaccharide, such as heparin (HP), heparan sulfate (HS), and dermatan sulfate (DS). CS is composed of a repeating disaccharide unit, [-4) GlcA (β1-3) GalNAc (β1-]n, where GlcA is glucuronate and GalNAc is N-acetylgalactosamine [1]. CS chains show structural diversity because sulfonate groups can be bound to variety positions of the disaccharide units, and its molecular weight (MW) is also highly variable due to different numbers of the disaccharide unit [2–4]. On the other hand, it is well known that CS and other GAGs form conjugates with the specific proteins called proteoglycans (PGs) and PGs are ubiquitously distributed in not only extracellular matrix but also on cell surface in animal tissues. PGs are thought to be participating in cell-cell communications such as a cell adhesion and signalling events via proteins, especially; HS chains can control a signalling by interacting with cell growth factors and HP has been widely used as an anticoagulant agent [5–7]. Correspondingly, CS has also a similar function and it has been reported that CS contributes to various bioactivities, for example, development of central
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