Rhodamine B and 2-acetamido-1,3,6-tri-O-acetyl-4-deoxy-4-fluoro-d-glucopyranose (F-GlcNAc) inhibit chondroitin/dermatan and keratan sulphate synthesis by different mechanisms in bovine chondrocytes

• Published in: Molecular genetics and metabolism 2012-06, Vol.106 (2), p.214-220
• Main Authors: Derrick-Roberts, Ainslie L.K., Marais, Wanda, Byers, Sharon
• Format: Article
• Language: English
• Subjects: Acetylglucosamine - analogs & derivatives; Acetylglucosamine - pharmacology; Animals; Cartilage - metabolism; Cattle; Chondrocytes - drug effects; Chondrocytes - metabolism; Chondroitin Sulfates - biosynthesis; Dermatan Sulfate - biosynthesis; F-GlcNAc; Gene Expression Profiling; Gene Expression Regulation - drug effects; Glycosaminoglycan; Glycosaminoglycans - biosynthesis; Keratan Sulfate - biosynthesis; Mucopolysaccharidoses - genetics; Mucopolysaccharidoses - metabolism; Mucopolysaccharidosis; Proteoglycans - metabolism; Rhodamine B; Rhodamines - pharmacology; Substrate deprivation therapy
• ISSN: 1096-7192; 1096-7206
• DOI: 10.1016/j.ymgme.2012.04.002

MPS disorders result from a deficiency or absence of glycosaminoglycan (GAG) degrading enzymes leading to an imbalance between the synthesis and degradation of GAGs and their subsequent accumulation in a range of cells. The inhibition of GAG synthesis using small chemical inhibitors has been proposed as a novel therapeutic approach to treatment. Several inhibitors have been shown to decrease heparan sulphate GAG synthesis and in this study we evaluated a novel fluorinated analog of N-acetylglucosamine (2-acetamido-1,3,6-tri-O-acetyl-4-deoxy-4-fluoro-d-glucopyranose (F-GlcNAc)) and rhodamine B for their ability to also inhibit the synthesis of chondroitin/dermatan and keratan sulphate GAGs present in bovine cartilage. Both inhibitors decreased GAG synthesis in chondrocyte monolayer culture and in cartilage chip explant culture in a dose dependent manner. Both inhibitors decreased the size of newly synthesised proteoglycans and in the case of F-GlcNAc this was due to a decrease in newly synthesised GAG chain size. Rhodamine B, however, did not affect GAG chain size, while both inhibitors decreased the amount of chondroitin/dermatan and keratan sulphate GAG equally. The expression of genes responsible for the initiation and elongation of chondroitin/dermatan sulphate and keratan sulphate GAGs were downregulated in the presence of rhodamine B but not in the presence of F-GlcNAc. Thus the 2 inhibitors appear to have differing effects on GAG synthesis, with F-GlcNAc inhibiting the epimerisation of UDP-GlcNAc to UDP-GalNAc thus decreasing the availability of monosaccharides for addition to the growing GAG chain, whereas rhodamine B is more likely to reduce the number of GAG chains. Together with previous data these 2 inhibitors are capable of non-specific inhibition of GAG synthesis, reducing the production of chondroitin/dermatan sulphate, keratan sulphate and heparan sulphate GAGs. As such they would be applicable to therapy in a range of MPS disorders. ► F-GlcNAc and rhodamine B inhibit CS/DS and KS GAG synthesis in bovine cartilage. ► Inhibitors decreased GAG synthesis in chondrocyte monolayer and cartilage chip explant culture. ► F-GlcNAc decreased GAG chain size whereas rhodamine B decreased amount of GAG chains. ► Both inhibitors decreased the amount of CS/DS and KS GAG equally. ► Rhodamine B and F-GlcNAc have different mechanisms of action in reducing GAG synthesis.