A synthetic glycosaminoglycan mimetic (RGTA) modifies natural glycosaminoglycan species during myogenesis

Abstract

Crucial events in myogenesis rely on the highly regulated spatiotemporal distribution of cell surface heparan sulfate proteoglycans to which are associated growth factors, thus creating a specific microenvironment around muscle cells. Most growth factors involved in control of myoblast growth and differentiation are stored in the extracellular matrix through interaction with specific sequences of glycosaminoglycan oligosaccharides, mainly heparan sulfate (HS). Different HS subspecies revealed by specific antibodies, have been shown to provide spatiotemporal regulation during muscle development. We have previously shown that glycosaminoglycan (GAG) mimetics called RGTA (ReGeneraTing Agent), stimulate muscle precursor cell growth and differentiation. These data suggest an important role of GAGs during myogenesis; however, little is yet known about the different species of GAGs synthesized during myogenesis and their metabolic regulation. We therefore quantified GAGs during myogenesis of C2.7 cells and show that the composition of GAG species was modified during myogenic differentiation. In particular, HS levels were increased during this process. In addition, the GAG mimetic RGTA, which stimulated both growth and differentiation of C2.7 cells, increased the total amount of GAG produced by these cells without significantly altering their rate of sulfation. RGTA treatment further enhanced HS levels and changed its sub-species composition. Although mRNA levels of the enzymes involved in HS biosynthesis were almost unchanged during myogenic differentiation, heparanase mRNA levels decreased. RGTA did not markedly alter these levels. Here we show that the effects of RGTA on myoblast growth and differentiation are in part mediated through an alteration of GAG species and provide an important insight into the role of these molecules in normal or pathologic myogenic processes.

Glycosaminoglycans and their synthetic mimetics inhibit RANTES-induced migration and invasion of human hepatoma cells

“…One of the molecular mechanisms of such an effect probably relies on a direct binding of glycosaminoglycan mimetics to RANTES/CCL5, which could result in an inhibition of the chemokine binding to glycosaminoglycans carried by proteoglycans expressed at the cell surface. However, it has been shown that the glycosaminoglycan mimetic cell treatment induced the changes in glycosaminoglycan amount, composition, and localization in C2.7 myoblast cultures undergoing differentiation (19). Therefore, it cannot be excluded that, in the present study, OTR4120 or OTR4131 could also interfere by this way with RANTES binding to the cells, resulting in the decreased chemotactic effects.…”

“…Synthetic polymers, called RGTA (for regenerating agent; refs. 19,20,30), have been engineered to mimic the stabilizing properties of glycosaminoglycans toward heparin-binding growth factors. OTR4120 and OTR4131 were tested in our assays.…”

A Synthetic Glycosaminoglycan Mimetic Binds Vascular Endothelial Growth Factor and Modulates Angiogenesis

“…Synthesis of OTR4120-OTR4120 is a carboxymethyl-dextran sulfate with a degree of substitution of 1.13 for sulfate residues and 0.46 for carboxymethyl residues, for a maximal degree of substitution of 3.0 (29,30). These substitution degrees are similar to those assayed in heparin used as a control.…”

Insights on a new path of pre-mitochondrial apoptosis regulation by a glycosaminoglycan mimetic

“…OTR4120 was covalently bound to BSA in a potassium phosphate buffer (0.2 M, pH 8) medium as previously described. 40 The protein concentration was determined by the kit BCA protein assay (Pierce). The surface of 96 flat-well were coated overnight at 41C with 100 ml of OTR4120-BSA (20 mg/ml of equivalent BSA), prepared complexes in 50 mM TrisHCl, pH 7.4, supplemented with 12.5 mM EDTA.…”