Post-translational alterations in newly synthesized cartilage proteoglycans induced by the glutamine analogue 6-diazo-5-oxo-l-norleucine. Time course of inhibition and recovery

Abstract

Incorporation of [35S]sulphate by cultures of matrix-free cells from chick embryo sterna in the presence of the glutamine analogue 6-diazo-5-oxo-L-norleucine (0.58 mM) was inhibited in a time-dependent manner to less than 15% of that in control cultures after 2 h. Characterization of the major cartilage proteoglycan synthesized under these conditions showed that it contained few, if any, normal-sized chondroitin sulphate chains and only about half of the normal complement of substituted serine residues. Subsequent addition of D-glucosamine hydrochloride (final concn. 2 mM) resulted in a time-dependent recovery of [35S]sulphate incorporation to 90% of control cultures after 2 h, but restored the chondroitin sulphate chains to normal size within 15 min. On the basis of these results, it is concluded that a 2 h preincubation is necessary to deplete the chondrocytes of the endogenous supply of UDP-N-acetyl-D-glucosamine required for optimal glycoconjugate synthesis, and that this situation results in the synthesis of a chondroitin sulphate proteoglycan with significantly altered properties, owing to the paucity of glycosaminoglycan chains; however, this condition is completely reversible if the D-glucosamine pool is repleted.

Calcium signaling is required for ultrasound‐stimulated aggrecan synthesis by rat chondrocytes

“…Molecular analysis and morphological studies have confirmed the phenotype and genotype of these cells in that they produce cartilage-specific genes, and continue doing so for many days in culture without de-differentiating [29]. Chondrocytes have been shown to respond to various mechanical stimuli by an increase in gene expression and/or a rise in the rate of proteoglycan and collagen synthesis [6,15] Calcium may have a role in mediating these mechanical effects, as the calcium signaling pathway is shown to be involved in the process of mechanotransduction when other types of mechanical stimuli are used. Brighton et al have demonstrated that calcium signaling mechanism mediates the proliferative response of calvarial bone cells to mechanical stimulus in the form of biaxial strain [ 11.…”