Ng C K, Handley C J, Mason R M, Robinson H C
Department of Biochemistry, Monash University, Clayton, Vic., Australia.
Biochem J. 1989 Nov 1;263(3):761-7. doi: 10.1042/bj2630761.
The synthesis and distribution of hyaluronate and proteoglycan were studied in bovine articular cartilage in short-term explant culture with [3H]acetate and H2(35)SO4 as precursors. The incorporation of [3H]acetate into hyaluronate and sulphated glycosaminoglycans was linear with time, except that hyaluronate synthesis showed a marked lag at the beginning of the incubation. [3H]Hyaluronate represented 4-7% of the total [3H]glycosaminoglycans synthesized over a 6 h period. However, the distributions of [3H]hyaluronate and 3H-labelled sulphated glycosaminoglycans were different: about 50% of the newly synthesized [3H]hyaluronate appeared in the medium, compared with less than 5% of the 3H-labelled sulphated proteoglycans. A pulse-chase experiment revealed that the release of newly synthesized [3H]hyaluronate from cartilage was rapid. No difference was observed in the distribution of [3H]hyaluronate between medium and tissue by cartilage from either the superficial layer or the deep layer of articular cartilage. When articular cartilage was incubated with 0.4 mM-cycloheximide, proteoglycan synthesis was markedly inhibited, whereas the synthesis of hyaluronate was only partially inhibited and resulted in more of the newly synthesized hyaluronate being released into the medium. Analysis of the hydrodynamic size of [3H]hyaluronate isolated from cartilage on Sephacryl-1000 revealed one population that was eluted as a broad peak (Kav. less than 0.7), compared with two populations (Kav. greater than 0.5 and less than 0.5) appearing in the medium of cultures. These data suggest that hyaluronate is synthesized in excess of proteoglycan synthesis and that the hyaluronate that is not complexed with proteoglycans is rapidly lost from the tissue.
以[3H]乙酸盐和H2(35)SO4作为前体，在短期外植体培养中研究了牛关节软骨中透明质酸盐和蛋白聚糖的合成与分布。[3H]乙酸盐掺入透明质酸盐和硫酸化糖胺聚糖的过程与时间呈线性关系，只是透明质酸盐合成在孵育开始时显示出明显的延迟。在6小时的时间段内，[3H]透明质酸盐占合成的总[3H]糖胺聚糖的4-7%。然而，[3H]透明质酸盐和3H标记的硫酸化糖胺聚糖的分布不同：新合成的[3H]透明质酸盐约50%出现在培养基中，而3H标记的硫酸化蛋白聚糖则不到5%。脉冲追踪实验表明，新合成的[3H]透明质酸盐从软骨中快速释放。关节软骨表层或深层的软骨在培养基和组织之间的[3H]透明质酸盐分布没有差异。当关节软骨与0.4 mM的环己酰亚胺一起孵育时，蛋白聚糖合成受到明显抑制，而透明质酸盐合成仅受到部分抑制，并导致更多新合成的透明质酸盐释放到培养基中。对从软骨中分离的[3H]透明质酸盐在Sephacryl-1000上的流体力学大小分析显示，有一个群体以宽峰形式洗脱（Kav小于0.7），而在培养物的培养基中出现两个群体（Kav大于0.5和小于0.5）。这些数据表明，透明质酸盐的合成超过蛋白聚糖的合成，并且未与蛋白聚糖复合的透明质酸盐会迅速从组织中流失。
