Sjöberg I, Fransson L A
Biochem J. 1977 Nov 1;167(2):383-92. doi: 10.1042/bj1670383.
Foetal human lung fibroblasts, grown in monolayer, were allowed to incorporate (35)SO(4) (2-) for various periods of time. (35)S-labelled macromolecular anionic products were isolated from the medium, a trypsin digest of the cells in monolayer and the cell residue. The various radioactive polysaccharides were identified as heparan sulphate and a galactosaminoglycan population (chondroitin sulphate and dermatan sulphate) by ion-exchange chromatography and by differential degradations with HNO(2) and chondroitinase ABC. Most of the heparan sulphate was found in the trypsin digest, whereas the galactosaminoglycan components were largely confined to the medium. Electrophoretic studies on the various (35)S-labelled galactosaminoglycans suggested the presence of a separate chondroitin sulphate component (i.e. a glucuronic acid-rich galactosaminoglycan). The (35)S-labelled galactosaminoglycans were subjected to periodate oxidation of l-iduronic acid residues followed by scission in alkali. A periodate-resistant polymer fraction was obtained, which could be degraded to disaccharides by chondroitinase AC. However, most of the (35)S-labelled galactosaminoglycans were extensively degraded by periodate oxidation-alkaline elimination. The oligosaccharides obtained were essentially resistant to chondroitinase AC, indicating that the iduronic acid-rich galactosaminoglycans (i.e. dermatan sulphate) were composed largely of repeating units containing sulphated or non-sulphated l-iduronic acid residues. The l-iduronic acid residues present in dermatan sulphate derived from the medium and the trypsin digest contained twice as much ester sulphate as did material associated with the cells. The content of d-glucuronic acid was low and similar in all three fractions. The relative distribution of glycosaminoglycans among the various fractions obtained from cultured lung fibroblasts was distinctly different from that of skin fibroblasts [Malmström, Carlstedt, Aberg & Fransson (1975) Biochem. J.151, 477-489]. Moreover, subtle differences in co-polymeric structure of dermatan sulphate isolated from the two cell types could be detected.
将人胎儿肺成纤维细胞以单层培养，使其在不同时间段内掺入（35）SO（4）（2-）。从培养基、单层细胞的胰蛋白酶消化物以及细胞残渣中分离出（35）S标记的大分子阴离子产物。通过离子交换色谱法以及用HNO（2）和软骨素酶ABC进行的差异降解，将各种放射性多糖鉴定为硫酸乙酰肝素和一组半乳糖胺聚糖（硫酸软骨素和硫酸皮肤素）。大部分硫酸乙酰肝素存在于胰蛋白酶消化物中，而半乳糖胺聚糖成分主要存在于培养基中。对各种（35）S标记的半乳糖胺聚糖进行的电泳研究表明存在一种单独的硫酸软骨素成分（即富含葡萄糖醛酸的半乳糖胺聚糖）。对（35）S标记的半乳糖胺聚糖进行高碘酸盐氧化l-艾杜糖醛酸残基，然后在碱中裂解。得到了一种抗高碘酸盐的聚合物级分，其可被软骨素酶AC降解为二糖。然而，大部分（35）S标记的半乳糖胺聚糖通过高碘酸盐氧化-碱消除被广泛降解。得到的寡糖基本上对软骨素酶AC有抗性，这表明富含艾杜糖醛酸的半乳糖胺聚糖（即硫酸皮肤素）主要由含有硫酸化或非硫酸化l-艾杜糖醛酸残基的重复单元组成。源自培养基和胰蛋白酶消化物的硫酸皮肤素中存在的l-艾杜糖醛酸残基所含的硫酸酯是与细胞相关物质的两倍。d-葡萄糖醛酸的含量较低，且在所有三个级分中相似。从培养的肺成纤维细胞获得的各种级分中糖胺聚糖的相对分布与皮肤成纤维细胞的明显不同[马尔姆斯特伦、卡尔施泰特、阿贝格和弗兰松（1975年）《生物化学杂志》151，477 - 489]。此外，还能检测到从两种细胞类型分离出的硫酸皮肤素在共聚物结构上的细微差异。
