Removal of glycosaminoglycans from cultures of human skin fibroblasts.
Removal of glycosaminoglycans from cultures of human skin fibroblasts.
复制标题
从人皮肤成纤维细胞培养物中去除糖胺聚糖。
DOI：
10.1042/bj1940299
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发表时间：
1981
期刊：
影响因子：
--
通讯作者：
J. Silbert
中科院分区：
文献类型：
--
作者：
P. J. Gill;J. Adler;C. Silbert;J. Silbert
早期传代的人皮肤成纤维细胞在含有[35 S]硫酸盐、[3 H]葡糖胺、[3 H]岩藻糖、[3 H]脯氨酸或[3 H]亮氨酸的最低必需培养基中以单层生长2-3天，以标记蛋白聚糖、糖蛋白或胶原蛋白和其他蛋白质。将从超声处理的冷冻干燥的黄肝素的上清液获得的粗酶制剂加入细胞单层。该处理去除了大部分35 S标记的糖胺聚糖，而没有明显去除3 H标记的蛋白质或3 H标记的糖蛋白。细胞作为单层保持附着和存活。用粗制F.肝素酶，然后加入含有[35 S]硫酸盐的新鲜生长培养基。食品药品管理肝素酶没有显著改变产生的35 S标记的糖胺聚糖的量或类型。因此F.肝素酶可用于提供耗尽表面糖胺聚糖的培养细胞单层。这些细胞保持附着、存活，随后合成正常数量和类型的糖胺聚糖。
Early-passage human skin fibroblasts were grown as monolayers for 2-3 days in minimum essential medium containing [35S]sulphate, [3H]glucosamine, [3H]fucose, [3H]proline or [3H]leucine to label proteoglycans, glycoproteins or collagen and other proteins. A crude enzyme preparation obtained from a supernatant from sonicated freeze-dried Flavobacter heparinum was added to the cell monolayers. This treatment removed most of the 35S-labelled glycosaminoglycans, with no appreciable removal of the 3H-labelled proteins or 3H-labelled glycoproteins. The cells remained attached and viable as a monolayer. The formation of 35S-labelled glycosaminoglycans was examined after pretreating cultures with crude F. heparinum enzyme, followed by addition of fresh growth medium containing [35S]sulphate. The F. heparinum enzyme did not significantly alter the amount or type of 35S-labelled glycosaminoglycans produced. Thus F. heparinum enzyme can be used to provide cultured-cell monolayers depleted of surface glycosaminoglycans. These cells remain attached, viable and subsequently synthesize normal amounts and type of glycosaminoglycans.