Heim H K, Oestmann A, Thiele H, Sewing K F
Abteilung Allgemeine Pharmakologie, Medizinische Hochschule, Hannover, BRD.
Digestion. 1989;44(1):26-35. doi: 10.1159/000199889.
Glycoprotein and protein production of isolated pig gastric mucosal cells were determined by the incorporation of N-acetyl-[14C]D-glucosamine ([14C]GlcNAc) and [3H]L-leucine ([3H]Leu) into acid-insoluble macromolecules (AIM). In four cell fractions (F1-F4), obtained by counterflow centrifugation, specific [14C]GlcNAc incorporation was greatest in the mucous cell-enriched F2. Tracer incorporation by F2 cells, proceeded linearly up to 20 h, was inhibited by cycloheximide or incubation at 0 degree C, and enhanced by PGE2 1 mumol/l. Gel chromatography of released AIM revealed that PGE2-stimulated [14C]GlcNAc incorporation was predominantly directed into high molecular weight (2 X 10(6) daltons) glycoproteins, whereas [3H]Leu incorporation was mainly related to proteins of albumin-like molecular weight. We conclude that incorporation of [14C]GlcNAc by enriched pig gastric mucous cells (F2), further analyzed by gel chromatography, is a suitable probe to study the production of high molecular weight gastric mucous glycoproteins in vitro.
通过将N-乙酰-[14C]D-葡萄糖胺（[14C]GlcNAc）和[3H]L-亮氨酸（[3H]Leu）掺入酸不溶性大分子（AIM）中，测定分离的猪胃黏膜细胞的糖蛋白和蛋白质产量。在通过逆流离心获得的四个细胞组分（F1-F4）中，富含黏液细胞的F2中[14C]GlcNAc的特异性掺入量最大。F2细胞的示踪剂掺入在长达20小时内呈线性进行，受环己酰亚胺或在0℃孵育的抑制，并被1μmol/L的前列腺素E2增强。对释放的AIM进行凝胶色谱分析表明，前列腺素E2刺激的[14C]GlcNAc掺入主要导向高分子量（2×10^6道尔顿）糖蛋白，而[3H]Leu掺入主要与白蛋白样分子量的蛋白质有关。我们得出结论，通过凝胶色谱进一步分析，富含猪胃黏液细胞（F2）对[14C]GlcNAc的掺入是研究体外高分子量胃黏液糖蛋白产生的合适探针。
