Permanent exposure of mucin-secreting HT-29 cells to benzyl-N-acetyl-α-D-galactosaminide induces abnormal O-glycosylation of mucins and inhibits constitutive and stimulated MUC5AC secretion
Permanent exposure of mucin-secreting HT-29 cells to benzyl-N-acetyl-α-D-galactosaminide induces abnormal O-glycosylation of mucins and inhibits constitutive and stimulated MUC5AC secretion
复制标题
DOI：
10.1042/bj3340283
复制
发表时间：
1998-08-15
影响因子：
4.1
通讯作者：
Huet, G
中科院分区：
文献类型：
--
作者：
Hennebicq-Reig, S;Lesuffleur, T;Huet, G
先前的工作表明，用苄基-N-乙酰基-α-D-氨基半乳糖苷处理HT-29甲氨蝶呤（MTX）细胞导致粘蛋白寡糖链发生深刻变化。为了深入分析这种药物的作用，我们首先测定了HT-29 MTX细胞合成的粘蛋白寡糖链的结构以及药物长期暴露引起的变化。来自未处理细胞的粘蛋白含有9个单唾液酸化结构（核心类型1、2、3和4）和4个双唾液酸化结构（类型1、2和4）。核心1结构占优势，特别是NeuAc α 2-3Gal β 1- 3GalNAc-醇。HT-29 MTX细胞在第2-21天暴露于苄基-N-乙酰基-α-D-氨基半乳糖苷导致细胞内粘蛋白及其唾液酸和半乳糖含量减少，T（Gal β 1-3GalNAc α-O-Ser/Thr）和Tn（GalNAc α-O-Ser/Thr）抗原性增加。检测到Gal β 1-3GalNAc α 2，3-唾液酸转移酶活性和mRNA表达均增加3倍。在超微结构水平上，T-抗原在对照细胞的粘蛋白液滴中检测不到，但在处理细胞的胞浆内囊泡中强烈表达。在这些细胞中，MUC 1和MUC 3转录物上调，而MUC 2、MUC 5 B和MUC 5AC下调。此外，组成型和促分泌素诱导的MUC 5AC分泌减少，没有检测到粘液层。总之，苄基-N-乙酰基-α-D-氨基半乳糖苷诱导异常O-糖基化和MUC 5AC分泌调节改变。
Previous work has shown that treatment of HT-29 methotrexate (MTX) cells with benzyl-N-acetyl-alpha-D-galactosaminide results in profound changes in mucin oligosaccharide chains. To analyse in depth the effect of this drug, we first determined the structure of mucin oligosaccharide chains synthesized by HT-29 MTX cells and the changes induced by permanent drug exposure. Mucins from untreated cells contained nine monosialylated structures (core types 1, 2, 3 and 4) and four disialylated structures (types 1, 2 and 4). Core 1 structures predominated, in particular NeuAc alpha 2-3Gal beta 1-3GalNAc-ol. Exposure of HT-29 MTX cells to benzyl-N-acetyl-alpha-D-galactosaminide from days 2-21 resulted in a decrease in intracellular mucins and both their sialic acid and galactose content, and an increased T (Gal beta 1-3GalNAc alpha-O-Ser/Thr) and Tn (GalNAc alpha-O-Ser/Thr) antigenicity. A 3-fold increase in both Gal beta 1-3GalNAc alpha 2,3-sialyltransferase activity and mRNA expression was detected. At the ultrastructural level, T-antigen was not detectable in mucin droplets in control cells, but was strongly expressed in intracytoplasmic vesicles in treated cells. In these cells, MUC1 and MUC3 transcripts were up-regulated, whereas MUC2, MUC5B and MUC5AC were down-regulated. Furthermore, constitutive and secretagogue-induced MUC5AC secretion was reduced and no mucus layer was detected. In conclusion, benzyl-N-acetyl-alpha-D-galactosaminide induces abnormal O-glycosylation and altered regulation of MUC5AC secretion.