Functional analysis of carbohydrate antigens in chemically induced tumor cells derived from beta-1,4-galactosyltransferase-I knockout mice

β -1,4-半乳糖基转移酶I （beta4GalT-I）是合成某些2型n -聚糖和核心2 - o -聚糖所必需的糖基转移酶。我们培育了β 4galt - i敲除（KO）小鼠来研究这些碳水化合物的多种体内功能。首先，我们从表型正常的β 4galt - i纯合突变和杂合突变小鼠中产生了胚胎成纤维细胞系。两种细胞系都通过重复传代实现了永生。小鼠β - 4galt - i表达载体转染β - 4galt - i空成纤维细胞系对细胞的永生化无影响。为了评估肿瘤特异性碳水化合物抗原（如sialyl Lewis抗原）在肿瘤发生中的作用，我们利用来自beta4GalT-I KO小鼠的化学诱导皮肤肿瘤细胞系对碳水化合物抗原进行了功能分析。转染小鼠β - 4galt - i表达载体的细胞与转染β - 4galt - i表达载体的细胞在细胞生长和对纤维连接蛋白的粘附能力方面无显著差异。当评估β 4galt - 1零细胞系通过纤维连接蛋白包被和/或基质包被的transwell的运动性和侵袭性时，观察到明显的迁移和侵袭。转染小鼠beta4GalT-I表达载体后，细胞的运动性和侵袭性呈表达水平依赖性下降。虽然将β - 4galt - i转染到β - 4galt - i细胞后未检测到唾液Lewis抗原，但RCA 120凝集素检测到β - a1,4-连锁中有显著的半乳糖残基表达。这些结果表明β - 4galt - i合成的碳水化合物调节肿瘤细胞的恶性。

Beta-1,4-galactosyltransferase I (beta4GalT-I) is an essential glycosyltransferase to synthesize some kinds of type 2 N-glycans and core 2 O-glycans. We have generated beta4GalT-I knockout (KO) mice to study the multiple in vivo function of these carbohydrates. First, we have generated embryonic fibroblast cell lines from both beta4GalT-I homozygously mutated and heterozygously mutated, phenotypically normal, mice. Both cell lines were immortalized by repetitive passages. Transfection of mouse beta4GalT-I expression vector to beta4GalT-I null fibroblast cell line have no effect on immortalization of cells. In order to evaluate tumor specific carbohydrate antigens such as sialyl Lewis antigens in tumorigenesis, we have conducted functional analysis of carbohydrate antigens using chemically induced skin tumor cell lines derived from beta4GalT-I KO mice. No significant differences in cellular growth and adherent abilities to fibronectin were observed between beta4GalT-I null cells and those which were transfected with mouse beta4GalT-I expression vector. When motility and invasiveness of beta4GalT-I null cell lines through fibronectin-coated and/or Matrigel-coated transwells were assessed, significant migrations and invasions were observed. Moreover, the cell motility and invasiveness were declined by the transfection of mouse beta4GalT-I expression vector in an expression level dependent manner. Though sialyl Lewis antigens have not be detected after beta4GalT-I transfections to beta4GalT-I null cell lines, significant expressions of galactose residuedetected by RCA 120 lectin in the beta1,4-linkage was observed. These results suggest that carbohydrates synthesized by beta4GalT-I regulate the malignancy of tumor cells.