Department of Veterinary Clinical Sciences, University of Minnesota, St. Paul, Minnesota, United States of America.
PLoS One. 2011 Jan 19;6(1):e16281. doi: 10.1371/journal.pone.0016281.
Similar to mechanisms of recruitment of activated leukocytes to inflamed tissues, selectins mediate adhesion and extravasation of circulating cancer cells. Our objective was to determine whether sialyl Lewis X modified core 2 O-glycans (C2-O-sLe(X)) present on colon and hepatic carcinoma cells promote their adhesion and invasion. We examined membrane expression of C2-O-sLe(X), selectin binding, invasion of human colon and hepatic carcinoma cell lines, and mRNA levels of alpha-2,3 fucosyltransferase (FucT-III) and core 2 beta-1,6 N-acetylglucosaminyltransferase (C2GnT1) genes, necessary for C2-O-sLe(X) synthesis, by quantitative reverse-transcriptase (RT) PCR. Synthesis of core 2 branched O-glycans decorated by sLe(X) is dependent on C2GnT1 function and thus we determined enzyme activity of C2GnT1. The cell lines that expressed C2GnT1 and FucT-III mRNA by quantitative RT-PCR were highly positive for C2-O-sLe(X) by flow cytometry, and colon carcinoma cells possessed highly active C2GnT1 enzyme. Cells bound avidly to E-selection but not to P- and L-selectin. Gene knock-down of C2GnT1 in colon and hepatic carcinoma cells using short hairpin RNAs (shRNA) resulted in a 40-90% decrease in C2-O-sLe(X) and a 30-50% decrease in E-selectin binding compared to control cells. Invasion of hepatic and colon carcinoma cells containing C2GnT1 shRNA was significantly reduced compared to control cells in Matrigel assays and C2GnT1 activity was down-regulated in the latter cells. The sLe(X) epitope was predominantly distributed on core 2 O-glycans on colon and hepatic carcinoma cells. Our findings indicate that C2GnT1 gene expression and the resulting C2-O-sLe(X) carbohydrates produced mediate the adhesive and invasive behaviors of human carcinomas which may influence their metastatic potential.
类似于激活的白细胞募集到炎症组织的机制，选择素介导循环癌细胞的黏附和渗出。我们的目的是确定结肠和肝癌细胞表面的唾液酸化路易斯 X 修饰的核心 2 O-糖链（C2-O-sLe(X)）是否促进它们的黏附和侵袭。我们通过定量逆转录聚合酶链反应（RT-PCR）检查了 C2-O-sLe(X)的膜表达、选择素结合、人结肠和肝癌细胞系的侵袭以及合成 C2-O-sLe(X）所必需的α-2,3 岩藻糖基转移酶（FucT-III）和核心 2 β-1,6 N-乙酰氨基葡萄糖基转移酶（C2GnT1）基因的 mRNA 水平。C2-O-sLe(X)的合成依赖于 C2GnT1 功能，因此我们测定了 C2GnT1 的酶活性。通过定量 RT-PCR 表达 C2GnT1 和 FucT-III mRNA 的细胞系通过流式细胞术对 C2-O-sLe(X)呈高度阳性，并且结肠癌细胞具有高度活跃的 C2GnT1 酶。细胞与 E-选择素强烈结合，但不与 P-和 L-选择素结合。用短发夹 RNA(shRNA) 对结肠和肝癌细胞中的 C2GnT1 进行基因敲低，与对照细胞相比，C2-O-sLe(X)减少了 40-90%，E-选择素结合减少了 30-50%。在 Matrigel 测定中，含有 C2GnT1 shRNA 的肝癌和结肠癌细胞的侵袭性明显低于对照细胞，并且在后一种细胞中 C2GnT1 活性下调。sLe(X)表位主要分布在结肠和肝癌细胞的核心 2 O-糖链上。我们的研究结果表明，C2GnT1 基因表达和由此产生的 C2-O-sLe(X)碳水化合物介导了人类癌症的黏附和侵袭行为，这可能影响它们的转移潜力。
