Dimitroff C J, Pera P, Dall'Olio F, Matta K L, Chandrasekaran E V, Lau J T, Bernacki R J
Department of Pharmacology and Therapeutics, Department of Gynecologic Oncology, Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, New York, 14263, USA.
Biochem Biophys Res Commun. 1999 Mar 24;256(3):631-6. doi: 10.1006/bbrc.1999.0388.
Sialoglycans on the cell surface of human colon cancer (HCC) cells have been implicated in cellular adhesion and metastasis. To clarify the role of N-acetylneuraminic acid (NeuAc) linked alpha2,3 to galactose (Gal) on the surface of HCC cells, we studied the intercellular adhesion of HCC cell lines expressing increasing NeuAcalpha2,3Gal-R. Our model system consisted of the HCC SW48 cell line, which inherently possesses low levels of cell surface alpha2,3 and alpha2,6 sialoglycans. To generate SW48 clonal variants with elevated cell surface NeuAcalpha2,3Gal-R linkages, we transfected the expression vector, pcDNA3, containing either rat liver cDNA encoding Galbeta1,3(4)GlcNAc alpha2,3 sialyltransferase (ST3Gal III) or human placental cDNA encoding Galbeta1,3GalNAc/Galbeta1,4GlcNAc alpha2,3 sialyltransferase (ST3Gal IV) into SW48 cells. Selection of neomycin-resistant clones (600 microgram G418/ml) having a higher percentage of cells expressing NeuAcalpha2,3Gal-R (up to 85% positive Maackia amurenis agglutinin staining compared with 30% for wild type cells) was performed. These ST3Gal III and ST3Gal IV clonal variants demonstrated increased adherence to IL-1beta-activated human umbilical vein endothelial cells (HUVEC) (up to 90% adherent cells compared with 63% for wild type cells). Interestingly, ST3Gal III and ST3Gal IV clonal variants also bound non-activated HUVEC up to 4-fold more effectively than wild type cells. Cell surface NeuAcalpha2,3Gal-R expression within the various SW48 clonal variants correlated directly with increased adhesion to HUVEC (r=0.84). Using HCC HT-29 cells, which express high levels of surface NeuAcalpha2,3Gal-R, addition of synthetic sialyl, sulfo or GalNAc Lewis X structures were found to specifically inhibit intercellular adhesion. At 1.0mM, NeuAcalpha2,3Galbeta1,3(Fucalpha1, 4)GlcNAc-OH and Galbeta1,4(Fucalpha1,3)GlcNAcbeta1,6(SE-6Galbeta1++ +, 3)GalNAcalpha1-O-methyl inhibited HT-29 cell adhesion to IL-1beta-stimulated HUVEC by 100% and 68%, respectively. GalNAcbeta1, 4(Fucalpha1,3)GlcNAcbeta1-O-methyl and GalNAcbeta1,4(Fucalpha1, 3)GlcNAcbeta1,6Manalpha1,6Manbeta1-0-C30H61, however, did not possess inhibitory activity. In conclusion, these studies demonstrated that cell surface NeuAcalpha2,3Gal-R expression is involved in HCC cellular adhesion to HUVEC. These specific carbohydrate-mediated intercellular adhesive events may play an important role in tumor angiogenesis, metastasis and growth control.
人类结肠癌细胞（HCC）表面的唾液酸聚糖与细胞黏附和转移有关。为了阐明HCC细胞表面N-乙酰神经氨酸（NeuAc）以α2,3连接到半乳糖（Gal）的作用，我们研究了表达量不断增加的NeuAcα2,3Gal-R的HCC细胞系的细胞间黏附。我们的模型系统由HCC SW48细胞系组成，该细胞系本身细胞表面α2,3和α2,6唾液酸聚糖水平较低。为了产生细胞表面NeuAcα2,3Gal-R连接增加的SW48克隆变体，我们将表达载体pcDNA3转染到SW48细胞中，该载体包含编码Galβ1,3(4)GlcNAc α2,3唾液酸转移酶（ST3Gal III）的大鼠肝脏cDNA或编码Galβ1,3GalNAc/Galβ1,4GlcNAc α2,3唾液酸转移酶（ST3Gal IV）的人胎盘cDNA。选择具有较高百分比表达NeuAcα2,3Gal-R的新霉素抗性克隆（600微克G418/毫升）（与野生型细胞的30%相比，高达85%的细胞对黑荆树凝集素染色呈阳性）。这些ST3Gal III和ST3Gal IV克隆变体显示出对IL-1β激活的人脐静脉内皮细胞（HUVEC）的黏附增加（与野生型细胞的63%相比，高达90%的黏附细胞）。有趣的是，ST3Gal III和ST3Gal IV克隆变体与未激活的HUVEC结合的效率也比野生型细胞高4倍。各种SW48克隆变体中的细胞表面NeuAcα2,3Gal-R表达与对HUVEC的黏附增加直接相关（r=0.84）。使用表达高水平表面NeuAcα2,3Gal-R的HCC HT-29细胞，发现添加合成的唾液酸、磺基或GalNAc Lewis X结构可特异性抑制细胞间黏附。在1.0毫摩尔时，NeuAcα$2,3Galβ1,3(Fucα1, 4)GlcNAc-OH和Galβ1,4(Fucα1,3)GlcNAcβ1,6(SE-6Galβ1++ +, 3)GalNAcα1-O-甲基分别将HT-29细胞对IL-1β刺激的HUVEC的黏附抑制了100%和68%。然而，GalNAcβ1, 4(Fucα1,3)GlcNAcβ1-O-甲基和GalNAcβ1,4(Fucα1, 3)GlcNAcβ1,6Manα1,6Manβ1-0-C30H61没有抑制活性。总之，这些研究表明细胞表面NeuAcα2,3Gal-R表达参与了HCC细胞与HUVEC的黏附。这些特定的碳水化合物介导的细胞间黏附事件可能在肿瘤血管生成、转移和生长控制中起重要作用。
