抑制HT29-Gal细胞粘蛋白O-型糖链的合成增加细菌-第三军医大学学报.doc

肠分化细胞粘蛋白O-型糖链合成抑制导致MUC2表达减低以及对细菌侵袭易感叶钧 宋丽丽 刘韵? 第三军医大学西南医院全军消化病研究所， 重庆，400038 【摘要】 目的 探讨O-型糖链合成的抑制对肠分化细胞内细菌侵袭数量和细胞内MUC2表达水平的影响。 方法 对肠分化细胞（HT-29-Gal）用O-型糖链抑制剂（benzyl-N-acetyl-α-D-galactosaminide，benzyl-α-GalNAc）处理，采用Real-time PCR和Western-blot方法检测MUC2 mRNA和蛋白的表达情况。并将HT-29-Gal细胞及benzyl-α-GalNAc 处理的HT-29-Gal细胞分别与肠致病性大肠杆菌（EPEC）和肠出血性大肠杆菌（EHEC O157:H7） 37°C孵育2h， 再加入100ug/ml的庆大霉素，杀灭细胞外及粘附于细胞表面的细菌。最后采用系列稀释克隆计数法观察benzyl-α-GalNAc 处理的HT-29-Gal细胞对细菌侵袭的影响。 结果 Real-time PCR和Western-blot检测发现经benzyl-α-GalNAc处理的HT-29-Gal细胞MUC2的mRNA和蛋白表达水平明显降低（p0.05）。侵袭入benzyl-α-GalNAc 处理的HT-29-Gal细胞的EPEC和EHEC O157:H7的数量较对照细胞显著增加（p0.05)。------- 结论 抑制HT-29-Gal细胞粘蛋白O-型糖链的合成导致侵袭入细胞内细菌数量增加和的表达降低。【关键词】O-型糖链; benzyl-α-GalNAc; HT-29-Gal 【中国图书资料分类号】nhibition of O-glycan Chains Synthesis in Intestinal Differentiated Epithelial Cells Leads to Less MUC2 Expression and Susceptibility to Bacteria Invasion Ye Jun, Song lili, Liu Yun, Tian Yin, Pan Qiong, Peng Zhihong, Wang Rongquan. Institute of Gastroenterology of PLA, Southwest Hospital, Third Military Medical University, Chongqing 400038, China. [ Abstract ] Objective To investigate the effect of inhibition of O-glycan chains synthesis in intestinal differentiated epithelial cells on the bacteria invasion into cells and MUC2 exxpression . Methods The intestinal differentiated cells (HT-29-Gal) were treated with O-glycan chains synthesis inhibitor ( benzyl-N-acetyl-α-D-galactosaminide, benzyl-α -GalNAc ) and levels of MUC2 mRNA and protein were detected by real-time PCR and Western-blot. The HT-29-Gal cells treated with or without benzyl-α -GalNAc were co-cultured with Enteropathogenic E.coli （EPEC） and enterohemorrhage E．Coli（EHEC O157:H7）for 2h, then gentamicin(100ug/ml) was added for 2h exclude extracellular and cellular surface-adherent bacteria after 3 times washing with PBS. Finally, use the serial dilution cloning count method to observe the numbers of invaded bacteria into the HT-29-Gal cells treated with or without benzyl-α-GalNAc. Results Levels of MU