Role of the glycocalyx in regulating access of microparticles to apical plasma membranes of intestinal epithelial cells: implications for microbial attachment and oral vaccine targeting.
Role of the glycocalyx in regulating access of microparticles to apical plasma membranes of intestinal epithelial cells: implications for microbial attachment and oral vaccine targeting.
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DOI：
10.1084/jem.184.3.1045
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发表时间：
1996-09-01
期刊：
影响因子：
--
通讯作者：
Neutra MR
中科院分区：
文献类型：
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作者：
Frey A;Giannasca KT;Weltzin R;Giannasca PJ;Reggio H;Lencer WI;Neutra MR
抗原和病原体通过M细胞跨上皮屏障的跨上皮转运可能是诱导肠粘膜免疫的先决条件。抗原和病原体的有效运输需要粘附于M细胞顶端表面。已提出将含抗原颗粒偶联至霍乱毒素（CTB）的五聚体结合亚基作为增加抗原摄取的手段，因为CTB受体神经节苷脂GM 1是存在于所有肠上皮细胞的顶端膜中的糖脂。为了测试肠上皮细胞和M细胞膜糖脂对病毒、细菌和颗粒粘膜疫苗的大小范围内的配体的可及性，我们分析了不同大小的CTB探针与兔派尔集合淋巴结上皮的结合。可溶性CTB-异硫氰酸荧光素（直径6.4 nm）与所有上皮细胞的顶膜结合。CTB耦合到14 nm胶体金（最终直径，28.8 nm）未能坚持肠上皮细胞，但坚持M细胞。CTB包被的荧光微粒（最终直径1.13微米）在体内不能粘附到肠上皮细胞或M细胞上，在体外不能粘附到分化良好的Caco-2肠上皮细胞上.然而，这些颗粒在体外特异性结合到BALB/c 3 T3成纤维细胞上的GM 1和缺乏刷状缘和糖萼的未分化Caco-2细胞。通过电子显微镜测量糖萼厚度表明，相对较薄（20 nm）的糖萼足以防止1微米微粒进入糖脂受体。因此，肠上皮细胞糖萼的屏障功能可能是重要的，在限制微生物粘附膜糖脂，并在CTB介导的靶向疫苗的M细胞和粘膜免疫系统。
Transepithelial transport of antigens and pathogens across the epithelial barrier by M cells may be a prerequisite for induction of mucosal immunity in the intestine. Efficient transport of antigens and pathogens requires adherence to M cell apical surfaces. Coupling of antigen-containing particles to the pentameric binding subunit of cholera toxin (CTB) has been proposed as a means for increasing antigen uptake because the CTB receptor, ganglioside GM1, is a glycolipid present in apical membranes of all intestinal epithelial cells. To test the accessibility of enterocyte and M cell membrane glycolipids to ligands in the size ranges of viruses, bacteria, and particulate mucosal vaccines, we analyzed binding of CTB probes of different sizes to rabbit Peyer's patch epithelium. Soluble CTB-fluorescein isothiocyanate (diameter 6.4 nm) bound to apical membranes of all epithelial cells. CTB coupled to 14 nm colloidal gold (final diameter, 28.8 nm) failed to adhere to enterocytes but did adhere to M cells. CTB- coated, fluorescent microparticles (final diameter, 1.13 microns) failed to adhere to enterocytes or M cells in vivo or to well- differentiated Caco-2 intestinal epithelial cells in vitro. However, these particles bound specifically to GM1 on BALB/c 3T3 fibroblasts in vitro and to undifferentiated Caco-2 cells that lacked brush borders and glycocalyx. Measurements of glycocalyx thickness by electron microscopy suggested that a relatively thin (20 nm) glycocalyx was sufficient to prevent access of 1-micron microparticles to glycolipid receptors. Thus, the barrier function of the intestinal epithelial cell glycocalyx may be important in limiting microbial adherence to membrane glycolipids, and in CTB-mediated targeting of vaccines to M cells and the mucosal immune system.