Pothoulakis C, Gilbert R J, Cladaras C, Castagliuolo I, Semenza G, Hitti Y, Montcrief J S, Linevsky J, Kelly C P, Nikulasson S, Desai H P, Wilkins T D, LaMont J T
Section of Gastroenterology, Boston University School of Medicine, Massachusetts, USA.
J Clin Invest. 1996 Aug 1;98(3):641-9. doi: 10.1172/JCI118835.
The intestinal effects of Clostridium difficile toxin A are inidated by toxin binding to luminal enterocyte receptors. We reported previously that the rabbit ileal brush border (BB) receptor is a glycoprotein with an alpha-d-galactose containing trisaccharide in the toxin-binding domain (1991. J. Clin. Invest. 88:119-125). In this study we characterized the rabbit ileal BB receptor for this toxin. Purified toxin receptor peptides of 19 and 24 amino acids showed 100% homology with rabbit sucrase-isomaltase (SI). Guinea pig receptor antiserum reacted in Western blots with rabbit SI and with the purified toxin receptor. Antireceptor IgG blocked in vitro binding of toxin A to rabbit ileal villus cell BB. Furthermore, anti-SI IgG inhibited toxin A-induced secretion (by 78.1%, P < 0.01), intestinal permeability (by 80.8%, P < 0.01), and histologic injury (P < 0.01) in rabbit ileal loops in vivo. Chinese hamster ovary cells transfected with SI cDNA showed increased intracellular calcium increase in response to native toxin (holotoxin) or to a recombinant 873-amino acid peptide representing the receptor binding domain of toxin A. These data suggest that toxin A binds specifically to carbohydrate domains on rabbit ileal SI, and that such binding is relevant to signal transduction mechanisms that mediate in vitro and in vivo toxicity.
艰难梭菌毒素A的肠道效应是由毒素与肠腔肠上皮细胞受体结合引发的。我们之前报道过，兔回肠刷状缘（BB）受体是一种糖蛋白，在毒素结合结构域含有一个含α-d-半乳糖的三糖（1991年。《临床研究杂志》88:119 - 125）。在本研究中，我们对该毒素的兔回肠BB受体进行了特性分析。纯化的19和24个氨基酸的毒素受体肽与兔蔗糖酶 - 异麦芽糖酶（SI）显示出100%的同源性。豚鼠受体抗血清在蛋白质免疫印迹中与兔SI及纯化的毒素受体发生反应。抗受体IgG在体外阻断了毒素A与兔回肠绒毛细胞BB的结合。此外，抗SI IgG在体内抑制了兔回肠肠袢中毒素A诱导的分泌（78.1%，P < 0.01）、肠道通透性（80.8%，P < 0.01）以及组织学损伤（P < 0.01）。用SI cDNA转染的中国仓鼠卵巢细胞对天然毒素（全毒素）或代表毒素A受体结合结构域的重组873个氨基酸的肽产生反应时，细胞内钙增加。这些数据表明，毒素A特异性结合兔回肠SI上的碳水化合物结构域，且这种结合与介导体外和体内毒性的信号转导机制相关。
