IL-13/Stat6-mediated sialylation mechanisms of mucins and biological role of sialylated mucins for protection against gastrointestinal nematode infection

我们以前曾报道，对小鼠胃肠道线虫感染的保护作用与IL-13/Stat6介导的杯状细胞唾液酸连接粘蛋白的产生有关。本项目的目标是研究1)IL-13/STAT6介导的粘蛋白唾液酸化的机制，以及2)唾液酸化粘蛋白对小鼠感染巴西奈瑟氏菌的保护作用。由于到目前为止还没有合适的小鼠产生粘蛋白的细胞系，我们使用了人结肠癌细胞株HT-29。在用过碘酸希夫反应证实HT-29产生粘蛋白的基础上，检测了与IL-4受体结合的外源IL-13和IL-4对粘蛋白的数量和性质的影响，特别是对粘蛋白唾液酸化的影响。根据粘蛋白的特性，从基因和蛋白质水平研究了信号转导分子的激活和唾液酸基转移酶等酶的表达。关于S…的影响用唾液酸寡糖进行的体内和体外实验均不能阻止新月形虫在肠道绒毛上的建立，提示唾液酸的作用与粘蛋白有关。此外，成虫植入实验表明，尽管粘液的靶标(S)尚不清楚，但粘液对已建立的成虫的驱虫作用不是通过结构上的，而是通过生理作用发生的。与此相关的是，从感染小鼠的小肠中分离粘液，以测定己糖和唾液酸化粘蛋白的含量。虽然该项目尚未完全完成，但该项目发表的结果将为Th2应答诱导的胃肠道线虫感染保护性反应的效应期机制提供重要信息。此外，该结果还将明确Th2细胞因子IL-13诱导的粘蛋白产生的修饰机制，这是目前变态反应研究的热点。较少

We previously reported that protection against gastrointestinal nematode, Nippostroagylus brasiliensis, infection in mice was associated with IL-13/Stat6-mediated sialic acid-linked mucin production in goblet cells. The goals of this project are to investigate 1)the mechanism of IL-13/Stat6-mediated sialylation of mucins, and 2)a role of sialylated mucins for protection against N.brasiliensis infection in mice. Since suitable cell lines of mucin producing cells derived from mouse are not available so far, we employed human colon cancer cell line, HT-29. After confirming the mucin production of HT-29 by Periodic Acid Schiff reaction, effects of exogenous IL-13 and IL-4,both bind to IL-4 receptor, on the amount and character of mucins, especially sialylation of mucins, were examined. Base on the characteristics of mucins, activation of signal transduction molecules and expression of enzymes, such as sialyltransferases, were examined in gene and protein levels. Concerning the effects of s … More ialylated mucins on N.brasiliensis, in vivo and in vitro experiments using sialylated oligosaccharides failed to block the establishment of N.brasiliensis on intestinal villi suggesting that sialic acid affects as being linked to mucins. Further, adult worm implantaion experiments suggested that worm expulsion occurred through not structural but physiological effects of mucus on established adult N.brasiliensis, although the target(s) of the mucus were remained unclear. Related to this, mucus were isolated from small intestine of infected mice to estimate the contents of hexose and sialylated mucins. Although the project has not been done completely, results published by this project will provide significant information about the mechanisms of effector phase of protective response induced by Th2 response against gastrointestinal nematode infection. Additionally, the results will clear the modification mechanisms of mucin production induced by Th2 cytokine, IL-13, which is now the highlight of allergy research. Less