INTESTINAL GOBLET CELLS--DIFFERENTIAION AND SECRETION

该项目的重点是确定调节 肠杯状细胞的分化和粘液分泌特性。 此前的调查一直受到无法维持 在体外任何条件下分化杯状细胞。 最近， 然而，发现人结肠癌细胞系HT 29表达 小肠的形态学和酶表型特征 在无葡萄糖培养基中生长时的上皮。 而且，克隆人， 命名为“N2”的HT29株系， 分化成一个纯粹的杯状细胞群体， 无葡萄糖培养基。 HT29和N2细胞系将被开发为模型系统，以研究 调节杯状细胞分化和粘液分泌。 杯状细胞的形态学、免疫化学和生化标志 分化过程将被定义。 细胞外基质组分 和可渗透的基质将被测试其对 HT 29和N2杯状细胞的增殖和分化。 间接 有证据表明，寄生虫感染大鼠的T淋巴细胞可能分泌 一种特异性调节杯状细胞生长的可溶性因子。 来自巴西日本圆线虫感染的大鼠的T淋巴细胞将被 体外暴露于抗原，并测试其条件培养基 在HT29中诱导杯状细胞增殖和分化的能力 和N2细胞系。 钙在介导基线和 粘液分泌的加速形式将通过使用 形态学、放射自显影和放射免疫测定技术来测量 不同浓度的细胞外钙的分泌率， 在钙通道阻滞剂（Co2+）的存在下。 佛波醇酯将 用于检验蛋白激酶C的激活是一种 粘液分泌颗粒复合胞吐作用的重要步骤。 从这些研究中获得的基本信息将提供新的见解 进入杯状细胞形成的过程，并将增强 了解粘膜如何保持其粘液的保护涂层。

This project is focused on defining the factors which regulate the differentiation and mucus-secretory properties of intestinal goblet cells. Previous investigations have been limited by the inability to maintain differentiated goblet cells under any conditions in vitro. Recently, however, a human colon cancer cell line, HT29, was found to express the morphological and enzymatic phenotypes characteristic of small intestinal epithelium when grown in glucose-free medium. Furthermore, a clone, designated "N2", has been isolated from the parent HT29 line, which differentiates into a pure population of goblet cells when grown in glucose-free medium. The HT29 and N2 cell lines will be developed as model systems to study the regulation of goblet cell differentiation and mucus secretion. Morphological, immunochemical and biochemical landmarks of the goblet cell differentiation process will be defined. Extracellular matrix components and permeable substrates will be tested for their influence on the proliferation and differentiation of HT29 and N2 goblet cells. Indirect evidence suggests that T-lymphocytes in parasite-infected rats may secrete a soluble factor which specifically modulates goblet cell growth. T-lymphocytes from Nippostrongylus brasiliensis-infected rats will be exposed to antigen in vitro and the conditioned medium tested for its ability to induce goblet cell proliferation and differentiation in the HT29 and N2 cell lines. The role of calcium in mediating the baseline and accelerated forms of mucus secretion will be established by using morphological, autoradiographic and radioimmune assay techniques to measure the secretory rates in varying concentrations of extracellular calcium or in the presence of a calcium channel blocker (Co2+). Phorbol esters will be used to test the hypothesis that activation of protein kinase C is an essential step in the compound exocytosis of mucous secretory granules. The basic information gained from these studies will provide new insights into the process of goblet cell formation, and will enhance the understanding of how the mucosa maintains its protective coating of mucus.