INTESTINAL GOBLET CELLS--DIFFERENTIAION AND SECRETION

这个项目的重点是定义调节 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的杯状细胞。最近， 然而，一种人类结肠癌细胞系HT29被发现表达这种 小肠的形态和酶表型特征 在无糖培养基中生长的上皮细胞。此外，克隆人， 已从亲本HT29系中分离到一株代号为“n2”的 在培养过程中分化为纯杯状细胞 无糖培养液。 HT29和N2细胞系将被开发为模型系统来研究 杯状细胞分化和粘液分泌的调节。 杯状细胞的形态、免疫化学和生化标志 将定义差异化过程。细胞外基质成分 和可渗透基质将测试它们对 HT29和N2杯状细胞的增殖和分化。间接法 有证据表明，寄生虫感染大鼠的T淋巴细胞可能会分泌 一种特殊调节杯状细胞生长的可溶性因子。 感染巴西拟青霉大鼠的T淋巴细胞将被 体外暴露于抗原和条件培养液中检测其 HT29诱导杯状细胞增殖和分化的能力 和N_2细胞系。钙在调节基线和血压变化中的作用 粘液分泌的加速形式将通过使用 形态、放射自显影和放射免疫分析技术 不同细胞外钙或钙离子浓度下的分泌速率 在钙通道阻滞剂(Co2+)存在的情况下。佛波醇酯将 用来检验这样的假设：蛋白激酶C的激活是一个 粘液分泌颗粒复合胞吐的必要步骤。 从这些研究中获得的基本信息将提供新的见解 进入杯状细胞的形成过程，并将增强 了解粘膜如何维持其粘液保护层。

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.