INTESTINAL SECRETORY MECHANISMS

肠道分泌机制

• 批准号: 3228717
• 负责人: KIERTISIN DHARMSATHAPHORN
• 金额: $ 26.54万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-07-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3228717
• 关键词: autoradiography; calcium metabolism; calcium transporting ATPase; calmodulin; cell membrane; chloride channels; cyclic AMP; diarrhea; disease /disorder prevention /control; drug metabolism; enzyme inhibitors; gastrointestinal absorption /transport; gastrointestinal epithelium; gastrointestinal hormones; ion transport; membrane permeability; passive transport; physical chemical interaction; potassium channel; prostaglandins; protein kinase; radiotracer; secretion; sodium potassium exchanging ATPase; transport proteins

Previously, our laboratory has identified a human colonic epithelial cell line, T84, as a suitable model for investigation of Cl- secretory mechanisms. The cell line, composed of only secretory cells and free of influence from paracrine, neurocrine or endocrine controls, is ideal for studies of regulatory controls of Cl- secretion at the cellular level. Our studies in the past four years identified transport pathways involved in various Cl- secretory processes and unmasked a synergistic phenomenon between cyclic nucleotide and Ca++ -related secretions. Recent studies suggest a more complicated interaction of secondary messengers in Ca++ - related secretory mechanisms and implicate involvement of phospholipid turnovers. A candidate messenger (phospholipid metabolite) which co- activates Cl- secretion with free cytosolic Ca++ is arachidonic acid. A candidate messenger which inhibits Cl- secretion is diacylglycerol. This renewal application proposes to sort out the contribution of secondary messengers, particularly phospholipid metabolites using cell monolayers and plasma membrane preparations. Three approaches will be utilized: 1) Correlative studies of the time courses and dose-responses on each secondary messenger and a biological response to identify potential candidates. Activators or inhibitors of phospholipases, diglyceride lipase, lipoxygenase, cyclooxygenase or G-proteins will also be used in the studies under varying [Ca++]i or [Ca++] (for plasma membrane). Initial studies will focus on the mechanism of action of carbachol, histamine, 4- Br-A23187, ionomycin, taurodeoxy-cholate and adenosine. 2) Reconstitution studies will be carried out after a better method to deliver fat-soluble substances to the cell or plasma membrane is developed. The biological actions of each candidate messenger will be directly tested both alone and in combination with others to mimic the responses of each secretagogue listed in correlative studies. Again, the use of specific enzymes and their inhibitors will complement the studies proposed under varying [Ca++]i or [Ca++]. 3) Comparative studies will be carried out in other epithelial cell lines that secrete Cl-, e.g., CaCO and HT-29 clones to test whether the same mechanism also applies to them. It is expected that some cell lines with a defective secondary messenger or a defective transport pathway will provide useful complementary insights into the role of that messenger on the transport pathway.

此前，我们的实验室已经确定了人类结肠上皮细胞 T84细胞系是研究Cl-分泌的合适模型 机制等 该细胞系仅由分泌细胞组成，不含 旁分泌、神经内分泌或内分泌控制的影响，是理想的， 在细胞水平上研究Cl-分泌的调节控制。 我们 过去四年的研究确定了 不同的Cl-分泌过程，并揭示了协同现象 环核苷酸和Ca++相关分泌物之间的关系。 最近的研究 提示Ca++ -Ca ~（2+）-Ca ~（2+）- 相关的分泌机制和牵连参与磷脂 失误。 一种候选信使（磷脂代谢物）， 激活Cl-分泌与游离胞质Ca++是花生四烯酸。 一 抑制Cl-分泌的候选信使是二酰基甘油。 这 续期申请建议整理中学供款 信使，特别是使用细胞单层的磷脂代谢物， 质膜制备。 将采用三种方法：1） 时间进程和剂量反应的相关性研究 第二信使和生物反应，以确定潜在的 候选人 磷脂酶激活剂或抑制剂，甘油二酯 脂肪酶、脂氧合酶、环氧合酶或G-蛋白也将用于本发明。 在不同的[Ca++]i或[Ca++]（对于质膜）下的研究。 初始 研究将集中在卡巴胆碱、组胺、4- Br-A23187、离子霉素、牛磺脱氧胆酸盐和腺苷。 2)复溶 研究将进行后，一个更好的方法来提供脂溶性 细胞膜或细胞质膜上的物质。 生物 每个候选信使的行动将被直接测试， 与其他物质结合以模拟每种促分泌素的反应 在相关研究中列出。 再次，使用特定的酶和 它们的抑制剂将补充在不同[Ca++]i下提出的研究。 或[Ca++]。 3)将在其他上皮细胞中进行比较研究。 分泌Cl-的细胞系，例如，CaCO和HT-29克隆，以测试是否 同样的机制也适用于他们。 预计一些细胞 具有缺陷的第二信使或缺陷的转运途径的细胞系 将提供有益的补充洞察力的作用， 在传输路径上。