MEMBRANE LIPID ORDER AND LYMPHOCYTE FUNCTION

磷脂组织的改变， 淋巴细胞质膜可影响多种淋巴细胞 功能协调发展的 本建议旨在评估以下方面的作用： 磷脂包装在淋巴细胞与 网状内皮细胞和细胞溶解性T细胞的目标 淋巴细胞（CTL）。 分离的未成熟和活化淋巴细胞 结合mercury 540（MC 540），一种荧光 亲脂性探针的松散包装的脂质，而淋巴细胞 与外周循环隔离的血液则不能。 这是否 磷脂包装的差异是导致 这些淋巴细胞与其他细胞的相互作用将 通过几种方式测试。 利用离心作用产生一种 控制和可再现的力，以区分粘附和 非贴壁细胞，外周血淋巴细胞与非贴壁细胞相互作用的强度， 血淋巴细胞、胸腺细胞和脾细胞与巨噬细胞 并测量内皮细胞。 使用流式细胞术， 用MC 540染色的胸腺细胞和脾细胞将被 根据荧光强度分级， 测定的级分的粘附性质。 的 细胞相互作用对脂质组织的依赖性将是 通过从循环中清除来进行体内检查 回输的淋巴细胞磷脂堆积不同。 两 生物物理方法将被用来研究机制 磷脂组织的改变会影响 坚持。 光漂白后的荧光恢复将是 用于评估是否增加了 胸腺细胞与松散包装的脂质的结果增加 膜受体的迁移率。 荧光转移法 将被用来确定是否松散包装的脂质在 活化的淋巴细胞伴随着局部聚集的 淋巴细胞粘附相关膜蛋白LFA-1。 最后， CTL中的磷脂组织和它们的靶点将是 通过MC 540染色评估，以评估这种细胞的潜在作用。 CTL对自身裂解的抗性和CTL对自身裂解的抗性中的参数。 杀死靶细胞的效率。

Alterations in the organization of the phospholipids in the lymphocyte plasma membrane may affect several lymphocyte functions. This proposal is designed to assess the role of phospholipid packing in the interactions of lymphocytes with reticuloendothelial cells and with the targets of cytolytic T lymphocytes (CTL). Immature and activated lymphocytes isolated from tissues bind merocyanine 540 (MC540), a fluorescent lipophilic probe for loosely-packed lipids, whereas lymphocytes isolated from the peripheral circulation do not. Whether this difference in phospholipid packing is responsible for differences in the interactions of these lymphocytes with other cells will be tested in several ways. Using centrifugation to produce a controlled and reproducible force to distinguish adherent from nonadherent cells, the strength of the interactions of peripheral blood lymphocytes, thymocytes and splenocytes with macrophages and endothelial cells will be measured. Using flow cytometry, thymocytes and splenocytes stained with MC540 will be fractionated according to fluorescence intensity and the adherence properties of the fractions determined. The dependence of cellular interactions on lipid organization will be examined in vivo by following clearance from the circulation of reinfused lymphocytes which differ in phospholipid packing. Two biophysical approaches will be used to investigate the mechanisms by which altered phospholipid organization could affect adherence. Fluorescence recovery after photobleaching will be employed to assess whether increases agglutinability of thymocytes with loosely-packed lipids results from increased mobility of membrane receptors. Fluorescence transfer methods will be used to determine whether the loose packing of lipids in activated lymphocytes is accompanied by local aggregation of a lymphocyte adhesion-related membrane protein, LFA-1. Finally, phospholipid organization in CTL and their targets will be assessed by MC540 staining to evaluate the potential role of this parameter in the resistance of CTL to self-lysis and in the efficiency of killing of target cells.