Direct demonstration of cytokine synthesis heterogeneity among human memory/effector T cells by flow cytometry.

The array of cytokines produced by T cells in effector sites is a primary means by which these cells mediate host defense. It is well recognized that cloned T cells are heterogeneous with regard to cytokine synthesis and, thus, in their ability to mediate specific immune responses, but the extent to which the patterns of cytokine secretion observed in cloned cells reflect actual populations of memory/effector T cells existing in vivo is largely unknown. Here, we report our findings using a multiparameter flow cytometric assay that allows simultaneous determination of an individual T-cell's ability to produce multiple cytokines and its phenotype after only short (4 to 8 hours) in vitro incubation with an activating stimulus and the secretion inhibitor Brefeldin A.

Coordinate expression of beta 1 and beta 2 integrin "activation" epitopes during T cell responses in secondary lymphoid tissue.

The monoclonal antibodies (mAb) 15/7 and 24 recognize unique activation-dependent, conformational epitopes on beta 1 and beta 2-integrins, respectively. The expression of both of these epitopes closely correlates with the ligand binding ability of their respective integrins, and thus serves as indicators of functional integrin "activation". Here, we have used six-parameter flow cytometry to examine the expression of these epitopes and conventional beta 1- and beta 2-integrin epitopes during human T cell activation in secondary lymphoid tissues in vivo, focusing particularly on the virgin to memory/effector cell transition.

Control of lymphocyte recirculation in man. II. Differential regulation of the cutaneous lymphocyte-associated antigen, a tissue-selective homing receptor for skin-homing T cells.

Recent work indicates that a novel homing receptor (HR)/endothelial ligand pair--the cutaneous lymphocyte-associated Ag (CLA) and E-selectin--is involved in targeting a unique skin-associated subpopulation of memory T cells to cutaneous sites of chronic inflammation. To investigate the regulatory mechanisms responsible for the generation of a memory T cell subset with skin-selective homing capability, we used multiparameter flow cytometry to assess the expression of CLA on T cells during initial T cell activation in secondary lymphoid tissues (the virgin to memory transition), and upon reactivation in skin. Our analyses indicate that in vivo induction of CLA (and E-selectin-binding ability) first occurs during the virgin to memory transition, and is regulated in a highly tissue-selective manner.

Control of lymphocyte recirculation in man. I. Differential regulation of the peripheral lymph node homing receptor L-selectin on T cells during the virgin to memory cell transition.

Conventional virgin lymphocytes of a given class show relatively homogeneous recirculation through secondary lymphoid tissues, whereas memory/effector populations are composed of distinct subsets with differential, often tissue-selective migratory capability. In keeping with these observations, CD45RAhigh/ROlow "virgin" T cells in human peripheral blood uniformly express the peripheral lymph node homing receptor (HR) L-selectin, whereas among the CD45RAlow/ROhigh "memory/effector" subset, the expression of this HR is bimodal. To investigate the mechanisms responsible for the generation of memory/effector T cell subsets with differential homing potential, we developed a multiparameter flow cytometric technique that defines a common pathway of post-thymic T cell differentiation in secondary lymphoid tissues.