Takata Hiroshi, Takiguchi Masafumi
Division of Viral Immunology, Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
J Immunol. 2006 Oct 1;177(7):4330-40. doi: 10.4049/jimmunol.177.7.4330.
Multicolor flow cytometric analysis for the expression of three effector molecules, i.e., perforin (Per), granzyme A (GraA), and granzyme B (GraB), in human CD8(+) T cells demonstrated that they included five subpopulations, implying the following pathway for the differentiation of CD8(+) T cells: Per(-)GraA(-)GraB(-)-->Per(-)GraA(+)GraB(-)-->Per(low)GraA(+)GraB(-)--> Per(low)GraA(+)GraB(+)-->Per(high)GraA(+)GraB(+). The analysis of the expression of these molecules in the subsets classified by the combination of the expression of CCR7 and CD45RA or by that of CD27, CD28, and CD45RA showed that functional CD8(+) T cell subsets could be partially identified by these phenotypic classifications. However, the functional subsets could be precisely identified by the classification using five cell surface markers or three cell surface markers and three cytolytic molecules. Per(-)GraA(-)GraB(-) and Per(-/low)GraA(+)GraB(-) cells were predominantly found in CCR5(-)CCR7(+) and CCR5(high/low)CCR7(-) subsets, respectively, of CD8(+) T cells expressing the CD27(+)CD28(+)CD45RA(-) phenotype, whereas Per(low)GraA(+)GraB(+) cells were found in the CCR5(low)CCR7(-) subset of those expressing this phenotype and in a part of the CCR5(-/low)CCR7(-) subset of those expressing the CD27(-/low)CD28(-)CD45RA(-/+) phenotype. Ex vivo EBV-specific CD8(+) T cells, which were Per(low/-)GraA(+)GraB(-/+) cells, hardly or very weakly killed the target cells, indicating that these were not effector T cells. These findings suggest that the Per(-)GraA(-)GraB(-), Per(-/low)GraA(+)GraB(-), and Per(low)GraA(+)GraB(+) cells were central memory, early effector memory, and late effector memory T cells, respectively. Per(-/low)GraA(+)GraB(-) cells gained GraB expression after TCR stimulation, indicating that early effector memory T cells could differentiate into late effector and effector T cells. The present study showed the existence of three memory subsets and the pathway for their differentiation.
通过多色流式细胞术分析人CD8(+) T细胞中三种效应分子即穿孔素（Per）、颗粒酶A（GraA）和颗粒酶B（GraB）的表达情况，结果表明它们包含五个亚群，这意味着CD8(+) T细胞的分化途径如下：Per(-)GraA(-)GraB(-)-->Per(-)GraA(+)GraB(-)-->Per(低)GraA(+)GraB(-)--> Per(低)GraA(+)GraB(+)-->Per(高)GraA(+)GraB(+)。根据CCR7和CD45RA表达的组合或CD27、CD28和CD45RA的表达对亚群进行分类，并分析这些分子在其中的表达情况，结果显示通过这些表型分类可以部分鉴定功能性CD8(+) T细胞亚群。然而，使用五个细胞表面标志物或三个细胞表面标志物和三个溶细胞分子进行分类可以精确鉴定功能亚群。Per(-)GraA(-)GraB(-)和Per(-/低)GraA(+)GraB(-)细胞分别主要存在于表达CD27(+)CD28(+)CD45RA(-)表型的CD8(+) T细胞的CCR5(-)CCR7(+)和CCR5(高/低)CCR7(-)亚群中，而Per(低)GraA(+)GraB(+)细胞存在于表达该表型的CCR5(低)CCR7(-)亚群以及表达CD27(-/低)CD28(-)CD45RA(-/+)表型的CCR5(-/低)CCR7(-)亚群的一部分中。体外EBV特异性CD8(+) T细胞为Per(低/-)GraA(+)GraB(-/+)细胞，几乎不或非常微弱地杀伤靶细胞，表明这些不是效应T细胞。这些发现提示Per(-)GraA(-)GraB(-)、Per(-/低)GraA(+)GraB(-)和Per(低)GraA(+)GraB(+)细胞分别为中枢记忆T细胞、早期效应记忆T细胞和晚期效应记忆T细胞。Per(-/低)GraA(+)GraB(-)细胞在TCR刺激后获得GraB表达，表明早期效应记忆T细胞可分化为晚期效应T细胞和效应T细胞。本研究显示了三种记忆亚群的存在及其分化途径。
