Differentiation of effector/memory Vdelta2 T cells and migratory routes in lymph nodes or inflammatory sites

Abstract

Vdelta2 T lymphocytes recognize nonpeptidic antigens without presentation by MHC molecules and mount both immediate effector functions and memory responses after microbial infection. However, how Vdelta2 T cells mediate different facets of a memory response remains unknown. Here, we show that the expression of CD45RA and CD27 antigens defines four subsets of human Vdelta2 T cells with distinctive compartmentalization routes. Naive CD45RA+CD27+ and memory CD45RA-CD27+ cells express lymph node homing receptors, abound in lymph nodes, and lack immediate effector functions. Conversely, memory CD45RA-CD27- and terminally differentiated CD45RA+CD27- cells, which express receptors for homing to inflamed tissues, are poorly represented in the lymph nodes while abounding at sites of inflammation, and display immediate effector functions. These observations and additional in vitro experiments indicate a lineage differentiation pattern for human Vdelta2 T cells that generates naive cells circulating in lymph nodes, effector/memory cells patrolling the blood, and terminally differentiated effector cells residing in inflamed tissues.

Figure 1.

Phenotypic characterization of Vδ2 T cell subsets. (A) Peripheral blood mononuclear cells were stained with antibodies to Vδ2, CD45RA, and CD27. Upon analysis on gated Vδ2⁺ cells, three subsets were identified. (B and C) Two-color FACS^® analysis of peripheral blood mononuclear cells stained with antibodies to Vδ2, CD3, and CD8. (D) Upon analysis in a healthy population, the proportion of cells within different subsets was variable among individuals, the variability being more pronounced in the CD45RA⁻CD27⁻ subset.

Figure 2.

Distribution of Vδ2 T cell subsets in lymph nodes. (A) Mononuclear cells were taken from inguinal lymph nodes of organ donors. Cells were stained with antibodies to Vδ2, CD45RA, and CD27. As a control, cells were double stained with antibodies to Vδ2 and Vγ9. (B) Analysis of the relative proportion of the CD45RA⁺CD27⁺, CD45RA⁻CD27⁺, and CD45RA⁻CD27⁻ subsets of Vδ2⁺ T cells in the lymph nodes of six donors. (C) Vδ2 T cell subsets were sorted and analyzed for the expression of different chemokine receptors and CD62L. Results similar to those shown here were obtained in a total of six donors.

Figure 3.

Distribution of Vδ2 T cell subsets at sites of inflammation. Mononuclear cells were collected from one ascitic (A) and two cerebrospinal fluids (B and C) from three patients affected by tuberculous peritonitis and meningitis, respectively. Cells were stained with antibodies to Vδ2, CD45RA, and CD27. (D–F) Vδ2 T cell subsets were sorted and analyzed for the expression of different chemokine receptor and CD62L.

Figure 4.

Different subsets of Vδ2 T cells display different effector functions. The four subsets of Vδ2 T cells were sorted according to the expression of CD45RA and CD27 and were tested for their ability to (A) proliferate, (B) produce IFNγ, and (C) release BLT-esterase (as a parameter of cytotoxic activity) upon in vitro stimulation with IPP. Comparable results were obtained in eight different healthy donors. (A) Triangles, CD45RA⁺CD27⁺; diamonds, CD45RA⁻ CD27⁺; squares, CD45RA⁻CD27⁻; and circles, CD45RA⁺CD27⁻. (B and C) Black bars, CD45RA⁺CD27⁺; hatched bars, CD45RA⁻CD27⁺; open bars, CD45RA⁻CD27⁻; and gray bars, CD45RA⁺CD27⁻. (D) The CD45RA⁺CD27⁺ and CD45RA⁻CD27⁺ subsets of Vδ2⁺ T cells were sorted and the expression of Vγ9 was assessed by FACS^® staining.

Figure 5.

Lineage developmental pattern of different subsets of Vδ2 T cells. CD45RA⁺CD27⁺, CD45RA⁻CD27⁺, and CD45RA⁻CD27⁻ subsets of Vδ2 T cells were sorted from peripheral blood mononuclear cells and cultured in vitro with IPP and IL-2 for 12 d. Cells were collected and tested for CD45RO and CD27 expression (A, C, and E) and for their ability to proliferate and produce IFNγ upon restimulation with IPP in vitro (B, D, and F). (G) The length of telomeres in CD45RA⁺CD27⁺ (lane 2), CD45RA⁻CD27⁺ (lane 3), and CD45RA⁻CD27⁻ (lane 4) represents subsets of peripheral blood Vδ2 T cells. Lane 1, molecular weight markers. kb, kilobases.