The kinetics of CD4+Foxp3+ T cell accumulation during a human cutaneous antigen-specific memory response in vivo
Milica Vukmanovic-Stejic, Elaine Agius, Nicola Booth, Padraic J. Dunne, Katie E. Lacy, John R. Reed, Toni O. Sobande, Steven Kissane, Mike Salmon, Malcolm H. Rustin, Arne N. Akbar
Milica Vukmanovic-Stejic, Elaine Agius, Nicola Booth, Padraic J. Dunne, Katie E. Lacy, John R. Reed, Toni O. Sobande, Steven Kissane, Mike Salmon, Malcolm H. Rustin, Arne N. Akbar
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Research Article

The kinetics of CD4+Foxp3+ T cell accumulation during a human cutaneous antigen-specific memory response in vivo

Abstract

Naturally occurring CD4+CD25hiFoxp3+ Tregs (nTregs) are highly proliferative in blood. However, the kinetics of their accumulation and proliferation during a localized antigen-specific T cell response is currently unknown. To explore this, we used a human experimental system whereby tuberculin purified protein derivative (PPD) was injected into the skin and the local T cell response analyzed over time. The numbers of both CD4+Foxp3– (memory) and CD4+Foxp3+ (putative nTreg) T cells increased in parallel, with the 2 populations proliferating at the same relative rate. In contrast to CD4+Foxp3– T cell populations, skin CD4+Foxp3+ T cells expressed typical Treg markers (i.e., they were CD25hi, CD127lo, CD27+, and CD39+) and did not synthesize IL-2 or IFN-γ after restimulation in vitro, indicating that they were not recently activated effector cells. To determine whether CD4+Foxp3+ T cells in skin could be induced from memory CD4+ T cells, we expanded skin-derived memory CD4+ T cells in vitro and anergized them. These cells expressed high levels of CD25 and Foxp3 and suppressed the proliferation of skin-derived responder T cells to PPD challenge. Our data therefore demonstrate that memory and CD4+ Treg populations are regulated in tandem during a secondary antigenic response. Furthermore, it is possible to isolate effector CD4+ T cell populations from inflamed tissues and manipulate them to generate Tregs with the potential to suppress inflammatory responses.

Authors

Milica Vukmanovic-Stejic, Elaine Agius, Nicola Booth, Padraic J. Dunne, Katie E. Lacy, John R. Reed, Toni O. Sobande, Steven Kissane, Mike Salmon, Malcolm H. Rustin, Arne N. Akbar

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Figure 2

The MT is a well-characterized model of a memory immune response.

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The MT is a well-characterized model of a memory immune response. Sample...
Samples were collected between 0 and 19 days after PPD injection. (A) Skin biopsies were collected for immunohistochemistry (top panel), and cutaneous lymphocytes were isolated from skin suction blisters that were induced over the sites of PPD injection at different time points (lower panel). (B) PPD-specific memory T cells accumulate during the course of MT. PBMCs and blister cells were stimulated with PPD for 15 hours in the presence of Brefeldin A and stained for intracellular expression of IFN-γ (right y axis, solid lines). The number of CD3+ T cells within dermal perivascular infiltrates was determined by indirect immunoperoxidase staining of 5-μm tissue sections. The 5 largest perivascular infiltrates were counted and data presented per unit area (UA). The mean ± SEM of 5 individuals per time point is shown (dashed line, left y axis). (C) Blister cells recovered from day 7 blisters were stimulated with PPD or with irrelevant antigen (HSV, CMV, tetanus) or left unstimulated and stained for intracellular expression of IFN-γ. Representative dot plots (n = 3 samples per antigen) are shown. Numbers denote the percentage of cells expressing IFN-γ. (D) IFN-γ production from day 7 blisters raised over MT response or the site of the control (saline) injection. Numbers denote the percentage of cells expressing IFN-γ. PBMCs and blister cells (PPD and control) from the same donor were stimulated with PPD overnight as described in Methods.