Immune responses to foreign and self-Ags can be controlled by regulatory T cells (Tregs) expressing CD4 and IL-2Rα chain (CD25). Defects in Tregs lead to autoimmunity, whereas induction of Ag-specific CD4⁺CD25⁺ Tregs restores tolerance. Ag-specific CD4⁺CD25⁺ FOXP3⁺Tregs activated by the T helper type 2 (Th2) cytokine, IL-4, and specific alloantigen promote allograft tolerance. These Tregs expressed the specific IL-5Rα and in the presence of IL-5 proliferate to specific but not third-party Ag. These findings suggest that recombinant IL-5 (rIL-5) therapy may promote Ag-specific Tregs to mediate tolerance. This study showed normal CD4⁺CD25⁺ Tregs cultured with IL-4 and an autoantigen expressed Il-5rα. Treatment of experimental autoimmune neuritis with rIL-5 markedly reduced clinical paralysis, weight loss, demyelination, and infiltration of CD4⁺ (Th1 and Th17) CD8⁺ T cells and macrophages in nerves. Clinical improvement was associated with expansion of CD4⁺CD25⁺FOXP3⁺ Tregs that expressed Il-5rα and proliferated only to specific autoantigen that was enhanced by rIL-5. Depletion of CD25⁺ Tregs or blocking of IL-4 abolished the benefits of rIL-5. Thus, rIL-5 promoted Ag-specific Tregs, activated by autoantigen and IL-4, to control autoimmunity. These findings may explain how Th2 responses, especially to parasitic infestation, induce immune tolerance. rIL-5 therapy may be able to induce Ag-specific tolerance in autoimmunity.

Resting and activated subpopulations of CD4⁺CD25⁺CD127^loT regulatory cells (Treg) and CD4⁺CD25⁺CD127⁺ effector T cells in MS patients and in healthy individuals were compared. Peripheral blood mononuclear cells isolated using Ficoll Hypaque were stained with monoclonal antibodies and analysed by flow cytometer. CD45RA and Foxp3 expression within CD4⁺ cells and in CD4⁺CD25⁺CD127^loT cells identified Population I; CD45RA⁺Foxp3⁺, Population II; CD45RA⁻Foxp3^hi and Population III; CD45RA⁻Foxp3⁺ cells. Effector CD4⁺CD127⁺ T cells were subdivided into Population IV; memory /effector CD45RA⁻ CD25⁻Foxp3⁻ and Population V; effector naïve CD45RA⁺CD25⁻Foxp3⁻CCR7⁺ and terminally differentiated RA⁺ (TEMRA) effector memory cells. Chemokine receptor staining identified CXCR3⁺Th1-like Treg, CCR6⁺Th17-like Treg and CCR7⁺ resting Treg. Resting Treg (Population I) were reduced in MS patients, both in untreated and treated MS compared to healthy donors. Activated/memory Treg (Population II) were significantly increased in MS patients compared to healthy donors. Activated effector CD4⁺ (Population IV) were increased and the naïve/ TEMRA CD4⁺ (Population V) were decreased in MS compared to HD. Expression of CCR7 was mainly in Population I, whereas expression of CCR6 and CXCR3 was greatest in Populations II and intermediate in Population III. In MS, CCR6⁺Treg were lower in Population III. This study found MS is associated with significant shifts in CD4⁺T cells subpopulations. MS patients had lower resting CD4⁺CD25⁺CD45RA⁺CCR7⁺ Treg than healthy donors while activated CD4⁺CD25^hiCD45RA⁻Foxp3^hiTreg were increased in MS patients even before treatment. Some MS patients had reduced CCR6⁺Th17-like Treg, which may contribute to the activity of MS.

In rat models, CD4+CD25+T regulatory cells (Treg) play a key role in the induction and maintenance of antigen specific transplant tolerance, especially in DA rats with PVG cardiac allografts(1, 2). We have previously described generation of alloantigen specific Treg (Ts1), by culture of naïve natural CD4+CD25+ Treg (nTreg) with specific alloantigen and IL-2 for 4 days. These cells express mRNA for IFN-γ receptor (ifngr) and suppress donor but not third party cardiac allograft rejection mediated by alloreactive CD4+T cells at ratios of We induced highly suppressive Th1-like Treg from naïve nTreg in 7 days by culture with alloantigen, first with rIL-2 then with rIL-12p70. These Th1-like Treg delayed specific-donor allograft rejection demonstrating therapeutic potential

The T helper (Th) 2 cytokine interleukin (IL)-4 has been implicated as a major regulatory cytokine for the induction of transplant tolerance, but few studies have examined the capacity of IL-4 to induce tolerance. The effect of IL-4 therapy alone or with low doses of anti-CD4 monoclonal antibody (mAb) therapy on survival of fully allogeneic PVG neonatal heart graft in adult DA rats was examined. Rat recombinant (r) IL-4 was given at 30 μg(10 4 U)/kg daily intraperitoneally for 10 days and MRC OX35 (anti-CD4, nondepleting) or MRC OX81 (anti-IL-4) was given intraperitoneally on days 0, 3, 7, and 10. Semiquantitative reverse transcriptase-polymerase chain reaction was used to assay mRNA for cytokine in the graft, regional node and spleen and fluorescence-activated cell sorting was used to assay alloantibody Ig isotypes. Grafts in rIL-4-treated rats survived a median period of 39 days(range, 28-52 days), significantly longer than in both untreated and nontransfected Chinese hamster ovary-K1 supernatant-treated controls (median, 14 days; range, 10-16 days, P =0.009). rIL-4 treatment with a suboptimal dose of anti-CD4 mAb prolonged median survival to 70 days(range, 63-80 days), which was longer than rIL-4 treatment alone or anti-CD4 mAb alone (median, 36 days; range, 30-55 days; P <0.0045). Combining MRC OX81 with MRC OX35 therapy led to earlier rejection at a median period of 26 days (range, 20-28 days); MRC OX81 alone had no effect on graft survival. Alloantibody titers, especially IgG1, were higher in rIL-4-treated animals and lower in anti-CD4 mAb-treated animals than in animals with normal rejection( P <0.05). IL-4 mRNA was increased in regional lymph nodes and spleen of the rIL-4-treated groups compared with all other groups, but there were no differences for IL-2, interferon-γ, or IL-10. rIL-4 therapy markedly prolonged neonatal cardiac allograft survival, and, with anti-CD4 therapy, it further prolonged survival. It induced IL-4 mRNA in lymphoid tissues and enhanced alloantibody production, especially IgG1, which demonstrated enhanced Th2 responses, but did not affect Th1 cytokines.

CD4⁺CD25⁺Foxp3⁺ T cells are regulatory/suppressor cells (Tregs) that include non-antigen (Ag)–specific as well as Ag-specific Tregs. How non–Ag-specific naive CD4⁺CD25⁺ Treg develop into specific Tregs is unknown. Here, we generated adaptive Tregs by culture of naive CD4⁺CD25⁺Foxp3⁺ T cells with allo-Ag and either interleukin-2 (IL-2) or IL-4. Within days, IL-2 enhanced interferon-γ receptor (Ifnγr) and Il-5 mRNA and IL-4 induced a reciprocal profile with de novo IL-5Rα and increased IFN-γ mRNA expression. Both IL-2– and IL-4–alloactivated CD4⁺CD25⁺ Tregs within 3 to 4 days of culture had enhanced capacity to induce tolerance to specific donor but not to third-party cardiac allografts. These hosts became tolerant as allografts functioned more than 250 days, with a physiologic ratio of less than 10% CD4⁺CD25⁺Foxp3⁺ T cells in the CD4⁺ population. CD4⁺CD25⁺ T cells from tolerant hosts given IL-2–cultured cells had increased Il-5 and Ifnγr mRNA. Those from hosts given IL-4–cultured cells had enhanced IL-5Rα mRNA expression and IL-5 enhanced their proliferation to donor but not third-party allo-Ag. Thus, IL-2 and IL-4 activated allo-Ag–specific Tregs with distinct phenotypes that were retained in vivo. These findings suggested that T-helper 1 (Th1) and Th2 responses activate 2 pathways of adaptive Ag-specific Tregs that mediate tolerance. We propose they be known as T-suppressor 1 (Ts1) and Ts2 cells.