Circulating T Regulatory Cells Are Persistently Reduced in Non-Severe Acquired Aplastic Anemia

Abstract

Objective

Acquired aplastic anemia (aAA) is characterized by an autologous immunological attack against hematopoietic stem and progenitor cells, and immunotolerance disruption is frequent, with reduced T regulatory cells (Tregs) frequencies and increased effector cytotoxic cells. Tregs are reduced in aAA and increase in number after successful immunosuppressive therapies.

Methods

In this retrospective study, we investigated the frequency of circulating Tregs by multiparametric flow cytometry immunophenotyping in non-severe aAA patients before and after immunosuppressive therapy. The samples were stained with the following antibodies: ECD anti-CD3, PE or PC5 anti-CD4, FITC anti-CD8, and PE anti-CD25, and Tregs were identified by first gating on linear parameters for lymphocyte identification and then for CD3 expression. In CD3⁺CD4⁺ cells, Tregs were further identified on the basis of CD25 and FOXP3 expression.

Results

Although the number of Tregs tended to increase after immunosuppressive treatments, their circulating frequency remained lower than that of healthy subjects, regardless of their responsiveness to therapies. Moreover, the relative frequency combined with absolute Treg counts might be more informative in the differential diagnosis of bone marrow failure syndromes.

Conclusions

The persistent decrease in circulating Tregs could be the result of immunosuppressive agents that could preferentially expand other T-cell subsets. At the same time, an imbalance in immunotolerance might persist, which is also favored by chronic antigen stimulation.

Keywords

Regulatory T cells (Tregs) / Acquired aplastic anemia (aAA) / Immnosuppressive therapy (IST) / Persistent reduction / Nonsevere aplastic anemia / FOXP3

References

[1] GiudiceV, SelleriC. Aplastic anemia: Pathophysiology. Semin Hematol., 2022, 59(1): 13-20.

[2] PatelBA, GiudiceV, YoungNS. Immunologic effects on the haematopoietic stem cell in marrow failure. Best Pract Res Clin Haematol., 2021, 342. 101276

[3] GiudiceV, FengX, LinZ, et al.. Deep sequencing and flow cytometric characterization of expanded effector memory CD8+CD57+ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia. Haematologica., 2018, 103(5): 759-769.

[4] GiudiceV, CardamoneC, TriggianiM, et al.. Bone marrow failure syndromes, overlapping diseases with a common cytokine signature. Int J Mol Sci., 2021, 222705.

[5] SuH, LonghiMS, WangP, et al.. Human CD4+CD25(high)CD127 (low/neg) regulatory T cells. Methods Mol Biol., 2012, 806: 287-299.

[6] ChenX, OppenheimJJ. Resolving the identity myth: key markers of functional CD4+FoxP3+ regulatory T cells. Int Immunopharmacol., 2011, 11(10): 1489-1496.

[7] VignaliDA, CollisonLW, WorkmanCJ. How regulatory T cells work. Nat Rev Immunol., 2008, 8(7): 523-532.

[8] de LatourRP, VisconteV, TakakuT, et al.. Th17 immune responses contribute to the pathophysiology of aplastic anemia. Blood., 2010, 116(20): 4175-4184.

[9] SolomouEE, RezvaniK, MielkeS, et al.. Deficient CD4+ CD25+ FOXP3+ T regulatory cells in acquired aplastic anemia. Blood., 2007, 110(5): 1603-1606.

[10] KordastiS, MarshJ, Al-KhanS, et al.. Functional characterization of CD4+ T cells in aplastic anemia. Blood., 2012, 119(9): 2033-2043.

[11] GiudiceV, BiancottoA, WuZ, et al.. Aptamer-based proteomics of serum and plasma in acquired aplastic anemia. Exp Hematol., 2018, 68: 38-50.

[12] ZhaoX, FengX, WuZ, et al.. Persistent elevation of plasma thrombopoietin levels after treatment in severe aplastic anemia. Exp Hematol., 2018, 58: 39-43.

[13] DeZernAE, ChurpekJE. Approach to the diagnosis of aplastic anemia. Blood Adv., 2021, 5(12): 2660-2671.

[14] GaleRP, HinterbergerW, YoungNS, et al.. What causes aplastic anaemia?. Leukemia., 2023, 37(6): 1191-1193.

[15] SerioB, GiudiceV, SelleriC. All roads lead to interferon-γ: from known to untraveled pathways in acquired aplastic anemia. Medicina (Kaunas)., 2023, 59122170.

[16] TeramuraM, KobayashiS, IwabeK, et al.. Mechanism of action of antithymocyte globulin in the treatment of aplastic anaemia: in vitro evidence for the presence of immunosuppressive mechanism. Br J Haematol., 1997, 96(1): 80-84.

[17] ShiJ, GeM, LuS, et al.. Intrinsic impairment of CD4(+)CD25(+) regulatory T cells in acquired aplastic anemia. Blood., 2012, 120(8): 1624-1632.

[18] LimSP, CostantiniB, MianSA, et al.. Treg sensitivity to FasL and relative IL-2 deprivation drive idiopathic aplastic anemia immune dysfunction. Blood., 2020, 136(7): 885-897.

[19] GurkanS, LuanY, DhillonN, et al.. Immune reconstitution following rabbit antithymocyte globulin. Am J Transplant., 2010, 10(9): 2132-2141.

[20] AggarwalN, ManleyAL, ChenJ, et al.. Effects of ruxolitinib on murine regulatory T cells are immune-context dependent. Exp Hematol., 2023, 125–126: 16-19.

[21] FurukawaA, WiselSA, TangQ. Impact of immune-modulatory drugs on regulatory T cell. Transplantation., 2016, 100(11): 2288-2300.

[22] CrepinT, CarronC, RoubiouC, et al.. ATG-induced accelerated immune senescence: clinical implications in renal transplant recipients. Am J Transplant., 2015, 15(4): 1028-1038.

[23] PiekarskaA, PawelecK, Szmigielska-KapłonA, et al.. The state of the art in the treatment of severe aplastic anemia: immunotherapy and hematopoietic cell transplantation in children and adults. Front Immunol., 2024, 151378432.

RIGHTS & PERMISSIONS

The Author(s), under exclusive licence to Huazhong University of Science and Technology