Elsevier

Human Immunology

Volume 81, Issue 8, August 2020, Pages 452-459
Human Immunology

Research article
T follicular helper cell-mediated IL-21 production suppresses FOXP3 expression of T follicular regulatory-like cells in diffuse large B cell lymphoma patients

https://doi.org/10.1016/j.humimm.2020.05.008Get rights and content

Abstract

Based on CD25 expression, T follicular helper cells (Tfh) could be divided into T follicular regulatory (Tfr)-like subset (CD25+CD4+CXCR5+) and CD25 Tfh subset (CD25CD4+CXCR5+). Patients with diffuse large B cell lymphoma (DLBCL) display high level of Tfr-like cells in blood and tumor. This Tfr-like subset could suppress CD8 T cell response while promote tumor cell proliferation. In this study, we investigated the transcription factors and regulatory elements associated with Tfr-like cells in DLBCL patients. Both circulating and tumor-infiltrating Tfr-like cells presented slightly higher Blimp-1 expression and significantly higher Foxp3 expression than the CD25 Tfh subset. As the IL-2 receptor, CD25 could be moderately upregulated in stimulated CD25 Tfh cells. However, stimulated CD25 Tfh cells could not upregulate Foxp3, indicating that the distinction between Foxp3-low CD25CXCR5+CD4+ T cells and Foxp3-high CD25+CXCR5+CD4+ T cells was not due to differences in stimulation status. Regarding cytokine production, while both Tfr-like and CD25 Tfh cells upregulated IL-21 and IL-10 during stimulation, the CD25 Tfh cells presented significantly higher IL-21 and lower IL-10 expression than the Tfr-like cells, and the TGF-β expression was only increased in Tfr-like cells. Interestingly, IL-21 secreted from CD25 Tfh cells negatively regulated the expression of Foxp3 and IL-10 of autologous Tfr-like cells. Together, these results demonstrated that the Tfr-like and CD25 Tfh subsets of circulating Tfh cells presented different functions and should be investigated separately.

Introduction

Interleukin (IL)-21 is a pleiotropic cytokine belonging to the IL-2 family [1]. The IL-21 receptor is a heterodimer comprised of the IL-21R chain and the common γ (γc) chain, and signals via JAK1/JAK3 to mediate STAT3, STAT1, and STAT5 phosphorylation [2]. IL-21 is produced by activated T follicular helper (Tfh) cells, T helper (Th)17 cells, natural killer T (NKT) cells, and CD8 T cells, and can act on a variety of immune cells to perform diverse effector functions [3], [4], [5]. In germinal center, Tfh-mediated IL-21 promotes the proliferation, immunoglobulin production, and plasma cell differentiation of B cells [6]. IL-21 is also involved in the differentiation, proliferation, and cytokine production of Th17 and Tfh cells [7], [8]. TGF-β promotes Treg differentiation, but in the presence of either IL-21 or IL-6, Th17 differentiation is favored. Additionally, IL-21 inhibits IL-2 production and decreases Treg viability [9]. In tumor, IL-21 potently enhances the survival and cytotoxicity of CD8 T cells, NK cells, and NKT cells [5], [10]. On the other hand, IL-21 increases the production of IL-10, a regulatory cytokine, in Th1, Th17, and B cells [11], [12]. IL-21 acts in opposition to GM-CSF and limits the number of conventional dendritic cells via STAT3-dependent and BIM-dependent induction of apoptosis [13].

Diffuse large B cell lymphoma (DLBCL) is an aggressive B cell lymphoma currently treated via a standard R-CHOP (anti-CD20 rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone) regimen [14], [15]. Heterogeneous response to R-CHOP is observed in DLBCL patients, with complete remission in some patients and partial remission in others. A subset of patients may develop relapsed disease following remission, which is harder to manage [16]. In addition, prediction of patient response is not yet possible, further complicating the treatment process. These difficulties are present, largely due to a lack of understanding in the complex interactions between various cellular and molecular mechanisms involved in the pathogenesis of DLBCL.

Several DLBCL cell lines are shown to express IL-21R, and upon receiving IL-21, a proapoptotic pathway involving the phosphorylation of STAT3 and upregulation of c-Myc is activated, resulting in the death of DLBCL but not healthy B cells [17]. However, the CXCR5+ circulating CD4 T cells, which normally express higher IL-21 than CXCR5- circulating CD4 T cells and are regarded as the peripheral counterpart of germinal center T follicular helper cells (Tfh) [18], expressed lower IL-21 and higher IL-10 in DLBCL patients than in healthy controls, and promoted primary DLBCL survival via IL-10-mediated effects [19]. We later showed that in DLBCL patients, a significantly higher proportion of CXCR5+ CD4 T cells expressed Foxp3, similar to the T follicular regulatory (Tfr) cells. This Tfr-like subset existed at even higher frequency in the tumor, and supported the proliferation of autologous CD19+ tumor cells. The effect of IL-21 on the regulation of Tfr cells remains unclear, and is investigated in this study.

Section snippets

Study participants

Twenty-seven subjects with de novo presentation of primary DLBCL and twenty-seven healthy subjects were included in this study. The patients presented stage III DLBCL at diagnosis, based on the Ann Arbor Classification System [20]. The patient group and the control group each included 15 females and 12 males between 40 and 70 years of age. All participants gave written informed consent. Ethical approval was obtained from the Ethics Committee of the First Affiliated Hospital of Xiamen University

Circulating CD25+CXCR5+CD4+ Tfr-like cells presented high Bcl-6, low Blimp-1, and high Foxp3 expression

Peripheral blood mononuclear cells (PBMCs) and tumor-infiltrating lymphocytes (TILs) were harvested from twenty-seven patients with newly diagnosed, untreated DLBCL. We previously showed that the Tfr cells were characterized by CD4+CD25+CXCR5+ surface marker expression [21]. Here, we sorted circulating CD4 T cells into CD25+CXCR5+, CD25CXCR5+, and CXCR5 fractions (Fig. 1A). Compared to healthy subjects, DLBCL patients presented higher frequencies of CD25+CXCR5+ CD4 T cells and lower

Discussion

In the current investigation, we demonstrated that the Tfh cells (CXCR5+CD4+ T cells) could be separated into the CD25+CXCR5+ subset (Tfr-like) and the CD25CXCR5+ subset (CD25 Tfh), which were different in the expression of transcription factors and cytokines. The Tfr-like cells (CD25+CXCR5+CD4+) presented slightly higher Blimp-1 and significantly higher Foxp3 than the CD25 Tfh cells (CD25CXCR5+CD4+). The cytokine expression was also distinctive in each subset. Most of the previous studies

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This work was supported by National Natural Science Foundation of China (No.81770126; No.81800163; No.81800196), and Foundation of health and family planning Commission of in Fujian Province of China (2017-2-99).

References (27)

  • C.G. Vinuesa et al.

    Blood relatives of follicular helper T cells

    Immunity

    (2011)
  • R. Spolski et al.

    Interleukin-21: basic biology and implications for cancer and autoimmunity

    Annu. Rev. Immunol.

    (2008)
  • J.M. Coquet et al.

    IL-21 is produced by NKT cells and modulates NKT cell activation and cytokine production

    J. Immunol.

    (2007)
  • Cited by (7)

    View all citing articles on Scopus
    1

    These authors contributed equally to this work.

    View full text