Many Faces of Regulatory T Cells: Heterogeneity or Plasticity?

Abstract

Regulatory T cells (Tregs) are essential for maintaining the immune balance in normal and pathological conditions. In autoimmune diseases and transplantation, they restrain the loss of self-tolerance and promote engraftment, whereas in cancer, an increase in Treg numbers is mostly associated with tumor growth and poor prognosis. Numerous markers and their combinations have been used to identify Treg subsets, demonstrating the phenotypic diversity of Tregs. The complexity of Treg identification can be hampered by the unstable expression of some markers, the decrease in the expression of a specific marker over time or the emergence of a new marker. It remains unclear whether such phenotypic shifts are due to new conditions or whether the observed changes are due to initially different populations. In the first case, cellular plasticity is observed, whereas in the second, cellular heterogeneity is observed. The difference between these terms in relation to Tregs is rather blurred. Considering the promising perspectives of Tregs in regenerative cell-based therapy, the existing confusing data on Treg phenotypes require further investigation and analysis. In our review, we introduce criteria that allow us to distinguish between the heterogeneity and plasticity of Tregs normally and pathologically, taking a closer look at their diversity and drawing the line between two terms.

Keywords: heterogeneity; phenotype switch; plasticity; regulatory T cells; treg-based therapy.

Figure 1

Heterogeneity in thymic development of Tregs. Two types of TregPs arise from CD4⁺ SP thymocytes. APCs present self-peptides to CD4⁺ SP thymocytes via the TCR, where intermediate levels of signaling give rise to the Treg developmental pathway. Strong signals result in negative selection. TCR- and co-stimulatory CD28-mediated signaling is required for both types of TregPs, with TCR signaling being stronger for CD25⁺ TregP cells. The development of CD25^–Foxp3^low TregP cells is also dependent on LFA-1, which enhances TCR signaling. The TCR-dependent stage induces the expression of TNFRSF members by CD25⁺ TregP cells. They increase their sensitivity to IL-2. In the TCR-independent stage, CD25⁺ TregP cells convert into mature CD25⁺Foxp3⁺ Tregs. This process is dependent on IL-2 or on the related common γ-chain cytokines IL-15 and IL-7. CD25^–Foxp3^low TregP cells require IL-15 at this stage. FoxP3 expression further upregulates the expression of other Treg-associated molecules. Mature Tregs derived from two types of TregPs show functional differences. Tregs from CD25⁺ TregP cells prevent experimental autoimmune encephalitis, whereas Tregs from FOXP3^low TregP cells are able to suppress colitis.

Figure 2

Plasticity of Tregs during differentiation. Thymic precursors give rise to nTregs and Tconv cells. In SLOs, they are activated by antigen presentation driven by APCs. Strong antigen stimulation leads to the differentiation of nTregs into eTregs, which enter the periphery. Tconvs that have received appropriate antigen stimulation also require TGF-β and IL-2 for further differentiation into Tregs. Under inflammatory conditions, they are activated and differentiate into eTregs. The differentiation of eTregs into cmTregs occurs with their retention in SLOs with the upregulation of homing molecules such as CD62L and CD44. emTregs are cells that have responded to antigens and that can survive for long periods in the absence of antigens in the peripheral circulation or in non-lymphoid tissues. Tissue homing of emTregs is associated with the expression of chemokine receptors and adhesion molecules that define the presence of specialized tissue-resident Treg subsets in the muscle, skin, lung, gut, central nervous system (CNS) and VAT.

Figure 3

Plasticity in autoimmune diseases. Conventional Tregs have the ability to convert into pro-inflammatory Th-like Tregs and Th17 under certain factors (shown in red). Restraining factors are shown in green. Th1-like Treg phenotype is observed in T1D, SLE, MS, experimental autoimmune encephalomyelitis (EAE) and psoriasis, Th2-like in systemic sclerosis (SSc) and collagen-induced arthritis (CIA), Th17-like in pSS, SSc, EAE, MS, RA, T1D and psoriasis.

Figure 4

Targeting Tregs in cancer. Blockades of CD25, CTLA-4, TIGIT and LAG-3 with mAbs, siRNA conjugates (anti-CTLA-4), ADCs and immunotoxins (anti-CD25) or cell-penetrating peptides (anti-FoxP3) lead to Treg depletion. Blockades of CCR8 and CCR4 with mAbs and immunotoxins result in inhibition of Treg recruitment to the TME and enhancement of anti-tumor responses. Inhibition of GITR with mAb promotes the differentiation of Tregs into Teffs and alleviates Treg-mediated suppression of the anti-tumor immune response. The effects of the PD-1 blockade in cancer are controversial.

Figure 5

Targeting Tregs in autoimmunity and transplantation. Low-dose IL-2 stimulation of expanded Tregs results in upregulation of CD25, GITR, CTLA-4, Ki67, Helios, CD39, CD45R0 and CCR7, making Tregs more suppressive. IL-33 treatment in transplantation results in a tolerogenic phenotype of Tregs with increased expression of CTLA-4, PD-1, CD39 and CD73. CD39 and CD73 generate adenosine, which binds to the adenosine receptor A2 on the surface of Tregs, increasing the intracellular concentration of cAMP in them and leading to impaired proliferation. Inhibition of Notch-1 with anti-Notch-1 mAbs is associated with increased expression of LAG-3, CTLA-4 and Ki67 and increased TGF-β production.