Th17/Treg-cell balance in the peripheral blood of pregnant females with a history of recurrent spontaneous abortion receiving progesterone or cyclosporine A

Abstract

A successful pregnancy requires the maternal immune system to accept a fetus expressing allogeneic paternal antigens and provide competent responses to infections. Accordingly, maternal-fetal immune abnormalities may have an important role in the development of recurrent spontaneous abortion (RSA). Ever since the establishment of the association between immunologic abnormalities and RSA, various types of immune therapy to restore normal immune homeostasis have been increasingly developed. Although previous studies have focused on the maternal-fetal interface, non-invasive examination is of great importance in clinical practice. The present study investigated the balance between type-17 T-helper (Th17) and T-regulatory (Treg) cells in the peripheral blood to improve the current understanding of the pathogenesis of RSA. Imbalances in Th17/Treg cells and associated molecular profiles were observed in patients with RSA. Furthermore, it was determined that the immunosuppressant cyclosporine A reduced the proportion of Th17 cells and promoted Treg-cell dominance by upregulating the expression of co-inhibitory molecules in pregnant females with a history of RSA. Progesterone, the traditional maternal-care drug, also had a certain immunomodulatory role through restoring the levels of several co-inhibitory molecules (including T-cell immunoglobulin mucin family member-3, programmed cell death-1 and cytotoxic T-lymphocyte associated protein-4) in the treatment of RSA. Changes in these immune molecules within the maternal peripheral blood may be indicators for monitoring pregnancy and prediction of RSA.

Keywords: Th17 cells; Treg cells; cyclosporine A; progesterone; recurrent spontaneous abortion.

Figure 1

Effects of P4 and CsA on the percentage of Treg and Th17 cells in peripheral blood. (A) Compared with that in the NP group, the subjects in the RSA group had a significantly lower percentage of Treg cells in the peripheral blood. CsA significantly increased the percentage of Treg cells. (B) Compared with that in the NP group, the subjects in the RSA group had a higher percentage of Th17 cells in the peripheral blood. CsA decreased the percentage of Th17 cells. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^**P<0.01, ^***P<0.001 vs. NP; ^#P<0.05 vs. RSA. CsA, cyclosporine A; Treg cells, T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; Fox, forkhead box; IL, interleukin; P4, progesterone.

Figure 2

CsA promotes TGF-β1 expression in peripheral blood cells of pregnant females with RSA. Compared with that in the NP group, the production of TGF-β1in (A) CD4⁺ T cells, (B) Treg cells and (C) Th17 cells in the peripheral blood of subjects in the RSA group was decreased. While CsA and P4 promoted the expression of TGF-β1 in Th17 cells, P4 had no effect on TGF-β1 production by CD4⁺ T cells and Treg cells. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^*P<0.05, ^**P<0.01, ^***P<0.001 vs. NP; ^##P<0.01, ^###P<0.001 vs. RSA. CsA, cyclosporine A; pTreg cells, peripheral T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; P4, progesterone; Fox, forkhead box; IL, interleukin; TGF, transforming growth factor; FSC, forward scatter.

Figure 3

CsA promotes IL-10 expression in peripheral blood cells of pregnant females with RSA. Compared with that in the NP group, the production of IL-10 in (A) CD4⁺ T cells, (B) Treg cells and (C) Th17 cells in the peripheral blood of subjects in the RSA group was decreased. While CsA promoted the production of IL-10, P4 had no effect on it. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^**P<0.01, ^***P<0.001 vs. NP; ^#P<0.05, ^###P<0.001 vs. RSA. CsA, cyclosporin A; pTreg cells, peripheral T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; P4, progesterone; IL, interleukin.

Figure 4

Effects of P4 and CsA on CTLA-4 expression in peripheral blood cells of pregnant females with a history of RSA. Compared with that in the NP group, the production of CTLA-4 in (A) CD4⁺ T cells, (B) Treg cells and (C) Th17 cells in the peripheral blood of subjects in the RSA group was decreased. CsA promoted the expression of CTLA-4 in all three indicated subsets of T cells. P4 also promoted CTLA-4 expression in total CD4⁺T and Th17 cells. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^*P<0.05, ^**P<0.01, ^***P<0.001 vs. NP; ^#P<0.05, ^##P<0.01, ^###P<0.001 vs. RSA. CsA, cyclosporine A; pTreg cells, peripheral T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; P4, progesterone; CTLA, cytotoxic T-lymphocyte-associated protein 4.

Figure 5

Effects of P4 and CsA on Tim-3 expression in peripheral blood cells of pregnant females with a history of RSA. Compared with that in the NP group, the production of Tim-3 in (A) CD4⁺ T cells and (B) Treg cells in the peripheral blood of subjects in the RSA group was decreased. (C) In Th17 cells in the peripheral blood, no significant difference in Tim-3 expression between the RSA and NP group was determined. P4 and CsA promoted Tim-3 expression in all three indicated subsets of T cells. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^***P<0.001 vs. NP; ^##P<0.01, ^###P<0.001 vs. RSA. CsA, cyclosporine A; pTreg cells, peripheral T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; P4, progesterone; TIM-3, T-cell immunoglobulin mucin family member-3.

Figure 6

Effects of P4 and CsA on PD-1 expression in peripheral blood cells of pregnant females with a history of RSA. Compared with that in the NP group, the production of PD-1 in (A) CD4⁺ T cells and (B) Treg cells in the peripheral blood of subjects in the RSA group was decreased. (C) In Th17 cells in the peripheral blood, no significant difference in PD-1 expression between the RSA and NP group was determined. While P4 and CsA had no effects on PD-1 expression in total CD4⁺ T cells and Th17 cells, P4 promoted PD-1 expression in Treg cells. Groups: NP, normal early pregnancy; RSA; patients with RSA; P4, pregnant females with a history of RSA treated with P4; CsA, pregnant females with a history of RSA treated with CsA. ^*P<0.05,^**P<0.01, ^***P<0.001 vs. NP; ^##P<0.01 vs. RSA. CsA, cyclosporine A; pTreg cells, peripheral T-regulatory cells; Th17 cells, type-17 T-helper cells; RSA, recurrent spontaneous abortion; P4, progesterone; PD-1, programmed cell death-1.