Increasing evidence indicates that Tfh cells are important in the pathogenesis of SLE. Tfh cells are recognized as a distinct T cell subset, which provides help for GC formation, B cell affinity maturation, and immunoglobulin class switching, as an indispensable part of adaptive immunity. Our previous work showed that patients with SLE have an increased number of peripheral Tfh cells, which positively correlates with autoantibody titers (anti-dsDNA antibodies) and disease activity, as measured by the SLEDAI. Others have reported that the aberrant expression of Tfh cells is a common feature in mouse models of SLE, suggesting its contribution in the development of autoimmune diseases [4, 5].
Besides Tfh cells, a subset of Treg cells, named Tfr cells, have been identified. These cells share common characteristics with Tfh and conventional Treg cells and can inhibit GC responses, regulating the number of Tfh and GC B cells [6,7,8]. Therefore, it is generally believed that Tfr cells constrain the B cell “help” provided by Tfh cells to maintain immune homeostasis. An aberrant or disordered Tfh/Tfr balance may result in the break of tolerance, excessive B cell proliferation, antibody production, and the development of autoimmune diseases. A recent study showed the importance of the Tfr/Tfh balance in autoimmune responses in BXD2 mice, which display spontaneous autoreactive GC formation [21]. In addition, intravenous immunoglobulin administration to mice with collagen-induced arthritis augments the number of Tfr cells and represses the subsequent maturation of GC B cells [22], which also supports the idea of a critical role for Tfr cells in autoimmune diseases.
There was no difference in absolute number of Tfh17 subsets between SLE and healthy controls. The proportion of Tfh17 in CD4 T cells was significantly higher in SLE than in HC, although there was no significant difference in Tfh17 proportion in lymphocytes. Therefore, in this study, the focus is on the Tfr/Tfh balance in the SLE.
In B cell co-culture, the Tfh2 and Tfh17 cells exceeded the Tfh1 cells to support antibody production [23]. A general trend of increasing proportions of Tfh2 and Tfh17 cells and decreasing proportions of Tfh1 cells was observed in SLE [24]. The numbers of Tfh1 and Tfh2 cells are relatively lower in SLE than in HC in our study. It is possible that Tfh2 cells do not play an important pathogenic role in this cohort of patients.
There was no significant difference between CXCR5+PD-1low Treg and CXCR5+PD-1high Treg in our study. In a study, it was found that the majority of Tfr cells in the lymph nodes express low levels of PD-1 and reside at the border between the T cell zone and B cell follicle, with very few found in the germinal centers (GCs) [9]. Although PD-1+ Tfr cells expressed higher levels of CD38, CTLA-4, and GARP than PD-1- Tfr cells, both potently suppressed antibody production in vitro. These results demonstrate the phenotypic diversity of human Tfr cells [9].
In our study, we found a deficiency of Tfr cell subsets, including CXCR5+PD-1lowTreg and CXCR5+PD-1high Treg, and increased Tfh cells in the peripheral blood of SLE patients. The shifted balance between circulating CXCR5+PD-1low Treg and Tfh cells correlated not only with reduced serum IL-2, IL-10, and increased IL-21 levels in patients but also with clinical SLE parameters, e.g., ESR, anti-dsDNA antibodies, and disease activity (SLEDAI Scores). These findings are consistent with previous studies in vitro, in which Tfh and CXCR5+PD-1low Treg or CXCR5+PD1high Treg cells can antagonize B cell function, production of high-affinity antibodies, and the memory B cell differentiation [22]. CXCR5+PD-1low Treg cells play an important immunosuppressive function by curbing self-reactive auto-antibodies development within the GC during an inflammatory immune response [5]. Therefore, deregulation of the Tfr and Tfh cell compartments is associated with disease severity, B cell frequency, and antibody production in SLE.
There have been several relatively successful attempts to reduce the severity of SLE in humans via blockade of Tfh-cell differentiation and activity. Studies using monoclonal antibodies against ICOSL inhibited the development of Tfh and GC B cells resulting in decreased anti-dsDNA antibodies and improved kidney function in both human and mouse [23]. For years, SLE therapy has relied on broad spectrum immunosuppressants; however, a growing body of work shows that a targeted increase of regulatory T cells may be a more attractive therapy [18, 19, 24, 25].
IL-2 is essential for the development and maintenance of Treg cells, which prevent the development of autoimmune disease. Low-dose IL-2 can promote Tregs by activating the transcription factor STAT5, which binds to the Foxp3 locus and promotes Foxp3 expression without activation of effector T cells. More recently, IL-2 has been shown to be essential for the inhibition of Tfh cell development. Thus, in this study, we asked if low-dose IL-2 therapy might also elevate the Tfr/Tfh ratio, exploring a novel concept for rational therapeutic design.
Our previous studies had proven a deficient Treg cell compartment and decreased IL-2 levels in circulation of SLE, and the efficacy of low-dose IL-2 treatment. But there was no study addressing the impact of low-dose IL-2 on Tfr:Tfh balance. After effective therapy, especially low-dose IL-2 therapy, the imbalanced Tfr and Tfh subsets were reversed accompanying improvement of disease activity. Furthermore, Tfr subsets were all increased regardless of output measurement; proportion and absolute number. Although we did not see a significant change in Th17 frequency, the ratios of CXCR5+PD-1low Treg/Tfh17 and CXCR5+PD-1high Treg/Tfh17 were significantly decreased compared to those in healthy controls. Besides, we did not see any obvious difference between CXCR5+PD-1low Treg and CXCR5+PD-1high Treg, perhaps reflecting a functional overlap of these two subsets.
The strength of our study was to systematically analyze the change in Tfr and Tfh subsets in SLE and its correlation with relevant clinical parameters. The main limitation of this article was a relatively small number of patients with a very heterogeneous disease, which may give rise to a caution about over-interpretation of the data. Another limitation is that we definite Treg cell as CD4+CD25highCD127low, but not CD4+CD25highCD127low Foxp3+, both stain methods should be applied in the future studies.