The impact of selenium on regulatory T cell frequency and immune checkpoint receptor expression in patients with diffuse large B cell lymphoma (DLBCL)

Abstract

For many decades, selenium (Se) has been known as a potential anti-cancer agent that can also improve the function of immune cells in a variety of solid tumors. However, there is no report on the role of Se on CD4⁺ T cell subsets like CD4⁺CD25⁺FOXP3⁺ regulatory T cells (Tregs) in lymphoma patients. In this randomized clinical trial, we investigated the effect of 3-month Se consumption on the frequency of CD4⁺CD25⁺FOXP3⁺ Tregs and the expression of immune checkpoint receptors in thirty-two non-Hodgkin lymphoma (NHL) patients (16 patients with Se (Se+) and 16 without Se (Se-) consumption) with diffuse large B-cell lymphoma (DLBCL) subtype at stable remission. The change in the frequency of Tregs and expression of immune checkpoint receptors including CTLA-4, LAG-3, TIM-3, and PD-L1 genes were evaluated after 3 months in both groups using flow cytometry and SYBR Green Real-time PCR method, respectively. The results showed that the frequency of CD4⁺CD25⁺FOXP3⁺ Tregs and expression of immune checkpoint receptors did not significantly change after 3-month Se consumption in DLBCL patients. However, alteration in the frequency of CD4⁺CD25^-FOXP3⁺ Treg subsets was positively correlated with change in CTLA-4, LAG-3, and TIM-3 expression in the Se+ group. Three-month Se supplementation did not prevent relapse in Se+ group. Taken together, Se supplementation alone did not affect the frequency of CD4⁺CD25⁺FOXP3⁺ Tregs, expression of checkpoint receptors, and prevention of relapse in DLBCL patients at stable remission phase but might influence the functional properties of other Treg subsets like CD4⁺CD25^-FOXP3⁺ Tregs.

Keywords: Diffuse large B-cell lymphoma (DLBCL); Immune checkpoint receptors; Regulatory T cells (Tregs); Selenium (Se).

Fig. 1

Flow cytometry analysis of Tregs. For Treg quantification, the lymphocytes (R1) were defined based on the forward vs. side scatter data acquisition plotting. Then, the CD4⁺ cells (R2) were first selected in the lymphocyte population (R1) followed by the gating of CD25⁺FOXP3⁺ cells (Q2 quadrant) within CD4⁺ lymphocytes

Fig. 2

Change in the frequency of Tregs and related subsets after 3 months in Se+ and Se− groups. Change in the frequency of Tregs and related subsets was calculated after 3 months in both Se+ and Se− groups. Data are presented as mean ± SD and analyzed by Graphpad Prism version 8. P value < 0.05 was considered as statistically significant. Se selenium, Tregs regulatory T cells

Fig. 3

Change in the expression of immune checkpoint receptors after 3 months in Se+ and Se− groups. Change in the mean expression level of immune checkpoint receptors (dCt) was defined after 3 months in both Se+ and Se− groups. Data are presented as mean ± SD and analyzed by Graphpad Prism version 8. P value < 0.05 was considered as statistically significant. Se selenium

Fig. 4

The relationship between change in the frequency of CD4⁺CD25⁻FOXP3⁺ Tregs and expression of immune checkpoint receptors in Se+ (a) and Se− (b) groups. Data are presented as mean ± SD and analyzed by Graphpad Prism version 8. P value < 0.05 was considered as statistically significant. Se selenium, r_s Spearman correlation coefficient