Targeting of myeloid-derived suppressor cells by all-trans retinoic acid as host-directed therapy for human tuberculosis

Abstract

Conventional anti-tuberculosis (TB) therapies comprise lengthy antibiotic treatment regimens, exacerbated by multi-drug resistant and extensively drug resistant mycobacterial strains. We assessed the ability of all-trans retinoic acid (ATRA), as repurposed compound serving as host-directed therapy (HDT), to counteract the suppressive effects of myeloid-derived suppressor cells (MDSCs) obtained from active TB cases (untreated or during week one of treatment) on T-cell responsiveness. We show for the first time that MDSCs suppress non-specific T-cell activation and production of interleukin (IL)-2, IL-4, IL-13 and GM-CSF via contact-dependent mechanisms. ATRA treatment decreases MDSC frequency, but fails to mature MDSCs to non-suppressive, terminally differentiated myeloid cells and does not restore T-cell function or cytokine production in the presence of MDSCs. The impact of ATRA treatment on improved immunity, using the concentration tested here, is likely to be minimal, but further identification and development of MDSC-targeting TB host-directed therapies are warranted.

Keywords: All-trans retinoic acid; Host-directed therapies; Mycobacterium tuberculosis; Myeloid-derived suppressor cells; Tuberculosis.

Fig. 1.

Effect of MDSC on non-specifically activated (anti-CD3CD28 beads) Total T cell and T cell subset responses. Frequency of Total T cells (CD3+) and T cell subsets from in vitro T cell co-culture as well as trans-well co-culture of allogeneic T cells (CD3+ cells) isolated from PBMC of a Quantiferon positive individual (8 replicates), and MDSC (HLA-CD33+ cells) isolated from PBMC of individuals with active TB disease (n = 8). Plots show QFT+ individual T cell replicates. Panels a-g show T cell frequency of various subsets and panel h shows cell proliferation as measure of T cell response. Kruskal-Wallis test with Dunns post-hoc test. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 2.

Effect of MDSC on Unstimulated Total T cell and T cell subset responses. Frequency of Total T cells (CD3+) and T cell subsets from in vitro T cell co-culture as well as trans-well co-culture of allogeneic T cells (CD3+ cells) isolated from PBMC of a Quantiferon positive individual (9 replicates), and MDSC (HLA-CD33+ cells) isolated from PBMC of individuals with active TB disease (n = 9). Plots show QFT+ individual T cell replicates. Panels a-g show T cell frequency of various subsets and panel h shows cell proliferation as measure of T cell response. Kruskal-Wallis test with Dunns post-hoc test. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 3.

Effect of MDSC on Total T cell cytokine production. Analyte concentration (pg/ml) was measured for various culture combinations (MDSC only, T cell only, MDSC-T cell coculture), under differing stimulation combinations (Unstimulated and anti-CD3CD28 beads) by multiplex analysis (n = 18). Plots show analyte concentration in coculture supernatant. Mixed model analysis, LS means and Type II decomposition analysis followed. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 4.

Effect of ATRA on the frequency of (a) Total MDSC, (b) M-MDSC, (c) PMN-MDSC, (d) E-MDSC from in vitro cultures of MDSC (HLA-CD33+ cells) isolated from PBMC of individuals with active TB disease (n = 18). Plots show individual TB patient data. Wilcoxon matched pairs signed rank test. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 5.

Effect of ATRA on MDSC maturation to non-suppressive mature myeloid cell subsets. Median Fluorescence Intensity of HLA-DR on isolated MDSC (HLA-CD33+) from ATRA-treated in vitro culture (n = 16 a&b and n = 18c&d). Plots show individual TB patient data. Wilcoxon matched pairs signed rank test. Median with interquartile range. P < 0.05 significant.

Fig. 6.

Effect of ATRA on restoration of non-specifically activated (anti-CD3CD28 beads) T cell responses in the presence of MDSC. ATRA treated in vitro co-culture of allogeneic T cells (CD3+ cells) isolated from PBMC of a Quantiferon positive individual (6 replicates), and MDSC (HLA-CD33+ cells) isolated from PBMC of individuals with active TB disease (n = 6). Plots show QFT+ individual T cell replicates. Panels a-g show T cell frequency of various subsets and panel h shows cell proliferation as measure of T cell response. Wilcoxon matched pairs signed rank test. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 7.

Effect of ATRA on restoration of Unstimulated T cell responses in the presence of MDSC. ATRA treated in vitro co-culture of allogeneic T cells (CD3+ cells) isolated from PBMC of a Quantiferon positive individual (8 replicates), and MDSC (HLA-CD33+ cells) isolated from PBMC of individuals with active TB disease (n = 8). Plots show QFT+ individual T cell replicates. Panels a-g show T cell frequency of various subsets and panel h shows cell proliferation as measure of T cell response. Wilcoxon matched pairs signed rank test. Median with interquartile range indicated on plots. P < 0.05 was considered significant.

Fig. 8.

Effect of ATRA on restoration of T cell responses in the presence of MDSC. Analyte concentration (pg/ml) was measured for various culture combinations (MDSC only, T cell only, MDSC-T cell coculture), under differing stimulation combinations (Unstimulated and anti-CD3CD28 beads) with or without ATRA treatment, by multiplex analysis. Plots show analyte concentration in coculture supernatant. Mixed model analysis, LS means and Type II decomposition analysis. Median with interquartile range indicated on plots. P < 0.05 was considered significant.