Expression of inhibitory regulators of innate immunity in patients with active tuberculosis

Abstract

Background: Tuberculosis (TB) is an important cause of morbidity and mortality worldwide. Toll-like-receptors (TLRs) are important for the recognition of the causative agent Mycobacterium tuberculosis. Negative regulation of TLRs is necessary to control deleterious inflammatory damage, but could provide a means of immune evasion by M. tuberculosis as well.

Methods: To obtain insight in the extent of expression of inhibitory regulators of immunity in patients with active TB, peripheral-blood-mononuclear-cells (PBMCs) and plasma were obtained from 54 TB patients and 29 healthy blood donors from Chittagong, Bangladesh. Bilateral alveolar macrophages were obtained from an infected versus a contralateral normal lung segment of 9 patients. Statistical analyses were performed using Mann-Whitney U and Wilcoxon matched pairs testing. Correlations were calculated using the Spearman rho test.

Results: PBMCs harvested from TB patients demonstrated increased mRNA expression of IL-1-receptor-associated-kinase-M, suppressor-of-cytokine-signalling-3 and Toll-interacting-protein. Flow cytometry revealed enhanced expression of IL-1-receptor-like-1 (ST2) on lymphocytes. Plasma soluble ST2 was elevated in patients with TB and correlated with established TB biomarkers, most strongly with soluble interleukin-2 receptor subunit α and interleukin-8. Alveolar macrophage mRNA expression of negative TLR regulators did not differ between the infected and contralateral lung side.

Conclusion: These results show enhanced expression of distinct negative regulators of innate immunity in PBMCs of patients with TB and identify plasma soluble ST2 as a potential novel biomarker for TB disease activity.

Figure 1

ST2 during active pulmonary tuberculosis. (A) ST2 mRNA in PBMCs; (B) ST2 surface expression measured on CD4 and CD8 positive lymphocytes and CD14 positive monocytes (representative histograms). FMO, fluorescence minus one. (C) Idem for individual subjects (geomean channel fluorescence intensity (MFI)). (D) Plasma sST2 concentrations. (E) ST2 mRNA in alveolar macrophages (AM) of TB patients originating from the TB infected lung (open squares) or contralateral lung (closed squares). Depicted mRNA levels are normalized to the house keeping gene β2-microglobulin. Depicted are dot plots with medians; open dots: TB patients, closed dots: healthy donors. ***P < 0.001.

Figure 2

SIGIRR during active pulmonary tuberculosis. (A) SIGIRR mRNA in PBMCs. (B) SIGIRR surface expression measured on CD4 and CD8 positive lymphocytes and CD14 positive monocytes (representative histograms). FMO, fluorescence minus one. (C) Idem for individual subjects (mean channel fluorescence intensity, MFI). (D) SIGIRR mRNA in alveolar macrophages (AM) of TB patients originating from the TB infected lung (open squares) or contralateral lung (closed squares). Depicted mRNAs are normalized to the house keeping gene β2-microglobulin. Depicted are dot plots with medians; open dots: TB patients, closed dots: healthy donors.

Figure 3

Intracellular negative regulators and mediators of TLR signaling. mRNA levels in PBMCs of TB patients (open dots) and healthy controls (closed dots) and TB patient AMs from the diseased (TB-positive) lung segment (open squares) and from the matching contralateral lung (closed squares). PBMC mRNA expression of IRAK-M (A), MKP-1 (C), SOCS-3 (E), TOLLIP (F) and A20 (I). AM mRNA expression of IRAK-M (B), MKP-1 (D), TOLLIP (G) and A20 (J). TOLLIP (H) expression levels in CD4 or CD8 positive lymphocytes and CD14 positive monocytes, as determined by flow cytometry (mean channel fluorescence intensity, MFI). mRNA expression is normalized to β2-microglobulin. Depicted are dot plots with medians; open dots: TB patients, closed dots: healthy donors. *P < 0.05, **P < 0.01.