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18 January 2017 : Laboratory Research  

Toll-Like Receptor 4 Signaling in High Mobility Group Box-1 Protein 1 Mediated the Suppression of Regulatory T-Cells

Chunyan Luo12ABCDF, Huiting Liu3DEF, Hu Wang2ADEF, Jiajun Wang12ABEFG

DOI: 10.12659/MSM.902081

Med Sci Monit 2017; 23:300-308

Abstract

BACKGROUND: Treg cells play a central role in the suppression of immune response, and their suppressive capacity can be modulated by toll-like receptor (TLR) ligands. However, the detailed pathway of TLR ligand modulation is still unknown. The present study aimed to evaluate the effect of the high mobility group box-1 protein 1 (HMGB1) and lipopolysaccharide (LPS) on Treg cells through TLR4 signaling.

MATERIAL AND METHODS: Treg cells were purified from healthy human peripheral blood mononuclear cells (PBMCs) by magnetic-bead activity cell sorting (MACS), blocked by anti-TLR4 monoclonal antibody, and then incubated with different concentration of LPS or HMGB1. The level of gene expression of IL-1β, IL-10, IFN-γ, and TGF-β were detected using quantitative real-time polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA), and the proliferation of Treg cells after treating by LPS and HMGB1 was analyzed by flow cytometry. The NF-κB expression in Treg cells was examined by Western blotting.

RESULTS: LPS treated CD4 CD25 Treg cells directly increased the expression of IL-1b and IL-10 and decreased the expression of IFN-γ and TGF-β. However, HMGB1 treatment resulted in a marked decreased expression of IL-1β, IL-10, IFN-γ, and TGF-β. The proliferation of CD4+ T cells was significantly inhibited by Treg cells in the LPS treatment group, but weaken in the HMGB1 treatment group. These data suggest that HMGB1 and LPS stimulation could downregulate the expression NF-κB p65 in cytoplasmic proteins and increase the expression in nuclear proteins, thus leading to modulation of IL-1β, IL-10, IFN-γ, and TGF-β expression; moreover, the suppressive function of Treg cells could be regulated by TLR4.

CONCLUSIONS: TLR4 signaling in HMGB1 mediated the suppressive function of Treg cells through the activation of the NF-κB pathway.

Keywords: High Mobility Group Proteins, T-Lymphocytes, Regulatory, Toll-Like Receptors

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750