Calycosin suppresses the activating effect of granulocyte-macrophage-colony-stimulating factor-producing T helper cells on macrophages in experimental atherosclerosis

TL;DR

Calycosin reduces the harmful effects of a specific type of T cell that worsens atherosclerosis by inhibiting its activation of macrophages.

Contribution

This study reveals a new mechanism by which calycosin protects against atherosclerosis by targeting ThGM cells.

Findings

Calycosin reduces the frequency and activity of ThGM cells in a mouse model of atherosclerosis.

Calycosin-treated ThGM cells show reduced macrophage activation and foam cell formation.

NR4A3 upregulation by calycosin is critical for inhibiting ThGM cell function.

Abstract

T cells are contributors to atherosclerosis pathogenesis. Granulocyte-macrophage-colony-stimulating factor (GM-CSF)-producing T helper (ThGM) cells, a specialized helper T cell subset that highly expresses GM-CSF but lacks other helper T cell markers, could exacerbate atherosclerosis development. Calycosin has been reported to suppress atherosclerosis progression. However, the effect of calycosin on ThGM cells is unknown. This study was designed to test the calycosin-induced impact on the pro-atherosclerotic function of ThGM cells in a mouse atherosclerosis model.

Apolipoprotein E knockout (ApoE−/−) mice were fed a high-fat diet and calycosin. The phenotype and cytokine expression of aortic ThGM cells were assessed by flow cytometry. Calycosin-derived influences on ThGM cell differentiation, proliferation, and function were determined by flow cytometry, quantitative RT-PCR, Immunoblotting, gene silencing assays, and co-culture with macrophages.

Aortic ThGM cell frequency was attenuated after calycosin administration. Live aortic ThGM cells, phenotypically featuring CD4+CCR6−CCR8−CXCR3−CCR10+, showed slower proliferation and weaker macrophage-activating capability in calycosin-treated mice. Besides, calycosin repressed in vitro ThGM cell differentiation and subsequently impaired ThGM cell-mediated macrophage activation, oxidized low-density lipoprotein (Ox-LDL) uptake, and foam cell formation. Importantly, calycosin upregulated nuclear receptor subfamily 4 group A member 3 (NR4A3) in ThGM cells. NR4A3 silencing partially restored the function of calycosin-treated ThGM cells.

Calycosin inhibits ThGM cell activity to suppress ThGM-cell-mediated activation of pro-atherosclerotic macrophages to ultimately ameliorate atherosclerosis progression. Therefore, we revealed a novel mechanism by which calycosin protects against atherosclerosis.

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