Bergenin, a PPARγ agonist, inhibits Th17 differentiation and subsequent neutrophilic asthma by preventing GLS1-dependent glutaminolysis.

Abstract

Bergenin is a natural PPARγ agonist that can prevent neutrophil aggregation, and often be used in clinics for treating respiratory diseases. Recent data show that Th17 cells are important for neutrophil aggregation and asthma through secreting IL-17A. In this study, we investigated the effects of bergenin on Th17 differentiation in vitro and subsequent neutrophilic asthma in mice. Naïve T cells isolated from mouse mesenteric lymph nodes were treated with IL-23, TGF-β, and IL-6 to induce Th17 differentiation. We showed that in naïve T cells under Th17-polarizing condition, the addition of bergenin (3, 10, 30 μM) concentration-dependently decreased the percentage of CD4+ IL-17A+ T cells and mRNA expression of specific transcription factor RORγt, and function-related factors IL-17A/F, IL-21, and IL-22, but did not affect the cell vitality and apoptosis. Furthermore, bergenin treatment prevented GLS1-dependent glutaminolysis in the progress of Th17 differentiation, slightly affected the levels of SLC1A5, SLC38A1, GLUD1, GOT1, and GPT2. Glutamine deprivation, the addition of glutamate (1 mM), α-ketoglutarate (1 mM), or GLS1 plasmid all significantly attenuated the above-mentioned actions of bergenin. Besides, we demonstrated that bergenin (3, 10, and 30 μM) concentration-dependently activated PPARγ in naïve T cells, whereas PPARγ antagonist GW9662 and siPPARγ abolished bergenin-caused inhibition on glutaminolysis and Th17 differentiation. Furthermore, we revealed that bergenin inhibited glutaminolysis by regulating the level of CDK1, phosphorylation and degradation of Cdh1, and APC/C-Cdh1-mediated ubiquitin-proteasomal degradation of GLS1 after activating PPARγ. We demonstrated a correlation existing among bergenin-affected GLS1-dependent glutaminolysis, PPARγ, "CDK1-APC/C-Cdh1" signaling, and Th17 differentiation. Finally, the therapeutic effect and mechanisms for bergenin-inhibited Th17 responses and neutrophilic asthma were confirmed in a mouse model of neutrophilic asthma by administration of GW9662 or GLS1 overexpression plasmid in vivo. In conclusion, bergenin repressed Th17 differentiation and then alleviated neutrophilic asthma in mice by inhibiting GLS1-dependent glutaminolysis via regulating the "CDK1-APC/C-Cdh1" signaling after activating PPARγ.