Regulatory T-cell and neutrophil extracellular trap interaction contributes to carcinogenesis in non-alcoholic steatohepatitis

Background & Aims: Regulatory T-cells (Tregs) impair cancer immunosurveillance by creating an immunosuppressive envi-ronment that fosters tumor cell survival. Our previous findings demonstrated that neutrophil extracellular traps (NETs), which are involved both in innate and adaptive immunity, are abundant in livers affected by non-alcoholic steatohepatitis (NASH). However, how NETs interact with Tregs in the development of NASH-associated hepatocellular carcinoma (NASH-HCC) is not known. Methods: A choline-deficient, high-fat diet+diethylnitrosamine mouse model and the stelic animal model were utilized for NASH-HCC and a western diet mouse model was used for NASH development. Treg depletion was achieved using FoxP3-DTR mice. RNA sequencing was used to explore the mechanism by which NETs could regulate Treg differentiation. Bioenergetic analyses of naive CD4+ T-cells were assessed by Seahorse. Results: Although the absolute number of CD4+ T-cells is lower in NASH livers, the Treg subpopulation is selectively increased. Depleting Tregs dramatically inhibits HCC initiation and pro-gression in NASH. There is a positive correlation between increased NET and hepatic Treg levels. RNA sequencing data re-veals that NETs impact gene expression profiles in naive CD4+ T -cells, with the most differentially expressed genes being those involved in mitochondrial oxidative phosphorylation. By facili-tating mitochondrial respiration, NETs can promote Treg differ-entiation. Metabolic reprogramming of naive CD4+ T-cells by NETs requires toll-like receptor 4. Blockade of NETs in vivo using Pad4-/-mice or DNase I treatment reduces the activity of Tregs. Conclusions: Tregs can suppress immunosurveillance in the premalignant stages of NASH. NETs facilitate the crosstalk be-tween innate and adaptive immunity in NASH by promoting Treg activity through metabolic reprogramming. Therapies targeting NETs and Treg interactions could offer a potential strategy for preventing HCC in patients with NASH. Lay summary: Regulatory T-cells (Tregs) can promote tumor development by suppressing cancer immunosurveillance, but their role in carcinogenesis during non-alcoholic steatohepatitis (NASH) progression is unknown. Herein, we discovered that selectively increased intrahepatic Tregs can promote an immu-nosuppressive environment in NASH livers. Neutrophil extra -cellular traps (NETs) link innate and adaptive immunity by promoting Treg differentiation via metabolic reprogramming of naive CD4+ T-cells. This mechanism could be targeted to prevent liver cancer in patients with NASH. (c) 2021 Published by Elsevier B.V. on behalf of European Association for the Study of the Liver.

Automated summary

The study shows an increased number of Tregs in NASH-HCC, and depleting Tregs can inhibit the initiation and progression of HCC. There is a positive correlation between increased NETs and hepatic Tregs levels, and NETs promote Treg activity through metabolic reprogramming, linking innate and adaptive immunity. Therapies targeting NETs and Treg interactions may offer a potential strategy for preventing HCC in NASH patients.