Hepatic Arachidonic Acid Dysregulation Impairs Double-Negative Regulatory T Cell-Mediated Immunoregulation via CD39 Downregulation and Drives MASLD Development.

Immune dysregulation is a key driver of metabolic dysfunction-associated steatotic liver disease (MASLD). Double-negative regulatory T (DNT) cells, which are essential for hepatic immune homeostasis, are functionally impaired in MASLD. CD39, an ectoenzyme that converts ATP to adenosine, sustains DNT cell viability and immunoregulatory function. However, the mechanisms of MASLD-induced CD39 dysregulation remain unknown. In this study, a transcriptomic analysis of hepatic DNT cells from MASLD mice revealed reduced ATP hydrolysis, as confirmed by flow cytometry, and lower CD39 level compared to normal control diet-fed mice. Similarly, single-cell transcriptomic data showed that CD39 was downregulated in hepatic DNT cells from MASLD patients, and higher CD39 expression was associated with enhanced cytotoxicity and viability. Furthermore, in vitro experiments confirmed that CD39 maintained the immunosuppressive function of DNT cells by hydrolyzing ATP and suppressing monocyte activation. Mechanistically, arachidonic acid (AA), which is enriched in the livers of MASLD, suppressed CD39 via hypoxia-inducible factor-1α (HIF-1α). This AA-HIF-1α-CD39 axis impaired DNT cell-mediated monocyte immunosuppression, accelerating immune imbalance and MASLD progression. These findings establish the AA-HIF-1α-CD39 axis as a key pathway driving DNT cell dysfunction in MASLD, highlighting potential therapeutic strategies targeting metabolic-immune crosstalk to restore hepatic immune balance.