DNER Promotes NOTCH1-Driven Preferential Pathogenic Th17.1 Cell Polarization and Metabolically Unhealthy Obesity

Notch signaling emerges as a driver of adipose tissue (AT) inflammation. Having previously shown that obese AT derived mesenchymal stem cells (Ob-ASC) promote Th17 and pathogenic Th17.1 cell polarization upon interaction with immune cells, here we identify DNER, predominantly expressed by Ob-ASC in human AT, as the preponderant NOTCH ligand upregulated upon Ob-ASC/immune cell interaction., and demonstrate its causal role in NOTCH1-driven pathogenic Th17.1 cell activation. Indeed, NOTCH1 pharmacological inhibition reversed the Ob-ASC-mediated skewing toward IL-17A/IFNγ double-secreting Th17.1 cells and blocked intranuclear translocation of its intra-cellular domain (NICD1) in T cells. Transcriptomic profiling of Ob-ASC/immune cell co-cultures confirmed DNER as the top-upregulated NOTCH ligand. Mechanistically, DNER silencing in Ob-ASC reduced IL-17A secretion, while gain-of-function experiments with recombinant Fc-DNER demonstrated a preferential skewing toward IL-17A and IFNg secretion, dependent on NOTCH1, whose activation was validated by NICD1 intra-nuclear translocation in the presence of Fc-DNER. STED microscopy further demonstrated DNER-NOTCH1 nanoscale proximity at the Ob-ASC/T cell interface. In vivo, Dner-deficient mice fed a hypercaloric diet were protected from weight gain, fat mass accretion, dyslipidemia, liver steatosis, and exhibited reduced AT macrophage and CD8+ T cell infiltration, with subcutaneous AT-specific downregulation of Il17a, Leptin, Notch1, its canonical ligands and targets. Ex-vivo Ob-ASC/splenocyte co-cultures confirmed that Ob-ASC drive preferential DNER-dependent Th17.1 activation via NICD1 translocation. Together, these findings establish DNER as a novel Ob-ASC expressed NOTCH1 ligand that orchestrates preferential pathogenic Th17.1 cell polarization and metabolic dysfunction in obesity, representing a promising therapeutic target.