Emig, R.; Robbe, Z. L.; Kley, C.; Smolgovsky, S.; Travers, J. G.; Blanton, R. M.; McKinsey, T. A.; Black, L. D.; Alcaide, P.

Background: Heart failure with preserved ejection fraction (HFpEF) is a major clinical challenge characterized by diastolic dysfunction. Left ventricular stiffening and inflammation are hallmarks of HFpEF, yet the contribution of extracellular matrix (ECM) stiffness and the immune-stromal mechanisms driving ECM stiffening in cardiometabolic HFpEF remain poorly understood. Methods: We used the murine '2-hit model' of cardiometabolic HFpEF, in which the combination of high fat diet and hypertension induced by L-NAME causes diastolic dysfunction. We evaluated diastolic function by echocardiography and ECM mechanics by uniaxial tensile testing of decellularized cardiac tissue. Functional in vivo studies included genetic depletion of T cells, interferon-{gamma} (IFN{gamma}) knockout mice, and pharmacological lysyl oxidase inhibition. We combined co-cultures of CD4 T cells and cardiac fibroblasts (CFB) with mechanical testing of cardiac ECM and molecular biology to elucidate cellular and molecular mechanisms. Results: Left ventricular ECM stiffness strongly correlated with impaired diastolic function in experimental cardiometabolic HFpEF. Cardiac CD4 T cell infiltration was required for ECM stiffening and upregulation of lysyl oxidase enzymes in CFB. CD4 T cell-derived IFN{gamma} was both necessary and sufficient to induce LOXL3 in CFB, which increased ECM stiffness in vitro. Mechanistically, IFN{gamma} signaling activated hypoxia-inducible factor-1 (HIF1) in CFB, driving LOXL3 expression and subsequent collagen crosslinking. Genetic or pharmacologic disruption of this IFN{gamma}-HIF1-LOXL3 axis in vivo attenuated adverse ECM remodeling and improved diastolic function. Conclusions: CD4 T cells promote pathological ECM stiffening in cardiometabolic HFpEF through IFN{gamma}-mediated, LOXL3-dependent ECM crosslinking by CFB. Targeting this immune-stromal pathway may offer a novel therapeutic strategy for HFpEF.