S100A9 inhibition ameliorates HFpEF by modulating mitochondrial fission and oxidative stress

Heart failure with preserved ejection fraction (HFpEF) is characterized by diastolic dysfunction and myocardial stiffness, with limited treatment options due to the unclear molecular mechanisms underlying the disease. In this study, we investigate the role of S100A9, an inflammatory mediator, in regulating mitochondrial dynamics in HFpEF. Using "two-hit" (high-fat diet and L-NAME) and db/db mouse models, we show that S100A9 is significantly elevated in both cardiac tissue and serum, correlating with impaired diastolic function, cardiac hypertrophy, and increased oxidative stress. Inhibition of S100A9 with Paquinimod (PAQ) improved diastolic function, reduced cardiac hypertrophy, and decreased S100A9-positive macrophage infiltration, while preventing M1 macrophage polarization. In vitro, S100A9 secreted by palmitic acid-stimulated RAW 264.7 macrophages promoted mitochondrial fission in AC16 cardiomyocytes by increasing p-Drp1 and Fis1 expression, similar to the effects observed with recombinant S100A9. Excessive mitochondrial fission, regulated by S100A9, is a key factor in HFpEF progression. Transcriptomic analysis revealed significant upregulation of pyruvate dehydrogenase kinase 4 (PDK4) in HFpEF mice. Mechanistically, S100A9 induced PDK4 expression via SPI1-mediated transcription, exacerbating oxidative stress and mitochondrial fragmentation. PAQ treatment or silencing PDK4/SPI1 in AC16 cells reversed these effects, restoring ATP levels and stabilizing mitochondrial membrane potential. Cardiomyocyte-specific PDK4 knockdown in vivo further ameliorated HFpEF progression without affecting systolic function. These findings highlight S100A9 inhibition as a promising therapeutic strategy for HFpEF by targeting mitochondrial dysfunction through the S100A9/SPI1/PDK4 axis.