Huo, Yu

Introduction: Endothelial cell (EC) dysfunction, driven by comorbidities of heart failure with preserved ejection fraction (HFpEF), has emerged as a potential contributor to disease progression, yet the EC specific molecular mediators in HFpEF remain poorly defined. Purpose: To identify EC-specific molecular mediators that are involved in HFpEF and determine their functional roles in disease pathogenesis. Hypothesis: Based on omics analysis of a murine HFpEF model, we hypothesized that EC plasminogen activator inhibitor-1 (PAI-1) is a key mediator of HFpEF pathogenesis. Methods: Male RiboTagEC (Cdh5CreERT2+/Rpl22HA/+) mice were used in a HFpEF model induced by hypertensive/metabolic stresses (60% calories from lard and L-NAME 0.5 g/L in water) or administrated with normal water and diet as controls. Hearts and skeletal muscle were collected for both bulk RNA sequencing (Bulk-seq) and EC-specific translatome analysis using translating ribosome affinity purification assay followed by RNA-sequencing (TRAP-seq). The functional roles of EC PAI-1 were tested using male tamoxifen-inducible, endothelial specific PAI-1 knockout (Cdh5CreERT2+/PAI-1flox/flox, PAI-1iEC KO) and control mice (PAI-1flox/flox, PAI-1iEC+/+) subjected to HFpEF induction. Cardiac function was assessed by echocardiography, pressure–volume (PV) loop analysis, and treadmill exercise testing. Student’s t-test was used (p < 0.05). Results: Bulk-seq and EC-specific TRAP-seq identified PAI-1 as one of the most upregulated genes in both heart and skeletal muscle EC in HFpEF. Bulk-seq demonstrated a 2.5-fold change (FC, log 2 FC=1.35) increases in the heart and 12.2-fold increases (log 2 FC = 3.61) in the skeletal muscle. TRAP-seq showed a 1.75-fold change (log 2 FC=0.81) increases in the heart and 4.76-fold increases (log 2 FC = 2.25) in the skeletal muscle. These consistent findings led us to investigate the functional role of EC PAI-1 in the pathogenesis of HFpEF. With similar body weight (Control: 40±4 g, n=10; KO: 38±6 g, n=11; p=0.319) and ejection fraction (Control: 54±6 %, n=10; KO: 54±5 %, n=12; p=0.686), we found that PAI-1iEC KO mice exhibited improved diastolic function after 10 weeks of HFpEF induction. Specifically, echocardiography demonstrated a significant reduction in E/e′ in KO mice (Control: 27±4, n=10; KO: 24±3, n=12; p=0.018). PV analysis confirmed this improvement with lower end-diastolic pressure (Control: 19±2 mmHg, n=8; KO: 13±5 mmHg, n=10; p=0.009), reduced Tau (Control: 9.7±1.2 ms, n=8; KO: 8.6±1.0 ms, n=10; p=0.047), and a reduced EDPVR constant β (Control: 0.20±0.05, n=6; KO: 0.10±0.03, n=6; p=0.017). KO mice also exhibited improved treadmill running distance (Control: 179±35 m, n=10; KO: 214±40 m, n=12; p=0.043). Conclusion: Endothelial PAI-1 contributes to diastolic dysfunction and reduced exercise capacity in HFpEF. These findings highlight endothelial PAI-1 as a potential therapeutic target for HFpEF. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.