Decoding heart failure through plasma proteomics: differential protein signatures across the ejection fraction spectrum with sex-specific insights

Abstract Introduction Heart failure with preserved ejection fraction (HFpEF) and reduced ejection fraction (HFrEF) involve distinct pathophysiological mechanisms.In HFpEF, left ventricular(LV) remodelling is linked to a systemic proinflammatory state driven by comorbidities, whereas HFrEF is primarily driven by cardiomyocyte loss.A major challenge in HFpEF is the lack of specific biomarkers for diagnosis,monitoring, and treatment.Sex differences exist in HF, yet the underlying mechanisms remain poorly understood. Aim To identify proteomic signatures of HF across the EF spectrum, with consideration of sex-specific differences. Methods Circulating plasma protein levels were analysed in 80 age-(70±8.5years)and sex-matched(70%men) HF patients(40 HFrEF,40 HFpEF) who underwent extensive clinical phenotyping.Proximity extension assay technology, where two antibodies bind to their target protein and are detected by next-generation sequencing, was used for protein quantification. After removing duplicate proteins and assays failing quality control, 2330 proteins were analysed per sample.Differential expression analysis compared protein levels between HF subgroups, followed by enrichment analysis to identify key protein functions.Sex-specific analyses were conducted within each HF subgroup.Proteins were considered differentially expressed at FDR-adjusted p-value0.05 and fold change≥1.5. Results HFpEF patients had higher BMI, better kidney function,and lower NT-proBNP than HFrEF patients, with similar comorbidities(Table). 195 proteins were differentially expressed between groups. Natriuretic peptides(NPPB,NPPC,NT-proBNP), neuropeptides(NPY,GH1,CGC),and MME(encoding neprilysin) were upregulated in HFrEF relative to HFpEF, reflecting pronounced neurohormonal activation(FigA). Conversely, cytoskeletal(RAB11FIP3, KIF22, CORO1A, MYO9B) and inflammatory proteins(PPP1CC, IKBK6, MAP3K, VAV3), were upregulated in HFpEF relative to HFrEF. Enrichment analysis identified cytoskeletal protein, GTPase, actin and kinase binding as key protein functions in HFpEF, while neuropeptides and hormone activity were distinct in HFrEF(FigB). Sex-specific differences were assessed via comparative heatmaps stratified by EF(FigC). In HFpEF, six proteins were uniquely overexpressed in men vs women, including MMP3 and NPPC, indicating extracellular matrix degradation(MMP3) and adverse LV remodelling(NPPC).In contrast, in women vs men with HFpEF, six proteins including APOC1 and GHRL were overexpressed, which are involved in fatty acid metabolism and LDL clearance. The HFrEF group showed differential expression limited to previously described sex-associated proteins. Conclusion Distinct proteomic signatures differentiate HFrEF and HFpEF, with sex-specific differences primarily observed in HFpEF.These findings highlight divergent molecular mechanisms underlying HF subtypes and the importance of sex-specific effects in HFpEF pathophysiology, which may influence HF progression and treatment response.Baseline characteristics table Figure A.B.C