Menopause exacerbates cardiovascular–kidney–metabolic–driven inflammatory and metabolic remodeling leading to diastolic dysfunction in a female swine model

Background Cardiovascular–kidney–metabolic (CKM) syndrome denotes the synergistic detrimental interplay between metabolic risk factors and chronic kidney disease, leading to heart failure with preserved ejection fraction (HFpEF). Particularly adipose tissue inflammation has recently been implicated as a driver of HFpEF. HFpEF disproportionately affects women, with incidence rising post-menopause, as hormonal loss promotes lipid dysregulation, adiposity and inflammation shifting to a more pro-inflammatory phenotype under hormonal and metabolic stress. Purpose Using integrated molecular and functional analyses to investigate how metabolic derangement (CKM) and menopause (CKMX) synergistically modify pericardial adipose tissue (PAT) and thereby contribute to the development of diastolic dysfunction. Methods Two-year-old female swine were assigned to healthy control (n=7), CKM (n=5), or CKMX (n=5). CKM and CKMX animals were exposed for 6 months to combined metabolic stress (western-style diet plus streptozotocin-induced hyperglycaemia) and chronic kidney disease (renal microembolization), while CKMX additionally underwent ovariectomy to induce menopause. LC/MS-based proteomics was performed on PAT and left ventricular myocardium. Functional assessment included pressure–volume loop analysis in all groups, whereas cardiomyocyte passive force, calcium sensitivity and high-resolution respirometry (OROBOROS) were measured in control and CKMX swine only. Results Proteomics showed differential expression of 541 (CKMX vs control) and 97 (CKM vs control) proteins in PAT, with enrichment of complement/coagulation pathways and reduced oxidative phosphorylation, indicating an inflamed and metabolically compromised PAT in CKMX. Cardiac proteomics revealed amplification of CKM-driven remodeling in CKMX, with increased cholesterol and lipoprotein metabolism and further suppression of branched-chain amino acid and NADP⁺ pathways. Functionally, CKM showed elevated end-diastolic pressure (EDP) only during afterload elevation, whereas CKMX showed elevated baseline EDP, with preserved systolic function (Fig1). Consistent with increased diastolic stiffness, CKMX isolated cardiomyocytes displayed greater passive stiffness and enhanced calcium sensitivity vs control (Fig2A). Mitochondrial dysfunction was observed in CKMX (Fig2B), with impaired complex-I driven respiration in CKMX swine, characterized by reduced OXPHOS capacity and maximal uncoupled respiration, while complex-II and leak respiration were preserved. Conclusion Menopause aggravates CKM-induced remodeling of PAT and myocardium. Proteomic and functional analyses imply a role for inflammation, myocardial metabolic dysfunction, and mitochondrial impairment which is associated with enhanced calcium sensitivity. Altogether, these factors enhance myocardial stiffness, and overt diastolic dysfunction, thereby identifying menopause as a key amplifier of CKM-associated HFpEF.