Cardiac proteomic screening of heart failure patients with preserved and reduced ejection fraction reveal common alterations in inflammation pathways and protein phosphorylation

Franziska Koser (Münster)1, M. Abdellatif (Graz)2, T. Bock (Cologne)3, M. Krüger (Cologne)3, S. Sedej (Graz)2, W. A. Linke (Münster)1

1University Hospital Münster Institute of Physiology II Münster, Deutschland; 2Medical University of Graz Division of Cardiology Graz, Österreich; 3University of Cologne Institute for Genetics Cologne, Deutschland


Background: The prevalence of heart failure with preserved ejection fraction (HFpEF) is likely to rise with our increasing longevity and the burden of comorbidities. Despite the high public-health importance, evidence-based therapies of HFpEF are limited and the pathomechanisms remain elusive, although low-grade systemic inflammation and endothelial dysfunction may be involved.

We aimed to elucidate differences in protein expression and phosphorylation in human HFpEF and heart failure with reduced ejection fraction (HFrEF) as compared to nonfailing patient hearts.

Methods & Results:
Left ventricular myocardial tissues of explanted or donor human hearts (Medical University Graz) were retrospectively classified into nonfailing control (Ctrl), HFpEF or HFrEF (mean EF (%): 63 (Ctrl), 62 (HFpEF), 24 (HFrEF); mean age 58.1±9.5; mean BMI 26.6±2.7) and analysed by label-free quantitative mass spectrometry (N=6-7/group). Cardiac proteome analysis identified 49 significantly regulated proteins (fold-change >1.5, p<0.05) in HFpEF and 161 in HFrEF versus Ctrl hearts, out of 1813 cardiac proteins detected. Although fewer proteins were significantly altered in HFpEF than in HFrEF, 41% were uniquely regulated in HFpEF versus Ctrl (e.g., ANXA3, DAPK3). Gene ontology enrichment analysis of the cardiac proteome revealed that proteins associated with “complement activation” (e.g., C6, C4B) and “innate immune response” (e.g., S100A8, STING1) were significantly upregulated in HFpEF and HFrEF versus Ctrl hearts. Proteins uniquely altered in HFrEF were mainly associated with coagulation and immune response terms. Thus, low-grade systemic inflammation seems to be a disease factor in both human HFpEF and HFrEF hearts. Phosphoproteome analysis revealed 83 phosphosites in cardiac proteins that were differentially expressed (fold-change >1.5, p<0.05) in HFpEF and 181 in HFrEF versus Ctrl hearts, out of 1045 different phosphosites mapped in cardiac proteins. Phosphoproteomic alterations in HFpEF and HFrEF versus Ctrl hearts concerned mainly sarcomeric proteins (e.g. titin, desmin, troponin I), which were mostly hyper-phosphorylated. Moreover, gene ontology enrichment analysis revealed that proteins associated with “protein kinase binding“ (e.g., protein phosphatase 1 regulatory subunits 12A, 12B and 12C) were hyperphosphorylated in HFpEF versus Ctrl. Interestingly, immunoblot-based analyses confirmed, in HFpEF versus Ctrl hearts the presence of low-grade systemic inflammation and oxidative stress, with significantly increased expression of ICAM1 and a trend to decreased expression of eNOS. Moreover, preliminary immunostaining of patient heart tissue shows almost exclusively S100A8 positive staining in HFpEF and HFrEF, but not in Ctrl hearts.

Low-grade systemic inflammation is present in both HFpEF and HFrEF patient hearts. There is no specific HFpEF inflammatory phenotype, at least in this cohort that could be used for diagnostics or HFpEF-specific therapy development. However, our data suggest a link between systemic inflammation and sarcomeric protein phosphorylation, which we will follow up in future work.
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