The human heart beats more than two billion times in an average life span and each contraction is precisely controlled by an intricate interplay between electrical signals and mechanical forces. At the anatomical level, it is composed of four cavities, four valves, large arteries, and veins, which act in concert to achieve proper filling, ejection, contraction, and overall pump function. The heart’s own blood supply is ensured by two coronary arteries. The human heart is composed of four major cell types—cardiac fibroblasts (CFs), cardiomyocytes, smooth muscle cells (SMCs), and endothelial cells (ECs)1. Their proportion with respect to number and volume, however, remains controversial. CFs are mesenchymal cells, which produce the extracellular matrix (ECM) scaffold of the heart and are thought to constitute more than half of all heart cells2. Cardiomyocytes are estimated to provide about 30% of the total cell number but account for over 70% of the total cardiac mass because of their large volume. In contrast, SMCs, which support the vascular system, and ECs, which form the interior lining of the heart, blood vessels, and cardiac valves, are generally believed to be much less abundant. However, these estimates have been challenged and a recent report claims that ECs are the largest cellular population within the heart3.
In common with other muscle tissues, the heart is dominated by a small number of proteins involved in the contractile apparatus. It employs tissue-specific isoforms such as cardiac troponins, which are used in the diagnosis of myocardial infarction. From a physiological and pathophysiological perspective, it would be desirable to gain deeper insights into the molecular characteristics of the heart at the spatial and cellular levels. In particular, characterization of the healthy state of the human heart would be an important starting point to investigate heart disease, which—despite major progress remains the leading cause of death in developed countries and is rapidly increasing in developing ones4.
Relatively little is known about the protein composition of the different regions and cell types of the heart. Previous studies have focused on defining differences between specific regions of the heart, or single-diseased heart compartments5, or from nonhuman, or subcellular material6,7. Phosphoproteomic studies have also been applied for the analysis of mammalian hearts8,9. Moreover, other studies use transcriptomic approaches10–12, which is an imperfect proxy for protein levels and their dynamics. However, proteins are the driving forces of the cellular machinery and they are involved in the control of virtually all physiologic events. The high dynamic range of the muscle proteome presents a formidable challenge to the comprehensive analysis of the heart at the level of expressed proteins. This is because very abundant proteins make it difficult to detect low abundant regulatory proteins in the same sample. The majority of studies only identified a few thousand proteins, and there is a paucity of studies of the human, nondiseased heart, because of the difficulty in obtaining the relevant tissue.
A global protein expression “footprint” of the healthy heart can be used as a reference library to compare against footprints of malfunctioning hearts in the search for biomarkers, therapeutic targets, or disease signatures. Recent advances in MS-based proteomics technology now allow the identification of very deep proteomes13,14. Our group has already established proteomics maps of the mouse liver and brain15,16 and analyzed skeletal muscle in considerable depth and sensitivity17,18. Here, we set out to generate a spatial and cell-type-resolved proteomic map of the healthy human heart. To this end, we measured 16 regions of three human hearts, as well as primary cell types. We employed high-sensitivity sample preparation, peptide fractionation, and an advanced label-free LC–MS workflow to quantify a total of more than 11,000 proteins. Our results establish proteomic differences between heart regions, suggest functional differences, and pinpoint potential cell-type markers. To illustrate the usefulness of the heart proteomic map, we apply it to define molecular changes in patients suffering from atrial fibrillation (AFib).
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