Metabolic Remodeling in Moderate Synchronous versus Dyssynchronous Pacing-Induced Heart Failure: Integrated Metabolomics and Proteomics Study

<div><p>Heart failure (HF) is accompanied by complex alterations in myocardial energy metabolism. Up to 40% of HF patients have dyssynchronous ventricular contraction, which is an independent indicator of mortality. We hypothesized that electromechanical dyssynchrony significantly affects metabolic remodeling in the course of HF. We used a canine model of tachypacing-induced HF. Animals were paced at 200 bpm for 6 weeks either in the right atrium (synchronous HF, SHF) or in the right ventricle (dyssynchronous HF, DHF). We collected biopsies from left ventricular apex and performed comprehensive metabolic pathway analysis using multi-platform metabolomics (GC/MS; MS/MS; HPLC) and LC-MS/MS label-free proteomics. We found important differences in metabolic remodeling between SHF and DHF. As compared to Control, ATP, phosphocreatine (PCr), creatine, and PCr/ATP (prognostic indicator of mortality in HF patients) were all significantly reduced in DHF, but not SHF. In addition, the myocardial levels of carnitine (mitochondrial fatty acid carrier) and fatty acids (12:0, 14:0) were significantly reduced in DHF, but not SHF. Carnitine parmitoyltransferase I, a key regulatory enzyme of fatty acid ß-oxidation, was significantly upregulated in SHF but was not different in DHF, as compared to Control. Both SHF and DHF exhibited a reduction, but to a different degree, in creatine and the intermediates of glycolysis and the TCA cycle. In contrast to this, the enzymes of creatine kinase shuttle were upregulated, and the enzymes of glycolysis and the TCA cycle were predominantly upregulated or unchanged in both SHF and DHF. These data suggest a systemic mismatch between substrate supply and demand in pacing-induced HF. The energy deficit observed in DHF, but not in SHF, may be associated with a critical decrease in fatty acid delivery to the ß-oxidation pipeline, primarily due to a reduction in myocardial carnitine content.</p></div

Metabolomic and proteomic profile of glucose metabolism.

<p>Data from Control, SHF, and DHF hearts. Detected metabolites and enzymes are indicated with bold font. Metabolome is presented in arbitrary units while proteome is presented as fold change compared to Control. Filled bars: metabolites. Open bars: proteins. G1-P: glucose 1-phosphate, G6-P: glucose 6-phosphate, F6-P: fructose 6-phosphate, F1,6-P: fructose 1,6-bisphosphate, GAP: glycealdehydo 3-phosphate, DHAP: dihydroxyacetone phosphate, 1,3-PG: 1,3-bisphosphoglycerate, 3-PG: 3-phosphoglycerate, 2-PG: 2-phosphoglycerate, PEP: phosphoenolpyruvate. P*<0.05.</p

Fatty acid catabolism.

<p>A schematic overview of fatty acid catabolism is presented with the levels of detected metabolites and relevant proteins in Control, SHF, and DHF. Detected metabolites and proteins are indicated with bold font. Blue color indicates enzymes involved in fatty acid oxidation. Metabolome is presented in arbitrary units while proteome is presented as fold change compared to Control. Filled bars: metabolites, Open bars: proteins. OCTN2: organic cation transporter novel type 2, FATP: fatty acid transport protein, FABP: fatty acid binding protein, CPT1: carnitine palmitoyltransferase I, CPT2: carnitine palmitoyltransferase II, CACT: carnitine O-acetyltransferase; DH: dehydrogenase; ETF: electron-transferring flavoprotein; ETFDH: electron transfer flavoprotein-ubiquinone oxidoreductase; CRAT: carnitine O-acetyltransferase. *P<0.05.</p

Heat map of selected metabolic proteome.

<p>Metabolism-related proteins detected by LC-MS/MS, and presented as fold change in SHF and DHF compared with those from Control. Green indicates a significant decrease, and read indicates a significant increase in the level of protein expression as compared to Control.</p

Heat map of myocardial metabolome.

<p>The data obtained by multi-platform metabolomics (GC/MS, MS/MS, HPLC) and presented as fold change in SHF and DHF as compared to Control. Green indicates a significant decrease, and read indicates a significant increase in the level of metabolite as compared to Control. BCAA: branched-chain amino acid, PPP: pentose phosphate pathway, GSH: glutathione, GC/MS: gas-chromatography/mass-spectrometry, MS/MS: tandem mass-spectrometry, HPLC: high performance liquid chromatography.</p

Quantitative analysis of myocardial carnitine and acylcarnitines.

<p>Majority of acylcarnitines were significantly reduced in both SHF and DHF as compared to Control (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118974#pone.0118974.s007" target="_blank">S3 Fig</a>. for the levels of plasma acylcarnitines). P*<0.05</p

Protein expression of creatine kinase (CK) isoforms.

<p>Both cytosolic CK-B type (<b>A</b>) and mitochondrial CK (<b>B</b>) are upregulated in SHF and DHF as compared to Control. *p<0.05.</p

Hemodynamics of SHF and DHF in pre- and post-tachypacing.

<p>*:p<0.05 (pre vs. post, paired <i>t</i>-test)</p><p>Hemodynamics of SHF and DHF in pre- and post-tachypacing.</p