Cardiomyocyte-specific deletion of PTP1B protects against HFD-induced cardiomyopathy through direct regulation of cardiac metabolic signaling

Background Heart failure is the number one cause of death worldwide and mortality is directly correlated with the high incidence of obesity and diabetes. Indeed, the epidemic phenomenon of obesity was projected to reach 50% in the US by the year 2030. However, the mechanisms linking metabolic dysfunction with heart disease are not clear. Protein Tyrosine Phosphatase 1B (PTP1B), a negative regulator of insulin signaling, is considered to be an emerging therapeutic target against the development of obesity, insulin resistance, and diabetes. Increased PTP1B levels and activity have been observed in brain, muscle and adipose tissues isolated from obese and/or diabetic animals, as well as in human obese human patients. Its role, however, and the mechanisms by which it modulates metabolic processes in the heart remain unknown. Method and Results We generated cardiomyocyte (CM)-specific PTP1B knock-out (PTP1Bfl/fl::ꭤMHCCre/+) mice to investigate the cardiomyocyte-specific role of PTP1B in response to high fat diet (HFD)-induced cardiac dysfunction. While we did not observe any physiological or functional cardiac differences at baseline, in response to HFD, we found that PTP1Bfl/fl::ꭤMHCCre/+ mice were protected against development of cardiac hypertrophy, mitochondrial dysfunction, and diminished cardiac steatosis. Metabolomics data revealed that hearts with CM-specific deletion of PTP1B had increased fatty acid oxidation and NAD+ metabolism, but reduced glucose metabolism; we further validated these findings by real-time qPCR analysis. Mechanistically, we identified a novel PTP1B PKM2-AMPK axis in the heart, which acts as a molecular switch to promote fatty acid oxidation. In this regard, we identified that hearts from PTP1Bfl/fl::ꭤMHCCre/+ mice had upregulated levels of nicotinamide adenine dinucleotide (NAD+) and NAD phosphate (NADPH), leading to higher levels of nicotinamide phosphoribosyl transferase (NAMPT), the rate-limiting step of the NAD+ salvage pathway and an enzyme associated with obesity and diabetes. Conclusions Together, these results suggest that CM-specific deletion of PTP1B mediates a substrate switch from glucose to fatty acid metabolism, protecting hearts against development of HFD-induced cardiac hypertrophy and dysfunction through mechanisms involving a novel PTP1B/PKM2/AMPK axis that is critical for the regulation of NAMPT and NAD+ biosynthesis. ### Competing Interest Statement The authors have declared no competing interest. * 18S : 18S ribosomal RNA ꭤMHC : alpha myosin heavy chain (MYH6) ACADVL : acyl-CoA dehydrogenase very long chain AGO2 : argonaute 2 AMP : adenosine monophosphate AMPK : AMP-activated protein kinase ANF : atrial natriuretic factor ATP : adenosine triphosphate CM : cardiomyocyte CPT1b : carnitine palmitoyl-transferase 1b CVD : cardiovascular disease EEF1A1 : elongation factor 1-alpha 1 ERK : extracellular signal-regulated kinase ESI : electrospray ionization ETC : electron transport chain FAO : fatty acid oxidation FCCP : carbonyl cyanide-p-trifluoromethoxyphenyl-hydrazon H&E : hematoxylin/eosin HADHB : acetyl-CoA acyltransferase HFD : high-fat diet HFpEF : heart failure with preserved ejection fraction HSL : hormone-sensitive lipase IR : insulin receptor IRS : insulin receptor substrate LC3B : microtubule-associated protein 1A/1B-light chain 3B LVIDd : end-diastolic internal dimensions of the left ventricle LVIDs : end-systolic internal dimensions of the left ventricle LVPW : left ventricular posterior wall thickness MAPK : Ras–mitogen-activated protein kinase MED13 : mediator complex subunit 13 mTOR : mammalian target of rapamycin MYH6 : α-myosin heavy chain MYH7 : b-myosin heavy chain NAD : nicotinamide adenine dinucleotide NADPH : nicotinamide adenine dinucleotide phosphate NAM : nicotinamide NAMPT : nicotinamide phosphoribosyl-transferase ND : normal diet NDUFB8 NADH : ubiquinone oxidoreductase subunit B8 NMN : nicotinamide ribonucleotide OCR : oxygen consumption rates OXPHO : oxidative phosphorylation PDH : pyruvate dehydrogenase PI3K : phosphatidylinositol 3-kinase PKB/AKT : protein kinase B PKM2 : pyruvate kinase muscle isozyme 2 POMC : pro-opiomelanocortin PTP1B : protein tyrosine phosphatase PTP1B+/+::ꭤMHCCre/+ : alpha myosin heavy chain promoter driven by expression of Cre in the background of the wildtype PTP1B allele PTP1Bfl/fl::ꭤMHCCre/+ : alpha myosin heavy chain promoter driving deletion of PTP1B specifically in cardiomyocytes ROS : reactive oxygen species RPL4 : ribosomal protein L4 RSD : relative standard deviation SDHB : succinate dehydrogenase subunit b SDS-PAGE : sodium dodecyl-sulfate polyacrylamide gel electrophoresis TCA : tricarboxylic acid TEM : transmission electron microscope VEGF : vascular endothelial growth factor VEGFR : vascular endothelial growth factor receptor WGA : wheat germ agglutinin