Basic Research in Cardiology最新文献

The ketone body 3-hydroxybutyrate (3-OHB) increases cardiac output and myocardial perfusion without affecting blood pressure in humans, but the cardiovascular sites of action remain obscure. Here, we test the hypothesis in rats that 3-OHB acts directly on the heart to increase cardiac contractility and directly on blood vessels to lower systemic vascular resistance. We investigate effects of 3-OHB on (a) in vivo hemodynamics using echocardiography and invasive blood pressure measurements, (b) isolated perfused hearts in Langendorff systems, and (c) isolated arteries and veins in isometric myographs. We compare Na-3-OHB to equimolar NaCl added to physiological buffers or injection solutions. At plasma concentrations of 2-4 mM in vivo, 3-OHB increases cardiac output (by 28.3±7.8%), stroke volume (by 22.4±6.0%), left ventricular ejection fraction (by 13.3±4.6%), and arterial dP/dt_max (by 31.9±11.2%) and lowers systemic vascular resistance (by 30.6±11.2%) without substantially affecting heart rate or blood pressure. Applied to isolated perfused hearts at 3-10 mM, 3-OHB increases left ventricular developed pressure by up to 26.3±7.4 mmHg and coronary perfusion by up to 20.2±9.5%. Beginning at 1-3 mM, 3-OHB relaxes isolated coronary (EC₅₀=12.4 mM), cerebral, femoral, mesenteric, and renal arteries as well as brachial, femoral, and mesenteric veins by up to 60% of pre-contraction within the pathophysiological concentration range. Of the two enantiomers that constitute racemic 3-OHB, D-3-OHB dominates endogenously; but tested separately, the enantiomers induce similar vasorelaxation. We conclude that increased cardiac contractility and generalized systemic vasorelaxation can explain the elevated cardiac output during 3-OHB administration. These actions strengthen the therapeutic rationale for 3-OHB in heart failure management.

Myocardial infarction (MI) is the leading cause of death worldwide. Glycogen synthase kinase-3 (GSK-3) has been considered to be a promising therapeutic target for cardiovascular diseases. GSK-3 is a family of ubiquitously expressed serine/threonine kinases. GSK-3 isoforms appear to play overlapping, unique, and even opposing functions in the heart. Previously, our group identified that cardiac fibroblast (FB) GSK-3β acts as a negative regulator of fibrotic remodeling in the ischemic heart. However, the role of FB-GSK-3α in MI pathology is not defined. To determine the role of FB-GSK-3α in MI-induced adverse cardiac remodeling, GSK-3α was deleted specifically in the residential fibroblast or myofibroblast (MyoFB) using tamoxifen (TAM) inducible Tcf21 or Periostin (Postn) promoter-driven Cre recombinase, respectively. Echocardiographic analysis revealed that FB- or MyoFB-specific GSK-3α deletion prevented the development of dilative remodeling and cardiac dysfunction. Morphometrics and histology studies confirmed improvement in capillary density and a remarkable reduction in hypertrophy and fibrosis in the KO group. We harvested the hearts at 4 weeks post-MI and analyzed signature genes of adverse remodeling. Specifically, qPCR analysis was performed to examine the gene panels of inflammation (TNFα, IL-6, IL-1β), fibrosis (COL1A1, COL3A1, COMP, Fibronectin-1, Latent TGF-β binding protein 2), and hypertrophy (ANP, BNP, MYH7). These molecular markers were essentially normalized due to FB-specific GSK-3α deletion. Further molecular studies confirmed that FB-GSK-3α could regulate NF-kB activation and expression of angiogenesis-related proteins. Our findings suggest that FB-GSK-3α plays a critical role in the pathological cardiac remodeling of ischemic hearts, therefore, it could be therapeutically targeted.

Cardiotoxicity is a major complication of anthracycline therapy that negatively impacts prognosis. Effective pharmacotherapies for prevention of anthracycline-induced cardiomyopathy (AICM) are currently lacking. Increased plasma levels of the neutrophil-derived enzyme myeloperoxidase (MPO) predict occurrence of AICM in humans. We hypothesized that MPO release causally contributes to AICM. Mice intravenously injected with the anthracycline doxorubicin (DOX) exhibited higher neutrophil counts and MPO levels in the circulation and cardiac tissue compared to saline (NaCl)-treated controls. Neutrophil-like HL-60 cells exhibited increased MPO release upon exposition to DOX. DOX induced extensive nitrosative stress in cardiac tissue alongside with increased carbonylation of sarcomeric proteins in wildtype but not in Mpo^-/- mice. Accordingly, co-treatment of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with DOX and MPO aggravated loss of hiPSC-CM-contractility compared to DOX treatment alone. DOX-treated animals exhibited pronounced cardiac apoptosis and inflammation, which was attenuated in MPO-deficient animals. Finally, genetic MPO deficiency and pharmacological MPO inhibition protected mice from the development of AICM. The anticancer efficacy of DOX was unaffected by MPO deficiency. Herein we identify MPO as a critical mediator of AICM. We demonstrate that DOX induces cardiac neutrophil infiltration and release of MPO, which directly impairs cardiac contractility through promoting oxidation of sarcomeric proteins, cardiac inflammation and cardiomyocyte apoptosis. MPO thus emerges as a promising pharmacological target for prevention of AICM.

While low concentrations of high-density lipoprotein-cholesterol (HDL-C) are widely accepted as an independent cardiovascular risk factor, HDL-C-rising therapies largely failed, suggesting the importance of both HDL functions and individual subspecies. Indeed HDL particles are highly heterogeneous, with small, dense pre-beta-HDLs being considered highly biologically active but remaining poorly studied, largely reflecting difficulties for their purification. We developed an original experimental approach allowing the isolation of sufficient amounts of human pre-beta-HDLs and revealing the specificity of their proteomic and lipidomic profiles and biological activities. Pre-beta-HDLs were enriched in highly poly-unsaturated species of phosphatidic acid and phosphatidylserine, and in an unexpectedly high number of proteins implicated in the inflammatory response, including serum paraoxonase/arylesterase-1, vitronectin and clusterin, as well as in complement regulation and immunity, including haptoglobin-related protein, complement proteins and those of the immunoglobulin class. Interestingly, amongst proteins associated with lipid metabolism, phospholipid transfer protein, cholesteryl ester transfer protein and lecithin:cholesterol acyltransferase were strongly enriched in, or restricted to, pre-beta-HDL. Furthermore, pre-beta-HDL potently mediated cellular cholesterol efflux and displayed strong anti-inflammatory activities. A correlational network analysis between lipidome, proteome and biological activities highlighted 15 individual lipid and protein components of pre-beta-HDL relevant to cardiovascular disease, which may constitute novel diagnostic targets in a pathological context of altered lipoprotein metabolism.

In the context of myocardial infarction, the burst of superoxide generated by reverse electron transport (RET) at complex I in mitochondria is a crucial trigger for damage during ischaemia/reperfusion (I/R) injury. Here we outline the necessary conditions for superoxide production by RET at complex I and how it can occur during reperfusion. In addition, we explore various pathways that are implicated in generating the conditions for RET to occur and suggest potential therapeutic strategies to target RET, aiming to achieve cardioprotection.

Activation of signal transducer and activator of transcription 3 (STAT3) has been identified as a key cardioprotective signal not only in animal studies but also in humans-in animals, STAT3 is causally involved in cardioprotection. In response to late ischemic conditioning, canonical function of STAT3 activation upregulates the expression of cardioprotective and anti-apoptotic proteins. In its non-canonical function, STAT3 is activated during ischemic conditioning and is part of the cardioprotective cytosolic survival activating factor enhancement pathway. Activated STAT3 is imported and localized to the mitochondria. Mitochondrial STAT3 stimulates the activity of mitochondrial electron transport chain complex I, reduces mitochondrial reactive oxygen species production and mitochondrial permeability transition pore opening. Finally, two novel aspects of STAT activation in cardioprotection are discussed: a genetic variance of the STAT encoding region as a potential primordial confounding variable for cardioprotection, and the cardioprotective potential of sodium-glucose cotransporter 2 inhibitors through STAT3 activation.

Pharmacological inhibition of factor Xa by rivaroxaban has been shown to mediate cardioprotection and is frequently used in patients with, e.g., atrial fibrillation. Rivaroxaban's anti-inflammatory actions are well known, but the underlying mechanisms are still incompletely understood. To date, no study has focused on the effects of rivaroxaban on the bone marrow (BM), despite growing evidence that the BM and its activation are of major importance in the development/progression of cardiovascular disease. Thus, we examined the impact of rivaroxaban on BM composition under homeostatic conditions and in response to a major cardiovascular event. Rivaroxaban treatment of mice for 7 days markedly diminished mature leukocytes in the BM. While apoptosis of BM-derived mature myeloid leukocytes was unaffected, lineage-negative BM cells exhibited a differentiation arrest at the level of granulocyte-monocyte progenitors, specifically affecting neutrophil maturation via downregulation of the transcription factors Spi1 and Csfr1. To assess whether this persists also in situations of increased leukocyte demand, mice were subjected to cardiac ischemia/reperfusion injury (I/R): 7 d pretreatment with rivaroxaban led to reduced cardiac inflammation 72 h after I/R and lowered circulating leukocyte numbers. However, BM myelopoiesis showed a rescue of the leukocyte differentiation arrest, indicating that rivaroxaban's inhibitory effects are restricted to homeostatic conditions and are mainly abolished during emergency hematopoiesis. In translation, ST-elevation MI patients treated with rivaroxaban also exhibited reduced circulating leukocyte numbers. In conclusion, we demonstrate that rivaroxaban attenuates neutrophil maturation in the BM, which may offer a therapeutic option to limit overshooting of the immune response after I/R.

The heterocellular nature of the heart has been receiving increasing attention in recent years. In addition to cardiomyocytes as the prototypical cell type of the heart, non-myocytes such as endothelial cells, fibroblasts, or immune cells are coming more into focus. The rise of single-cell sequencing technologies enables  identification of ever more subtle differences and has reignited the question of what defines a cell's identity. Here we provide an overview of the major cardiac cell types, describe their roles in homeostasis, and outline recent findings on non-canonical functions that may be of relevance for cardiology. We highlight modes of biochemical and biophysical interactions between different cardiac cell types and discuss the potential implications of the heterocellular nature of the heart for basic research and therapeutic interventions.

During myocardial ischemia and reperfusion (IR) injury matrix metalloproteinase-2 (MMP-2) is rapidly activated in response to oxidative stress. MMP-2 is a multifunctional protease that cleaves both extracellular and intracellular proteins. Oxidative stress also impairs mitochondrial function which is regulated by different proteins, including mitofusin-2 (Mfn-2), which is lost in IR injury. Oxidative stress and mitochondrial dysfunction trigger the NLRP3 inflammasome and the innate immune response which invokes the de novo expression of an N-terminal truncated isoform of MMP-2 (NTT-MMP-2) at or near mitochondria. We hypothesized that MMP-2 proteolyzes Mfn-2 during myocardial IR injury, impairing mitochondrial function and enhancing the inflammasome response. Isolated hearts from mice subjected to IR injury (30 min ischemia/40 min reperfusion) showed a significant reduction in left ventricular developed pressure (LVDP) compared to aerobically perfused hearts. IR injury increased MMP-2 activity as observed by gelatin zymography and increased degradation of troponin I, an intracellular MMP-2 target. MMP-2 preferring inhibitors, ARP-100 or ONO-4817, improved post-ischemic recovery of LVDP compared to vehicle perfused IR hearts. In muscle fibers isolated from IR hearts the rates of mitochondrial oxygen consumption and ATP production were impaired compared to those from aerobic hearts, whereas ARP-100 or ONO-4817 attenuated these reductions. IR hearts showed higher levels of NLRP3, cleaved caspase-1 and interleukin-1β in the cytosolic fraction, while the mitochondria-enriched fraction showed reduced levels of Mfn-2, compared to aerobic hearts. ARP-100 or ONO-4817 attenuated these changes. Co-immunoprecipitation showed that MMP-2 is associated with Mfn-2 in aerobic and IR hearts. ARP-100 or ONO-4817 also reduced infarct size and cell death in hearts subjected to 45 min ischemia/120 min reperfusion. Following myocardial IR injury, impaired contractile function and mitochondrial respiration and elevated inflammasome response could be attributed, at least in part, to MMP-2 activation, which targets and cleaves mitochondrial Mfn-2. Inhibition of MMP-2 activity protects against cardiac contractile dysfunction in IR injury in part by preserving Mfn-2 and suppressing inflammation.

Endothelial dysfunction is an early event in coronary microvascular disease. Integrin-linked kinase (ILK) prevents endothelial nitric oxide synthase (eNOS) uncoupling and, thus, endothelial dysfunction. However, the specific role of endothelial ILK in cardiac function remains to be fully elucidated. We hypothesised that endothelial ILK plays a crucial role in maintaining coronary microvascular function and contractile performance in the heart. We generated an endothelial cell-specific ILK conditional knock-out mouse (ecILK cKO) and investigated cardiovascular function. Coronary endothelial ILK deletion significantly impaired cardiac function: ejection fraction, fractional shortening and cardiac output decreased, whilst left ventricle diastolic internal diameter decreased and E/A and E/E' ratios increased, indicating not only systolic but also diastolic dysfunction. The functional data correlated with extensive extracellular matrix remodelling and perivascular fibrosis, indicative of adverse cardiac remodelling. Mice with endothelial ILK deletion suffered early ischaemic-like events with ST elevation and transient increases in cardiac troponins, which correlated with fibrotic remodelling. In addition, ecILK cKO mice exhibited many features of coronary microvascular disease: reduced cardiac perfusion, impaired coronary flow reserve and arterial remodelling with patent epicardial coronary arteries. Moreover, endothelial ILK deletion induced a moderate increase in blood pressure, but the antihypertensive drug Losartan did not affect microvascular remodelling whilst only partially ameliorated fibrotic remodelling. The plasma miRNA profile reveals endothelial-to-mesenchymal transition (endMT) as an upregulated pathway in endothelial ILK conditional KO mice. Our results show that endothelial cells in the microvasculature in endothelial ILK conditional KO mice underwent endMT. Moreover, endothelial cells isolated from these mice and ILK-silenced human microvascular endothelial cells underwent endMT, indicating that decreased endothelial ILK contributes directly to this endothelial phenotype shift. Our results identify ILK as a crucial regulator of microvascular endothelial homeostasis. Endothelial ILK prevents microvascular dysfunction and cardiac remodelling, contributing to the maintenance of the endothelial cell phenotype.