Palmitate induces cardiomyocyte death via inositol requiring enzyme-1 (IRE1)-mediated signaling independent of X-box binding protein 1 (XBP1).

Overloading of the saturated fatty acid (SFA) palmitate induces cardiomyocyte death. The purpose of this study is to elucidate signaling pathways contributing to palmitate-induced cardiomyocyte death. Palmitate-induced cardiomyocyte death was induced in Toll-like receptor 2/4 double-knockdown cardiomyocytes to a similar extent as wild-type cardiomyocytes, while cardiomyocyte death was canceled out by triacsin C, a long-chain acyl-CoA synthetase inhibitor. These results indicated that palmitate induced cytotoxicity after entry and conversion into palmitoyl-CoA. Palmitoyl-CoA is not only degraded by mitochondrial oxidation but also taken up as a component of membrane phospholipids. Palmitate overloading causes cardiomyocyte membrane fatty acid (FA) saturation, which is associated with the activation of endoplasmic reticulum (ER) unfolded protein response (UPR) signaling. We focused on the ER UPR signaling as a possible mechanism of cell death. Palmitate loading activates the UPR signal via membrane FA saturation, but not via unfolded protein overload in the ER since the chemical chaperone 4-phenylbutyrate failed to suppress palmitate-induced ER UPR. The mammalian UPR relies on three ER stress sensors named inositol requiring enzyme-1 (IRE1), PKR-like endoplasmic reticulum kinase (PERK), and activating transcription factor 6 (ATF6). Palmitate loading activated only IRE1 and PERK. Knockdown of PERK did not affect palmitate-induced cardiomyocyte death, while knockdown of IRE1 suppressed palmitate-induced cardiomyocyte death. However, knockdown of X-box binding protein 1 (XBP1), the downstream effector of IRE1, did not affect palmitate-induced cardiomyocyte death. These results were validated by pharmacological inhibitor experiments. In conclusion, we identified that palmitate-induced cardiomyocyte death was triggered by IRE1-mediated signaling independent of XBP1.

Adrenal cortex hypoxia modulates aldosterone production in heart failure.

Plasma aldosterone concentration increases in proportion to the severity of heart failure, even during treatment with renin-angiotensin system inhibitors. This study investigated alternative regulatory mechanisms of aldosterone production that are significant in heart failure. Dahl salt-sensitive rats on a high-salt diet, a rat model of heart failure with cardio-renal syndrome, had high plasma aldosterone levels and elevated ß3-adrenergic receptor expression in hypoxic zona glomerulosa cells. In H295R cells (a human adrenocortical cell line), hypoxia-induced ß3-adrenergic receptor expression. Hypoxia-mediated ß3-adrenergic receptor expression augmented aldosterone production by facilitating hydrolysis of lipid droplets though ERK-mediated phosphorylation of hormone-sensitive lipase, also known as cholesteryl ester hydrolase. Hypoxia also accelerated the synthesis of cholesterol esters by acyl-CoA:cholesterol acyltransferase, thereby increasing the cholesterol ester content in lipid droplets. Thus, hypoxia enhanced aldosterone production by zona glomerulosa cells via promotion of the accumulation and hydrolysis of cholesterol ester in lipid droplets. In conclusion, hypoxic zona glomerulosa cells with heart failure show enhanced aldosterone production via increased catecholamine responsiveness and activation of cholesterol trafficking, irrespective of the renin-angiotensin system.

Pressure overload inhibits glucocorticoid receptor transcriptional activity in cardiomyocytes and promotes pathological cardiac hypertrophy.

Glucocorticoid receptor (GR) is abundantly expressed in cardiomyocytes. However, the role of GR in regulating cardiac hypertrophy and heart failure in response to pressure overload remains unclear. Cardiomyocyte-specific GR knockout (GRcKO) mice, mineralocorticoid receptor (MR) knockout (MRcKO), and GR and MR double KO (GRMRdcKO) mice were generated using the Cre-lox system. In response to pressure overload, GRcKO mice displayed worse cardiac remodeling compared to control (GRf/f) mice, including a greater increase in heart weight to body weight ratio with a greater increase in cardiomyocytes size, a greater decline in left ventricular contractility, and higher reactivation of fetal genes. MRcKO mice showed a comparable degree of cardiac remodeling compared to control (MRf/f) mice. The worse cardiac remodeling in pressure overloaded GRcKO mice is not due to compensatory activation of cardiomyocyte MR, since pressure overloaded GRMRdcKO mice displayed cardiac remodeling to the same extent as GRcKO mice. Pressure overload suppressed GR-target gene expression in the heart. Although plasma corticosterone levels and subcellular localization of GR (nuclear/cytoplasmic GR) were not changed, a chromatin immunoprecipitation assay revealed that GR recruitment onto the promoter of GR-target genes was significantly suppressed in response to pressure overload. Rescue of the expression of GR-target genes to the same extent as sham-operated hearts attenuated adverse cardiac remodeling in pressure-overloaded hearts. Thus, GR works as a repressor of adverse cardiac remodeling in response to pressure overload, but GR-mediated transcription is suppressed under pressure overload. Therapies that maintain GR-mediated transcription in cardiomyocytes under pressure overload can be a promising therapeutic strategy for heart failure.

Sirt1 counteracts decrease in membrane phospholipid unsaturation and diastolic dysfunction during saturated fatty acid overload.

BACKGROUND: The fatty acid (FA) composition of membrane phospholipid reflects at least in part dietary fat composition. Saturated FA (SFA) suppress Sirt1 activity, while monounsaturated FA (MUFA) counteract this effect. OBJECTIVE: We explored a role of Sirt1 in homeostatic control of the fatty acid composition of membrane phospholipid in the presence of SFA overload. METHODS AND RESULTS: Sirt1 deficiency in cardiomyocytes decreased the expression levels of liver X receptor (LXR)-target genes, particularly stearoyl-CoA desaturase-1 (Scd1), a rate-limiting enzyme in the cellular synthesis of MUFA from SFA, increased membrane SFA/MUFA ratio, and worsened left ventricular (LV) diastolic function in mice fed an SFA-rich high fat diet. In cultured cardiomyocytes, Sirt1 knockdown (KD) exacerbated the palmitate overload-induced increase in membrane SFA/MUFA ratio, which was associated with decrease in the expression of LXR-target genes, including Scd1. Forced overexpression of Scd1 in palmitate-overloaded Sirt1KD cardiomyocytes lowered the SFA/MUFA ratio. Nicotinamide mononucleotide (NMN) increased Sirt1 activity and Scd1 expression, thereby lowering membrane SFA/MUFA ratio in palmitate-overloaded cardiomyocytes. These effects of NMN were not observed for Scd1KD cardiomyocytes. LXRα/ßKD exacerbated palmitate overload-induced increase in membrane SFA/MUFA ratio, while LXR agonist T0901317 alleviated it. NMN failed to rescue Scd1 protein expression and membrane SFA/MUFA ratio in palmitate-overloaded LXRα/ßKD cardiomyocytes. The administration of NMN or T0901317 showed a dramatic reversal in membrane SFA/MUFA ratio and LV diastolic function in SFA-rich HFD-fed mice. CONCLUSION: Cardiac Sirt1 counteracted SFA overload-induced decrease in membrane phospholipid unsaturation and diastolic dysfunction via regulating LXR-mediated transcription of the Scd1 gene.

Adventitial CXCL1/G-CSF expression in response to acute aortic dissection triggers local neutrophil recruitment and activation leading to aortic rupture.

RATIONALE: In-hospital outcomes are generally acceptable in patients with type B dissection; however, some patients present with undesirable complications, such as aortic expansion and rupture. Excessive inflammation is an independent predictor of adverse clinical outcomes. OBJECTIVE: We have investigated the underlying mechanisms of catastrophic complications after acute aortic dissection (AAD) in mice. METHODS AND RESULTS: When angiotensin II was administered in lysyl oxidase inhibitor-preconditioned mice, AAD emerged within 24 hours. The dissection was initiated at the proximal site of the descending thoracic aorta and propagated distally into an abdominal site. Dissection of the aorta caused dilatation, and ≈70% of the mice died of aortic rupture. AAD triggered CXCL1 and granulocyte-colony stimulating factor expression in the tunica adventitia of the dissected aorta, leading to elevation of circulating CXCL1/granulocyte-colony stimulating factor levels. Bone marrow CXCL12 was reduced. These chemokine changes facilitated neutrophil egress from bone marrow and infiltration into the aortic adventitia. Interference of CXCL1 function using an anti-CXCR2 antibody reduced neutrophil accumulation and limited aortic rupture post AAD. The tunica adventitia of the expanded dissected aorta demonstrated high levels of interleukin-6 (IL-6) expression. Neutrophils were the major sources of IL-6, and CXCR2 neutralization significantly reduced local and systemic levels of IL-6. Furthermore, disruption of IL-6 effectively suppressed dilatation and rupture of the dissected aorta without any influence on the incidence of AAD and neutrophil mobilization. CONCLUSIONS: Adventitial CXCL1/granulocyte-colony stimulating factor expression in response to AAD triggers local neutrophil recruitment and activation. This leads to adventitial inflammation via IL-6 and results in aortic expansion and rupture.

Cardiac Sirt1 mediates the cardioprotective effect of caloric restriction by suppressing local complement system activation after ischemia-reperfusion.

Caloric restriction (CR) confers cardioprotection against ischemia-reperfusion (I/R) injury. We previously found the essential roles of endothelial nitric oxide synthase in the development of CR-induced cardioprotection and Sirt1 activation during CR (Shinmura K, Tamaki K, Ito K, Yan X, Yamamoto T, Katsumata Y, Matsuhashi T, Sano M, Fukuda K, Suematsu M, Ishii I. Indispensable role of endothelial nitric oxide synthase in caloric restriction-induced cardioprotection against ischemia-reperfusion injury.Am J Physiol Heart Circ Physiol 308: H894-H903, 2015). However, the exact mechanism by which Sirt1 in cardiomyocytes mediates the cardioprotective effect of CR remains undetermined. We subjected cardiomyocyte-specific Sirt1 knockout (CM-Sirt1(-/-)) mice and the corresponding control mice to either 3-mo ad libitum feeding or CR (-40%). Isolated perfused hearts were subjected to 25-min global ischemia, followed by 60-min reperfusion. The recovery of left ventricle function after I/R was improved, and total lactate dehydrogenase release into the perfusate during reperfusion was attenuated in the control mice treated with CR, but a similar cardioprotective effect of CR was not observed in the CM-Sirt1(-/-)mice. The expression levels of cardiac complement component 3 (C3) at baseline and the accumulation of C3 and its fragments in the ischemia-reperfused myocardium were attenuated by CR in the control mice, but not in the CM-Sirt1(-/-)mice. Resveratrol treatment also attenuated the expression levels of C3 protein in cultured neonatal rat ventricular cardiomyocytes. Moreover, the degree of myocardial I/R injury in conventional C3 knockout (C3(-/-)) mice treated with CR was similar to that in the ad libitum-fed C3(-/-)mice, although the expression levels of Sirt1 were enhanced by CR. These results demonstrate that cardiac Sirt1 plays an essential role in CR-induced cardioprotection against I/R injury by suppressing cardiac C3 expression. This is the first report suggesting that cardiac Sirt1 regulates the local complement system during CR.

Lung natural killer cells play a major counter-regulatory role in pulmonary vascular hyperpermeability after myocardial infarction.

RATIONALE: Natural killer (NK) cells are lymphocytes of the innate immune system that play specialized and niche-specific roles in distinct organs. OBJECTIVE: We investigated the possible function of NK cells in the pathogenesis of congestive heart failure after myocardial infarction. METHODS AND RESULTS: Depletion of NK cells from mice had little effect on cytokine expression (tumor necrosis factor-α, interleukin [IL]-6, and IL-1ß), neutrophil and macrophage infiltration into infarcted myocardium, or left ventricular remodeling after myocardial infarction. However, these mice exhibited severe respiratory distress associated with protein-rich, high-permeability alveolar edema accompanied by neutrophil infiltration. In addition, there were 20-fold more NK cells in the mouse lungs than in heart, and these cells were accumulated around the vasculature. CD107a-positive and interferon-γ-positive cell populations were unchanged, whereas IL-10-positive populations increased. Adoptive transfer of NK cells from wild-type mice, but not from IL-10 knockout mice, into the NK cell-depleted mice rescued the respiratory phenotype. IL-1ß-mediated dextran leakage from a lung endothelial cell monolayer was also blocked by coculture with NK cells from wild-type mice but not from IL-10 knockout mice. CONCLUSIONS: This study is the first to identify a critical role for lung NK cells in protecting lung from the development of cardiogenic pulmonary edema after myocardial infarction.

Activation of pyruvate dehydrogenase by dichloroacetate has the potential to induce epigenetic remodeling in the heart.

Dichloroacetate (DCA) promotes pyruvate entry into the Krebs cycle by inhibiting pyruvate dehydrogenase (PDH) kinase and thereby maintaining PDH in the active dephosphorylated state. DCA has recently gained attention as a potential metabolic-targeting therapy for heart failure but the molecular basis of the therapeutic effect of DCA in the heart remains a mystery. Once-daily oral administration of DCA alleviates pressure overload-induced left ventricular remodeling. We examined changes in the metabolic fate of pyruvate carbon (derived from glucose) entering the Krebs cycle by metabolic interventions of DCA. (13)C6-glucose pathway tracing analysis revealed that instead of being completely oxidized in the mitochondria for ATP production, DCA-mediated PDH dephosphorylation results in an increased acetyl-CoA pool both in control and pressure-overloaded hearts. DCA induces hyperacetylation of histone H3K9 and H4 in a dose-dependent manner in parallel to the dephosphorylation of PDH in cultured cardiomyocytes. DCA administration increases histone H3K9 acetylation in in vivo mouse heart. Interestingly, DCA-dependent histone acetylation was associated with an up-regulation of 2.3% of genes (545 out of 23,474 examined). Gene ontology analysis revealed that these genes are highly enriched in transcription-related categories. This evidence suggests that sustained activation of PDH by DCA results in an overproduction of acetyl-CoA, which exceeds oxidation in the Krebs cycle and results in histone acetylation. We propose that DCA-mediated PDH activation has the potential to induce epigenetic remodeling in the heart, which, at least in part, forms the molecular basis for the therapeutic effect of DCA in the heart.

Indispensable role of endothelial nitric oxide synthase in caloric restriction-induced cardioprotection against ischemia-reperfusion injury.

Caloric restriction (CR) confers cardioprotection against ischemia-reperfusion injury (IRI). We previously found that treatment with N(G)-nitro-l-arginine methyl ester completely abrogates CR-induced cardioprotection and increases nuclear sirtuin 1 (Sirt1) expression. However, it remains unclear whether endothelial nitric oxide (NO) synthase (eNOS) plays a role in CR-induced cardioprotection and Sirt1 activation. We subjected eNOS-deficient (eNOS(-/-)) mice to either 3-mo ad libitum (AL) feeding or CR (-40%). Isolated perfused hearts were subjected to 25-min global ischemia followed by 60-min reperfusion. The degree of myocardial IRI in AL-fed eNOS(-/-) mice was more severe than that in AL-fed wild-type mice. Furthermore, CR did not exert cardioprotection in eNOS(-/-) mice. eNOS(-/-) mice exhibited elevated blood pressure and left ventricular hypertrophy compared with wild-type mice, although they underwent CR. Although nuclear Sir1 content was increased, the increases in cardiac Sirt1 activity with CR was absent in eNOS(-/-) mice. In eNOS(-/-) mice treated with hydralazine, blood pressure and left ventricular weight became comparable with CR-treated wild-type mice. However, CR-induced cardioprotection was not observed. Resveratrol enhanced cardiac Sirt1 activity but failed to mimic CR-induced cardioprotection in eNOS(-/-) mice. Finally, combination therapy with resveratrol and hydralazine attenuated myocardial IRI and reduced infarct size in eNOS(-/-) mice, and their effects were comparable with those observed in CR-treated wild-type mice. These results demonstrate the essential roles of eNOS in the development of CR-induced cardioprotection and Sirt1 activation during CR. The combination of a relatively low dose of resveratrol with an adequate vasodilator therapy might be useful for managing patients with endothelial dysfunction associated with impaired NO bioavailability.

Temporal dynamics of cardiac immune cell accumulation following acute myocardial infarction.

Acute myocardial infarction (MI) causes sterile inflammation, which is characterized by recruitment and activation of innate and adaptive immune system cells. Here we delineate the temporal dynamics of immune cell accumulation following MI by flow cytometry. Neutrophils increased immediately to a peak at 3 days post-MI. Macrophages were numerically the predominant cells infiltrating the infarcted myocardium, increasing in number over the first week post-MI. Macrophages are functionally heterogeneous, whereby the first responders exhibit high expression levels of proinflammatory mediators, while the late responders express high levels of the anti-inflammatory cytokine IL-10; these macrophages can be classified into M1 and M2 macrophages, respectively, based on surface-marker expression. M1 macrophages dominated at 1-3 days post-MI, whereas M2 macrophages represented the predominant macrophage subset after 5 days. The M2 macrophages expressed high levels of reparative genes in addition to proinflammatory genes to the same levels as in M1 macrophages. The predominant subset of dendritic cells (DCs) was myeloid DC, which peaked in number on day 7. Th1 and regulatory T cells were the predominant subsets of CD4(+) T cells, whereas Th2 and Th17 cells were minor populations. CD8(+) T cells, γδT cells, B cells, natural killer (NK) cells and NKT cells peaked on day 7 post-MI. Timely reperfusion reduced the total number of leukocytes accumulated in the post-MI period, shifting the peak of innate immune response towards earlier and blunting the wave of adaptive immune response. In conclusion, these results provide important knowledge necessary for developing successful immunomodulatory therapies.

Biphasic time course of the changes in aldosterone biosynthesis under high-salt conditions in Dahl salt-sensitive rats.

OBJECTIVE: The comorbidity of excess salt and elevated plasma aldosterone has deleterious effects in cardiovascular disease. We evaluated the mechanisms behind the paradoxical increase in aldosterone biosynthesis in relation to dietary intake of salt. METHODS AND RESULTS: Dahl salt-sensitive (Dahl-S) and salt-resistant (Dahl-R) rats were fed a high-salt diet, and plasma and tissue levels of aldosterone in the adrenal gland and heart were quantified by liquid chromatography-electrospray ionization-tandem mass spectrometry. In Dahl-S rats, we found that the delayed and paradoxical increase in aldosterone biosynthesis after the initial and appropriate response to high salt. The late rise in aldosterone biosynthesis was accompanied by upregulation of CYP11B2 expression in the zona glomerulosa and increased adrenal angiotensin II levels and renin-angiotensin system components. It preceded the appearance of left ventricular systolic dysfunction and renal insufficiency. Blockade of angiotensin AT(1) receptors reversed the paradoxical increase in aldosterone biosynthesis. In contrast, Dahl-R rats maintained the initial suppression of aldosterone biosynthesis. Aldosterone levels in the heart closely paralleled those in the plasma and adrenal gland and disappeared after bilateral adrenalectomy. CONCLUSIONS: Chronic salt overload in Dahl-S rats stimulates aberrant aldosterone production via activation of the local renin-angiotensin system in the adrenal gland, thereby creating the comorbidity of excess salt and elevated plasma aldosterone.

PGD2-CRTH2 pathway promotes tubulointerstitial fibrosis.

Urinary excretion of lipocalin-type PGD(2) synthase (L-PGDS), which converts PG H(2) to PGD(2), increases in early diabetic nephropathy. In addition, L-PGDS expression in the tubular epithelium increases in adriamycin-induced nephropathy, suggesting that locally produced L-PGDS may promote the development of CKD. In this study, we found that L-PGDS-derived PGD(2) contributes to the progression of renal fibrosis via CRTH2-mediated activation of Th2 lymphocytes. In a mouse model, the tubular epithelium synthesized L-PGDS de novo after unilateral ureteral obstruction (UUO). L-PGDS-knockout mice and CRTH2-knockout mice both exhibited less renal fibrosis, reduced infiltration of Th2 lymphocytes into the cortex, and decreased production of the Th2 cytokines IL-4 and IL-13. Furthermore, oral administration of a CRTH2 antagonist, beginning 3 days after UUO, suppressed the progression of renal fibrosis. Ablation of IL-4 and IL-13 also ameliorated renal fibrosis in the UUO kidney. Taken together, these data suggest that blocking the activation of CRTH2 by PGD(2) might be a strategy to slow the progression of renal fibrosis in CKD.

A new preventive strategy for hypoglycemia incorporating added food diet in patients with type 2 diabetes who received sitagliptin therapy.

There has been concern as to whether dipeptidyl peptidase-4 (DPP-4) inhibitors can be used safely in patients with relatively good glycemic control. This study, approved by the institutional review board of Hanzoumon Diabetes City Atlas Clinic, examined whether DPP-4 inhibitor sitagliptin could safely achieve good glycemic control without severe hypoglycemia by employing the "added food" concept. The subjects were 60 patients (46 men and 14 women) with type 2 diabetes who started sitagliptin therapy during a 1-month period from December 15, 2009 to January 15, 2010. They were recommended to have added food between meals to prevent hypoglycemia, while maintaining the same daily calorie intake. HbA(1c) decreased from 7.1 ± 1.2% to 6.5 ± 0.6% after 6 months of sitagliptin treatment (p < 0.001). In patients with a baseline HbA(1c) <7%, it decreased from 6.5 ± 0.3% to 6.1 ± 0.4% (p < 0.001). Systolic blood pressure was significantly reduced from 127.7 ± 17.0 to 122.7 ± 17.9 mmHg in the patients with a baseline HbA(1c) < 7% (p = 0.018). However, body weight increased by approximately 900 g and high-density lipoprotein cholesterol decreased significantly from 1.57 ± 0.46 to 1.43 ± 0.35 mmol/L (p < 0.01) in the patients concomitantly receiving sulfonylureas with sitagliptin. Excellent glycemic control was achieved by sitagliptin treatment together with the added food concept. However, combined use of sitagliptin with sulfonylureas requires attention to weight gain and the lipid profile. Further clinical studies will elucidate whether sitagliptin can decrease cardiovascular events as well as normalizing blood glucose and lowering the blood pressure.

Metabolic remodeling induced by mitochondrial aldehyde stress stimulates tolerance to oxidative stress in the heart.

RATIONALE: Aldehyde accumulation is regarded as a pathognomonic feature of oxidative stress-associated cardiovascular disease. OBJECTIVE: We investigated how the heart compensates for the accelerated accumulation of aldehydes. METHODS AND RESULTS: Aldehyde dehydrogenase 2 (ALDH2) has a major role in aldehyde detoxification in the mitochondria, a major source of aldehydes. Transgenic (Tg) mice carrying an Aldh2 gene with a single nucleotide polymorphism (Aldh2*2) were developed. This polymorphism has a dominant-negative effect and the Tg mice exhibited impaired ALDH activity against a broad range of aldehydes. Despite a shift toward the oxidative state in mitochondrial matrices, Aldh2*2 Tg hearts displayed normal left ventricular function by echocardiography and, because of metabolic remodeling, an unexpected tolerance to oxidative stress induced by ischemia/reperfusion injury. Mitochondrial aldehyde stress stimulated eukaryotic translation initiation factor 2alpha phosphorylation. Subsequent translational and transcriptional activation of activating transcription factor-4 promoted the expression of enzymes involved in amino acid biosynthesis and transport, ultimately providing precursor amino acids for glutathione biosynthesis. Intracellular glutathione levels were increased 1.37-fold in Aldh2*2 Tg hearts compared with wild-type controls. Heterozygous knockout of Atf4 blunted the increase in intracellular glutathione levels in Aldh2*2 Tg hearts, thereby attenuating the oxidative stress-resistant phenotype. Furthermore, glycolysis and NADPH generation via the pentose phosphate pathway were activated in Aldh2*2 Tg hearts. (NADPH is required for the recycling of oxidized glutathione.) CONCLUSIONS: The findings of the present study indicate that mitochondrial aldehyde stress in the heart induces metabolic remodeling, leading to activation of the glutathione-redox cycle, which confers resistance against acute oxidative stress induced by ischemia/reperfusion.

4-hydroxy-2-nonenal protects against cardiac ischemia-reperfusion injury via the Nrf2-dependent pathway.

Reactive oxygen species (ROS) attack polyunsaturated fatty acids of the membrane and trigger lipid peroxidation, which results in the generation of alpha,beta-unsaturated aldehydes, such as 4-hydroxy-2-nonenal (4-HNE). There is compelling evidence that high concentrations of aldehydes are responsible for much of the damage elicited by cardiac ischemia-reperfusion injury, while sublethal concentrations of aldehydes stimulate stress resistance pathways, to achieve cardioprotection. We investigated the mechanism of cardioprotection mediated by 4-HNE. For cultured cardiomyocytes, 4-HNE was cytotoxic at higher concentrations (>or=20 microM) but had no appreciable cytotoxicity at lower concentrations. Notably, a sublethal concentration (5muM) of 4-HNE primed cardiomyocytes to become resistant to cytotoxic concentrations of 4-HNE. 4-HNE induced nuclear translocation of transcription factor NF-E2-related factor 2 (Nrf2), and enhanced the expression of gamma-glutamylcysteine ligase (GCL) and the core subunit of the Xc(-) high-affinity cystine transporter (xCT), thereby increasing 1.45-fold the intracellular GSH levels. Cardiomyocytes treated with either Nrf2-specific siRNA or the GCL inhibitor l-buthionine sulfoximine (BSO) were less tolerant to 4-HNE. Moreover, the cardioprotective effect of 4-HNE pretreatment against subsequent glucose-free anoxia followed by reoxygenation was completely abolished in these cells. Intravenous administration of 4-HNE (4 mg/kg) activated Nrf2 in the heart and increased the intramyocardial GSH content, and consequently improved the functional recovery of the left ventricle following ischemia-reperfusion in Langendorff-perfused hearts. This cardioprotective effect of 4-HNE was not observed for Nrf2-knockout mice. In summary, 4-HNE activates Nrf2-mediated gene expression and stimulates GSH biosynthesis, thereby conferring on cardiomyocytes protection against ischemia-reperfusion injury.

Decrease in membrane phospholipids unsaturation correlates with myocardial diastolic dysfunction.

Increase in saturated fatty acid (SFA) content in membrane phospholipids dramatically affects membrane properties and cellular functioning. We sought to determine whether exogenous SFA from the diet directly affects the degree of membrane phospholipid unsaturation in adult hearts and if these changes correlate with contractile dysfunction. Although both SFA-rich high fat diets (HFDs) and monounsaturated FA (MUFA)-rich HFDs cause the same degree of activation of myocardial FA uptake, triglyceride turnover, and mitochondrial FA oxidation and accumulation of toxic lipid intermediates, the former induced more severe diastolic dysfunction than the latter, which was accompanied with a decrease in membrane phospholipid unsaturation, induction of unfolded protein response (UPR), and a decrease in the expression of Sirt1 and stearoyl-CoA desaturase-1 (SCD1), catalyzing the conversion of SFA to MUFA. When the SFA supply in the heart overwhelms the cellular capacity to use it for energy, excess exogenous SFA channels to membrane phospholipids, leading to UPR induction, and development of diastolic dysfunction.

The histone 3 lysine 9 methyltransferase inhibitor chaetocin improves prognosis in a rat model of high salt diet-induced heart failure.

Histone acetylation has been linked to cardiac hypertrophy and heart failure. However, the pathological implications of changes in histone methylation and the effects of interventions with histone methyltransferase inhibitors for heart failure have not been fully clarified. Here, we focused on H3K9me3 status in the heart and investigated the effects of the histone H3K9 methyltransferase inhibitor chaetocin on prognoses in Dahl salt-sensitive rats, an animal model of chronic heart failure. Chaetocin prolonged survival and restored mitochondrial dysfunction. ChIP-seq analysis demonstrated that chronic stress to the heart induced H3K9me3 elevation in thousands of repetitive elements, including intronic regions of mitochondria-related genes, such as the gene encoding peroxisome proliferator-activated receptor-gamma coactivator 1 alpha. Furthermore, chaetocin reversed this effect on these repetitive loci. These data suggested that excessive heterochromatinization of repetitive elements of mitochondrial genes in the failing heart may lead to the silencing of genes and impair heart function. Thus, chaetocin may be a potential therapeutic agent for chronic heart failure.

Basal-Supported Oral Therapy with Sitagliptin Counteracts Rebound Hyperglycemia Caused by GLP-1 Tachyphylaxis.

Introduction. Treatment with a glucagon-like peptide 1 (GLP-1) analog fails in some patients due to rebound hyperglycemia caused by tachyphylaxis (GLP-1 tachyphylaxis). We investigated the efficacy of basal-supported oral therapy (BOT) with insulin glargine and sitagliptin for counteracting GLP-1 tachyphylaxis. Materials and Methods. The subjects were 12 men and 3 women aged 59.9 ± 10.0 years who had been treated with GLP-1 analogs. All of them had developed rebound hyperglycemia caused by GLP-1 tachyphylaxis. Their GLP-1 analog-based therapy was switched to BOT with insulin glargine plus sitagliptin and other medications. The primary outcomes were whether switching of therapy was associated with a change of hemoglobin A1c (HbA1c) and whether weight gain occurred. Results. Baseline HbA1c was 8.0 ± 0.9%. It decreased to 7.3 ± 0.9% at 3 months after switching (P < 0.01) and to 7.2 ± 0.9% at 4 months (P < 0.05). Weight gain was 1.1 kg after 1 month (P < 0.01) and 2.3 kg after 5 months (P < 0.01). Conclusion. Switching to BOT with insulin glargine and sitagliptin improved glycemic control. The significant decrease of HbA1c demonstrated that this combination can counteract deterioration of glycemic control due to rebound hyperglycemia secondary to GLP-1 tachyphylaxis. However, weight gain remains a problem.

Endogenous prostaglandin D2 and its metabolites protect the heart against ischemia-reperfusion injury by activating Nrf2.

We recently demonstrated that glucocorticoids markedly upregulate the expression of cyclooxygenase-2 in cardiomyocytes and protect hearts from ischemia-reperfusion (I/R) injury by activating lipocalin-type prostaglandin D (PGD) synthase (L-PGDS)-derived PGD(2) biosynthesis. We examined a downstream mechanism of cardioprotection elicited by PGD(2) biosynthesis. Acute PGD(2) treatment did not protect hearts against I/R injury. We then speculated that PGD(2) and its metabolite 15-deoxy-Δ12,14-PGJ(2) activate gene expression networks to mediate the glucocorticoid-mediated cardioprotection. Using an unbiased approach, we identified that glucocorticoids induce a number of well-known erythroid-derived 2-like 2 (Nrf2) target genes in the heart in an L-PGDS-dependent manner and that the cardioprotective effect of glucocorticoids against I/R injury was not seen in Nrf2-knockout hearts. We showed relatively low expression of PGD(2) receptors (ie, DP1 and DP2) in the heart but abundant expression of PGF(2α) receptor (FP), which binds PGF(2α) and PGD(2) with equal affinity. Glucocorticoids also failed to induce the expression of L-PGDS-dependent Nrf2 target genes in FP-knockout hearts. PGD(2) acted through its metabolite 15-deoxy-Δ12,14-PGJ(2) in the heart as evidenced by the glucocorticoid-mediated activation of peroxisome proliferator-activated receptor-γ. In turn, glucocorticoids failed to induce the expression of L-PGDS-dependent Nrf2 target genes in hearts pretreated with peroxisome proliferator-activated receptor-γ antagonist GW9662, and glucocorticoid-mediated cardioprotection against I/R injury was compromised in FP-knockout mice and GW9662-treated mice. In conclusion, PGD(2) protects heart against I/R injury by activating Nrf2 predominantly via FP receptor. In addition, we propose activation of peroxisome proliferator-activated receptor-γ by the dehydrated metabolite of PGD(2) (15-deoxy-Δ12,14-PGJ(2)) as another mechanism by which glucocorticoids induce cardioprotection.