Dysfunction In Heart Failure With Preserved Ejection Fraction Research Articles

Geniposide alleviates heart failure with preserved ejection fraction in mice by regulating cardiac oxidative stress via MMP2/SIRT1/GSK3β pathway.

Heart failure with preserved ejection fraction (HFpEF) is a complex clinical syndrome with cardiac dysfunction, fluid retention and reduced exercise tolerance as the main manifestations. Current treatment of HFpEF is using combined medications of related comorbidities, there is an urgent need for a modest drug to treat HFpEF. Geniposide (GE), an iridoid glycoside extracted from Gardenia Jasminoides, has shown significant efficacy in the treatment of cardiovascular, digestive and central nervous system disorders. In this study we investigated the therapeutic effects of GE on HFpEF experimental models in vivo and in vitro. HFpEF was induced in mice by feeding with HFD and L-NAME (0.5 g/L) in drinking water for 8 weeks, meanwhile the mice were treated with GE (25, 50 mg/kg) every other day. Cardiac echocardiography and exhaustive exercise were performed, blood pressure was measured at the end of treatment, and heart tissue specimens were collected after the mice were euthanized. We showed that GE administration significantly ameliorated cardiac oxidative stress, inflammation, apoptosis, fibrosis and metabolic disturbances in the hearts of HFpEF mice. We demonstrated that GE promoted the transcriptional activation of Nrf2 by targeting MMP2 to affect upstream SIRT1 and downstream GSK3β, which in turn alleviated the oxidative stress in the hearts of HFpEF mice. In H9c2 cells and HL-1 cells, we showed that treatment with GE (1 μM) significantly alleviated H2O2-induced oxidative stress through the MMP2/SIRT1/GSK3β pathway. In summary, GE regulates cardiac oxidative stress via MMP2/SIRT1/GSK3β pathway and reduces cardiac inflammation, apoptosis, fibrosis and metabolic disorders as well as cardiac dysfunction in HFpEF. GE exerts anti-oxidative stress properties by binding to MMP2, inhibiting ROS generation in HFpEF through the SIRT1/Nrf2 signaling pathway. In addition, GE can also affect the inhibition of the downstream MMP2 target GSK3β, thereby suppressing the inflammatory and apoptotic responses in HFpEF. Taken together, GE alleviates oxidative stress/apoptosis/fibrosis and metabolic disorders as well as HFpEF through the MMP2/SIRT1/GSK3β signaling pathway.

Statins improve cardiac endothelial function to prevent heart failure with preserved ejection fraction through upregulating circRNA-RBCK1

Heart failure with preserved ejection fraction (HFpEF) is associated with endothelial dysfunction. We have previously reported that statins prevent endothelial dysfunction through inhibition of microRNA-133a (miR-133a). This study is to investigate the effects and the underlying mechanisms of statins on HFpEF. Here, we show that statins upregulate the expression of a circular RNA (circRNA-RBCK1) which is co-transcripted with the ring-B-box-coiled-coil protein interacting with protein kinase C-1 (RBCK1) gene. Simultaneously, statins increase activator protein 2 alpha (AP-2α) transcriptional activity and the interaction between circRNA-RBCK1 and miR-133a. Furthermore, AP-2α directly interacts with RBCK1 gene promoter in endothelial cells. In vivo, lovastatin improves diastolic function in male mice under HFpEF, which is abolished by loss function of endothelial AP-2α or circRNA-RBCK1. This study suggests that statins upregulate the AP-2α/circRNA-RBCK1 signaling to suppress miR-133a in cardiac endothelial cells and prevent diastolic dysfunction in HFpEF.

Retinal Vascular Changes in Heart Failure with Preserved Ejection Fraction Using Optical Coherence Tomography Angiography.

Background: Systemic microvascular regression and dysfunction are considered important underlying mechanisms in heart failure with preserved ejection fraction (HFpEF), but retinal changes are unknown. Methods: This prospective study aimed to investigate whether retinal microvascular and structural parameters assessed using optical coherence tomography angiography (OCT-A) differ between patients with HFpEF and control individuals (i.e., capillary vessel density, thickness of retina layers). We also aimed to assess the associations of retinal parameters with clinical and echocardiographic parameters in HFpEF. HFpEF patients, but not controls, underwent echocardiography. Macula-centered 6 × 6 mm volume scans were computed of both eyes. Results: Twenty-two HFpEF patients and 24 controls without known HFpEF were evaluated, with an age of 74 [68-80] vs. 68 [58-77] years (p = 0.027), and 73% vs. 42% females (p = 0.034), respectively. HFpEF patients showed vascular degeneration compared to controls, depicted by lower macular vessel density (p < 0.001) and macular ganglion cell-inner plexiform layer thickness (p = 0.025), and a trend towards lower total retinal volume (p = 0.050) on OCT-A. In HFpEF, a lower total retinal volume was associated with markers of diastolic dysfunction (septal e', septal and average E/e': R2 = 0.38, 0.36, 0.25, respectively; all p < 0.05), even after adjustment for age, sex, diabetes mellitus, or atrial fibrillation. Conclusions: Patients with HFpEF showed clear levels of retinal vascular changes compared to control individuals, and retinal alterations appeared to be associated with markers of more severe diastolic dysfunction in HFpEF. OCT-A may therefore be a promising technique for monitoring systemic microvascular regression and cardiac diastolic dysfunction.

Indole-3-Propionic Acid Protects Against Heart Failure With Preserved Ejection Fraction.

Heart failure with preserved ejection fraction (HFpEF) is a common but poorly understood form of heart failure, characterized by impaired diastolic function. It is highly heterogeneous with multiple comorbidities, including obesity and diabetes, making human studies difficult. Metabolomic analyses in a mouse model of HFpEF showed that levels of indole-3-propionic acid (IPA), a metabolite produced by gut bacteria from tryptophan, were reduced in the plasma and heart tissue of HFpEF mice as compared with controls. We then examined the role of IPA in mouse models of HFpEF as well as 2 human HFpEF cohorts. The protective role and therapeutic effects of IPA were confirmed in mouse models of HFpEF using IPA dietary supplementation. IPA attenuated diastolic dysfunction, metabolic remodeling, oxidative stress, inflammation, gut microbiota dysbiosis, and intestinal epithelial barrier damage. In the heart, IPA suppressed the expression of NNMT (nicotinamide N-methyl transferase), restored nicotinamide, NAD+/NADH, and SIRT3 (sirtuin 3) levels. IPA mediates the protective effects on diastolic dysfunction, at least in part, by promoting the expression of SIRT3. SIRT3 regulation was mediated by IPA binding to the aryl hydrocarbon receptor, as Sirt3 knockdown diminished the effects of IPA on diastolic dysfunction in vivo. The role of the nicotinamide adenine dinucleotide circuit in HFpEF was further confirmed by nicotinamide supplementation, Nnmt knockdown, and Nnmt overexpression in vivo. IPA levels were significantly reduced in patients with HFpEF in 2 independent human cohorts, consistent with a protective function in humans, as well as mice. Our findings reveal that IPA protects against diastolic dysfunction in HFpEF by enhancing the nicotinamide adenine dinucleotide salvage pathway, suggesting the possibility of therapeutic management by either altering the gut microbiome composition or supplementing the diet with IPA.

Ultrasound Findings with Bedside Ultrasound of Heart Failure with Preserved LVEF

Heart failure with preserved ejection fraction (HFpEF), formerly termed diastolic heart failure, constitutes nearly half the heart failure cases. Patients may exhibit acute heart failure symptoms, yet bedside echocardiograms might indicate normal left ventricular systolic function, potentially leading to the premature exclusion of heart failure as a differential diagnosis. This review delves into fundamental echocardiographic principles associated with diastolic dysfunction, crucial for recognition in HFpEF patients. Specifically, it explores the VALVE protocol, a stepwise approach tailored for the exigent nature of emergency departments. Acquiring accurate imaging windows in acutely ill patients poses a challenge, emphasizing the significance of precise measurements to avoid errors. Techniques such as E-point septal separation (EPSS) must be meticulously positioned for ejection fraction determinations. The discussion focuses on key metrics outlined in the VALVE protocol, encompassing ventricular function, annular tissue Doppler imaging (TDI) measurements, left atrium volume, mitral valve velocities, and the E/e0 ratio as indicators of diastolic dysfunction in HFpEF.

Abstract 15316: Neonatal Mesenchymal Stem Cells Ameliorate Cardiac Fibrosis and Diastolic Dysfunction in HFpEF via Expanding CD4 + CD25 + Foxp3 + T regs

Introduction: We have demonstrated that neonatal mesenchymal stem cells (nMSCs) extracted from human neonatal cardiac tissues and their exosomes, secretome, potently decrease cardiac fibrosis and improve cardiac function in the rat model of myocardial infarction. CD4 + CD25 + Foxp3 + (forkhead box 3) regulatory T cells (T regs ) are recently shown to regulate the formation of cardiac fibrosis in myocardial infarction. How nMSCs and secretome affect cardiac fibrosis and T regs of animals with heart failure with preserved ejection fraction (HFpEF) is unknown. Hypothesis: We hypothesized that nMSCs and secretome ameliorate cardiac fibrosis and diastolic dysfunction in HFpEF by regulating cardiac T regs . Methods: C57BL/6 mice were fed 60% high-fat diet and L-NAME to induce HFpEF or normal chow as a control. Left ventricular morphology and function were evaluated using an echocardiography. After the mice developed HFpEF, the animals were injected 10 6 nMSCs, 10 mg/kg of secretome, or equivalent amounts of vehicle as a control via tail vein. Myocardial fibrosis was quantified in trichrome-stained heart sections. Circulating and cardiac T regs were measured on a FACSCanto II cytometer. Fingolimod at a dose of 3 mg/kg was intraperitoneally injected daily for 18 days. Results: C57BL/6 mice fed high-fat diet and L-NAME for 5 weeks developed obesity, myocardial fibrosis, left ventricular hypertrophy and diastolic dysfunction (increases in mitral E/e’ ratio), and exercise intolerance with &gt; 50% of left ventricular ejection fraction. Compared to the HFpEF mice receiving vehicle, either nMSCs or secretome significantly decreased cardiac interstitial fibrosis and mitral E/e’ ratio and increased cardiac CD4 + CD25 + FoxP3 + T regs from 1 to 4 weeks post treatment. Importantly, either nMSCs or secretome enhanced the exercise speed and distance of the HFpEF mice. Fingolimod significantly decreased circulating and cardiac T regs in the HFpEF mice, which blocked the beneficial effects of nMSCs and secretome on cardiac fibrosis and diastolic function. Conclusions: nMSCs and secretome ameliorate cardiac fibrosis and diastolic dysfunction by expanding CD4 + CD25 + FoxP3 + T reg s in HFpEF. These results suggest that nMSCS and secretome have high therapeutic potential for HFpEF.

Abstract 13767: Cardiac Reprogramming Reduces Fibrosis and Improves Diastolic Dysfunction in Heart Failure With Preserved Ejection Fraction

Introduction: Heart failure with preserved ejection fraction (HFpEF) is a disease associated with high morbidity and mortality. Cardiac fibroblasts (CFs) contribute to HFpEF pathogenesis through interstitial/perivascular fibrosis and diastolic dysfunction; however, little is known regarding the mechanisms of the fibrosis, and therapies targeting it are lacking. Cardiac reprogramming is a promising approach for myocardial infarction by overexpressing Mef2c/Gata4/Tbx5/Hand2 (MGTH) in CFs, however, it remains undetermined whether cardiac reprogramming is effective in HFpEF. Aims: This study aims to determine the effectiveness of cardiac reprogramming in HFpEF. Methods: We generated a novel Tcf21iCre/Tomato/MGTH2A transgenic mouse system, where cardiac reprogramming can be induced in a druggable manner with lineage tracing. We first tested whether this system could improve the HFpEF mouse model. To determine the efficacy of cardiac reprogramming and the mechanisms of cardiac fibrosis in HFpEF, we performed multi-omics, including single cell RNA-sequencing (scRNA-seq), spatial transcriptomics, and epigenome analysis. Finally, we generated mouse lines that induce each of the reprogramming factors and investigated whether overexpression of a single factor alone could improve HFpEF, which we also examined in human CF. Results: We found that cardiac reprogramming reduced fibrosis and improved diastolic dysfunction in HFpEF. scRNA-seq and spatial transcriptomics revealed that multiple CF clusters upregulated fibrotic profiles to induce diffuse interstitial fibrosis, whereas the distinct profibrotic CF cluster contributed to perivascular fibrosis, which were reversed with cardiac reprogramming. Mechanistically, cardiac reprogramming changed the epigenetic landscape by recruiting Gata4 to loci. Notably, overexpression of only Gata4 improved HFpEF by reducing fibrosis and suppressed profibrotic signatures in human CFs without generating cardiomyocytes. Conclusions: Cardiac reprogramming may be a promising approach for HFpEF via anti-fibrosis.

Dapagliflozin Attenuates Heart Failure With Preserved Ejection Fraction Remodeling and Dysfunction by Elevating β-Hydroxybutyrate-activated Citrate Synthase.

Heart failure with preserved ejection fraction (HFpEF) is highly prevalent, accounting for 50% of all heart failure patients, and is associated with significant mortality. Sodium-glucose cotransporter subtype inhibitor (SGLT2i) is recommended in the AHA and ESC guidelines for the treatment of HFpEF, but the mechanism of SGLT2i to prevent and treat cardiac remodeling and dysfunction is currently unknown, hindering the understanding of the pathophysiology of HFpEF and the development of novel therapeutics. HFpEF model was induced by a high-fat diet (60% calories from lard) + N [w] -nitro- l -arginine methyl ester ( l -NAME-0.5 g/L) (2 Hit) in male Sprague Dawley rats to effectively recapture the myriad phenotype of HFpEF. This study's results showed that administration of dapagliflozin (DAPA, SGLT2 inhibitor) significantly limited the 2-Hit-induced cardiomyocyte hypertrophy, apoptosis, inflammation, oxidative stress, and fibrosis. It also improved cardiac diastolic and systolic dysfunction in a late-stage progression of HFpEF. Mechanistically, DAPA influences energy metabolism associated with fatty acid intake and mitochondrial dysfunction in HFpEF by increasing β-hydroxybutyric acid (β-OHB) levels, directing the activation of citrate synthase, reducing acetyl coenzyme A (acetyl-CoA) pools, modulating adenosine 5'-triphosphate production, and increasing the expression of mitochondrial oxidative phosphorylation system complexes I-V. In addition, following clinical DAPA therapy, the blood levels of β-OHB and citrate synthase increased and the levels of acetyl-CoA in the blood of HFpEF patients decreased. SGLT2i plays a beneficial role in the prevention and treatment of cardiac remodeling and dysfunction in HFpEF model by attenuating cardiometabolic dysregulation.

Abstract P2084: Adipose Tissues Thermogenesis Provides Cardioprotection Against Diastolic Dysfunction Developed In An Obesity-Induced HFpEF Model

Background: Heart failure with preserved ejection fraction (HFpEF) is a major source of morbidity/ mortality in the United States. A systemic inflammatory state established through underlying comorbidities, such as obesity, is thought to develop cardiac dysfunction in HFpEF. Clinical observations associate adiposity with diastolic dysfunction. Herein we demonstrate that adipose tissue is a direct source of diastolic dysfunction and reveal a cardioprotective role of adipose tissue thermogenesis in obesity related HFpEF. Methods and Results: Mice were treated with a β3-AR agonist, CL-316243 (CL), to induce thermogenesis, followed by HFpEF treatment consisting of HFD and L-NAME. Echocardiography revealed that this protected against HFpEF-induced diastolic dysfunction. Whole heart extracts and AT were then evaluated for the signature of β3-AR signaling. In the heart, no signature of β3-AR signaling was observed. In AT, though, various aspects of β3-AR induced thermogenesis were observed. Histological analysis revelated that CL treatment conferred resistance against obesity-induced whitening of interscapular brown adipose tissue (iBAT). Gene and protein expression changes of thermogenic regulators and lipid metabolism enzymes suggested greater thermogenic activity in CL-treated mice. Accordingly, indirect calorimetry showed enhanced energy expenditure, while regression analysis revealed that AT uniquely contributed to this. Transplantation experiments involving CL-treated donor AT and wild-type donors followed by echocardiographic measurements confirmed that the effects of CL were mediated through AT. This approach was complemented with a genetic model we previously used ( Ucp1 -Cre ERT2 ; Cdkn2a fl/fl ) to promote the expansion of beige AT. Here, beige AT expansion protected against diastolic dysfunction and cardiomyocyte hypertrophy developed during HFpEF. Conclusion: Our study indicates enhanced energy expenditure in AT is cardioprotective in obesity related HFpEF. Furthermore, identifying obese AT as a direct source of diastolic dysfunction highlights the therapeutic potential of targeting AT metabolism as a relevant source of diastolic dysfunction.

PO-01-172 METABOLIC SHIFT CONTRIBUTES TO SINOATRIAL NODE DYSFUNCTION IN HEART FAILURE WITH PRESERVED EJECTION FRACTION

Cardiometabolic alterations are frequent in patients with heart failure with preserved ejection fraction (HFpEF). We recently demonstrated sinoatrial node (SAN) dysfunction limits the chronotropic response in two distinct animal models of HFpEF; however, our new findings suggest a relationship between metabolism and SAN dysfunction. To identify metabolically-driven changes that facilitate SAN dysfunction in HFpEF. Male C57Bl6 mice fed with high fat (HFD) plus nitric oxide inhibitor (L-NAME) diet or regular chow served as HFpEF and controls, respectively. Transcriptomics, metabolomics, targeted quantitative proteomics of mitochondria, and optical mapping of SAN preparations from control and HFpEF-verified mice were performed. SAN from HFpEF-verified animals after 20 weeks of HFD+LNAME exhibited prolonged SAN recovery time (100%; P<0.05). A combination of multi-omics studies revealed changes in metabolic gene clusters suggesting a metabolic shift towards glycolysis as a source of energy. Supporting these findings, isolated mitochondria from HFpEF SANs showed depressed maximal mitochondrial respiration compared to controls (-42%; P<0.05). In addition, isolated pacemaker cells from HFpEF animals showed increased mitochondrial reactive oxygen species production compared to controls (+70%; P<0.05). Moreover, acute mitochondrial uncoupling induced by FCCP elicited a pronounced prolongation of SAN recovery time in HFpEF compared to controls (165%; P<0.05). Our results suggest that SAN dysfunction in HFpEF is closely associated with metabolic remodeling and energetic substrate shift. However, further studies are needed to establish the causality between metabolic changes and SAN dysfunction in HFpEF.

Catestatin Protects Against Diastolic Dysfunction by Attenuating Mitochondrial Reactive Oxygen Species Generation.

Background Catestatin has been reported as a pleiotropic cardioprotective peptide. Heart failure with preserved ejection fraction (HFpEF) was considered a heterogeneous syndrome with a complex cause. We sought to investigate the role of catestatin in HFpEF and diastolic dysfunction. METHODS AND RESULTS Administration of recombinant catestatin (1.5 mg/kg/d) improved diastolic dysfunction and left ventricular chamber stiffness in transverse aortic constriction mice with deoxycorticosterone acetate pellet implantation, as reflected by Doppler tissue imaging and pressure-volume loop catheter. Less cardiac hypertrophy and myocardial fibrosis was observed, and transcriptomic analysis revealed downregulation of mitochondrial electron transport chain components after catestatin treatment. Catestatin reversed mitochondrial structural and respiratory chain component abnormality, decreased mitochondrial proton leak, and reactive oxygen species generation in myocardium. Excessive oxidative stress induced by Ru360 abolished catestatin treatment effects on HFpEF-like cardiomyocytes in vitro, indicating the beneficial role of catestatin in HFpEF as a mitochondrial ETC modulator. The serum concentration of catestatin was tested among 81 patients with HFpEF and 76 non-heart failure controls. Compared with control subjects, serum catestatin concentration was higher in patients with HFpEF and positively correlated with E velocity to mitral annular e' velocity ratio, indicating a feedback compensation role of catestatin in HFpEF. Conclusions Catestatin protects against diastolic dysfunction in HFpEF through attenuating mitochondrial electron transport chain-derived reactive oxygen species generation. Serum catestatin concentration is elevated in patients with HFpEF, probably as a relatively insufficient but self-compensatory mechanism.

Vascular dysfunction in HFpEF: Potential role in the development, maintenance, and progression of the disease.

Heart failure with preserved ejection fraction (HFpEF) is a complex, heterogeneous disease characterized by autonomic imbalance, cardiac remodeling, and diastolic dysfunction. One feature that has recently been linked to the pathology is the presence of macrovascular and microvascular dysfunction. Indeed, vascular dysfunction directly affects the functionality of cardiomyocytes, leading to decreased dilatation capacity and increased cell rigidity, which are the outcomes of the progressive decline in myocardial function. The presence of an inflammatory condition in HFpEF produced by an increase in proinflammatory molecules and activation of immune cells (i.e., chronic low-grade inflammation) has been proposed to play a pivotal role in vascular remodeling and endothelial cell death, which may ultimately lead to increased arterial elastance, decreased myocardium perfusion, and decreased oxygen supply to the tissue. Despite this, the precise mechanism linking low-grade inflammation to vascular alterations in the setting of HFpEF is not completely known. However, the enhanced sympathetic vasomotor tone in HFpEF, which may result from inflammatory activation of the sympathetic nervous system, could contribute to orchestrate vascular dysfunction in the setting of HFpEF due to the exquisite sympathetic innervation of both the macro and microvasculature. Accordingly, the present brief review aims to discuss the main mechanisms that may be involved in the macro- and microvascular function impairment in HFpEF and the potential role of the sympathetic nervous system in vascular dysfunction.

Abstract 10273: Proteomic Analysis of Chronic Kidney Disease (CKD) in Heart Failure With Preserved Ejection Fraction (HFpEF)

Introduction: CKD is highly prevalent in HFpEF and is associated with increased morbidity and mortality. The biologic correlates of CKD in HFpEF are not well understood. We conducted a plasma proteomics analysis to assess biologic pathways associated with CKD in HFpEF. Methods: We utilized biosamples from TOPCAT for de novo measurements of 4776 plasma proteins using the SomaScan (version 4) proteomic assay, among 218 participants enrolled in the Americas. Linear regression and pathway analyses were used to study the relationship of proteins and pathways to the estimated glomerular filtration rate (eGFR). All regression estimates are standardized (per unit increase in z score) to facilitate comparisons. Results: After correction for multiple comparisons, we identified 73 proteins associated with eGFR. The strongest positively associated proteins were contactin-4, contactin-1, laminin-2, tenascin-X, and neuropilin-2. Proteins with the strongest negative association were transmembrane emp24 domain-containing protein 10, desmocollin-2, beta-2-microglobulin, neutrophil gelatinase-associated lipocalin, and cystatin C ( Figure 1 ). Biologic pathways associated with eGFR included complement system, acute phase response signaling, apoptosis, lysosomal expression and regulation, inflammation, fibrosis, chondroitin and dermatan sulfate degradation, notch signaling, and gluconeogenesis ( Figure 2 ). Conclusions: Using a broad proteomics approach, we identified novel biomarkers and pathways associated with renal dysfunction in HFpEF. Further research is required to assess whether targeting these pathways can ameliorate renal dysfunction in HFpEF patients.

Proteomic and phosphoproteomic profiling in heart failure with preserved ejection fraction (HFpEF)

Abstract Background The lack of therapies for HFpEF patients is a major unmet need, thus identifying cardiac specific pathways in HFpEF is a priority. Purpose Identify molecular features of protein and phosphoprotein in a murine model of HFpEF Methods Studies followed the principles of laboratory animal care (NIH Pub no. 85–23 revised 1985). HFpEF (N=4) was induced by NaCl drinking water, unilateral nephrectomy, and chronic aldosterone infusion (SAUNA) or saline (Sham; N=4) for 4 wks. Mice were euthanized and LV tissue was collected. Samples were homogenized, proteins extracted and subjected to digestion followed by phosphopeptide enrichment for proteomics and phosphoproteomics profiling. Results HFpEF mice had moderate hypertension (137.8±7.0 vs. Sham; 115.4±6.0mmHg; P&amp;lt;0.05), lung congestion (4.5±0.1 vs. 4.0±0.1; P&amp;lt;0.01), and LVH (3.7±0.1 vs. 3.3±0.1mg/g; P&amp;lt;0.05). ECHO showed normal LVEF with evidence of diastolic dysfunction in HFpEF mice. Mitral E velocity was reduced (1347±154 vs. 1971±284mm/s; P&amp;lt;0.05) and IVRT increased (24.3±2.6 vs. 14.4±1.6ms; P&amp;lt;0.05) vs. Sham. Protein and protein phosphosites from the LV were quantified by mass spectrometry. Analysis of the proteomics datasets revealed marked changes in sarcomere proteins, such as skeletal alpha (α)-actin (ACTA1; P=0.000039), beta (β)-myosin heavy chain (MYH7; P=0.006963) and myosin heavy chain 9 (MYH9; P=0.000408); the mitochondria-related proteins mitofusin 1 (MFN1; P=0.001059), mitochondrial dynamin like GTPase (aka optic atrophy protein 1, OPA1; P=0.046441) and transcription factor A mitochondrial (TFAM; P=0.005837); and the NAD-dependent protein deacetylase sirtuin-3 (SIRT3; P=0.000914). There was also a reduction in proteins involved in the oxidation of free fatty acid, pyruvate, and ketone bodies in the LV from HFpEF vs. Sham. Phosphoproteomics analysis also showed aberrant protein phosphorylation patterns linked to disparate subcellular compartments, ranging from sarcomere proteins (LIM domain-binding protein 3, LDB3; myozenin 2, Myoz2; titin, TTN), to nuclear-localized proteins (BAG family molecular chaperone regulator 3, BAG3; high mobility group protein HMG-I/HMG-Y, HMGA1) with known links to contractile dysfunction, LVH and/or cardiomyopathy. Additional GSEA analysis revealed the most relevant and enriched biological annotations in LV from HFpEF, to be processes involving immune system modulation and muscle contraction. Downregulated pathways were mainly related to multitude of GO terms associated with mitochondrial metabolism. Conclusion(s) This study presents the systematic, quantitative proteomics and phosphoproteomic analysis of the LV from the SAUNA HFpEF mice. We observed profound changes in proteins related to mitochondrial metabolism and function and heart contractile dysfunction in HFpEF which may be mediated by Sirtuin 3. Additional studies are warranted to investigate the specific role of Sirtuin 3 in mitochondrial metabolism and heart contractile dysfunction in HFpEF. Funding Acknowledgement Type of funding sources: Public grant(s) – EU funding. Main funding source(s): National Institute of Heath (NIH/ NHLBI)

Tubulin expression and modification in heart failure with preserved ejection fraction (HFpEF)

Diastolic dysfunction in heart failure with preserved ejection fraction (HFpEF) is characterised by increased left ventricular stiffness and impaired active relaxation. Underpinning pathomechanisms are incompletely understood. Cardiac hypertrophy and end stage heart disease are associated with alterations in the cardiac microtubule (MT) network. Increased amounts and modifications of α-tubulin associate with myocardial stiffness. MT alterations in HFpEF have not been analysed yet. Using ZSF1 obese rats (O-ZSF1), a validated HFpEF model, we characterised MT-modifying enzymes, quantity and tyrosination/detyrosination pattern of α-tubulin at 20 and 32 weeks of age. In the left ventricle of O-ZSF1, α-tubulin concentration (20 weeks: 1.5-fold, p = 0.019; 32 weeks: 1.7-fold, p = 0.042) and detyrosination levels (20 weeks: 1.4-fold, p = 0.013; 32 weeks: 1.3-fold, p = 0.074) were increased compared to lean ZSF1 rats. Tyrosination/α-tubulin ratio was lower in O-ZSF1 (20 weeks: 0.8-fold, p = 0.020; 32 weeks: 0.7-fold, p = 0.052). Expression of α-tubulin modifying enzymes was comparable. These results reveal new alterations in the left ventricle in HFpEF that are detectable during early (20 weeks) and late (32 weeks) progression. We suppose that these alterations contribute to diastolic dysfunction in HFpEF and that reestablishment of MT homeostasis might represent a new target for pharmacological interventions.

Abstract P3024: Mitochondrial Dysfunction Underlies Sinoatrial Node Dysfunction In Heart Failure With Preserved Ejection Fraction

Background: Metabolic comorbidities are frequent in patients with heart failure with preserved ejection fraction (HFpEF). We recently demonstrated that chronotropic incompetence and exercise intolerance are associated with intrinsic sinoatrial (SAN) dysfunction in animal models of HFpEF. However, there are no studies investigating whether metabolic dysfunction in HFpEF can lead to mitochondrial dysfunction in the SAN. Hypothesis: Our recent findings uncovering SAN pacemaker dysfunction in HFpEF upon metabolic stress led us to test the hypothesis that mitochondrial dysfunction may underlie this condition. Methods: Male C57Bl6 mice fed high fat diet (HFD) plus nitric oxide inhibitor (L-NAME) or regular chow served as HFpEF and control, respectively. Optical mapping, transcriptomics, targeted quantitative proteomics of mitochondrial proteins, mitochondrial respiration and reactive oxygen species (ROS) assays were conducted using explanted mouse SAN tissue or isolated pacemaker cells. Results: SAN from HFpEF-verified animals after 20 weeks of HFD+LNAME exhibited prolonged SAN recovery time after pacing (100%; P&lt;0.05). RNA sequencing revealed augmentation of gene clusters related to metabolism (i.e. ucp and pdk4 ) and extracellular matrix expansion (i.e. col, postn and Cilp1 ). Targeted proteomics validated SAN RNA-sequencing findings, showing upregulation of proteins related to ROS scavenging, fatty acid transport and TCA/OXPHOS (i.e. SOD, CPT2 and DHSA/B). Depressed maximal mitochondrial respiration was seen in isolated mitochondria from SAN of HFpEF animals compared to controls (-42%; P&lt;0.05). This was supported by increased generation of mitochondrial ROS in single pacemaker cells from HFpEF SAN (70%; P&lt;0.05). Acute mitochondrial dysfunction induced by a mitochondrial uncoupler (FCCP) elicited a pronounced prolongation of SAN recovery time in HFpEF compared to controls (165%; P&lt;0.05). Conclusion: Our results indicate that mitochondrial function is depressed in SAN from HFpEF animals, and is associated with SAN pacemaker dysfunction. Further studies are required to test cause-and-effect and to evaluate potential therapeutic strategies targeting mitochondrial pathways for HFpEF associated abnormalities of sinus rhythm.

The relationship between ambulatory arterial stiffness index and left ventricular diastolic dysfunction in HFpEF: a prospective observational study

BackgroundThe relationship between ambulatory arterial stiffness index (AASI) and left ventricular diastolic dysfunction (LVDD) in patients with heart failure with preserved ejection fraction (HFpEF) is unknown. We aimed to investigate the association between the AASI and LVDD in HFpEF.MethodsWe prospective enrolled consecutive patients with HFpEF in Chongqing, China. Twenty-four-hour ambulatory blood pressure monitoring (24 h-ABPM) and echocardiography were performed in each patient. AASI was obtained through individual 24 h-ABPM. The relationship between AASI and LVDD was analyzed.ResultsA total of 107 patients with HFpEF were included. The mean age was 68.45 ± 14.02 years and 63 (59%) were women. The patients were divided into two groups according to the upper normal border of AASI (0.55). AASI > 0.55 group were more likely to be older, to have higher mean systolic blood pressure and worsen left ventricular diastolic function than AASI group ≤ 0.55. AASI was closely positive related to the diastolic function parameters, including mean E/e′ (r = 0.307, P = 0.001), septal E/e′ (r = 0.290, P = 0.002), lateral E/e′ (r = 0.276, P = 0.004) and E (r = 0.274, P = 0.004). After adjusting for conventional risk factors, AASI was still an independent risk factors of mean E/e′ > 10 in patients with HFpEF (OR: 2.929, 95%CI: 1.214–7.064, P = 0.017), and the association between AASI and mean E/e′ > 14 was reduced (OR: 2.457, 95%CI: 1.030–5.860, P = 0.043). AASI had a partial predictive value for mean E/e′ > 10 (AUC = 0.691, P = 0.002), while the predictive value for mean E/e′ > 14 was attenuated (AUC = 0.624, P = 0.034).ConclusionAASI was positive related to E/e′ in HFpEF and might be an independent risk factor for the increase of mean E/e′.

3D quantitative characterization of mouse coronary capillary network in heart failure with preserved ejection fraction

Heart failure with preserved ejection fraction (HFpEF) is a cardiovascular disease characterized by telediastolic dysfunction. Main risks factors include advanced age (> 67 years old), sex (women being more susceptible to develop the disease), and several comorbidities, in particular obesity and diabetes. HFpEF has been shown to be associated with the rarefaction of the coronary small vessels. The aim of our study was to characterize quantitatively the 3D topological reorganization of the coronary microvasculature of the capillary coronary network in a mouse model of HFpEF. Experiments were done on 14-week old female C57BL/Ks Leprdb/db mice, accumulating risk factors of human HFpEF ( n = 7). Leprdb/db mice develop early obesity and type 2 diabetes, and C57BL/Ks mice have an increased risk of renal dysfunction. Eight-week old male C57BL/6J mice were used as normal control ( n = 7). 3D imaging was done by light-sheet microscopy on iDISCO+ optical cleared hearts after lectin-labeling of the capillaries. Image processing consisted of skeletonization and distance mapping of greyscale segmented images using ImageJ software. Capillary network in the left ventricle (LV), right ventricle (RV), and septum (S) was characterized by the volume network density, the fractal dimension, the average length, diameter, and tortuosity of the segments, and (normalized per mm 3 of cardiac tissue) the numbers of segments and nodes and the total network length. Data are given as mean ± SD. Differences were significant if P < 0.05. Compared to controls, Leprdb/db mice LV showed a significantly reduced volume density (40 ± 6 vs. 49 ± 9%), number of segments (402,000 ± 68,000 vs. 518,000 ± 110,000), number of nodes (220,000 ± 39,000 vs. 280,000 ± 60,000) and total length (55 ± 1.2 vs. 54 ± 1.7 m). LV capillaries were significantly more tortuous than control ones. Leprdb/db mice RV showed a significantly reduced fractal dimension (2.4 ± 0.03 vs. 2.4 ± 0.02), number of segments (339,000 ± 24,000 vs. 401,000 ± 43,000), number of nodes (180,000 ± 14,000 vs. 220,000 ± 25,000) and total length (53 ± 0.6 vs. 54 ± 1.3 m). RV capillaries were significantly larger than control ones. S capillary network was not different from control ( Fig. 1 ). Our study showed that LV capillary network presented signs of rarefaction and structural alterations that may contribute to LV dysfunction in HFpEF. It also evidenced alterations of the RV capillary network architecture, suggesting a possible impairment of RV function.

Proinflammatory T Cells with Downregulated Unfolded Protein Response Genes Contribute to Experimental Heart Failure with Preserved Ejection Fraction (HFpEF)

IntroductionHeart Failure with Preserved Ejection Fraction (HFpEF) is an uncurable widespread syndrome characterized by impaired cardiac relaxation (diastolic function), preserved cardiac contractility (systolic function), and the concordance of several comorbidities, such as obesity and hypertension. While T cell inflammation is known to contribute to HF with reduced cardiac contractility (HF with reduced EF), whether T cell inflammation occurs in HFpEF remains elusive. Recently, downregulation of the unfolded protein response (UPR) in cardiomyocytes was identified as a cardiometabolic HFpEF‐unique molecular signature. Interestingly, downregulation of the UPR has been reported to improve T cell anti‐tumor inflammatory responses. We hypothesized T cells contribute to diastolic dysfunction in HFpEF following T cell UPR downregulation in response to metabolic and nitrosative stress.MethodsWe modeled cardiometabolic HFpEF in male C57/Bl6 (WT), Nur77‐GFP mice, reporter mice for T cell antigen engagement, and T cell receptor‐a (Tcra−/−) mice using high‐fat diet (HFD) and L‐NAME‐supplemented drinking water to mimic obesity and hypertension, respectively. Mice fed standard chow (STD) were used as controls. Invasive hemodynamic analyses were used to measure cardiac function. Immune populations in the heart, mediastinal lymph nodes (MdLN), and spleen were characterized using flow cytometry. CD4+ T cells from MdLN and spleen were isolated using magnetic‐assisted cell sorting, and UPR gene expression was assessed using qPCR.ResultsWe observed significant increases in cardiac CD4+ T cells in WT mice fed HFD/L‐NAME, concordant with diastolic dysfunction and preserved EF, compared to STD mice. We found increased effector CD4+CD62LloCD44hiIFNγ+ T cells in spleens and MdLN isolated from WT HFD/L‐NAME mice compared to STD controls. Nur77‐GFP mice revealed no antigen recognition by CD4+ T cells in the heart following HFD/L‐NAME. Tcra−/−mice fed HFD/L‐NAME did not develop diastolic dysfunction or cardiomyocyte hypertrophy. Strikingly, qPCR analysis of splenic T cells of WT HFD/L‐NAME mice revealed significantly decreased expression of spliced X box‐binding protein 1 (XBP1s), compared to controls, with no significant changes in unspliced XBP1 or activating transcription factor 4 (ATF4). ATF6 and C/EBP homologous protein (CHOP) gene expression was also reduced compared to T cells from control mice.ConclusionOur data demonstrate that diastolic dysfunction and cardiomyocyte hypertrophy are T cell dependent in preclinical cardiometabolic HFpEF and reveal downregulation of two branches of the T cell UPR as a potential novel mechanism that contributes to T cell inflammation in HFpEF.

EN-510-03 SINOATRIAL NODE DYSFUNCTION IN HEART FAILURE WITH PRESERVED EJECTION FRACTION

Under normal conditions, the human heart beats over 2 billion times during an average lifetime due to rhythmic electrical impulses initiated in the sinoatrial node (SAN). In disease conditions, such as heart failure with preserved ejection fraction (HFpEF), failure to maintain and regulate the heart rate due to SAN dysfunction can occur. We recently identified latent SAN dysfunction in animal models of cardiometabolic HFpEF. However, little is known about the metabolic imbalances underlying SAN dysfunction in HFpEF. To identify metabolically-driven functional and transcriptomic changes in HFpEF SAN. Male C57Bl6 mice fed with high fat (HFD) plus nitric oxide inhibitor (L-NAME) diet or regular chow served as HFpEF and control, respectively. RNA sequencing and optical mapping of SAN preparations from control and HFpEF-verified mice were performed. After 20 weeks of HFD+L-NAME, the sinus node recovery time was significantly prolonged in HFpEF mice compared to controls (162 ± 25 vs 88 ± 19 ms; p<0.05). Transcriptome profiling of SAN tissue revealed a significant enhancement of multiple disease-associated genes, with striking changes in extracellular matrix genes and metabolic pathways. In concordance with the observed fibrotic remodeling, conduction velocity (CV) was significantly lower in SAN of HFpEF mice compared to controls (6.4 ± 0.5 vs 13.1 ± 1.7 cm/s; p<0.05). Moreover, although β-adrenergic receptor stimulation accelerated the CV in control SAN, failed to do so in HFpEF animals (20 ± 3.1 vs 10.2 ± 1.1 cm/s; p<0.05). Our results using a cardiometabolic HFpEF model indicate that SAN dysfunction is closely associated with molecular changes in metabolic pathways and extracellular matrix remodeling associated genes. Understanding the metabolic control of the SAN may open new therapeutic targets for HFpEF associated SAN dysfunction.