Overload-induced Heart Failure In Mice Research Articles

S14G-humanin confers cardioprotective effects against chronic adrenergic and pressure overload-induced heart failure in mice

S14G-humanin (HNG), an analog of the mitochondria-derived peptide humanin, has demonstrated protective effects against various cardiovascular diseases. However, the specific pharmacological effects of HNG in heart failure (HF) have not been previously reported. Therefore, in this study, we aimed to investigate the potential protective effect of HNG in HF using a mouse model. HF was induced in mice through intraperitoneal injection of isoproterenol or transverse aortic constriction, followed by separate administration of HNG to assess its therapeutic impact. Our results revealed that HNG treatment significantly delayed the onset of cardiac dysfunction and structural remodeling in the HF mouse model. Furthermore, HNG administration was associated with reduced infiltration of inflammatory cells, improved myocardial fibrosis, and attenuation of cardiomyocyte apoptosis in the treated cardiac tissues. Additionally, we identified the involvement of the transforming growth factor-beta signaling pathway in the beneficial effects of HNG in isoproterenol-induced HF mice. Collectively, these findings underscore the therapeutic potential of HNG in preventing the progression of HF, as demonstrated in two distinct HF mouse models.

Asiatic acid protects against pressure overload-induced heart failure in mice by inhibiting mitochondria-dependent apoptosis

BackgroundMitochondrial dysfunction and subsequent cardiomyocyte apoptosis significantly contribute to pressure overload-induced heart failure (HF). A highly oxidative environment leads to mitochondrial damage, further exacerbating this condition. Asiatic acid (AA), a proven antioxidant and anti-hypertrophic agent, might provide a solution, but its role and mechanisms in chronic pressure overload-induced HF remain largely unexplored. MethodsWe induced pressure overload in mice using transverse aortic constriction (TAC) and treated them with AA (100 mg/kg/day) or vehicle daily by oral gavage for 8 weeks. The effects of AA on mitochondrial dysfunction, oxidative stress-associated signaling pathways, and overall survival were evaluated. Additionally, an in vitro model using hydrogen peroxide-exposed neonatal rat cardiomyocytes was established to further investigate the role of AA in oxidative stress-induced mitochondrial apoptosis. ResultsAA treatment significantly improved survival and alleviated cardiac dysfunction in TAC-induced HF mice. It preserved mitochondrial structure, reduced the LVW/BW ratio by 20.24%, mitigated TAC-induced mitochondrial-dependent apoptosis by significantly lowering the Bax/Bcl-2 ratio and cleaved caspase-9/3 levels, and attenuated oxidative stress. AA treatment protected cardiomyocytes from hydrogen peroxide-induced apoptosis, with concurrent modulation of mitochondrial-dependent apoptosis pathway-related proteins and the JNK pathway. ConclusionsOur findings suggest that AA effectively combats chronic TAC-induced and hydrogen peroxide-induced cardiomyocyte apoptosis through a mitochondria-dependent mechanism. AA reduces cellular levels of oxidative stress and inhibits the activation of the JNK pathway, highlighting its potential therapeutic value in the treatment of HF.

The Aryl Hydrocarbon Receptor Ligand FICZ Improves Left Ventricular Remodeling and Cardiac Function at the Onset of Pressure Overload-Induced Heart Failure in Mice.

Adverse ventricular remodeling is the heart’s response to damaging stimuli and is linked to heart failure and poor prognosis. Formyl-indolo [3,2-b] carbazole (FICZ) is an endogenous ligand for the aryl hydrocarbon receptor (AhR), through which it exerts pleiotropic effects including protection against inflammation, fibrosis, and oxidative stress. We evaluated the effect of AhR activation by FICZ on the adverse ventricular remodeling that occurs in the early phase of pressure overload in the murine heart induced by transverse aortic constriction (TAC). Cardiac structure and function were evaluated by cardiac magnetic resonance imaging (CMRI) before and 3 days after Sham or TAC surgery in mice treated with FICZ or with vehicle, and cardiac tissue was used for biochemical studies. CMRI analysis revealed that FICZ improved cardiac function and attenuated cardiac hypertrophy. These beneficial effects involved the inhibition of the hypertrophic calcineurin/NFAT pathway, transcriptional reduction in pro-fibrotic genes, and antioxidant effects mediated by the NRF2/NQO1 pathway. Overall, our findings provide new insight into the role of cardiac AhR signaling in the injured heart.

Abstract P106: Development Of An Oral Drug Delivery System For The Cardio-protective Neuropeptide Alpha-calcitonin Gene Related Peptide

Alpha-calcitonin gene related peptide (α-CGRP) is a promising neuropeptide for the treatment of cardiovascular disease. We have developed an alginate biomaterial-based delivery system which increases the peptide’s bioavailability and have shown that subcutaneous delivery of alginate-CGRP microcapsules significantly improved cardiac function in pressure overload-induced heart failure in mice. The goal of this study is to develop efficient alginate microcapsule formulations for oral delivery of α-CGRP. An electrospray method was used to prepare four different formulations of alginate-CGRP microcapsules all of 200 μm diameter- i)- alginate-CGRP microcapsules, ii)- alginate-CGRP microcapsules with UV-light exposure, iii)- poly-L-ornithine (PLO) coated alginate-CGRP microcapsules, and iv)- PLO alginate-CGRP microcapsules with UV-light exposure. The stability of the microcapsules in the digestive tract was evaluated in deionized water, simulated gastric fluid (SGF; pH 1.2), and simulated intestinal fluid (SIF; pH 6.8). Over time, the size of all four microcapsule formulations remained almost unchanged in SGF, however all four of the microcapsule formulations swelled in presence of SIF. Compared to deionized water, the size of alginate-CGRP microcapsules after 6 h incubation in SIF increased by 1.7 fold. Since all four formulations yielded similar results, we chose to further study the alginate-CGRP microcapsules in vivo . To determine the bioactive nature of the released peptide, alginate microcapsules containing α-CGRP in doses of 1, 3, and 10 (mg/kg b.wt) were fed to male C57BL/6 mice via oral gavage. Systolic blood pressure (SBP) was subsequently measured by a tail-cuff method. α-CGRP microcapsules reduced SBP in a time-dependent manner. Alginate-CGRP at 1 and 3 mg/kg lowered the SBP by 25 mmHg for up to 4 h and 48 h, respectively. However, 10 mg/kg alginate-CGRP initially reduced SBP to undetectable levels which ultimately returned to baseline level by day 7. These studies indicate that alginate microcapsules can withstand the low pH of the stomach and the release of the peptide is bioactive in vivo . Thus, alginate microcapsules may provide an ideal formulation to deliver α-CGRP orally for the long-term treatment of cardiac diseases.

Abstract 14834: Protective Role of Cardiomyocyte-derived Ddt in Ischemic Cardiomyopathy

Background: D-dopachrome tautomerase (DDT), the only homolog of macrophage migration inhibitory factor (MIF), is a cytokine highly expressed in cardiomyocytes and exerts autocrine-paracrine effects by signaling through the CD74 receptor. Endogenous DDT and MIF prevent acute ischemia-reperfusion injury and pressure overload-induced heart failure in mice. This study investigated whether endogenous cardiomyocyte DDT has a role in ischemic cardiomyopathy (ICM). Methods: LV tissue was obtained from patients with ICM during heart transplantation and from non-transplanted donor hearts. Plasma DDT concentrations were measured in heart failure outpatients with ICM. Cardiomyocyte-specific DDT knockout (cKO) and littermate control (CON) mice underwent MI or sham surgery. Serial echocardiography was performed to assess LV remodeling after MI or sham surgery. Tissue from the non-infarct region was analyzed 3 days and 4 weeks after MI or sham surgery for histology and molecular studies. Results: Cardiac DDT mRNA and protein expression were reduced in LV from patients transplanted for ICM (n=8). Plasma DDT concentrations below the median value were associated with worse survival in ICM outpatients (p<0.05, n=32). In mice, baseline LV function was similar in DDT cKO and CON after sham surgery and 3 days post-MI. However, DDT cKO mice developed more rapid LV dilatation and decreased LV ejection fraction and stroke volume as early as 1-week post-MI (n=4-6/group, all P<0.05). The DDT cKO mice had smaller cardiomyocyte cross-sectional area 4 weeks after MI (p <0.05), as well as early diminished phosphorylation of mTOR and S6-kinase (3 days post-MI). They also showed increased apoptosis 3 days post-MI and an early increase in p38 MAP kinase activation. Conclusion: Cardiomyocyte-derived DDT prevents adverse cardiac remodeling in ICM, potentially through modulating mTOR/S6 kinase (adaptive hypertrophy) and p38 MAP kinase (limiting apoptosis). Down-regulation of DDT in patients with ICM may contribute to the pathogenesis of advanced heart failure.

Stimulation of exosome biogenesis by adiponectin, a circulating factor secreted from adipocytes.

Adiponectin is an adipocyte-derived circulating factor that protects various organs and tissues. Such a pleiotropic action mechanism has not yet been fully explained. Clinically important multimer adiponectin existing in serum bound to cells expressing T-cadherin, a glycosylphosphatidylinositol-anchored cadherin, but not to the cells expressing other known receptors, AdipoRs or calreticulin. Adiponectin bound to the cell-surface, accumulated inside of multivesicular bodies through T-cadherin, and increased exosome biogenesis and secretion from the cells. Such increased exosome production accompanied the reduction of cellular ceramides in endothelial cells and mouse aorta, and enhanced skeletal muscle regeneration. Significantly lower plasma exosome levels were found in mice genetically deficient in either adiponectin or T-cadherin. Therapeutic effects of mesenchymal stem cells (MSCs) for a pressure overload-induced heart failure in mice required the presence of adiponectin in plasma, T-cadherin expression and exosome biogenesis in MSCs themselves, accompanying an increase of plasma exosomes. Essentially all organs seem to have MSCs and/or their related somatic stem cells expressing T-cadherin. Our recent studies suggested the importance of exosome-stimulation by multimer adiponectin in its well-known pleiotropic organ protections.

Modulation of P2Y6R expression exacerbates pressure overload-induced cardiac remodeling in mice

Cardiac tissue remodeling caused by hemodynamic overload is a major clinical outcome of heart failure. Uridine-responsive purinergic P2Y6 receptor (P2Y6R) contributes to the progression of cardiovascular remodeling in rodents, but it is not known whether inhibition of P2Y6R prevents or promotes heart failure. We demonstrate that inhibition of P2Y6R promotes pressure overload-induced sudden death and heart failure in mice. In neonatal cardiomyocytes, knockdown of P2Y6R significantly attenuated hypertrophic growth and cell death caused by hypotonic stimulation, indicating the involvement of P2Y6R in mechanical stress-induced myocardial dysfunction. Unexpectedly, compared with wild-type mice, deletion of P2Y6R promoted pressure overload-induced sudden death, as well as cardiac remodeling and dysfunction. Mice with cardiomyocyte-specific overexpression of P2Y6R also exhibited cardiac dysfunction and severe fibrosis. In contrast, P2Y6R deletion had little impact on oxidative stress-mediated cardiac dysfunction induced by doxorubicin treatment. These findings provide overwhelming evidence that systemic inhibition of P2Y6R exacerbates pressure overload-induced heart failure in mice, although P2Y6R in cardiomyocytes contributes to the progression of cardiac fibrosis.

Molecular Mechanism of HSF1-Upregulated ALDH2 by PKC in Ameliorating Pressure Overload-Induced Heart Failure in Mice.

Evidences abound that HSF1 and ALDH2 are of cardioprotective effect, yet there is still no report on whether HSF1 can regulate ALDH2 to delay the occurrence of heart failure. We first established the pressure overload-induced heart failure model of mice by transverse aortic constriction (TAC) and discovered that, in the forming period of heart failure, changes of HSF1 and ALDH2 expression recorded the consistent trend. When HSF1 was upregulated/downregulated to delay/promote the occurrence of heart failure, PKC and ALDH2 also showed increased/decreased expression. And when ALDH2 was upregulated/downregulated, the role of HSF1 in delaying the occurrence of heart failure strengthened/weakened. Next, we used mechanical stretch to establish a pressure-stimulated myocardial hypertrophy model and discovered an increased expression of both HSF1 and ALDH2. When HSF1 was upregulated/downregulated to increase/decrease the expression of myocardial hypertrophy gene beta-MHC, PKC and ALDH2 recorded an increased/decreased expression. When an inhibitor was used to downregulate the expression of PKC in cardiomyocytes, we found that the role of HSF1 in upregulating ALDH2 beta-MHC weakened. These findings suggest that HSF1 can upregulate the expression of ALDH2 via PKC to promote pressure-stimulated myocardial compensatory hypertrophy, which is an important molecular pathway for HSF1 to ameliorate heart failure.

The Synthetic Curcumin Analogue GO-Y030 Effectively Suppresses the Development of Pressure Overload-induced Heart Failure in Mice

Curcumin is a naturally occurring p300-histone acetyltransferase (p300-HAT) inhibitor that suppresses cardiomyocyte hypertrophy and the development of heart failure in experimental animal models. To enhance the therapeutic potential of curcumin against heart failure, we produced a series of synthetic curcumin analogues and investigated their inhibitory activity against p300-HAT. The compound with the strongest activity was further evaluated to determine its effects on cardiomyocyte hypertrophy and pressure overload-induced heart failure in mice. We synthesised five synthetic curcumin analogues and found that a compound we have named GO-Y030 most strongly inhibited p300-HAT activity. Furthermore, 1 μM GO-Y030, in a manner equivalent to 10 µM curcumin, suppressed phenylephrine-induced hypertrophic responses in cultured cardiomyocytes. In mice undergoing transverse aortic constriction surgery, administration of GO-Y030 at a mere 1% of an equivalently-effective dose of curcumin significantly attenuated cardiac hypertrophy and systolic dysfunction. In addition, this low dose of GO-Y030 almost completely blocked histone H3K9 acetylation and eliminated left ventricular fibrosis. A low dose of the synthetic curcumin analogue GO-Y030 effectively inhibits p300-HAT activity and markedly suppresses the development of heart failure in mice.

P66Shc Deletion Ameliorates Oxidative Stress and Cardiac Dysfunction in Pressure Overload-Induced Heart Failure

p66Shc is a redox enzyme that plays an important role in the response of oxidative stress and the p53-dependent apoptosis. The expression level of p66Shc has a negative correlation with the resistance of oxidative stress in vivo and in vitro. We aim to demonstrate the function of p66Shc in pressure overload-induced heart failure. The pressure overload-induced heart failure was induced in mice by transverse aortic constriction (TAC). Cardiac dysfunction was shown by transthoracic echocardiography. Western blot was used to check the protein levels of phosphodiesterase type 5 (PDE5) and ventricular oxidative stress markers. Superoxide dismutase (SOD) mimetic M40401 and PDE5 inhibitor sildenafil were used in the treatment of mice. The deletion of p66Shc attenuated cardiac dysfunction and oxidative stress in pressure overload-induced heart failure. p66Shc deletion also decreased the expression of ventricular oxidative stress markers and enhanced PKG signaling by promoting the expression of PDE5. M40401 and sildenafil attenuated the TAC-induced cardiac dysfunction and oxidative stress in p66Shc overexpression mice. Our findings suggest that p66Shc participates in the regulation of cardiac dysfunction and oxidative stress in TAC-derived pressure overload-induced heart failure in mice, and SOD and PDE5 are molecules downstream of p66Shc in this regulatory process.

Cholesterol lowering attenuates pressure overload-induced heart failure in mice with mild hypercholesterolemia.

Epidemiological studies support a strong association between non-high-density lipoprotein cholesterol levels and heart failure incidence. The objective of the current study was to evaluate the effect of selective cholesterol lowering adeno-associated viral serotype 8 (AAV8)-mediated low-density lipoprotein receptor (LDLr) gene transfer on cardiac remodelling and myocardial oxidative stress following transverse aortic constriction (TAC) in female C57BL/6 LDLr-/- mice with mild hypercholesterolemia. Cholesterol lowering gene transfer resulted in a 65.9% (p<0.0001) reduction of plasma cholesterol levels (51.2 ± 2.2 mg/dl) compared to controls (150 ± 7 mg/dl). Left ventricular wall area was 11.2% (p<0.05) lower in AAV8-LDLr TAC mice than in control TAC mice. In agreement, pro-hypertrophic myocardial proteins were potently decreased in AAV8-LDLr TAC mice. The degree of interstitial fibrosis and perivascular fibrosis was 31.0% (p<0.001) and 29.8% (p<0.001) lower, respectively, in AAV8-LDLr TAC mice compared to control TAC mice. These structural differences were associated with improved systolic and diastolic function and decreased lung congestion in AAV8-LDLr TAC mice compared to control TAC mice. Cholesterol lowering gene therapy counteracted myocardial oxidative stress and preserved the potential for myocardial fatty acid oxidation in TAC mice. In conclusion, cholesterol lowering gene therapy attenuates pressure overload-induced heart failure in mice with mild hypercholesterolemia.

Abstract 255: Novel Role for Free Fatty Acid Receptor 4 in Response to Pathologic Pressure Overload-Induced Heart Failure in Mice

Free fatty acid receptor 4 (Ffar4, GPR120) is a GPCR for long chain fatty acids, including omega-3 polyunsaturated fatty acids (ω3-PUFAs). Previously, we demonstrated that eicosapentaenoic acid (EPA), an ω3-PUFA, prevents interstitial fibrosis and contractile dysfunction in pressure overload heart failure (transverse aortic constriction, TAC), but EPA was not incorporated into cardiac myocytes or fibroblasts, the traditional mechanism of action. Therefore, we hypothesized that Ffar4 is necessary for the anti-fibrotic and cardioprotective effects of EPA. In fact, we previously found that in primary cardiac fibroblasts, Ffar4 was sufficient and necessary to prevent TGFβ1-induced fibrosis. Here, we were surprised to find that Ffar4 also has important ω3-independent effects in cardiac myocytes in vivo . In mice with systemic deletion of Ffar4 (Ffar4KO mice) on a standard, corn oil diet (no EPA supplementation), TAC induced significantly more cardiac hypertrophy and more systolic (ejection fraction) and diastolic dysfunction (E/A ratio), but without excess fibrosis after 4 weeks. Mechanistically, transcriptome analysis in cardiac myocytes 3 days post-TAC indicated a deficiency of transcription in Ffar4KO myocytes. Sorting based on gene ontology indicated that genes functionally categorized as cell death and inflammation, including ilrn1 , hmox1 , Il33 , and ptgs2 (cox2) were not induced in the Ffar4KO myocytes. Ffar4 also activates cytoplasmic phospholipase A2 (cPLA2) inducing the production of oxylipins, and systemic deletion of cPLA2 induces exaggerated hypertrophy during development and following TAC. TAC increased cPLA2 expression in WT, but not Ffar4KO cardiac myocytes and oxylipin production was reduced in Ffar4KO mice. Finally, we confirmed that Ffar4 is expressed in human heart and down-regulated in heart failure. In summary, our data identify a novel, ω3-independent cardioprotective role for Ffar4 in cardiac myocytes.

Administration of a TLR9 Inhibitor Attenuates the Development and Progression of Heart Failure in Mice

Mitochondrial deoxyribonucleic acid, containing the unmethylated cytidine-phosphate-guanosine motif, stimulates Toll-like receptor 9 to induce inflammation and heart failure. A small chemical, E6446 [(6-[3-(pyrrolidin-1-yl)propoxy)-2-(4-(3-(pyrrolidin-1-yl)propoxy)phenyl]benzo[d]oxazole)], is a specific Toll-like receptor 9 inhibitor in cardiomyocytes. In this study, we showed that E6446 exerts beneficial effects for the prevention and treatment of pressure overload-induced heart failure in mice. When administered before the operation andchronically thereafter, E6446 prevented the development of left ventricular dilatation as well as cardiac dysfunction, fibrosis, and inflammation. Furthermore, when administered after the manifestation of cardiac dysfunction, E6446 slowed progression of cardiac remodeling. Thus, the inhibitor may be a novel therapeutic agent for treating patients with heart failure.

Loss of Endogenous HMGB2 Promotes Cardiac Dysfunction and Pressure Overload-Induced Heart Failure in Mice.

The rapid increase in the number of heart failure (HF) patients in parallel with the increase in the number of older people is receiving attention worldwide. HF not only increases mortality but decreases quality of life, creating medical and social problems. Thus, it is necessary to define molecular mechanisms underlying HF development and progression. HMGB2 is a member of the high-mobility group superfamily characterized as nuclear proteins that bind DNA to stabilize nucleosomes and promote transcription. A recent in vitro study revealed that HMGB2 loss in cardiomyocytes causes hypertrophy and increases HF-associated gene expression. However, it's in vivo function in the heart has not been assessed. Methods and Results: Western blotting analysis revealed increased HMGB2 expression in heart tissues undergoing pressure overload by transverse aorta constriction (TAC) in mice. Hmgb2 homozygous knockout (Hmgb2-/-) mice showed cardiac dysfunction due to AKT inactivation and decreased sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2a activity. Compared to wild-type mice, Hmgb2-/- mice had worsened cardiac dysfunction after TAC surgery, predisposing mice to HF development and progression. This study demonstrates that upregulation of cardiac HMGB2 is an adaptive response to cardiac stress, and that loss of this response could accelerate cardiac dysfunction, suggesting that HMGB2 plays a cardioprotective role.

Alpha-Lipoic Acid Protects Against Pressure Overload-induced Heart Failure &lt;i&gt;via&lt;/i&gt; ALDH2-Dependent Nrf1-FUNDC1 Signaling

Background: Acetaldehyde dehydrogenase 2 (ALDH2) could protect the heart against pressure overload-induced heart failure in mice. Alpha-lipoic acid (α-LA), a well-known antioxidant, was capable of activating ALDH2. However, impact of α-LA on pressure overload-induced heart failure remains unknown now. Methods: α-LA on pressure overload-induced heart failure were assessed using ALDH2 knockout (ALDH2-/-) mice and primary neonatal rat cardiomyocytes (NRCMs). TAC-induced left ventricular hypertrophy and contractile function were assessed by echocardiography and cell shortening/relengthening system. Heart histological measurement were using H&E, WGA, Masson’s trichrome, DHE and TUNEL. Mitochondrial mass and morphology were visualized using TEM. The target proteins and their interactions was investigated by bioinformatic analysis, luciferase reporter and chromatin immunoprecipitation (ChIP) assays. Findings: In vivo, we found that supplement of α-LA significantly reduced transverse aortic constriction (TAC)-induced LV hypertrophy and dysfunction in wild-type mice, as evidenced by preserved LV ejection fraction, a more favorable histologic change and restoration of mitochondrial normality. Whereas, α-LA did not affect TAC induced LV hypertrophy and dysfunction in ALDH2 gene knockout (KO) mice. In molecular level, α-LA significantly restored ALDH2 activity and expression as well as upregulated the expression of FUN14 domain containing 1 (FUNDC1), an activator of hypoxia-induced mitophagy, in wild type TAC mice, again, these effects were not observed in ALDH2 gene KO mice undergoing TAC. In angiotensin II –induced hypertrophy in vitro model of neonatal rat cardiomyocytes (NRCMs), we confirmed that α-LA could activate ALDH2 and nuclear respiratory factor 1 (Nrf1) - FUNDC-1 cascade. Interpretation: These data supported the notion that α-LA treatment could protect the heart against adverse effects of chronic pressure overload via ALDH2-dependent Nrf1-FUNDC1 signaling pathway. Funding Statement: This work was supported by the National Science Fund for Distinguished Young Scholars to AS (81725002) and a grant of Innovative Research Groups of the National Natural Science Foundation of China (81521001). Declaration of Interests: The authors have declared that no conflict of interest exists. Ethics Approval Statement: All of the mice experiments were performed under protocols approved by the Institutional Animal Care and Use Committee of the Fudan University.

Abstract 526: Free Fatty Acid Receptor 4 is Required for an Adaptive Response to Pathologic Pressure Overload-induced Heart Failure in Mice

Background: Our previous results indicate that eicosapentaenoic acid (EPA) prevents interstitial fibrosis and pathologic remodeling in the transverse aortic constriction (TAC) model of pressure overload induced heart failure in mice. However, EPA supplementation did not lead to incorporation into cardiac myocytes or fibroblasts in vivo , a traditional mechanism of action. Interestingly, we found that free fatty acid receptor 4 (Ffar4), a G-protein coupled receptor for long-chain fatty acids, was expressed in cardiac myocytes and fibroblasts. In primary cultures of cardiac fibroblasts, Ffar4 was sufficient and required to prevent TGFβ1-induced fibrosis, suggesting Ffar4 might mediate EPA cardioprotection. Methods: To understand the function of Ffar4 in the heart in vivo , we examined the response to TAC in male and female Ffar4 knockout mice (Ffar4KO) under basal conditions without EPA supplementation. Results: In male Ffar4KO mice, TAC induced a worsened dilated cardiomyopathy relative to wild-type (WT) mice. Specifically, in male Ffar4KO mice, TAC produced more hypertrophy (Heart weight: WT: 175±8 mg, n=14; Ffar4KO: 223±14 mg, n=10; P &lt;0.05), and generated more severe systolic and diastolic dysfunction (Ejection Fraction: WT: 42±3%, n=14; Ffar4KO: 28±4%, n=10; P &lt;0.05) (E/A ratio: WT: 2.0±0.2, n=12; Ffar4KO: 3.7±0.5, n=9; P &lt;0.05). However, TAC induced similar levels of interstitial fibrosis in male Ffar4 KO and WT mice (WT: 9.6±1.8%, n=14; Ffar4KO: 7.4±1.1%, n=10; ns ). While TAC worsened outcomes in male Ffar4KO mice, TAC in Ffar4 KO female mice produced no significant differences in the degree of hypertrophy or contractile dysfunction relative to WT mice. Discussion: Our results indicate that Ffar4 is required for an adaptive response to pathologic pressure overload in male mice. The lack of an exaggerated fibrotic response was surprising given our previous results indicating that in cardiac fibroblasts, Ffar4 prevents TGFβ1-induced fibrosis. This might indicate that Ffar4 signaling is minimal in fibroblasts under conditions of low omega-3 fatty acid levels. This result also suggests that the worsened contractile dysfunction in Ffar4KO mice is unrelated to fibrosis, but involves a direct effect in cardiac myocytes.

Abstract 418: Perm1 is a Novel Regulator of Mitochondrial Energetics in Cardiomyocytes

Mitochondrial oxidative metabolism and energy transduction pathways are critical for cardiac function. The expression of genes important for mitochondrial energetics is under the control of members of the peroxisome proliferator-activated receptor coactivator 1 (PGC-1) family and the estrogen-related receptor (ERR) subfamily. Perturbations in PGC-1 and/or ERR activities have been associated with impaired mitochondrial function and the development of heart failure. The mechanisms by which PGC-1 and ERR proteins regulate muscle specific transcriptional programs is not fully understood. P GC-1 and E RR-induced r egulator in m uscle 1 (Perm1) was recently identified in skeletal muscle as a muscle-specific key regulator of mitochondrial energetics. Although overexpression of Perm1 enhances oxidative capacity and fatigue resistance in mouse skeletal muscle, the underlying mechanisms remain to be elucidated. Perm1 is highly expressed in mouse and human hearts, and its expression level was reduced in pressure overload-induced heart failure in mice (49.9 ± 12.1% of sham, n=4, p&lt;0.05) and in patients with heart failure (45.5 ± 12.1% of donors, n=5, p&lt;0.05). Moreover, Perm1 is expressed in both the nucleus and the cytoplasm in primary cultured neonatal rat ventricular myocytes (NRVMs) and phenylephrine-induced cellular hypertrophy led to downregulation of Perm1 in both fractions (56.7 ± 1.7% of control). RNAseq followed by bioinformatic analysis revealed that siRNA-mediated Perm1 knockdown (KD) in NRVMs led to global downregulation of ERRα/PGC1α-target genes involved in oxidative phosphorylation, including the electron transport chain and TCA cycle ( Ndufs8; Ndufb8; Cox5a; Cox6a1; Coq2; Cox5a; Cox8a; Coq10b; Sdhb; Sdhd) . Basal respiration and ATP production rates were significantly reduced in Perm1-KD H9c2 rat ventricular myocyte-derived cells (60.4 ± 4.2% and 35.2 ± 8.8% of control cells that were transfected with scrambled-siRNA, both p&lt;0.05). This study demonstrates a novel role of Perm1 in cardiomyocytes that provides an additional layer of mitochondrial energetics regulation, presumably in an ERRα- and PGC1α-mediated manner, and suggests that downregulation of Perm1 may contribute to mitochondrial impairment during heart failure.

Role of heat shock transcription factor 1(HSF1)-upregulated macrophage in ameliorating pressure overload-induced heart failure in mice

In order to explore the role of macrophages in HSF1-mediated alleviation of heart failure, mice model of pressure overload-induced heart failure was established using transverse aortic constriction (TAC). Changes in cardiac function and morphology were studied in TAC and SHAM groups using ultrasonic device, tissue staining, electron microscopy, real-time quantitative polymerase chain reaction (RT-QPCR), and Western blotting. We found that mice in the TAC group showed evidence of impaired cardiac function and aggravation of fibrosis on ultrasonic and histopathological examination when compared to those in the SHAM group. The expressions of HSF1, LC3II/LC3I, Becline-1 and HIF-1, as well as autophagosome formation in TAC group were greater than that in SHAM group. On sub-group analyses in the TAC group, improved cardiac function and alleviation of fibrosis was observed in the HSF1 TG subgroup as compared to that in the wild type subgroup. Expressions of LC3II/LC3I, Becline-1 and HIF-1, too showed an obvious increase; and increased autophagosome formation was observed on electron microscopy. Opposite results were observed in the HSF1 KO subgroup. These results collectively suggest that in the pressure overload heart failure model, HSF1 promoted formation of macrophages by inducing upregulation of HIF-1 expression, through which heart failure was ameliorated.

Blockade of receptor for advanced glycation end products protects against systolic overload-induced heart failure after transverse aortic constriction in mice

Heart failure is the consequence of sustained, abnormal neurohormonal and mechanical stress and remains a leading cause of death worldwide. The aim of this work was to identify whether blockade of receptor for advanced glycation end products (RAGE) protected against systolic overload-induced heart failure and investigate the possible underlying mechanism. It was found that RAGE mRNA and protein expression was up-regulated in cardiac tissues from mice subjected to pressure overload by transverse aortic constriction (TAC). Importantly, inhibition of RAGE by treatment with soluble RAGE (sRAGE) or FPS-ZM1 (a high-affinity RAGE-specific inhibitor) for 8 weeks attenuated cardiac remodeling (including cardiac hypertrophy and fibrosis), and dysfunction in mice exposed to TAC. Furthermore, treatment of TAC mice with sRAGE or FPS-ZM1 enhanced phosphorylation of AMPK and reduced phosphorylation of mTOR and protein expression of NFκB p65 in cardiac tissues. In addition, treatment of TAC mice with sRAGE or FPS-ZM1 abated oxidative stress, attenuated endoplasmic reticulum stress, and suppressed inflammation in cardiac tissues. These data demonstrated the benefits of blocking RAGE on the progression of systolic overload-induced heart failure in mice, which was possibly through modulating AMPK/mTOR and NFκB pathways.

Endothelial deletion of protein tyrosine phosphatase-1B protects against pressure overload-induced heart failure in mice.

Cardiac angiogenesis is an important determinant of heart failure. We examined the hypothesis that protein tyrosine phosphatase (PTP)-1B, a negative regulator of vascular endothelial growth factor (VEGF) receptor-2 activation, is causally involved in the cardiac microvasculature rarefaction during hypertrophy and that deletion of PTP1B in endothelial cells prevents the development of heart failure. Cardiac hypertrophy was induced by transverse aortic constriction (TAC) in mice with endothelial-specific deletion of PTP1B (End.PTP1B-KO) and controls (End.PTP1B-WT). Survival up to 20 weeks after TAC was significantly improved in mice lacking endothelial PTP1B. Serial echocardiography revealed a better systolic pump function, less pronounced cardiac hypertrophy, and left ventricular dilation compared with End.PTP1B-WT controls. Histologically, banded hearts from End.PTP1B-KO mice exhibited increased numbers of PCNA-positive, proliferating endothelial cells resulting in preserved cardiac capillary density and improved perfusion as well as reduced hypoxia, apoptotic cell death, and fibrosis. Increased relative VEGFR2 and ERK1/2 phosphorylation and greater eNOS expression were present in the hearts of End.PTP1B-KO mice. The absence of PTP1B in endothelial cells also promoted neovascularization following peripheral ischaemia, and bone marrow transplantation excluded a major contribution of Tie2-positive haematopoietic cells to the improved angiogenesis in End.PTP1B-KO mice. Increased expression of caveolin-1 as well as reduced NADPH oxidase-4 expression, ROS generation and TGFβ signalling were observed and may have mediated the cardioprotective effects of endothelial PTP1B deletion. Endothelial PTP1B deletion improves cardiac VEGF signalling and angiogenesis and protects against chronic afterload-induced heart failure. PTP1B may represent a useful target to preserve cardiac function during hypertrophy.