Ventricle Tissue Research Articles

Abstract 4121970: Early to Mid-Gestational Testosterone Excess Sex-specifically Reprograms Cardiac Transcriptome and Metabolome Profiles of Sheep Fetuses

Exposure of sheep to excess testosterone (T) from days 30-90 of gestation (term 147 days) perturbs maternal systemic lipidome in late gestation, induces fetal growth restriction and sexually dimorphic perturbation in the cardiac left ventricular (LV) transcriptome in gestational (GD) day 90 fetuses culminating in cardiometabolic dysfunction in adult female offspring. We hypothesized that the detrimental impact of prenatal T excess on fetal cardiac transcriptome is a programmed event that will persist even after cessation of excess T exposure and will be accompanied by changes in fetal metabolome. Pregnant ewes were injected with 100 mg intramuscular T propionate or vehicle (C), twice weekly from GD 30-90. On GD120±5, 30 days after cessation of T treatment fetal weights were recorded, left ventricle (LV) tissue and fetal plasma processed for transcriptomics by RNA sequencing and metabolomics analysis (Female C (FC) n=6, T (FT) n=7; Male C (MC) n=6, T (MT) n=7). LV transcriptome-based pathways enrichment analysis revealed 1) downregulation of pathways associated with oxidative phosphorylation, citric acid cycle, pyruvate metabolism; 2) upregulation of TGF-β signaling pathway in T fetuses compared to C (p<0.05, FDR<0.1); 3) selective downregulation of the glycolytic pathway in MT relative to MC as opposed to downregulation of mitochondria-related pathways in FT vs FC (p<0.05, FDR<0.1). LV lipidomics analyses found glycerophospholipids and sphingolipids as main class lipid metabolites to be enriched in FT relative to FC fetuses (p<0.05, ER>1). Fetal systemic lipidome main class metabolites enriched (p<0.05, ER>2) in T vs C fetuses included 1) sphingomyelins in both sexes 2) fatty acyls metabolites only in MT vs MC as opposed to glycerophospholipids only in FT vs FC. LV polar metabolites enriched (p<0.05, ER>2) included 1) arginine&proline metabolism pathways in T vs C in both sexes 2) nicotinate and nicotinamide metabolism only in MT vs MC (p<0.05, ER>3), and porphyrin, riboflavin metabolism and N-glycan biosynthesis pathways only in FT vs FC. Systemic polar metabolites enriched (p<0.05, ER>2) included 1) D-amino acid metabolism pathways in T vs. C, 2) valine, leucine, and isoleucine pathways only in FT vs FC and glycerophospholipid and sphingolipid metabolism only in MT vs MC. These results suggest persistence of sex-specific perturbation in fetal transcriptome and systemic and cardiac metabolome in late gestational fetuses exposed to prenatal T excess.

Mercury monitoring in farmed Pacific bluefin tuna (Thunnus orientalis) using liquid asymmetric-electrode plasma optical emission spectroscopy: Ventricle tissue as a potential indicator

Tuna muscle is known to contain high levels of mercury, necessitating the monitoring of mercury concentrations in farmed fish at production sites. There is currently no straightforward in on-site method for testing whether the mercury levels in farmed tuna exceed regulatory limits. This study investigated the potential application of liquid asymmetric-electrode plasma optical emission spectroscopy (LAEP-OES) to estimate mercury concentrations in the tissues of cultured Pacific bluefin tuna at aquaculture farms. No significant differences in mercury concentrations were observed between the ventricle tissue and biopsied tissue from tuna bleeding site (TTBS), which can be harvested without reducing the market value of farmed tuna (Hg concentration of the ventricle tissue and the TTBS, median: 0.41 and 0.40 mg/kg, respectively). In addition, the relationship between mercury concentrations in the ventricle tissue and in the edible portions of three tuna fish was examined. The findings showed that total mercury level in the edible tissues varied, with the ratio of the mercury level in edible tissues to that in the ventricle ranging from 0.39 to 1.08. A positive correlation was observed between the average total mercury levels in edible tissues and ventricle tissue. These results suggest that determining the total mercury level in the ventricle, which is typically discarded at processing sites, may serve as a convenient proxy for quality control and monitoring purposes using the LAEP-OES method. However, larger sample sizes are needed to validate the proposed approach more robustly.

Garlic Powder Supplementary Diet Prevents Pulmonary Hypertension Syndrome by Reducing Apoptosis in Broiler Chickens

Background: Pulmonary hypertension syndrome (PHS) is a metabolic disorder presenting with cardiopulmonary symptoms. However, garlic powder (GP) is used to treat many diseases, such as heart and lung diseases. Objectives: This study investigated the effects of GP supplementary diet on apoptosis and right ventricle hypertrophy in broiler chickens with PHS. Methods: Ninety meat-type chickens were randomly divided into three groups: Sham, PHS and PHS+GP. The PHS was induced by triiodothyronine (T3) in broiler chickens, and GP was added to the ration after week 1 of rearing (PHS+GP). Then, lung and ventricle tissues were collected at 21 and 49 days of age. PHS was calculated at 21 and 49 days based on the ratio of the right ventricle to the total ventricles (RV/TV) weight index. Gene expression of caspase1 (CASP1) and caspase1 (CASP2) were evaluated by semi-quantitative RT-PCR in the lungs and right heart ventricles. Results: The findings showed that right ventricular hypertrophy increased at day 49 in the PHS group compared to sham (P<0.001), while garlic consumption decreased this ratio to the control level (P<0.05). Also, the expression of CASP1 and CASP2 in the lungs elevated on day 49 (P<0.001), and the GP diet prevented this increase (P<0.05). Moreover, in the right ventricle tissue, PHS affected CASP1 on days 21 and 49, although the expression of CASP2 significantly increased just at day 49 in the PHS group (P<0.001). This increase in both times was weakened by garlic consumption (P<0.05). Conclusion: Consuming garlic as a dietary supplement could prevent PHS by influencing apoptosis in the lung and heart of broilers.

Atrial fibrillation in transthyretin amyloidotic cardiomyopathy: prevalence and echocardiographic predictors.

Conduction disorders and arrhythmias frequently accompany cardiac amyloidosis (CA), with atrial fibrillation (AF) being the most prevalent manifestation. The prevalence of AF varies across different types of CA, with transthyretin (TTR) type showing the highest prevalence upon diagnosis. A retrospective, observational analysis was conducted to evaluate the prevalence of AF and to identify echocardiographic predictors related to the development of AF in our population of patients with transthyretin cardiac amyloidosis (TTR-CA). A total of 99 patients with TTR-CA were identified, with a median age of 82 (75-85) years, a median ejection fraction of 50% (43-60) and 97 of them wild type. At the time of cardiomyopathy diagnosis, 55% had AF, and during follow-up, 43% developed new AF. Among the latter group, there was a non-significant tendency to have a smaller diastolic diameter, lower left ventricular ejection fraction, increased septal thickness, higher pulmonary pressure, and lower tissue velocities, with statistical significance only found in the right ventricular S wave velocity: 8.5 cm/s (7.7-9) vs. 9.7 cm/s (8.4-10) (P=0.046). The high prevalence and incidence of AF in TTR-CA is demonstrated in our series. Doppler echocardiography might help to identify patients with signs of more advanced cardiomyopathy, such as lower right ventricle tissue velocity, who might be at higher risk of developing AF and gain the benefit of prompt diagnosis and treatment.

Evolution of translational control and the emergence of genes and open reading frames in human and non-human primate hearts.

Evolutionary innovations can be driven by changes in the rates of RNA translation and the emergence of new genes and small open reading frames (sORFs). In this study, we characterized the transcriptional and translational landscape of the hearts of four primate and two rodent species through integrative ribosome and transcriptomic profiling, including adult left ventricle tissues and induced pluripotent stem cell-derived cardiomyocyte cell cultures. We show here that the translational efficiencies of subunits of the mitochondrial oxidative phosphorylation chain complexes IV and V evolved rapidly across mammalian evolution. Moreover, we discovered hundreds of species-specific and lineage-specific genomic innovations that emerged during primate evolution in the heart, including 551 genes, 504 sORFs and 76 evolutionarily conserved genes displaying human-specific cardiac-enriched expression. Overall, our work describes the evolutionary processes and mechanisms that have shaped cardiac transcription and translation in recent primate evolution and sheds light on how these can contribute to cardiac development and disease.

P164 SOLUBLE GUANYLATE CYCLASE STIMULATORS: AN EMERGING OPTION IN THE TREATMENT OF PRESSURE AND VOLUME OVERLOAD INDUCED CHRONIC HEART FAILURE

Pressure and volume overload-induced chronic heart failure (HF) is associated with characteristic cardiac remodeling and ventricular myocardial proteomic alterations, that can lead to the formation of a substrate for arrhythmias. Antiarrhythmic drug therapy is still suboptimal and current therapies are not efficacious enough. The purpose of the current study was therefore to examine the efficiency of nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine monophosphate (cGMP) stimulator. We used as an experimental model male hypertensive Ren-2 transgenic rats (TGR) (n = 16), normotensive Hannover Sprague–Dawley rats (HSD, n = 17), and a volume-overload HF rat model induced by creating the aortocaval fistula (ACF) in rats of both strains (n = 27). Rats were after the induction of ACF treated by sGC stimulator BAY41-8543 (3 mg/kg/day for 30 weeks) alone or combined with angiotensin-converting enzyme inhibitor (ACEi) Trandolapril (0.25 mg/kg/day for 30 weeks). The age of the animals at the end of the experiment was 40 weeks. Left ventricle tissue and plasma samples were used for further analyses. Blood pressure (BP) was measured by telemetry. Combined treatment with sGC stimulator and ACEi was the most potent in lowering BP. Moreover, supplementation by sGC stimulator alone significantly decreased mortality and increased levels of the antioxidant enzyme superoxide dismutase 1, matrix metalloproteinase 2, total and phosphorylated connexin 43 as well as protein kinase C epsilon, implicated in remodeling of extracellular matrix and intercellular communication, in the left ventricle of TGR rats. Application of the sGC stimulator also slightly normalized elevated collagen deposition and hydroxyproline content in the left ventricular tissue of TGR rats. Unfortunately, we did not observe any effect of the sGC stimulator in the TGR ACF group. These results support the hypothesis that sGC stimulators might represent a class of drugs suitable for the prevention and treatment of related disorders in HF. This work was supported by APVV-21-0410, VEGA 2/0006/23, and EXCELES LX22NPO5104. Supported by APVV-21-0410, VEGA 2/0006/23, and EXCELES LX22NPO5104.

Oral supplementation of inositols effectively recovered lithium-induced cardiac dysfunctions in mice

This study investigates the effects of inositol (INO) supplementation on cardiac changes caused by Li in mice. The study involved 4 groups of C57BL6 mice (n=10 each): (i) mice orally administered with Li2CO3 for 8 weeks, then 4 additional weeks without (Li_group) or (ii) with INO supplementation (Li_INOdelayed_group) (total of 12 weeks); (iii) mice given Li2CO3 and INO supplementation concurrently for 12 weeks (Li+INO_group); (iv) one group left untreated (C–group). The INO was administered as a mixture of myo-inositol and d-chiro-inositol (80:1) in drinking water. The mice were characterised for heart morphology, function, electrical activity, arrhythmogenic susceptibility, and multiorgan histopathology (heart, liver and kidney). Cardiomyocyte size, protein expression of key signalling pathways related to hypertrophy, and transcription levels of ion channel subunits and hypertrophy markers were evaluated in the ventricle tissue. The study found that INO supplementation reduced the Li-induced cardiac adverse effects, including systolic impairment and increased susceptibility to arrhythmias. The positive effect on arrhythmias might be attributed to the restored expression levels of the potassium channel subunit Kv 1.5. Additionally, INO improved cardiomyocyte hypertrophy, possibly by inhibiting the Li-induced activation of the ERK1/2 signalling pathway and by restoring the normal expression level of BNP, and alleviated injury in the liver and kidney. The effect was preventive if INO supplementation was taken concurrently with Li and therapeutic if INO was administered after Li-induced cardiac impairments were established. These results provide new insights into the cardioprotective effect of INO and suggest a potential treatment approach for Li-induced cardiac disease.

Repeat administration of human umbilical cord mesenchymal stem cells improves left ventricular diastolic function in mice with heart failure with preserved ejection fraction

Currently, no therapy is proven to effectively improve heart failure with preserved ejection fraction (HFpEF). Although stem cell therapy has demonstrated promising results in treating ischemic heart disease, the effectiveness of treating HFpEF with human umbilical cord mesenchymal stem cells (hucMSCs) remains unclear. To answer this question, we administered hucMSCs intravenously (i.v.), either once or repetitively, in a mouse model of HFpEF induced by a high-fat diet and NG-nitroarginine methyl ester hydrochloride. hucMSC treatment improved left ventricular diastolic dysfunction, reduced heart weight and pulmonary edema, and attenuated cardiac modeling (inflammation, interstitial fibrosis, and hypertrophy) in HFpEF mice. Repeat hucMSC administration had better outcomes than a single injection. In vitro, hucMSC culture supernatants reduced maladaptive remodeling in neonatal-rat cardiomyocytes. Ribonucleic acid sequencing and protein level analysis of left ventricle (LV) tissues suggested that hucMSCs activated the protein kinase B (Akt)/forkhead box protein O1 (FoxO1) signaling pathway to treat HFpEF. Inhibition of this pathway reversed the efficacy of hucMSC treatment. In conclusion, these findings indicated that hucMSCs could be a viable therapeutic option for HFpEF.

Abstract Mo031: Tissue deficiency and altered eicosanoids levels in rodent and human heart failure.

Introduction: The role of eicosanoids, especially epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid with cardio-renal activity, in heart failure (HF) remains unclear. Hypothesis: Altered eicosanoids turnover, both in rodent and human heart failure, is a part of the pathophysiology of HF and contributes to poor prognosis. Methods: We measured active EETs and their biologically inactive metabolites, dihydroxyeicosatrienoic acids (DHETs) in the left ventricle (LV) and renal tissue obtained from a rodent heart failure model induced by aortocaval fistula (ACF). We measured protein expression of CYP2C23 and CYP2J3 in the kidney cortex and LV tissue by Western blot analysis to evaluate EETs production and mRNA expression of soluble epoxide hydrolase (sEH), the enzyme responsible for the degradation of EETs to DHETs. In the translational part, firstly, we enrolled 50 patients with HF to measure plasma 14,15-epoxyeicosatrienoic acid (14,15-EET) and 14,15-dihydroxyicosatrienoic acid (14,15-DHET) levels using commercial ELISA kits and compared them with 25 age- and sex-matched controls. Secondly, 11 consecutive patients with HFrEF indicated to cardiac resynchronization therapy (CRT) were enrolled to measure plasmatic 14,15-EET and 14,15-DHET levels in venous, arterial, and coronary sinus (CS) blood samples and correlate them with N-terminal pro-B-type natriuretic peptide (NT-proBNP) from the same compartments. Results: ACF rats exhibited tissue deficiency of EETs in the LV and kidney compared to controls, probably mainly caused by increased EETs degradation by sEH (unchanged expressions of CYP2C23 and CYP2J3, increased mRNA of sEH and EETs/DHETs ratio). In humans, both of the measured eicosanoids were significantly higher in the HF group: 14,15-EET (91.3 ±25.7 vs. 64.95 ±35.4 ng/ml) and 14,15-DHET (10.58 ±2.06 vs. 9.07 ±1.60 ng/ml), p for both < 0.001. HF patients with 14,15-EET levels less than 60 ng/ml had worse short term mortality compared to those above. In CS, NT-proBNP levels negatively correlated with plasmatic 14,15-EET levels (r = -0.63, p = 0.03) and positively with the DHET/EET ratio (r = 0.73, p = 0.02). Plasmatic 14,15-EET nor 14,15-DHET levels in measured compartments did not differ statistically (p = 0.21, p = 0,64, respectively). In individual patients, the levels of both eicosanoids correlated across all compartments. Conclusions: Tissue deficiency of active eicosanoids and their altered turnover may play an important role in HF pathophysiology.

Abstract Mo069: Phospholamban Acetylation Enhances Cardiomyocyte Calcium Cycling Under Conditions of High-Fat Feeding

Background: Diet-induced obesity (DIO) is an increasing driver of cardiovascular disease and cardiac dysfunction. The breakdown of excessive fat and sugar intake increases the presence of intracellular acetyl-CoA, which promotes the acetylation of available lysines, altering the stability and activity of these proteins. While numerous proteins involved in cardiac health have been identified as targets of acetylation, the impact of acetylation on calcium handling sarcoplasmic reticulum (SR) proteins, particularly the SERCA2a regulator phospholamban, remains poorly understood. Hypothesis: Phospholamban acetylation is upregulated by exposure to high fat conditions, in turn driving SERCA2a activity and promoting faster calcium SR reuptake. Approach: Left ventricle (LV) tissue from DIO mice and controls, and human LV tissue from control and diabetic patients, was collected and analyzed using immunoblotting and acetylation was quantified by immunoprecipitation with acetyl-lysine antibodies.Isolated working hearts from DIO and control mice were assessed before and after treatment with acute HDAC activator ITSA-1 and SIRT1 activator SRT1720 to promote deacetylation. Adult ventricular cardiomyocytes were isolated from DIO and control mice, and DIO cells were incubated in media containing excess lipid substrates (1% Lipid Mix 1, Millipore Sigma) and ITSA-1 orSRT1720. Changes in stimulated calcium dynamics were assessed using Fura-2AM and Fluo-4AM imaging. Acetylmimetic/acetylation incompetent PLN mutants were transfected into cultured cells to evaluate SERCA2a binding, oligomerization, and FRET efficiency. Results: Phospholamban acetylation significantly increased in LV tissue from both DIO mice and human diabetic patients. Acute treatment of isolated hearts and isolated cardiomyocytes from DIO mice with HDAC and SIRT1 activators significantly reduced contractility, relaxation, and SR calcium reuptake rates. Mutations in PLN mimicking or preventing lysine acetylation altered oligomerization and SERCA2a binding. Conclusions: Obesity increases the acetylation of phospholamban and enhances the reuptake of calcium into the SR, while deacetylation reverses these effects. These data directly tie nutritive status to a novel phospholamban post-translational modification that may directly impact myocardial calcium handling and contractile function.

A comparison of fixation and immunofluorescence protocols for successful reproducibility and improved signal in human left ventricle cardiac tissue.

Immunohistochemistry (IHC) and immunofluorescence (IF) are crucial techniques for studying cardiac physiology and disease. The accuracy of these techniques is dependent on various aspects of sample preparation and processing. However, standardised protocols for sample preparation of tissues, particularly for fresh-frozen human left ventricle (LV) tissue, have yet to be established and could potentially lead to differences in staining and interpretation. Thus, this study aimed to optimise the reproducibility and quality of IF staining in fresh-frozen human LV tissue by systematically investigating crucial aspects of the sample preparation process. To achieve this, we subjected fresh-frozen human LV tissue to different fixation protocols, primary antibody incubation temperatures, antibody penetration reagents, and fluorescent probes. We found that neutral buffered formalin fixation reduced image artefacts and improved antibody specificity compared to both methanol and acetone fixation. Additionally, incubating primary antibodies at 37°C for 3 h improved fluorescence intensity compared to the commonly practised 4°C overnight incubation. Furthermore, we found that DeepLabel, an antibody penetration reagent, and smaller probes, such as fragmented antibodies and Affimers, improved the visualisation depth of cardiac structures. DeepLabel also improved antibody penetration in CUBIC cleared thick LV tissue fragments. Thus, our data underscores the importance of standardised protocols in IF staining and provides various means of improving staining quality. In addition to contributing to cardiac research by providing methodologies for IF, the findings and processes presented herein also establish a framework by which staining of other tissues may be optimised.

Periostin regulates lysyl oxidase through ERK1/2 MAPK-dependent serum response factor in activated cardiac fibroblasts.

Collagen crosslinking, mediated by lysyl oxidase, is an adaptive mechanism of the cardiac repair process initiated by cardiac fibroblasts postmyocardial injury. However, excessive crosslinking leads to cardiac wall stiffening, which impairs the contractile properties of the left ventricle and leads to heart failure. In this study, we investigated the role of periostin, a matricellular protein, in the regulation of lysyl oxidase in cardiac fibroblasts in response to angiotensin II and TGFβ1. Our results indicated that periostin silencing abolished the angiotensin II and TGFβ1-mediated upregulation of lysyl oxidase. Furthermore, the attenuation of periostin expression resulted in a notable reduction in the activity of lysyl oxidase. Downstream of periostin, ERK1/2 MAPK signaling was found to be activated, which in turn transcriptionally upregulates the serum response factor to facilitate the enhanced expression of lysyl oxidase. The periostin-lysyl oxidase association was also positively correlated in an in vivo rat model of myocardial infarction. The expression of periostin and lysyl oxidase was upregulated in the collagen-rich fibrotic scar tissue of the left ventricle. Remarkably, echocardiography data showed a reduction in the left ventricular wall movement, ejection fraction, and fractional shortening, indicative of enhanced stiffening of the cardiac wall. These findings shed light on the mechanistic role of periostin in the collagen crosslinking initiated by activated cardiac fibroblasts. Our findings signify periostin as a possible therapeutic target to reduce excessive collagen crosslinking that contributes to the structural remodeling associated with heart failure.

Soluble guanylate cyclase stimulator as an emerging therapeutic option in the management of arrhythmias in heart failure rat model

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): This research was supported by VEGA grant no. 2/0002/20 and 2/0006/23, the Slovak Research and Development Agency under contract no. 21-0410, European Regional Development Fund: ITMS2014+: 313011AVG3. This research was also partially supported by the Ministry of Health of the Czech Republic within the project for the development of the research organization [grant number 00023001 (IKEM)—institutional support] and also supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Program EXCELES, Project No. LX22NPO5104)—Funded by the European Union—Next Generation EU Heart failure (HF) is a complex clinical syndrome characterized by a decrease in the efficiency of the heart pump, leading to a decline in hemodynamic status and cardiac remodelling, that can contribute to the formation of a substrate for arrhythmias. Antiarrhythmic drug therapy is still suboptimal and current therapies are not efficacious enough. The purpose of the current study was therefore to examine the efficiency of nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine monophosphate (cGMP) stimulator. We used as an experimental model male hypertensive Ren-2 transgenic rats (TGR, n = 16), normotensive Hannover Sprague–Dawley rats (HSD, n = 17), and a rat model of HF surgically induced by aortocaval fistula (ACF) in rats of both strains (n = 27). After ACF induction were rats treated by sGC stimulator BAY41-8543 (3 mg/kg/day for 30 weeks) alone or combined with angiotensin-converting enzyme inhibitor (ACEi) Trandolapril (0.25 mg/kg/day for 30 weeks). Age of the animals at the end of the experiment was 40 weeks. Left ventricle tissue and plasma samples were used for further analyses. Treatment by sGC stimulator significantly decreased rats' mortality, and systolic blood pressure, and significantly increased levels of antioxidant enzyme superoxide dismutase 1, matrix metalloproteinase 2, total and phosphorylated connexin 43 as well as protein kinase C epsilon, implicated in remodelling of extracellular matrix and intercellular communication, in the left ventricle of TGR rats. Application of sGC stimulator slightly normalized elevated collagen deposition and hydroxyproline content in the left ventricular tissue of TGR rats. Unfortunately, we did not observe any effect of the sGC stimulator in the TGR ACF group. These results support the hypothesis that sGC stimulators might represent a class of drugs suitable for combating heart failure with partially anti-arrhythmic potential, but further studies are needed.

Berberine Alleviates Right Heart Dysfunction in MCT-Induced PAH Rats by Targeting Myocardial Mitochondrial Oxidative Phosphorylation

Background: Right ventricular failure (RVF) stands as a leading cause of mortality among pulmonary hypertension (PAH) patients. The underlying mechanisms contributing to Right Ventricular (RV) dysfunction remain enigmatic, with a conspicuous absence of effective treatments. RV functional deterioration is influenced by mitochondria oxidative phosphorylation and fatty acid oxidation. Berberine (BBR), a natural active compound extracted from Berberis vulgaris rhizomes, has shown promise for treating cardiovascular diseases due to its interactions with multiple intracellular targets. This study aimed to investigate the applicability and therapeutic effcacy of BBR in RVF and explore the mechanisms involved in its beneficial effects in monocrotaline (MCT)-induced PAH rats. Methods: Right ventricular hypertrophy (RVH) and RVF were induced via a single injection of MCT (60 mg/kg, ip) in Sprague-Dawley rats. BBR (4 mg/kg body weight per day) was administered through oral gavage. RVH and RVF were confirmed through transthoracic echocardiography (TTE). Transmission electron microscopy (TEM) was utilized to observe changes in cardiomyocyte and mitochondrial ultrastructure within the right ventricle tissue. Comparisons were made among Normal control (NC), RVH, RVH+BBR, RVF, and RVF+ BBR groups. RNA sequencing (RNA-seq) of right ventricle tissue was performed for both the NC (treated with normal saline, ip) and right ventricle dysfunction (RVH and RVF) groups. Gene Ontology and KEGG enrichment analyses were conducted using the R-package “Clusterprofiler.” RV transcriptome results were validated through qPCR and Western blot analysis. Results: Echocardiography and elevated ANP and BNP levels confirmed the presence of RVF following the development of MCT-induced PAH. TEM imaging revealed significant disruptions in mitochondrial ultrastructure, particularly in a subset of interfibrillar mitochondria, which exhibited swelling, disorganization, and reduced cristae density in RVH rats. These disruptions were even more pronounced in RVF rats. Transcriptomic profiles of RV tissue from RVH and RVF identified a total of 175 differentially expressed genes (DEGs). Gene Ontology and KEGG enrichment analyses pinpointed 38 genes associated with oxidative phosphorylation. qPCR demonstrated downregulation of mitochondrial oxidative phosphorylation-related genes (NDUFA2, NDUFA13, COX7B, COX6C, GSTK1, ECH1) in RVF. BBR ameliorated right heart function, leading to decreased RV internal diameter at end-diastole, a reduced RV hypertrophy index, and an increased RV fractional area change. This was accompanied by the recovery of ANP and BNP levels. Furthermore, after BBR application, the mitochondrial structure within RV tissue exhibited marked improvement. Lastly, Western blot analysis revealed that fatty acid oxidation-related proteins (Decr1, Acadm, Acacb, Hadh, Hadhb, Prkag3, Acaa2) were downregulated in right heart dysfunction, and BBR effectively mitigated the decline in oxidation-related protein levels in both RVH and RVF. Conclusion: In summary, our results underscore the critical role of BBR in preserving cardiac function from RVF in MCT-induced PAH rats. This preservation appears to occur through the upregulation of mitochondrial oxidative phosphorylation levels, ultimately enhancing energy metabolism within the mitochondria of RV cardiomyocytes in the right heart of PAH. This work is supported by Natural Science Foundation of Shandong Province(ZR2022MH052), China Postdoctoral Fund (NO.2021M691944) to XMY. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Cardiorenal Effects of Chronic High Phosphate Diet in Female Rats

Cardiorenal Syndrome Type 4 (CRS4) refers to the development of cardiovascular dysfunction as a result of chronic kidney disease. The accumulation of circulating phosphate in the later stages of chronic kidney disease (CKD) has been positively associated with cardiac hypertrophy and dysfunction. In our previous work, we found that male Sprague Dawley rats that underwent 5/6 nephrectomy (5/6Nx) to induce CKD phenotypes also developed cardiac pathology when serum phosphorus levels became elevated. Small RNA sequencing of left ventricle tissue revealed upregulated microRNA-21-5p and -3p expression in 5/6Nx rats, when compared to sham-operated controls, and suppression of miR-21-5p attenuated left ventricle pathology in male 5/6Nx rats. We hypothesize that hyperphosphatemia, independent of CKD, can impair cardiac function and that miR-21 may mediate these effects. It has been recently reported that females exhibit higher urinary phosphate excretion in response to high dietary phosphate intake than males. In this study, hyperphosphatemia was induced by placing 9-week old female wild-type (WT) Sprague Dawley and global miR-21 knockout (miR-21−/−; lacking both miR-21-5p and -3p rats) (n=4-9/group) on either high phosphate (2% Pi) or control diet (0.6% Pi) for 8 weeks. Echocardiography and renal ultrasound were performed under anesthesia to assess cardiac function and renal cross-sectional area, respectively, prior to tissue collection at endpoint. Rats of either genotype fed 2% Pi diet exhibited increased left kidney weight, left kidney/body weight, and left kidney cross-sectional area (2-way ANOVA, main effect of diet; p<0.05). In animals fed 2% Pi, miR-21−/− rats had significantly higher left ventricle fractional shortening than WT rats (57.06 ± 3.32 vs. 43.96 ± 1.34; 2-way ANOVA with Tukey multiple comparisons; p<0.05). This study allowed the cardiovascular effects of chronically elevated phosphate to be isolated from other uremic toxins elevated in CKD. We find that loss of miR-21 augments cardiac function, supporting a role for miR-21 in mediating phosphate-related pathology. Ongoing studies are focused on identifying specific molecular mechanisms driving these changes. This study was supported by T32HL007852 and R01HL128332. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Role of bioenergetic hypoxia in the morphological transformation of the myocardium during vibration disease

BACKGROUND: Analysis of literature on the structural changes in the heart in patients with vibration disease using echocardiographic research methods revealed a concentric type of remodeling of the left ventricular chambers, which is associated with a high risk of cardiovascular complications, including sudden cardiac death, in people of working age.
 AIM: To determine the role of bioenergetic hypoxia in the development of morphological transformation of the myocardium to substantiate the efficacy of pharmacotherapy for vibration disease.
 MATERIALS AND METHODS: The energy production activity of cellular systems of heart tissue in vitro was analyzed by the polarographic method using a closed galvanic-type oxygen sensor (Clark electrode). The stressful effects of vibration were confirmed by the dynamics of the morphohistological picture of changes in the myocardial tissue of the left ventricle in the apical region after standard alcohol–paraffin wiring and staining of histological preparations with hematoxylin and eosin.
 RESULTS: Evaluation of the morphometric and bioenergetic parameters of cardiomyocytes under various experimental vibration modes (7, 21, and 56 sessions with a frequency of 8 and 44 Hz) confirmed the relationship between the provision of tissue with energy potential and morphological signs of pathological structural changes in the myocardial tissue, such as hypertrophy of cardiomyocytes, development of fibrosis, restructuring of the vascular bed, and necrosis.
 CONCLUSION: Analysis of the relationship between energy metabolism and morphohistological transformation of heart tissue allows us to resolve the role of universal and specific mechanisms in cardiac remodeling in the presence of vibration and pathogenetically substantiate the choice of drugs that not only have a vibration-protective effect but also inhibit pathological structural changes in the myocardial tissue.

Mixture of Doxycycline, ML-7 and L-NAME Restores the Pro- and Antioxidant Balance during Myocardial Infarction-In Vivo Pig Model Study.

The restoration of blood flow to the ischemic myocardium inflicts ischemia/reperfusion (I/R) heart injury (IRI). The main contributors to IRI are increased oxidative stress and subsequent excessive production of ROS, increased expression of NOS and peroxinitate, activation of MMPs, and enhanced posttranslational modifications of contractile proteins, which make them more susceptible to proteolytic degradation. Since the pathophysiology of IRI is a complex issue, and thus, various therapeutic strategies are required to prevent or reduce IRI and microvascular dysfunction, in the current study we proposed an innovative multi-drug therapy using low concentrations of drugs applied intracoronary to reach microvessels in order to stabilize the pro- and antioxidant balance during a MI in an in vivo pig model. The ability of a mixture of doxycycline (1 μM), ML-7 (0.5 μM), and L-NAME (2 μM) to modulate the pro- and antioxidative balance was tested in the left ventricle tissue and blood samples. Data showed that infusion of a MIX reduced the total oxidative status (TOS), oxidative stress index (OSI), and malondialdehyde (MDA). It also increased the total antioxidant capacity, confirming its antioxidative properties. MIX administration also reduced the activity of MMP-2 and MMP-9, and then decreased the release of MLC1 and BNP-26 into plasma. This study demonstrated that intracoronary administration of low concentrations of doxycycline in combination with ML-7 and L-NAME is incredibly efficient in regulating pro- and antioxidant balance during MI.

Establishment of a Biaxial Testing System for Characterization of Right Ventricle Viscoelasticity Under Physiological Loadings.

Prior studies have indicated an impact of cardiac muscle viscoelasticity on systolic and diastolic functions. However, the studies of ventricular free wall viscoelasticity, particularly for that of right ventricles (RV), are limited. Moreover, investigations on ventricular passive viscoelasticity have been restricted to large animals and there is a lack of data on rodent species. To fill this knowledge gap, this study aims to develop a biaxial tester that induces high-speed physiological deformations to characterize the passive viscoelasticity of rat RVs. The biaxial testing system was fabricated so that planar deformation of rat ventricle tissues at physiological strain rates was possible. The testing system was validated using isotropic polydimethylsiloxane (PDMS) sheets. Next, viscoelastic measurements were performed in healthy rat RV free walls by equibiaxial cyclic sinusoidal loadings and stress relaxation. The biaxial tester's consistency, accuracy, and stability was confirmed from the PDMS samples measurements. Moreover, significant viscoelastic alterations of the RV were found between sub-physiological (0.1Hz) and physiological frequencies (1-8Hz). From hysteresis loop analysis, we found as the frequency increased, the elasticity and viscosity were increased in both directions. Interestingly, the ratio of storage energy to dissipated energy (Wd/Ws) remained constant at 0.1-5Hz. We did not observe marked differences in healthy RV viscoelasticity between longitudinal and circumferential directions. This work provides a new experimental tool to quantify the passive, biaxial viscoelasticity of ventricle free walls in both small and large animals. The dynamic mechanical tests showed frequency-dependent elastic and viscous behaviors of healthy rat RVs. But the ratio of dissipated energy to stored energy was maintained between frequencies. These findings offer novel baseline information on the passive viscoelasticity of healthy RVs in adult rats.

A specific inflammatory suppression fibroblast subpopulation characterized by MHCII expression in human dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is a significant cause of heart failure that requires heart transplantation. Fibroblasts play a central role in the fibro-inflammatory microenvironment of DCM. However, their cellular heterogeneity and interaction with immune cells have not been well identified. An integrative analysis was conducted on single-cell RNA sequencing (ScRNA-Seq) data from human left ventricle tissues, which comprised 4 hearts from healthy donors and 6 hearts with DCM. The specific antigen-presenting fibroblast (apFB) was explored as a subtype of fibroblasts characterized by expressing MHCII genes, the existence of which was confirmed by immunofluorescence staining of 3 cardiac tissues from DCM patients with severe heart failure. apFB highly expressed the genes that response to IFN-γ, and it also have a high activity of the JAK-STAT pathway and the transcription factor RFX5. In addition, the analysis of intercellular communication between apFBs and CD4+T cells revealed that the anti-inflammatory ligand-receptor pairs TGFB-TGFR, CLEC2B-KLRB1, and CD46-JAG1 were upregulated in DCM. The apFB signature exhibited a positive correlation with immunosuppression and demonstrated diagnostic and prognostic value when evaluated using a bulk RNA dataset comprising 166 donors and 166 DCM samples. In conclusion, the present study identified a novel subpopulation of fibroblasts that specifically expresses MHCII-encoding genes. This specific apFBs can suppress the inflammation occurring in DCM. Our findings further elucidate the composition of the fibro-inflammatory microenvironment in DCM, and provide a novel therapeutic target.

E3 Ubiquitin Ligase ASB14 Inhibits Cardiomyocyte Proliferation by Regulating MAPRE2 Ubiquitination.

The ubiquitin proteasome system is a highly specific and selective protein regulatory system that plays an essential role in the regulation of the cell cycle. Despite its significance, the role of ubiquitination in cardiomyocyte proliferation remains largely unclear. This study aimed to investigate the potential impact of E3 ubiquitin ligase ASB14 (Ankyrin Repeat And SOCS Box Containing 14) on cardiac regeneration. We conducted a microarray analysis of apical resection ventricle tissues, and our findings revealed that ASB14 was down-regulated during the cardiac regenerative response. Subsequently, we examined the effect of ASB14 silencing on cardiomyocyte nuclear proliferation both in vitro and in vivo. Our results indicated that ASB14 silencing promoted cardiomyocyte nuclear proliferation, suggesting that ASB14 may play a role in regulating cardiac regeneration. To further investigate the potential therapeutic implications of ASB14 deficiency, we examined the cardiac function of mice with ASB14 deficiency in response to ischemic injury. Our findings showed that mice with ASB14 deficiency exhibited preserved cardiac function and a therapeutic effect in response to ischemic injury, which was attributed to the enhancement of cardiomyocyte nuclear proliferation. To elucidate the underlying mechanisms, we investigated the effect of ASB14 on microtubule-associated protein RP/EB family member 2 (MAPRE2) protein degradation. Our results indicated that the loss of ASB14 decreased the degradation of MAPRE2 protein, subsequently promoting cardiomyocyte nuclear proliferation and enhancing cardiac repair after myocardial infarction (MI). In conclusion, our study provides evidence that inhibition of ASB14-mediated MAPRE2 ubiquitination promotes cardiomyocyte nuclear proliferation, which may serve as a potential target for treating heart failure induced by MI injury.