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Background: Type 2 diabetes mellitus is characterized by hyperglycemia. Hyperglycemia increases free radicals and oxidative stress that damage heart muscle cells. Nuclear factor kappa B (NFkB) plays an important role in inflammation and cell damage. Radish leaves (Raphanus sativus) are known to have compounds that can control NFkB expression. Objective: To measure the effect of radish leaves on the expression of NFkB in heart muscle. Methods: The study design was experimental laboratory post test control group design. The subjects were diabetic male Wistar rats, weight 150-300 grams, 3-4 months old. Rats were divided into four groups and given treatment orally for 28 days. The 1st group received placebo. The 2nd group received glibenclamide 5 mg/kg BW/day. The 3rd and 4th groups received 50% and 100% ethanol extract of radish leaves. Heart NFkB was measured using immunohistochemistry. Results: Radish leaves ethanol extract 50% and 100% reduced NFkB in the heart muscle better than placebo and glibenclamide 5 mg/kg BW ((p=0.000). There was no difference between radish leaf extract 50% and 100% in reducing heart NFkB expression (p=0.876). Conclusion: Radish leaf extract 50% and 100% reduce NFkB expression better than placebo and glibenclamide 5 mg/kg BW.

The binding of calcium and magnesium ions to proteins is crucial for regulating heart contraction. However, other divalent cations, including xenobiotics, can accumulate in the myocardium and enter cardiomyocytes, where they can bind to proteins. In this article, we summarized the impact of these cations on myosin ATPase activity and EF-hand proteins, with special attention given to toxic cations. Optimal binding to EF-hand proteins occurs at an ionic radius close to that of Mg2+ and Ca2+. In skeletal Troponin C, Cd2+, Sr2+, Pb2+, Mn2+, Co2+, Ni2+, Ba2+, Mg2+, Zn2+, and trivalent lanthanides can substitute for Ca2+. As myosin ATPase is not a specific MgATPase, Ca2+, Fe2+, Mn2+, Ni2+, and Sr2+ could support myosin ATPase activity. On the other hand, Zn2+ and Cu2 significantly inhibit ATPase activity. The affinity to various divalent cations depends on certain proteins or their isoforms and can alter with amino acid substitution and post-translational modification. Cardiac EF-hand proteins and the myosin ATP-binding pocket are potential molecular targets for toxic cations, which could significantly alter the mechanical characteristics of the heart muscle at the molecular level. [ABSTRACT FROM AUTHOR]

ObjectiveFennel in animal diets for various purposes including improving weight gain, oxidative quality of meat, digestion and growth efficiency, closed cell volume, small intestine length and weight, carcass efficiency, feed intake, red blood cells and hemoglobin count, feed conversion, efficiency and health status and reduction of the total number of bacteria have been used. On the other hand, the DLK1 gene plays an important role in controlling various processes throughout the fetus and adulthood. Therefore, this study aimed to investigate the effect of fennel on DLK1 gene expression in Kermani sheep heart using real-time PCR.Materials and Methods In this study, 30 male Kermani lambs (6-month-old) with almost the same weight were selected and randomly divided into three experimental groups (10 in each group). Experimental animals were fed freely and separately with three levels of fennel (0, 1, and 2%) for three months. After slaughtering, the heart weight and fat around the heart were measured and the heart tissue was sampled. RNA extraction and cDNA synthesis were performed using standard kits. The expression of DLK1 genes (target gene) and beta-actin gene (reference gene) was performed using proper primers using real-time PCR method and the results were analyzed with the appropriate software.ResultsThe results of the quality assay of extracted RNAs on agarose gel and at A260 / A280 wavelength showed the good quality of total RNA. The results of real-time PCR and electrophoresis of PCR products on agarose gel showed that the DLK1 gene was expressed in heart tissue. Adding fennel to the diet did not significantly change the weight of the heart and the weight of fat around the heart compared to the control diet. The results showed that fennel nutrition at 1% and 2% levels significantly (P

We report a neonatal patient with hypertrophic cardiomyopathy (HCM), lactic acidosis and isolated complex I deficiency. Using a customized next-generation sequencing panel, we identified a novel hemizygous variant c.338G>A in the X-linked NDUFB11 gene that encodes the NADH: ubiquinone oxidoreductase subunit B11 of the mitochondrial respiratory chain (MRC) complex I (CI). Molecular and functional assays performed in the proband's target tissues—skeletal and heart muscle—showed biochemical disturbances of the MRC, suggesting a pathogenic role for this variant. In silico analyses initially predicted an amino acid missense change p.(Arg113Lys) in the NDUFB11 CI subunit. However, we showed that the molecular effect of the c.338G>A variant, which is located at the last nucleotide of exon 2 of the NDUFB11 gene in the canonical 'short' transcript (sized 462 bp), instead causes a splicing defect triggering the up-regulation of the expression of an alternative 'long' transcript (sized 492 bp) that can also be detected in the control individuals. Our results support the hypothesis that the canonical 'short' transcript is required for the proper NDUFB11 protein synthesis, which is essential for optimal CI assembly and activity, whereas the longer alternative transcript seems to represent a non-functional, unprocessed splicing intermediate. Our results highlight the importance of characterizing the molecular effect of new variants in the affected patient's tissues to demonstrate their pathogenicity and association with the clinical phenotypes. [ABSTRACT FROM AUTHOR]

Computer simulations, which are performed at a single wavelength at a time, have been traditionally used to estimate the optical properties of tissues. The results of these simulations need to be interpolated. For a broadband estimation of tissue optical properties, the use of computer simulations becomes time consuming and computer demanding. When spectral measurements are available for a tissue, the use of the photon diffusion approximation can be done to perform simple and direct calculations to obtain the broadband spectra of some optical properties. The additional estimation of the reduced scattering coefficient at a small number of discrete wavelengths allows to perform further calculations to obtain the spectra of other optical properties. This study used spectral measurements from the heart muscle to explain the calculation pipeline to obtain a complete set of the spectral optical properties and to show its versatility for use with other tissues for various biophotonics applications., (© 2024 Wiley‐VCH GmbH.)

The binding of calcium and magnesium ions to proteins is crucial for regulating heart contraction. However, other divalent cations, including xenobiotics, can accumulate in the myocardium and enter cardiomyocytes, where they can bind to proteins. In this article, we summarized the impact of these cations on myosin ATPase activity and EF-hand proteins, with special attention given to toxic cations. Optimal binding to EF-hand proteins occurs at an ionic radius close to that of Mg2+ and Ca2+. In skeletal Troponin C, Cd2+, Sr2+, Pb2+, Mn2+, Co2+, Ni2+, Ba2+, Mg2+, Zn2+, and trivalent lanthanides can substitute for Ca2+. As myosin ATPase is not a specific MgATPase, Ca2+, Fe2+, Mn2+, Ni2+, and Sr2+ could support myosin ATPase activity. On the other hand, Zn2+ and Cu2 significantly inhibit ATPase activity. The affinity to various divalent cations depends on certain proteins or their isoforms and can alter with amino acid substitution and post-translational modification. Cardiac EF-hand proteins and the myosin ATP-binding pocket are potential molecular targets for toxic cations, which could significantly alter the mechanical characteristics of the heart muscle at the molecular level.

Objective Fennel in animal diets forvarious purposes including improving weight gain, oxidative quality of meat, digestion and growth efficiency, closed cell volume, small intestine length and weight, carcass efficiency, feed intake, red blood cells and hemoglobin count, feed conversion, efficiencyand health status and reduction of the total number of bacteria have been used. On the other hand, DLKl gene plays an important role in controlling various processes throughout the fetus andadulthood. Therefore, the aim of this study was to investigate the effect of fennel on DLKl geneexpression in Kermani sheep heart using real-time PCR.Materials and MethodsIn this study, 30 male Kermani lambs (6-month-old) with almost the same weight were selectedand randomly divided into three experimental groups (10 in each group). Experimental animalswere fed freely and separately with three levels of fennel (0, 1 and 2%) for three months. After slaughtering, the heart weight and fat around the heart were measured and the heart tissue wassampled. RNA extraction and cDNA synthesis were performed using standard kits. Theexpression of DLKl genes (target gene) and beta-actin gene (reference gene) was performed usingproper primers using real-time PCR method and the results were analyzed with the appropriatesoftware. Results The results of quality assay of extracted RNAs on agarose gel and at A26O / A28O wavelengthshowed good quality of total RNA. The results of real-time PCR and electrophoresis of PCRproducts on agarose gel showed that DLKl gene was expressed in heart tissue. Adding fennel tothe diet did not significantly change the weight of the heart and the weight of fat around the heartcompared to the control diet. The results showed that fennel nutrition at 1% and 2% levelssignificantly (P <0.05) increased DLKl gene expression in heart tissue. Conclusions Based on the results, it can be concluded that fennel can be used in sheep diet to improve heartstructure through positive effects on DLKl gene expression. Since fennel has increased theexpression level of DLKl gene in heart tissue, it can be considered to improve heart ftnction. Itcan be concluded that fennel can be used for different purposes in livestock, but for each effect ineach tissue, more research should be done considering different genetic, epigenetic andphysiological conditions to reach the distinct conclusion. [ABSTRACT FROM AUTHOR]

Aging affects the energy metabolism differently in the cardiac and skeletal muscles. The study aim was to assess the effects of short-term calorie restriction (SCR) and refeeding on the expression of genes involved in the control of cardiac and skeletal muscle energy metabolism in old vs. young male rats. Young (4 mo) and old (24 mo) rats were subjected to 60% SCR for 30 days, and refed ad libitum for 2 or 4 days. In the cardiac (CM) and skeletal muscles (SM) we compared the gene expression (qPCR) of carnitine palmitoyltransferase-I (Cpt-I), peroxisome proliferator-activated receptor beta/delta (Ppar-β/δ), glucose transporter 4 (Glut4), peroxisome proliferator-activated receptor-γ coactivator-1α (Pgc-1α), and sirtuin 3 (Sirt3). In CM, aging increased Cpt-I expression but did not affect the other genes. In SM, Cpt-I, Glut4, Pgc-1α, and Sirt3 mRNA levels were lower in old than young rats. In CM of only young rats SCR increased Cpt-I expression which remained elevated after refeeding. Upon SCR, the expression of Ppar-β/δ, Glut4, Pgc-1α, and Sirt3 in CM increased in young but not old rats, and refeeding re-established control levels. In SM of young rats SCR increased Ppar-β/δ and Pgc-1α, and decreased Sirt3 expression, whereas refeeding generally decreased these mRNA levels. In SM of old rats SCR decreased only Pgc-1α expression. The adaptive response to SCR and subsequent refeeding is muscle tissue-specific and differs in young and old male rats. SCR appears to increase the efficiency of glucose and fatty acid utilization in the cardiac muscle of young, but not old male rats. [ABSTRACT FROM AUTHOR]

Objective: Physical exercise—especially at high intensity—is known to impose cardiac stress, as mirrored by, e.g., increased blood levels of cardiac stress biomarkers such as cardiac Troponin T (cTnT) and NT-proBNP. We examined healthy young participants to determine whether a few nights of short sleep duration alter the effects of acute exercise on these blood biomarkers. Methods: Sixteen men participated in a randomized order in a crossover design, comprising three consecutive nights of a) normal sleep duration (NS, 8.5 h of sleep/night) and b) sleep restriction (SR, 4.25 h of sleep/night). Blood was repeatedly sampled for determination of NT-proBNP and cTnT serum levels before and after a high-intensity exercise protocol (i.e., 75% VO2maxReserve cycling on an ergometer). Results: Under pre-exercise sedentary conditions, blood levels of cTnT and NT-proBNP did not significantly differ between the sleep conditions (P > 0.10). However, in response to exercise, the surge of circulating cTnT was significantly greater following SR than NS (+37–38% at 120–240 min post-exercise, P ≤ 0.05). While blood levels of NT-proBNP rose significantly in response to exercise, they did not differ between the sleep conditions. Conclusion: Recurrent sleep restriction may increase the cardiac stress response to acute high-intensity exercise in healthy young individuals. However, our findings must be further confirmed in women, older subjects and in patients with a history of heart disease.

Poor maternal nutrition during gestation can negatively affect offspring growth, development, and health pre- and post-natally. Overfeeding during gestation or maternal obesity (MO) results in altered metabolism and imbalanced endocrine hormones in animals and humans which will have long-lasting and detrimental effects on offspring growth and health. In this study, we examined the effects of overnutrition during gestation on autophagy associated pathways in offspring heart muscles at two gestational and one early postnatal time point (n = 5 for treated and untreated male and female heart respectively at each time point). Two-way ANOVA was used to analyze the interaction between treatment and sex at each time point. Our results revealed significant interactions of maternal diet by developmental stages for offspring autophagy signaling. Overfeeding did not affect the autophagy signaling at mid-gestation day 90 (GD90) in both male and female offspring while the inflammatory cytokines were increased in GD90 MO male offsrping; however, overfeeding during gestation significantly increased autophagy signaling, but not inflammation level at a later developmental stage (GD135 and day 1 after birth) in both males and females. We also identified a sexual dimorphic response in which female progeny were more profoundly influenced by maternal diet than male progeny regardless of developmental stages. We also determined the cortisol concentrations in male and female hearts at three developmental stages. We did not observe cortisol changes between males and females or between overfeeding and control groups. Our exploratory studies imply that MO alters autophagy associated pathways in both male and female at later developmental stages with more profound effects in female. This finding need be confirmed with larger sample numbers in the future. Our results suggest that targeting on autophagy pathway could be a strategy for correction of adverse effects in offspring of over-fed ewes. [ABSTRACT FROM AUTHOR]

New Findings: What is the central question of this study?Intracellular energetic processes in quiescent cardiac muscle release 'basal' heat; during contraction, a much larger amount of 'active' heat is also produced. Previously, measurement challenges have constrained researchers to assume that basal heat rate remains constant during contraction and shortening. Is this assumption correct?What is the main finding and its importance?We show that basal heat rate is modulated by the extent and velocity of muscle shortening. Their relative contributions are muscle specific. We apply a method with which researchers can now disentangle, for each experiment, changes in basal heat from active heat production, providing more precise measures of the individual energetic processes underlying cardiac muscle contraction. Separating the variations in cardiac basal heat rate from variations in active heat rate is necessary to determine cardiac muscle energy consumption accurately during the performance of active work. By developing a model of cardiac muscle basal heat rate, we aimed to investigate changes in basal heat rate when cardiac muscle performs work. Experiments were conducted on 10 isolated rat cardiac trabeculae subjected to both active (work‐loops) and quiescent (length‐change and velocity) interventions. Muscle force, length and heat output rate were measured simultaneously in a flow‐through work‐loop calorimeter. Quiescent muscle characteristics were used to parameterize muscle‐specific models of change in basal heat rate, thereby to predict dynamic changes in basal heat rate during active work‐loop contraction. Our data showed that the quiescent heat characteristics of cardiac muscle varied between samples, displaying dependence on both the extent and the rate of change in muscle length. We found a moderate correlation between muscle dimensions (cross‐sectional area and volume) and the length‐dependent basal heat parameter (P = 0.0330 and P = 0.0242, respectively), but no correlation with the velocity‐dependent parameter. These findings lead us to conclude that the heat output of cardiac muscle at quiescence varies with both the extent and the velocity of shortening, to an extent that is muscle specific, and that this variation must be measured and accounted for in each specimen when assessing active energetics. [ABSTRACT FROM AUTHOR]

Background: Autophagy is a mechanism that could be inhibited or activated through exercise. FOXO3a and Beclin-1 proteins are important proteins in autophagy regulation. Therefore, this study aimed to investigate the effect of 4 weeks high intensity interval training (HIIT) on the content of FOXO3a and Beclin-1 proteins in the left ventricular heart tissue with type 2 diabetic rats. Materials and Methods: In this experimental study, 12 two-month-old Sprague-Dawley rats with a mean weight of 270±20 g were selected. After diabetic induction with STZ and Nicotinamide, rats were randomly assigned to two groups, diabetic training (6 heads) and diabetic control (6 heads). The training group trained for 4 days a week in accordance with the training program for 4 weeks, while the control group did not have any training program. Also, the rats did not receive any insulin treatment during the study period. The protein content of the present study was measured by Western blotting. Data were analyzed by SPSS software version 19 and independent t-test. Results: Four weeks of HIIT training resulted in a significant decrease in FOXO3a (P=0.03) and Beclin-1 (P=0.0001) proteins content in the diabetic training group compared to the diabetic control group. Conclusion: Four weeks of HIIT training would decrease the content of FOXO3a and Beclin-1 proteins which leads to inhibition of autophagy in the left ventricle of subjects with type 2 diabetes. Therefore, HIIT training could be a new therapeutic approach in diabetics.

Poor maternal nutrition during gestation can negatively affect offspring growth, development, and health pre- and post-natally. Overfeeding during gestation or maternal obesity (MO) results in altered metabolism and imbalanced endocrine hormones in animals and humans which will have long-lasting and detrimental effects on offspring growth and health. In this study, we examined the effects of overnutrition during gestation on autophagy associated pathways in offspring heart muscles at two gestational and one early postnatal time point (n = 5 for treated and untreated male and female heart respectively at each time point). Two-way ANOVA was used to analyze the interaction between treatment and sex at each time point. Our results revealed significant interactions of maternal diet by developmental stages for offspring autophagy signaling. Overfeeding did not affect the autophagy signaling at mid-gestation day 90 (GD90) in both male and female offspring while the inflammatory cytokines were increased in GD90 MO male offsrping; however, overfeeding during gestation significantly increased autophagy signaling, but not inflammation level at a later developmental stage (GD135 and day 1 after birth) in both males and females. We also identified a sexual dimorphic response in which female progeny were more profoundly influenced by maternal diet than male progeny regardless of developmental stages. We also determined the cortisol concentrations in male and female hearts at three developmental stages. We did not observe cortisol changes between males and females or between overfeeding and control groups. Our exploratory studies imply that MO alters autophagy associated pathways in both male and female at later developmental stages with more profound effects in female. This finding need be confirmed with larger sample numbers in the future. Our results suggest that targeting on autophagy pathway could be a strategy for correction of adverse effects in offspring of over-fed ewes.

We report a neonatal patient with hypertrophic cardiomyopathy (HCM), lactic acidosis and isolated complex I deficiency. Using a customized next-generation sequencing panel, we identified a novel hemizygous variant c.338G>A in the X-linked NDUFB11 gene that encodes the NADH: ubiquinone oxidoreductase subunit B11 of the mitochondrial respiratory chain (MRC) complex I (CI). Molecular and functional assays performed in the proband’s target tissues—skeletal and heart muscle—showed biochemical disturbances of the MRC, suggesting a pathogenic role for this variant. In silico analyses initially predicted an amino acid missense change p.(Arg113Lys) in the NDUFB11 CI subunit. However, we showed that the molecular effect of the c.338G>A variant, which is located at the last nucleotide of exon 2 of the NDUFB11 gene in the canonical ‘short’ transcript (sized 462 bp), instead causes a splicing defect triggering the up-regulation of the expression of an alternative ‘long’ transcript (sized 492 bp) that can also be detected in the control individuals. Our results support the hypothesis that the canonical ‘short’ transcript is required for the proper NDUFB11 protein synthesis, which is essential for optimal CI assembly and activity, whereas the longer alternative transcript seems to represent a non-functional, unprocessed splicing intermediate. Our results highlight the importance of characterizing the molecular effect of new variants in the affected patient’s tissues to demonstrate their pathogenicity and association with the clinical phenotypes.

Based on recent success in using modified RNA in clinical applications, we tested the safety, feasibility, and efficacy of direct delivery of citrate-saline-formulated mRNA into an hibernating ischemic heart muscle using an electromechanical mapping and injection catheter system (NOGA/Myostar) in a porcine chronic myocardial ischemia model. Chronic ischemia was induced in domestic pigs (n = 24) using a bottleneck stent placed in the left anterior descending coronary artery. Low (1 mg) and high (7.5 mg) doses of citrate-saline-formulated vascular endothelial growth factor (VEGF)-A165 mRNA were administered in the study. LacZ mRNA and citrate-saline buffer were used as controls. Ten intramyocardial injections (200 μL each) of the mRNAs or citrate-saline buffer were given endovascularly into the hibernating ischemic myocardium using the NOGA catheter. Positron emission tomography 15O-radiowater imaging was performed 7 days after the induction of ischemia and 28 days after the mRNA delivery to measure quantitative myocardial blood perfusion. Coronary angiography, left ventricular function measurements, and clinical chemistry were obtained at each time point. Thirty-five days after the mRNA transfers, pigs were sacrificed, and infarct size and general histology were analyzed. LacZ mRNA pigs were sacrificed 24 h after the transduction. Citrate-saline-formulated mRNA delivery into the ischemic myocardium with endovascular injection catheter did not lead to meaningful transduction with the translation of VEGF-A165, nor therapeutic effects in the heart. VEGF-A165 mRNA showed no statistically significant improvements in left ventricular ejection fraction (LVEF), cardiac output, myocardial perfusion, infarct size, collateral growth, or capillary area in the study groups. However, there was a trend in the high-dose group toward an improved LVEF and cardiac output at rest. No significant adverse effects were observed. In conclusion, the NOGA/Myostar injection catheter system is ineffective in delivering citrate-saline-formulated mRNAs into the heart muscle with the doses and methods used in a porcine chronic myocardial ischemia model. [ABSTRACT FROM AUTHOR]

Despite global efforts, it took 7 months between the proclamation of global SARS-CoV-2 pandemic and the first FDA-approved treatment for COVID-19. During this timeframe, clinicians focused their efforts on repurposing drugs, such as hydroxychloroquine (HCQ) or azithromycin (AZM) to treat hospitalized COVID-19 patients. While clinical trials are time-consuming, the exponential increase in hospitalizations compelled the FDA to grant an emergency use authorization for HCQ and AZM as treatment for COVID-19, although there was limited evidence of their combined efficacy and safety. The authorization was revoked 4 months later, giving rise to controversial political and scientific debates illustrating important challenges such as premature authorization of potentially ineffective or unsafe therapeutics, while diverting resources from screening of effective drugs. Here we report on a preclinical drug screening platform, a cardiac microphysiological system (MPS), to rapidly identify clinically relevant cardiac liabilities associated with HCQ and AZM. The cardiac MPS is a microfabricated fluidic system in which cardiomyocytes derived from human induced pluripotent stem cells self-arrange into a uniaxially beating tissue. The drug response was measured using outputs that correlate with clinical measurements such as action potential duration (proxy for clinical QT interval) and drug-biomarker pairing. The cardiac MPS predicted clinical arrhythmias associated with QT prolongation and rhythm instabilities in tissues treated with HCQ. We found no change in QT interval upon acute exposure to AZM, while still observing a significant increase in arrhythmic events. These results suggest that this MPS can not only predict arrhythmias, but it can also identify arrhythmias even when QT prolongation is absent. When exposed to HCQ and AZM polytherapy, this MPS faithfully reflected clinical findings, in that the combination of drugs synergistically increased QT interval when compared to single drug exposure, while not worsening the overall frequency of arrhythmic events. The high content cardiac MPS can rapidly evaluate the cardiac safety of potential therapeutics, ultimately accelerating patients’ access to safe and effective treatments.

Despite global efforts, it took 7 months between the proclamation of global SARS-CoV-2 pandemic and the first FDA-approved treatment for COVID-19. During this timeframe, clinicians focused their efforts on repurposing drugs, such as hydroxychloroquine (HCQ) or azithromycin (AZM) to treat hospitalized COVID-19 patients. While clinical trials are time-consuming, the exponential increase in hospitalizations compelled the FDA to grant an emergency use authorization for HCQ and AZM as treatment for COVID-19, although there was limited evidence of their combined efficacy and safety. The authorization was revoked 4 months later, giving rise to controversial political and scientific debates illustrating important challenges such as premature authorization of potentially ineffective or unsafe therapeutics, while diverting resources from screening of effective drugs. Here we report on a preclinical drug screening platform, a cardiac microphysiological system (MPS), to rapidly identify clinically relevant cardiac liabilities associated with HCQ and AZM. The cardiac MPS is a microfabricated fluidic system in which cardiomyocytes derived from human induced pluripotent stem cells self-arrange into a uniaxially beating tissue. The drug response was measured using outputs that correlate with clinical measurements such as action potential duration (proxy for clinical QT interval) and drug-biomarker pairing. The cardiac MPS predicted clinical arrhythmias associated with QT prolongation and rhythm instabilities in tissues treated with HCQ. We found no change in QT interval upon acute exposure to AZM, while still observing a significant increase in arrhythmic events. These results suggest that this MPS can not only predict arrhythmias, but it can also identify arrhythmias even when QT prolongation is absent. When exposed to HCQ and AZM polytherapy, this MPS faithfully reflected clinical findings, in that the combination of drugs synergistically increased QT interval when compared to single drug exposure, while not worsening the overall frequency of arrhythmic events. The high content cardiac MPS can rapidly evaluate the cardiac safety of potential therapeutics, ultimately accelerating patients' access to safe and effective treatments. [ABSTRACT FROM AUTHOR]

There is an urgent need for ventricular assist devices (VADs) that can assist in complex biological functions such as the contraction of heart muscle. Although the design improvements implemented with each generation of VADs have helped to increase patient's wellbeing. But the portability, implantability due to the bulkiness, and the lack of pulsatility are continuing to be clinical challenges. In addition, contact between blood and artificial surfaces remains, necessitating long‐term blood‐thinning medications for patients with VADs. Herein, a concept of new VADs is demonstrated that may provide an improved power system with miniaturization and pulsatility control. This work will examine this new generation of electrically contractile polymer‐based actuators for positive inotropic support for both the left and right ventricle (high‐ and low‐pressure system) as cardiomyoplasty. A silicone coated electrothermal actuator is fabricated to engineer VAD, which is able to mimic the pressure on a model. This represents the first successful study demonstrating that artificial muscle can be a decent alternative to support patients with heart muscle weaknesses. [ABSTRACT FROM AUTHOR]

Myosin-based regulation in the heart muscle modulates the number of myosin motors available for interaction with calcium-regulated thin filaments, but the signaling pathways mediating the stronger contraction triggered by stretch between heartbeats or by phosphorylation of the myosin regulatory light chain (RLC) remain unclear. Here, we used RLC probes in demembranated cardiac trabeculae to investigate the molecular structural basis of these regulatory pathways. We show that in relaxed trabeculae at near-physiological temperature and filament lattice spacing, the RLC-lobe orientations are consistent with a subset of myosin motors being folded onto the filament surface in the interacting-heads motif seen in isolated filaments. The folded conformation of myosin is disrupted by cooling relaxed trabeculae, similar to the effect induced by maximal calcium activation. Stretch or increased RLC phosphorylation in the physiological range have almost no effect on RLC conformation at a calcium concentration corresponding to that between beats. These results indicate that in near-physiological conditions, the folded myosin motors are not directly switched on by RLC phosphorylation or by the titin-based passive tension at longer sarcomere lengths in the absence of thin filament activation. However, at the higher calcium concentrations that activate the thin filaments, stretch produces a delayed activation of folded myosin motors and force increase that is potentiated by RLC phosphorylation. We conclude that the increased contractility of the heart induced by RLC phosphorylation and stretch can be explained by a calcium-dependent interfilament signaling pathway involving both thin filament sensitization and thick filament mechanosensing. [ABSTRACT FROM AUTHOR]

A mathematical method is introduced for quantifying full field deformation images of right ventricle (RV) of the heart. These images are acquired from the RV of the heart during open-heart surgery and analysed using digital image correlation (DIC). The high degree of complexity of the deformation of the heart, especially for the patients who require heart surgery emphasises the importance of a method to analyse the visible section of the surface of the RV. This is difficult with conventional heart-monitoring methods, which rely on describing the overall deformation of the area of interest by measuring the deformation between two points only and discarding all other relevant information contained in the area of interest. In this work, we decomposed the full field deformation images of the visible section of RV into shape descriptor vectors and used the Euclidian distance between two shape descriptor vectors obtained from the reference image and the analysed image to quantify the full field deformation by a single number. The Euclidian distance was used to compare the motion and deformation state of the heart at various stages during the operation. We demonstrate that the Euclidian distance is a more robust indicator describing the overall function of the heart than an individual strain value, especially in case of poor-quality images from which the strain is derived. [ABSTRACT FROM AUTHOR]

Purpose: The objective of this study was to reprogram human adipogenic mesenchymal stem cells (hADMSCs) to form Purkinje cells and to use the reprogrammed Purkinje cells to bioprint Purkinje networks. Methods: hADMSCs were reprogrammed to form Purkinje cells using a multi-step process using transcription factors ETS2 and MESP1 to first form cardiac progenitor stem cells followed by SHOX2 and TBX3 to form Purkinje cells. A novel bioprinting method was developed based on Pluronic acid as the sacrificial material and type I collagen as the structural material. The reprogrammed Purkinje cells were used in conjunction with the novel bioprinting method to bioprint Purkinje networks. Printed constructs were evaluated for retention of functional protein connexin 40 (Cx40) and ability to undergo membrane potential changes in response to physiologic stimulus. Results: hADMSCs were successfully reprogrammed to form Purkinje cells based on the expression pattern of IRX3, IRX5, SEMA and SCN10. Reprogrammed purkinje cells were incorporated into a collagen type-1 bioink and the left ventricular Purkinje network was printed using anatomical images of the bovine Purkinje system as reference. Optimization studies demonstrated that 1.8 mg/mL type-I collagen at a seeding density of 300,000 cells per 200 µL resulted in the most functional bioprinted Purkinje networks. Furthermore, bioprinted Purkinje networks formed continuous syncytium, retained expression of vital functional gap junction protein Cx40 post-print, and exhibited membrane potential changes in response to electric stimulation and acetylcholine evaluated by DiBAC4(5), an electrically responsive dye. Conclusion: Based on the results of this study, hADMSCs were successfully reprogrammed to form Purkinje cells and bioprinted to form Purkinje networks. [ABSTRACT FROM AUTHOR]

Myosin-based mechanisms are increasingly recognized as supplementing their better-known actin-based counterparts to control the strength and time course of contraction in both skeletal and heart muscle. Here we use synchrotron small-angle X-ray diffraction to determine the structural dynamics of local domains of the myosin filament during contraction of heart muscle. We show that, although myosin motors throughout the filament contribute to force development, only about 10% of the motors in each filament bear the peak force, and these are confined to the filament domain containing myosin binding protein-C, the “C-zone.” Myosin motors in domains further from the filament midpoint are likely to be activated and inactivated first in each contraction. Inactivated myosin motors are folded against the filament core, and a subset of folded motors lie on the helical tracks described previously. These helically ordered motors are also likely to be confined to the C-zone, and the associated motor conformation reforms only slowly during relaxation. Myosin filament stress-sensing determines the strength and time course of contraction in conjunction with actin-based regulation. These results establish the fundamental roles of myosin filament domains and the associated motor conformations in controlling the strength and dynamics of contraction in heart muscle, enabling those structures to be targeted to develop new therapies for heart disease. [ABSTRACT FROM AUTHOR]

Purpose: The objective of this study was to bioengineer 3D patches from cardiac myocytes that have been reprogrammed from human adipogenic mesenchymal stem cells (hADMSCs). Methods: Human adipogenic mesenchymal stem cells (hADMSCs) were reprogrammed to form cardiac myocytes using transcription factors ETS2 and MESP1. Reprogrammed cardiac myocytes were cultured in a fibrin gel to bioengineer 3D patch patches. The effect of initial plating density (1–25 million cells per patch), time (28-day culture period) and treatment with 1 μM isoproterenol and 1 μM epinephrine were evaluated. Results: 3D patches were fabricated using cardiac myocytes that have been reprogrammed from hADMSCs. Based on optimization studies, it was determined that 10 million cells were needed to bioengineer a single patch, that measured 2 × 2 cm2. Furthermore, 3D patches fabricated 10 million cells were stable in culture for up to 28 days. Treatment of 3D patches with 1 μM isoproterenol and 1 μM epinephrine resulted in an increase in the electrical properties, as measured by electrical impulse amplitude and frequency. An increase in the expression of mTOR, KCNV1, GJA5, KCNJ16, CTNNT2, KCNV2, MYO3, FOXO1 and KCND2 was noted in response to treatment of 3D patches with isoproterenol and epinephrine. Conclusion: Based on the results of this study, there is evidence to support the successful fabrication of a highly functional 3D patches with measurable electrical activity using cardiac myocytes reprogrammed from hADMSCs. 3D patches fabricated using optimal conductions described in this study can be used to improve the functional properties of failing hearts. Predominantly, in case of the infarcted hearts with partial loss of electrical activity, the electrical properties of the 3D patches may restore the electrical activity of the heart. [ABSTRACT FROM AUTHOR]

Digital Image Correlation (DIC) was used for studying the anisotropic behavior of the thin walled right ventricle of the human heart. Strains measured with Speckle Tracking Echocardiography (STE) were compared with the DIC data. Both DIC and STE were used to measure longitudinal strains of the right ventricle in the beginning of an open-heart surgery as well as after the cardiopulmonary bypass. Based on the results, the maximum end-systolic strains obtained with the DIC and STE change similarly during the surgery with less than 10% difference. The difference is largely due to the errors in matching the longitudinal direction in the two methods, sensitivity of the measurement to the positioning of the virtual extensometer of in both STE and DIC, and physiological difference of the measurements as the DIC measures the top surface of the heart whereas the STE obtains the data from below. The anisotropy of the RV was measured using full field principal strains acquired from the DIC displacement fields. The full field principal strains cover the entire region of interest instead of just two points as the virtual extensometer approach used by the STE. The principal strains are not direction dependent measures, and therefore are more independent of the anatomy of the patient and the exact positioning of the virtual strain gage or the STE probe. The results show that the longitudinal strains alone are not enough to fully characterize the behavior of the heart, as the deformation of the heart can be very anisotropic, and the anisotropy changes during the surgery, and from patient to patient. [ABSTRACT FROM AUTHOR]

Abstract To date, the mechanical loads imposed on isolated cardiac muscle tissue in vitro have been oversimplified. Researchers typically applied loads that are time‐invariant, resulting in either isometric and auxotonic contractions, or flat‐topped (isotonic shortening) work‐loops. These contraction types do not fully capture the dynamic response of contracting tissues adapting to a variable load, such as is experienced by ventricular tissue in vivo. In this study, we have successfully developed a loading system that presents a model‐based, time‐varying, continuously updated, load to cardiac tissue preparations. We combined a Windkessel model of vascular fluid impedance together with Laplace's Law and encoded it in a real‐time hardware‐based force‐length control system. Experiments were carried out on isolated rat left ventricular trabeculae; we directly compare the work‐loops arising from this protocol with those of a typical simplified isotonic shortening work‐loop system. We found that, under body conditions, cardiac trabeculae achieved greater mechanical work output against our new loading system, than with the simplified isotonic work‐loop protocol. We further tested whether loading the tissue with a mechanical impedance defined by “diseased” Windkessel model parameters had an effect on the performance of healthy trabeculae. We found that trabecula shortening decreased when applying the set of Windkessel parameters describing the hypertensive condition, and increased in the hypotensive state. Our implementation of a real‐time model of arterial characteristics provides an improved, physiologically derived, instantly calculated load for use in studying isolated cardiac muscle, and is readily applicable to study various disease conditions.

With the development of advanced focusing optics for x-rays, we can now use x-ray beams with spot sizes in the micro- or nanometer range to scan cells and large areas of tissues and continuously record the diffraction signals. From this data, x-ray scattering maps or so-called x-ray darkfield images are computed showing how different types of cells or regions of tissues differ in their diffraction intensity. At the same time a diffraction pattern is available for each scan point which encodes the local nanostructure, averaged over many contributing constituents illuminated by the beam. In this work we have exploited these new capabilities of scanning x-ray diffraction to investigate cardiac muscle cells as well as cardiac tissue. We give examples of how cardiac cells, especially living, cultured cells, can be prepared to be compatible with the instrumentation constraints of nano- or micro-diffraction instruments. Furthermore, we show how the developmental stage, ranging from neonatal to adult cells, as well as the final preparation state of the cardiomyocytes influences the recorded scattering signal and how these diffraction signals compare to the structure of a fully developed cardiac muscle. [ABSTRACT FROM AUTHOR]

Force enhancement (FE) is a phenomenon that is present in skeletal muscle. It is characterized by progressive forces upon active stretching—distinguished by a linear rise in force—and enhanced isometric force following stretching (residual FE (RFE)). In skeletal muscle, non-cross-bridge (XB) structures may account for this behaviour. So far, it is unknown whether differences between non-XB structures within the heart and skeletal muscle result in deviating contractile behaviour during and after eccentric contractions. Thus, we investigated the force response of intact cardiac trabeculae during and after isokinetic eccentric muscle contractions (10% of maximum shortening velocity) with extensive magnitudes of stretch (25% of optimum muscle length). The different contributions of XB and non-XB structures to the total muscle force were revealed by using an actomyosin inhibitor. For cardiac trabeculae, we found that the force–length dynamics during long stretch were similar to the total isometric force–length relation. This indicates that no (R)FE is present in cardiac muscle while stretching the muscle from 0.75 to 1.0 optimum muscle length. This finding is in contrast with the results obtained for skeletal muscle, in which (R)FE is present. Our data support the hypothesis that titin stiffness does not increase with activation in cardiac muscle. [ABSTRACT FROM AUTHOR]

Easily assembled organotypic co-cultures have long been sought in medical research. In vitro tissue constructs with faithful representation of in vivo tissue characteristics are highly desirable for screening and characteristic assessment of a variety of tissue types. Cardiac tissue analogs are particularly sought after due to the phenotypic degradation and difficulty of culture of primary cardiac myocytes. This study utilized magnetic nanoparticles and primary cardiac myocytes in order to levitate and culture multicellular cardiac aggregates (MCAs). Cells were isolated from 2 day old Sprague Dawley rat hearts and subsequently two groups were incubated with either C1: 33 µL nanoshell/million cells or C2: 50 µL nanoshell/million cells. Varying numbers of cells for each concentration were cultured in a magnetic field in a 24 well plate and observed over a period of 12 days. Constructs generally formed spherical structures. Masson’s trichrome staining of a construct shows the presence of extracellular matrix protein, indicating the presence of functional fibroblasts. Many constructs exhibited noticeable contraction after 4 days of culture and continued contracting noticeably past day 9 of culture. Noticeable contractility indicates the presence of functional primary cardiac myocytes in culture. Phenotypic conservation of cardiac cells was ascertained using IHC staining by α-actinin and collagen. CD31 and fibrinogen were probed in order to assess localization of fibroblasts and endothelial cells. The study verifies a protocol for the use of magnetic levitation in order to rapidly assemble 3D cardiac like tissue with phenotypic and functional stability.

Exercise induces changes in muscle fibers and the extracellular matrix that may depend on elastin content and the activity of proteolytic enzymes. We investigated the influence of endurance training on the gene expression and protein content and/or activity of elastin, elastase, cathepsin K, and plasmin in skeletal and heart muscles and in the aorta. Healthy rats were randomly divided into untrained (n=10) and trained (n=10; 6 weeks of endurance training with increasing load) groups. Gene expression was evaluated via qRT-PCR. Elastin content was measured via enzyme-linked immunosorbent assay and enzyme activity was measured fluorometrically. Elastin content was significantly higher in skeletal (P=0.0014) and heart muscle (P=0.000022) from trained rats versus untrained rats, but not in the aorta. Although mRNA levels in skeletal muscle did not differ between groups, the activities of elastase (P=0.0434), cathepsin K (P=0.0343) and plasmin (P=0.000046) were higher in trained rats. The levels of cathepsin K (P=0.0288) and plasminogen (P=0.0005) mRNA were higher in heart muscle from trained rats, but enzyme activity was not. Enzyme activity in the aorta did not differ between groups. Increased elastin content in muscles may result in better adaption to exercise, as may remodeling of the extracellular matrix in skeletal muscle.

Introduction: Serum cardiac troponin-I is a specific biomarker and raised significantly after Myocardial Infarction. Saliva is a non-interventional bio-fluid which contains portion of serum constituents and can be key in diagnosis of MI. Aim: To estimate and correlate the level of cardiac Troponin-I (cTnI) in un-stimulated whole saliva and serum in acute MI patients and control group. Materials and Methods: Total of 60 individuals were enrolled and equally divided in to study (group I) and control group (Group II). Informed consent form was taken from all the subjects. Saliva and blood samples were obtained from patients with ECG features suggestive of acute MI within 24 hours. Serum and saliva samples were processed further for cTnI. The results obtained were then statistically analysed. Results: The mean cTnI level in serum of group I and group II were found to be 4.27±1.79 mg/L and 0.158±0.05 mg/L respectively. The mean cTnI level in saliva of group I and group II was found to be 0.67±0.10ng/L and 0.160±0.05 ng/L respectively. On analysing serum values and saliva values, p-value was found to be significant’. Conclusion: The saliva levels of cTnI were directly associated with serum levels demonstrating a highly significant strong positive relation and confirms the diagnostic ability of saliva for detection of cTnI.

Introduction: Serum cardiac troponin-I is a specific biomarker and raised significantly after Myocardial Infarction. Saliva is a non-interventional bio-fluid which contains portion of serum constituents and can be key in diagnosis of MI. Aim: To estimate and correlate the level of cardiac Troponin-I (cTnI) in un-stimulated whole saliva and serum in acute MI patients and control group. Materials and Methods: Total of 60 individuals were enrolled and equally divided in to study (group I) and control group (Group II). Informed consent form was taken from all the subjects. Saliva and blood samples were obtained from patients with ECG features suggestive of acute MI within 24 hours. Serum and saliva samples were processed further for cTnI. The results obtained were then statistically analysed. Results: The mean cTnI level in serum of group I and group II were found to be 4.27±1.79 mg/L and 0.158±0.05 mg/L respectively. The mean cTnI level in saliva of group I and group II was found to be 0.67±0.10ng/L and 0.160±0.05 ng/L respectively. On analysing serum values and saliva values, p-value was found to be significant'. Conclusion: The saliva levels of cTnI were directly associated with serum levels demonstrating a highly significant strong positive relation and confirms the diagnostic ability of saliva for detection of cTnI. [ABSTRACT FROM AUTHOR]

The transcription factors GATA4, GATA5 and GATA6 are important regulators of heart muscle differentiation (cardiomyogenesis), which function in a partially redundant manner. We identified genes specifically regulated by individual cardiogenic GATA factors in a genome-wide transcriptomics analysis. The genes regulated by gata4 are particularly interesting because GATA4 is able to induce differentiation of beating cardiomyocytes in Xenopus and in mammalian systems. Among the specifically gata4- regulated transcripts we identified two SoxF family members, sox7 and sox18 . Experimental reinstatement of gata4 restores sox7 and sox18 expression, and loss of cardiomyocyte differentiation due to gata4 knockdown is partially restored by reinstating sox7 or sox18 expression, while (as previously reported) knockdown of sox7 or sox18 interferes with heart muscle formation. In order to test for conservation in mammalian cardiomyogenesis, we confirmed in mouse embryonic stem cells (ESCs) undergoing cardiomyogenesis that knockdown of Gata4 leads to reduced Sox7 (and Sox18 ) expression and that Gata4 is also uniquely capable of promptly inducing Sox7 expression. Taken together, we identify an important and conserved gene regulatory axis from gata4 to the SoxF paralogs sox7 and sox18 and further to heart muscle cell differentiation. [ABSTRACT FROM AUTHOR]