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Electrical and dynamic stimulation are commonly employed to enhance the maturation of engineered cardiac tissue (ECT) derived from human induced pluripotent stem cells (iPSCs), reflecting the physiological environment of the heart. While electrical stimulation mimics natural bioelectrical signals and dynamic stimulation replicates mechanical forces, the combined effects of these stimuli on ECT maturation have not been thoroughly explored. We hypothesized that simultaneous electro-dynamic stimulation would enhance ECT maturation and function more effectively than either stimulus alone. Human iPSC-derived cardiovascular cells were co-cultured with Collagen I and Matrigel for 2 weeks, followed by a comparative analysis of four groups: no stimulation, dynamic stimulation, electrical stimulation, and simultaneous electro-dynamic stimulation. The functionality of ECTs was assessed by measuring contractile capacity and calcium indicators, and histological assessments examined structural maturation. Our results demonstrated that simultaneous electro-dynamic stimulation significantly increased the CM component, elevated TNNT2 mRNA expression levels, and enhanced calcium transient capacity. Additionally, ECTs subjected to simultaneous stimulation exhibited a positive force-frequency relationship in contractility and an elevation in peak calcium flux, indicative of advanced tissue maturation. Moreover, simultaneous stimulation promoted vascular network formation within the ECTs, suggesting improved structural organization. These findings underscore the importance of simultaneous stimulation for developing effective cardiac tissue engineering strategies., Competing Interests: Declaration of competing interest This research was supported by RIKEN BDR Organoid Project (to H.M.), and Grants-in-Aid for Scientific Research from the Ministry of Education, Science, Sports, and Culture of Japan (to H.M.) (#23K24420). W.M., K.Murata and H.M. are inventors of engineered cardiac tissue-related patents. All authors declare that they have no financial conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Cardiac biomarkers, especially high-sensitivity cardiac troponin C or I (hs-cTnC or hs-cTnI, respectively), are vital for diagnosing acute myocardial infarction (AMI). Despite the specificity of hs-cTn as a biomarker, the creatine kinase-myocardial band (CK-MB) is commonly used alongside hs-cTn in emergency departments (EDs). We analyzed 23,771 simultaneous hs-cTn (hs-cTnT or hs-cTnI) and CK-MB requests for 17,185 patients in tertiary hospital ED in 2022. The objective of this study was to assess their practical value in diagnosing AMI in real-world settings. Among all 17,185 patients tested, 98.0% underwent hs-cTnT and CK-MB tests, and substantially fewer underwent hs-cTnI testing. We observed concordance between the initial hs-cTn and CK-MB results in 71.3% of patients. Of 131 AMI cases, 57 were positive for both biomarkers, 63 for hs-cTn only, and none for CK-MB alone. CK-MB positivity was often found in the absence of AMI. Discrepancies between the hs-cTnT and hs-cTnI results occurred in 30.0% of patients. Indiscriminate CK-MB testing for diagnosing AMI in EDs should be reconsidered. Efficient use of CK-MB is important for reducing costs and ensuring optimal patient care.

Background: Impaired left ventricular relaxation, high filling pressures, and dysregulation of Ca 2+ homeostasis are common findings contributing to diastolic dysfunction in hypertrophic cardiomyopathy (HCM). Studies have shown that impaired relaxation is an early observation in the sarcomere-gene-positive preclinical HCM cohort, which suggests the potential involvement of myofilament regulators in relaxation. A molecular-level understanding of mechanism(s) at the level of the myofilament is lacking. We hypothesized that mutation-specific, allosterically mediated, changes to the cTnC (cardiac troponin C)-cTnI (cardiac troponin I) interface can account for the development of early-onset diastolic dysfunction via decreased PKA accessibility to cTnI., Methods: HCM mutations R92L-cTnT (cardiac troponin T; Arg92Leu) and Δ160E-cTnT (Glu160 deletion) were studied in vivo, in vitro, and in silico via 2-dimensional echocardiography, Western blotting, ex vivo hemodynamics, stopped-flow kinetics, time-resolved fluorescence resonance energy transfer, and molecular dynamics simulations., Results: The HCM-causative mutations R92L-cTnT and Δ160E-cTnT result in different time-of-onset diastolic dysfunction. R92L-cTnT demonstrated early-onset diastolic dysfunction accompanied by a localized decrease in phosphorylation of cTnI. Constitutive phosphorylation of cTnI (cTnI-D 23 D 24 ) was sufficient to recover diastolic function to non-Tg levels only for R92L-cTnT. Mutation-specific changes in Ca 2+ dissociation rates associated with R92L-cTnT reconstituted with cTnI-D 23 D 24 led us to investigate potential involvement of structural changes in the cTnC-cTnI interface as an explanation for these observations. We probed the interface via time-resolved fluorescence resonance energy transfer revealing a repositioning of the N-terminus of cTnI, closer to cTnC, and concomitant decreases in distance distributions at sites flanking the PKA consensus sequence. Implementing time-resolved fluorescence resonance energy transfer distances as constraints into our atomistic model identified additional electrostatic interactions at the consensus sequence., Conclusions: These data show that the early diastolic dysfunction observed in a subset of HCM is attributable to allosterically mediated structural changes at the cTnC-cTnI interface that impair accessibility of PKA, thereby blunting β-adrenergic responsiveness and identifying a potential molecular target for therapeutic intervention., Competing Interests: None.

Background: We aimed to investigate the effect and potential mechanism of enhancing Neuregulin1 (NRG1)/v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4 (ErbB4) expression on the differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes., Methods: We utilized CRISPR-CAS9 technology to knock in ErbB4 and obtained a single-cell clone IPSN-AAVS1-CMV-ErbB4 (iPSCs-ErbB4). Subsequently, we induced the differentiation of iPSCs into cardiomyocytes and quantified the number of beating embryoid bodies. Furthermore, quantitative real-time PCR assessed the expression of cardiomyocyte markers, including ANP (atrial natriuretic peptide), Nkx2.5 (NK2 transcription factor related locus 5), and GATA4 (GATA binding protein 4). On the 14th day of differentiation, we observed the α-MHC (α-myosin heavy chain)-positive area using immunofluorescent staining and conducted western blotting to detect the expression of cTnT (cardiac troponin) protein and PI3K/Akt signaling pathway-related proteins. Additionally, we intervened the iPSCs-ErbB4 + NRG1 group with the PI3K/Akt inhibitor LY294002 and observed alterations in the expression of cardiomyocyte differentiation-related genes., Results: The number of beating embryoid bodies increased after promoting the expression of NRG1/ErbB4 compared to the iPSCs control group. Cardiomyocyte markers ANP, Nkx2.5, and GATA4 significantly increased on day 14 of differentiation, and the positive area of α-MHC was three times that of the iPSCs control group. Moreover, there was a marked increase in cTnT protein expression. However, there was no significant difference in cardiomyocyte differentiation between the iPSCs-ErbB4 group and the iPSCs control group. Akt phosphorylation was significantly increased in the iPSCs-ErbB4 + NRG1 group. LY294002 significantly reversed the enhancing effect of NRG1/ErbB4 overexpression on Akt phosphorylation as well as the increase in α-MHC and cTnT expression., Conclusions: In conclusion, promoting the expression of NRG1/ErbB4 induced the differentiation of iPSC into cardiomyocytes, possibly through modulation of the PI3K/Akt signaling pathway., (© 2024. The Author(s).)

This study aimed to explore the effects of S-nitrosylation on caspase-3 modification and its subsequent effects on beef myofibril degradation in vitro. Recombinant caspase-3 was reacted with different concentrations of S-nitrosoglutathione (GSNO, nitric oxide donor) at 37 °C for 30 min and subsequently incubated with purified myofibrillar protein from bovine semimembranosus muscle. Results indicated that the activity of caspase-3 was significantly reduced after GSNO treatments ( P < 0.05) and showed a dose-dependent inhibitory effect, which was attributed to the increased S-nitrosylation extent of caspase-3. LC-MS/MS analysis revealed that caspase-3 was S-nitrosylated at cysteine sites 116, 170, 184, 220, and 264. Moreover, the degradation of desmin and troponin-T was notably suppressed by S-nitrosylated caspase-3 ( P < 0.05). To conclude, protein S-nitrosylation could modify the cysteine residues of caspase-3, which accounts for the reduced caspase-3 activity and further represses its proteolytic ability on beef myofibrillar protein.

The binding of Ca 2+ ions within the troponin core of the cardiac thin filament (CTF) regulates normal contraction and relaxation. Mutations within the troponin complexes are known to alter normal functions and result in the eventual development of cardiomyopathy. However, despite the importance of the problem, detailed microscopic knowledge of the mechanism of pathogenic effect of point mutations and their effects on the conformational free energy surface of CTF remains elusive. Mutations are known to transmit their effects hundreds of angstroms along this protein complex and between different component proteins. To explore the impact of point mutations on the conformational free energy barrier between the closed and blocked state of CTF, and to understand the transmission of mutation, we have carried out metadynamics simulations for the wild-type (WT) and two mutants (cardiac troponin T Arg92Trp (R92W) and Arg92Leu (R92L)). Specifically, we have investigated the conformational modification of the tropomyosin (Tm) and the troponin (Tn) complex during the closed-to-blocked state transition for both the WT and two hypertrophic cardiomyopathy causing mutations. Our calculations demonstrated that mutations within the cardiac troponin T (cTnT) protein alter conformational properties of the Tm and the other proteins of the Tn complex as well as the Ca 2+ binding affinity of the cTnC protein through the indirect mediation of cardiac troponin I (cTnI). Importantly, the data revealed a significant influence of the mutations on the conformational transition free energy barriers for both the Tm and cTnC proteins. Furthermore, we found both mutations independently alter the free energy barrier of transitions of cTnT. Such alteration in the free energy upon mutation of one protein in a complex, allosterically affects the others through structural and dynamical changes, leading to a pathogenic effect on the function of the thin filament., (© 2024 The Protein Society.)

Background: Clinically, most patients with lung cancer (LC) die from tumor spread and metastasis. Specific metastasis-related molecules can provide reference for clinical prediction of efficacy, evaluation of prognosis, and search for the best treatment plan. Troponin T1 (TNNT1) is highly expressed in various cancer tissues, which affects malignant behavior of tumor cells and is related to patients' survival and prognosis. However, the role and molecular mechanism of TNNT1 in LC invasion and metastasis have not yet been investigated., Methods: Gene expression profiling interactive analysis (GEPIA) online analysis was used to analyze TNNT1 expression in LC tissues. Quantitative real-time-polymerase chain reaction (qRT-PCR) or western blot were performed to measure TNNT1 or epithelial-to-mesenchymal transition (EMT)-related and Wnt/β-catenin pathway-related protein expression in LC cells. After TNNT1 knockdown, cell scratch healing and transwell assays were introduced to assess cell migration and invasion, respectively., Results: TNNT1 expression in LC tissues and cells was increased. TNNT1 knockdown notably impaired LC cell migration, invasion and EMT. TNNT1 knockdown inhibited Wnt/β-catenin pathway of LC cells. Lithium chloride (LiCl) addition partially restored the inhibition of TNNT1 knockdown on migration, invasion, EMT and Wnt/β-catenin of LC cells., Conclusion: TNNT1 knockdown attenuated LC migration, invasion and EMT, possibly through Wnt/β-catenin signaling., (© 2024 The Author(s). Thoracic Cancer published by John Wiley & Sons Australia, Ltd.)

Dilated cardiomyopathy (DCM) is one of the main causes of sudden cardiac death and heart failure and is the leading indication for cardiac transplantation worldwide. Mutations in dozens of cardiac genes have been connected to the development of DCM including the Troponin T2 gene (TNNT2). Here, we generated a human induced pluripotent stem cells (hiPSCs) from a DCM patient with a familial history that carries a missense mutation in TNNT2. The hiPSCs show typical morphology of pluripotent stem cells, expression of pluripotency markers, normal karyotype, and in vitro capacity to differentiate into all three germ layers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Colorectal cancer (CRC) is one of the most common digestive tract tumors in humans. At present, many scholars believe that the primary site of the tumor has a direct and profound impact on its curative effect. There are significant differences in the expression of many genes, tumor microenvironment, and prognosis between the left and right colon. However, there is a lack of detailed studies on whether the differentially expressed genes in the left and right colon significantly impact the prognosis of patients with CRC. Troponin T1 ( TNNT1 ) is an important gene that affects the prognosis difference between left and right colon cancer screening from "The Cancer Genome Atlas" database. By analyzing the differential gene expression data and clinical data of the left and right hemicolons in the database, the online prognostic database was used to screen the key molecules that significantly affect the tumor immune microenvironment and patient prognosis and to predict their functions and pathways. Quantitative reverse transcription-polymerase chain reaction was used to verify the expression difference of TNNT1 in CRC cell lines SW480 and HCT116, and normal human colorectal epithelial cell line FHC. The relationship between TNNT1 expression in 88 pairs of CRC samples and clinical information and pathologic parameters of patients with CRC was analyzed to judge the impact of TNNT1 expression on patient survival. Database analysis showed that TNNT1 was significantly overexpressed in CRC, and TNNT1 was one of the main differential genes between left colon cancer (LCC) and right colon cancer (RCC). The expression of TNNT1 was significantly increased in RCC, which could lead to poor prognosis of patients. Quantitative reverse transcription-polymerase chain reaction indicated that the expression of TNNT1 was significantly up-regulated in CRC cell lines SW480 and HCT116. Eighty-eight immunohistochemistry (IHC) of CRC tissues and adjacent tissues suggested that the expression of TNNT1 in CRC was significantly higher than that in normal adjacent tissues. By analyzing the clinical information and pathologic indicators matched with these clinical samples, we found that high TNNT1 expression in the primary tumor location (right colon) and high N stage (N2, N3) were unfavorable factors affecting the prognosis of patients with CRC. Multivariate Cox regression analysis suggested that high expression of TNNT1 may be an independent risk factor for the prognosis of patients with CRC. As one of the main differential genes between LCC and RCC, TNNT1 is representative to some extent. Its high expression may be one of the reasons why the prognosis of patients with RCC is worse than that of patients with LCC., Competing Interests: The authors declare no conflict of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Global ischemia has been shown to induce cardiac regenerative response in animal models. One of the suggested mechanisms behind cardiac regeneration is dedifferentiation of cardiomyocytes. How human adult cardiomyocytes respond to global ischemia is not fully known. In this study, biopsies from the left ventricle (LV) and the atrioventricular junction (AVj), a potential stem cell niche, were collected from multi-organ donors with cardiac arrest (N = 15) or without cardiac arrest (N = 6). Using immunohistochemistry, we investigated the expression of biomarkers associated with stem cells during cardiomyogenesis; MDR1, SSEA4, NKX2.5, and WT1, proliferation markers PCNA and Ki67, and hypoxia responsive factor HIF1α. The myocyte nuclei marker PCM1 and cardiac Troponin T were also included. We found expression of cardiac stem cell markers in a subpopulation of LV cardiomyocytes in the cardiac arrest group. The same cells showed a low expression of Troponin T indicating remodeling of cardiomyocytes. No such expression was found in cardiomyocytes from the control group. Stem cell biomarker expression in AVj was more pronounced in the cardiac arrest group. Furthermore, co-expression of PCNA and Ki67 with PCM1 was only found in the cardiac arrest group in the AVj. Our results indicate that a subpopulation of human cardiomyocytes in the LV undergo partial dedifferentiation upon global ischemia and may be involved in the cardiac regenerative response together with immature cardiomyocytes in the AVj., (© 2024. The Author(s).)

Soluble growth stimulation expressed gene 2 protein (sST2) is a myocardial protein induced by biomechanical stress. sST2 is widely present in the serum of patients with heart failure and is recommended as an important indicator to predict adverse outcomes in these patients. However, no postmortem biochemical analysis of sST2 in forensic practice has been reported. The present pilot study aimed to investigate the expression of sST2 in the pericardial fluid of patients with sudden cardiac death (SCD) caused by ischemic heart disease (IHD). In addition, to explore the relationship of sST2 with CK-MB, cTnT, and NT-proBNP, which have been proven to be auxiliary biomarkers for the diagnosis of SCD, we analyzed CK-MB, cTnT, NT-proBNP, and sST2 levels in twenty-one pericardial fluid samples from the Center of Forensic Investigation, China Medical University, with a Roche cobas e 411 electrochemiluminescence automatic immunoassay system and ST2/IL-33R Valukine™ enzyme-linked immunosorbent assay kit. The levels of sST2 in the pericardial fluid of patients with SCD caused by IHD were significantly increased (P < 0.01) and positively correlated with CK-MB and NT-proBNP (P < 0.0001). Receiver operating characteristic curve analysis indicated that the combined measurement of sST2 and NT-proBNP has a higher diagnostic value for SCD caused by IHD than the measurement of either indicator alone. This study preliminarily demonstrated that sST2 in the pericardial fluid was significantly increased in patients with SCD caused by IHD and might be used as a novel auxiliary biomarker for postmortem diagnosis of SCD in forensic practice., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Troponin T1 (TNNT1) plays a crucial role in muscle contraction but its role in cancer, particularly in kidney renal clear cell carcinoma (KIRC), is not well-understood. This study explores the expression, clinical significance and biological functions of TNNT1 in various cancers, with an emphasis on its involvement in KIRC. We analysed TNNT1 expression in cancers using databases like TCGA and GTEx, assessing its prognostic value, mutation patterns, methylation status and functional implications. The study also examined TNNT1's effect on the tumour microenvironment and drug sensitivity in KIRC, complemented by in vitro TNNT1 knockdown experiments in KIRC cells. TNNT1 is overexpressed in several cancers and linked to adverse outcomes, showing frequent upregulation mutations and abnormal methylation. Functionally, TNNT1 connects to muscle and cancer pathways, affects immune infiltration and drug responses, and its overexpression in KIRC is associated with advanced disease and reduced survival. Knocking down TNNT1 curbed KIRC cell growth. TNNT1's aberrant expression plays a significant role in tumorigenesis and immune modulation, highlighting its value as a prognostic biomarker and a potential therapeutic target in KIRC and other cancers. Further studies are essential to understand TNNT1's oncogenic mechanisms in KIRC., (© 2024 The Author(s). Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Importance: Clonal hematopoiesis of indeterminate potential (CHIP) may contribute to the risk of atrial fibrillation (AF) through its association with inflammation and cardiac remodeling., Objective: To determine whether CHIP was associated with AF, inflammatory and cardiac biomarkers, and cardiac structural changes., Design, Setting, and Participants: This was a population-based, prospective cohort study in participants of the Atherosclerosis Risk in Communities (ARIC) study and UK Biobank (UKB) cohort. Samples were collected and echocardiography was performed from 2011 to 2013 in the ARIC cohort, and samples were collected from 2006 to 2010 in the UKB cohort. Included in this study were adults without hematologic malignancies, mitral valve stenosis, or previous mitral valve procedure from both the ARIC and UKB cohorts; additionally, participants without hypertrophic cardiomyopathy and congenital heart disease from the UKB cohort were also included. Data analysis was completed in 2023., Exposures: CHIP (variant allele frequency [VAF] ≥2%), common gene-specific CHIP subtypes (DNMT3A, TET2, ASXL1), large CHIP (VAF ≥10%), inflammatory and cardiac biomarkers (high-sensitivity C-reactive protein, interleukin 6 [IL-6], IL-18, high-sensitivity troponin T [hs-TnT] and hs-TnI, N-terminal pro-B-type natriuretic peptide), and echocardiographic indices., Main Outcome Measure: Incident AF., Results: A total of 199 982 adults were included in this study. In ARIC participants (4131 [2.1%]; mean [SD] age, 76 [5] years; 2449 female [59%]; 1682 male [41%]; 935 Black [23%] and 3196 White [77%]), 1019 had any CHIP (24.7%), and 478 had large CHIP (11.6%). In UKB participants (195 851 [97.9%]; mean [SD] age, 56 [8] years; 108 370 female [55%]; 87 481 male [45%]; 3154 Black [2%], 183 747 White [94%], and 7971 other race [4%]), 11 328 had any CHIP (5.8%), and 5189 had large CHIP (2.6%). ARIC participants were followed up for a median (IQR) period of 7.0 (5.3-7.7) years, and UKB participants were followed up for a median (IQR) period of 12.2 (11.3-13.0) years. Meta-analyzed hazard ratios for AF were 1.12 (95% CI, 1.01-1.25; P = .04) for participants with vs without large CHIP, 1.29 (95% CI, 1.05-1.59; P = .02) for those with vs without large TET2 CHIP (seen in 1340 of 197 209 [0.67%]), and 1.45 (95% CI, 1.02-2.07; P = .04) for those with vs without large ASXL1 CHIP (seen in 314 of 197 209 [0.16%]). Large TET2 CHIP was associated with higher IL-6 levels. Additionally, large ASXL1 was associated with higher hs-TnT level and increased left ventricular mass index., Conclusions and Relevance: Large TET2 and ASXL1, but not DNMT3A, CHIP was associated with higher IL-6 level, indices of cardiac remodeling, and increased risk for AF. Future research is needed to elaborate on the mechanisms driving the associations and to investigate potential interventions to reduce the risk.

Despite progress in generating cardiomyocytes from pluripotent stem cells, these populations often include non-contractile cells, necessitating cardiomyocyte selection for experimental purpose. This study explores a novel cardiomyocyte enrichment mechanism: low-adhesion culture selection. The cardiac cells derived from human induced pluripotent stem cells were subjected to a coating-free low-adhesion culture using bovine serum albumin and high molecular weight dextran sulfate. This approach effectively increased the population of cardiac troponin T-positive cardiomyocytes. Similar results were obtained with commercially available low-adhesion culture dishes. Subsequently, we accessed the practicality of selection of cardiomyocytes using this phenomenon by comparing it with established methods such as glucose-free culture and selection based on puromycin resistance genes. The cardiomyocytes enriched through low-adhesion culture selection maintained autonomous pulsation and responsiveness to beta-stimuli. Moreover, no significant differences were observed in the expression of genes related to subtype commitment and maturation when compared to other selection methods. In conclusion, cardiomyocytes derived from pluripotent stem cells were more low-adhesion culture resistant than their accompanying non-contractile cells, and low-adhesion culture is an alternative method for selection of pluripotent stem cell-derived cardiomyocytes., (© 2024. The Author(s).)

Aim: In patients with end-stage kidney disease (ESKD), it is unclear whether an imbalance between myocardial oxygen supply and demand leads to myocardial injury (MI). This study clarifies the association between the balance of the rate pressure product (RPP), consisting of the systolic blood pressure multiplied by the pulse rate (PR), a marker for myocardial oxygen demand, and hemoglobin (Hb), a marker for oxygen supply, with MI., Methods: A total of 283 consecutive unselected patients for hemodialysis were enrolled in this retrospective, cross-sectional study, and were divided into four groups according to Hb levels (high or low) and RPP. Potential imbalances between myocardial oxygen supply and demand were defined as patients with simultaneous high RPP and low Hb levels. The odds ratio (OR) for MI, defined as cardiac troponin T (cTnT) of ≥ 0.15 ng/mL was investigated using logistic regression analysis between the four patient groups., Results: The mean age was 68.7 years, 71.3% were men, and 52.6% had diabetes. The mean Hb level was 9.0 g/dL, and 20.5% of patients were latently diagnosed with MI. The median RPP and cTnT level was 12,144 and 0.083 ng/mL, respectively. When exposed to simultaneous high RPP with low Hb, OR significantly increased compared with that of the well-balanced group (RPP ＜12,500 and Hb ≥ 9.0 g/dL; OR 3.63, p＜0.05). Similar results were obtained in multivariate analysis after adjusting for confounding variables. These associations were enhanced or weakened when the Hb cut-off level became lower (Hb=8 g/dL) or higher (Hb=10 g/dL)., Conclusions: As the myocardial oxygen supply and demand balance in patients with ESKD is potentially associated with MI, appropriate management for blood pressure, PR, and anemia may prevent MI.

Zebrafish larvae have emerged as a valuable model for studying heart physiology and pathophysiology, as well as for drug discovery, in part thanks to its transparency, which simplifies microscopy. However, in fluorescence-based optical mapping, the beating of the heart results in motion artifacts. Two approaches have been employed to eliminate heart motion during calcium or voltage mapping in zebrafish larvae: the knockdown of cardiac troponin T2A and the use of myosin inhibitors. However, these methods disrupt the mechano-electric and mechano-mechanic coupling mechanisms. We have used ratiometric genetically encoded biosensors to image calcium in the beating heart of intact zebrafish larvae because ratiometric quantification corrects for motion artifacts. In this study, we found that halting heart motion by genetic means (injection of tnnt2a morpholino) or chemical tools (incubation with para -aminoblebbistatin) leads to bradycardia, and increases calcium levels and the size of the calcium transients, likely by abolishing a feedback mechanism that connects contraction with calcium regulation. These outcomes were not influenced by the calcium-binding domain of the gene-encoded biosensors employed, as biosensors with a modified troponin C (Twitch-4), calmodulin (mCyRFP1-GCaMP6f), or the photoprotein aequorin (GFP-aequorin) all yielded similar results. Cardiac contraction appears to be an important regulator of systolic and diastolic Ca 2+ levels, and of the heart rate.

Integration of multiple data sources presents a challenge for accurate prediction of molecular patho-phenotypic features in automated analysis of data from human model systems. Here, we applied a machine learning-based data integration to distinguish patho-phenotypic features at the subcellular level for dilated cardiomyopathy (DCM). We employed a human induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) model of a DCM mutation in the sarcomere protein troponin T (TnT), TnT-R141W, compared to isogenic healthy (WT) control iPSC-CMs. We established a multimodal data fusion (MDF)-based analysis to integrate source datasets for Ca 2+ transients, force measurements, and contractility recordings. Data were acquired for three additional layer types, single cells, cell monolayers, and 3D spheroid iPSC-CM models. For data analysis, numerical conversion as well as fusion of data from Ca 2+ transients, force measurements, and contractility recordings, a non-negative blind deconvolution (NNBD)-based method was applied. Using an XGBoost algorithm, we found a high prediction accuracy for fused single cell, monolayer, and 3D spheroid iPSC-CM models (≥92 ± 0.08 %), as well as for fused Ca 2+ transient, beating force, and contractility models (>96 ± 0.04 %). Integrating MDF and XGBoost provides a highly effective analysis tool for prediction of patho-phenotypic features in complex human disease models such as DCM iPSC-CMs., (© 2024 Walter de Gruyter GmbH, Berlin/Boston.)

Background: Vaccine-related acute myocarditis is recognized as a rare and specific vaccine complication following mRNA-based COVID-19 vaccinations. The precise mechanisms remain unclear. We hypothesized that natural killer (NK) cells play a central role in its pathogenesis., Methods: Samples from 60 adolescents with vaccine-related myocarditis were analyzed, including pro-inflammatory cytokines, cardiac troponin T, genotyping, and immunophenotyping of the corresponding activation subsets of NK cells, monocytes, and T cells. Results were compared with samples from 10 vaccinated individuals without myocarditis and 10 healthy controls., Findings: Phenotypically, high levels of serum cytokines pivotal for NK cells, including interleukin-1β (IL-1β), interferon α2 (IFN-α2), IL-12, and IFN-γ, were observed in post-vaccination patients with myocarditis, who also had high percentage of CD57 + NK cells in blood, which in turn correlated positively with elevated levels of cardiac troponin T. Abundance of the CD57 + NK subset was particularly prominent in males and in those after the second dose of vaccination. Genotypically, killer cell immunoglobulin-like receptor (KIR) KIR2DL5B(-)/KIR2DS3(+)/KIR2DS5(-)/KIR2DS4del(+) was a risk haplotype, in addition to single-nucleotide polymorphisms related to the NK cell-specific expression quantitative trait loci DNAM-1 and FuT11, which also correlated with cardiac troponin T levels in post-vaccination patients with myocarditis., Conclusion: Collectively, these data suggest that NK cell activation by mRNA COVID-19 vaccine contributed to the pathogenesis of acute myocarditis in genetically and epidemiologically vulnerable subjects., Funding: This work was funded by the Hong Kong Collaborative Research Fund (CRF) 2020/21 and the CRF Coronavirus and Novel Infectious Diseases Research Exercises (reference no. C7149-20G)., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objective: We aimed to determine the role of Troponin T1 (TNNT1) in paclitaxel (PTX) resistance and tumor progression in breast cancer (BC)., Methods: Differentially expressed genes were obtained from the GSE4298 and GSE90564 datasets. Hub genes were isolated from protein-protein interaction networks and further validated by real-time quantitative polymerase chain reaction. The effect of TNNT1 on PTX resistance was determined using cell counting kit-8, 5-ethynyl-2'-deoxyuridine, wound healing, transwell, flow cytometry assays, and subcutaneous xenografted tumor model. Western blotting was used to detect proteins associated with PTX resistance, apoptosis, migration, invasion, and other key pathways. Hematoxylin-eosin and immunohistochemical staining were used to evaluate the role of TNNT1 in tumors., Results: After comprehensive bioinformatic analysis, we identified CCND1, IGF1, SFN, INHBA, TNNT1, and TNFSF11 as hub genes for PTX resistance in BC. TNNT1 plays a key role in BC and is upregulated in PTX-resistant BC cells. TNNT1 silencing inhibited PTX resistance, proliferation, migration, and invasion while promoting apoptosis of PTX-resistant BC cells. Tumor xenograft experiments revealed that TNNT1 silencing suppresses PTX resistance and tumor development in vivo. In addition, TNNT1 silencing inhibited the expression of proteins in the rat sarcoma virus (RAS)/rapidly accelerated fibrosarcoma1 (RAF1) pathway in vivo. Treatment with a RAS/RAF1 pathway activator reversed the inhibitory effect of TNNT1 silencing on proliferation, migration, and invasion while promoting apoptosis of PTX resistance BC cells., Conclusion: Silencing of TNNT1 suppresses PTX resistance and BC progression by inhibiting the RAS/RAF1 pathway, which is a promising biomarker and therapeutic target for drug resistance in BC., (© 2023 Wiley Periodicals LLC.)

It is generally accepted that oxidative stress plays a key role in the development of ischemia-reperfusion injury in ischemic heart disease. However, the mechanisms how reactive oxygen species trigger cellular damage are not fully understood. Our study investigates redox state and highly reactive substances within neonatal and adult cardiomyocytes under hypoxia conditions. We have found that hypoxia induced an increase in H 2 O 2 production in adult cardiomyocytes, while neonatal cardiomyocytes experienced a decrease in H 2 O 2 levels. This finding correlates with our observation of the difference between the electron transport chain (ETC) properties and mitochondria amount in adult and neonatal cells. We demonstrated that in adult cardiomyocytes hypoxia caused the significant increase in the ETC loading with electrons compared to normoxia. On the contrary, in neonatal cardiomyocytes ETC loading with electrons was similar under both normoxic and hypoxic conditions that could be due to ETC non-functional state and the absence of the electrons transfer to O 2 under normoxia. In addition to the variations in H 2 O 2 production, we also noted consistent pH dynamics under hypoxic conditions. Notably, the pH levels exhibited a similar decrease in both cell types, thus, acidosis is a more universal cellular response to hypoxia. We also demonstrated that the amount of mitochondria and the levels of cardiac isoforms of troponin I, troponin T, myoglobin and GAPDH were significantly higher in adult cardiomyocytes compared to neonatal ones. Remarkably, we found out that under hypoxia, the levels of cardiac isoforms of troponin T, myoglobin, and GAPDH were elevated in adult cardiomyocytes, while their level in neonatal cells remained unchanged. Obtained data contribute to the understanding of the mechanisms of neonatal cardiomyocytes' resistance to hypoxia and the ability to maintain the metabolic homeostasis in contrast to adult ones., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Introduction: The comorbidities of ischemic heart disease (IHD) and diabetes mellitus (DM) compromise the protection of the diabetic heart from ischemia/reperfusion (I/R) injury. We hypothesized that manipulation of reperfusion injury salvage kinase (RISK) and survivor activating factor enhancement (SAFE) pathways might protect the diabetic heart, and intervention of these pathways could be a new avenue for potentially protecting the diabetic heart., Methods: All hearts were subjected to 30-min ischemia and 30-min reperfusion. During reperfusion, hearts were exposed to molecules proven to protect the heart from I/R injury. The hemodynamic data were collected using suitable software. The infarct size, troponin T levels, and protein levels in hearts were evaluated., Results: Both cyclosporine-A and nitric oxide donor (SNAP) infusion at reperfusion protected 4-week diabetic hearts from I/R injury. However, 6-week diabetic hearts were protected only by SNAP, but not cyclosporin-A. These treatments significantly (p &lt; 0.05) improved cardiac hemodynamics and decreased infarct size., Conclusions: The administration of SNAP to diabetic hearts protected both 4- and 6-week diabetic hearts; however, cyclosporine-A protected only the 4-week diabetic hearts. The eNOS/GLUT-4 pathway executed the SNAP-mediated cardioprotection., (© 2024 S. Karger AG, Basel.)

Myogenesis is a highly orchestrated process whereby muscle precursor cells, myoblasts, develop into muscle fibers to form skeletal muscle during embryogenesis and regenerate adult muscle. Here, we studied the RNA-binding protein FUS (fused in sarcoma), which has been implicated in muscular and neuromuscular pathologies but is poorly characterized in myogenesis. Given that FUS levels declined in human and mouse models of skeletal myogenesis, and that silencing FUS enhanced myogenesis, we hypothesized that FUS might be a repressor of myogenic differentiation. Interestingly, overexpression of FUS delayed myogenesis, accompanied by slower production of muscle differentiation markers. To identify the mechanisms through which FUS inhibits myogenesis, we uncovered RNA targets of FUS by ribonucleoprotein immunoprecipitation (RIP) followed by RNA-sequencing (RNA-seq) analysis. Stringent selection of the bound transcripts uncovered Tnnt1 mRNA, encoding troponin T1 (TNNT1), as a major effector of FUS influence on myogenesis. We found that in myoblasts, FUS retained Tnnt1 mRNA in the nucleus, preventing TNNT1 expression; however, reduction of FUS during myogenesis or by silencing FUS released Tnnt1 mRNA for export to the cytoplasm, enabling TNNT1 translation and promoting myogenesis. We propose that FUS inhibits myogenesis by suppressing TNNT1 expression through a mechanism of nuclear Tnnt1 mRNA retention.

Introduction: Heart failure with preserved ejection fraction (HFpEF) is a common syndrome with high morbidity and mortality but without available evidence-based therapies. It is essential to investigate changes in gene expression profiles in preclinical HFpEF animal models, with the aim of searching for novel therapeutic targets., Methods: Wild-type male C57BL/6J mice were administrated with a combination of high-fat diet (HFD) and inhibition of constitutive nitric oxide synthase using N-nitro-l-arginine methyl ester (l-NAME) for 5 and 7 weeks. RNA sequencing was conducted to detect gene expression profiles, and bioinformatic analysis was performed to identify the core genes, pathways, and biological processes involved., Results: A total of 1,347 genes were differentially expressed in the heart at week 5 and 7 post-intervention. Gene Ontology enrichment analysis indicated that these greatly changed genes were involved mainly in cell adhesion, neutrophil chemotaxis, cell communication, and other functions. Using hierarchical cluster analysis, these differentially expressed genes were classified into 16 profiles. Of these, three significant profiles were ultimately identified. Gene co-expression network analysis suggested troponin T type 1 (Tnnt1) directly regulated 31 neighboring genes and was considered to be at the core of the associated gene network., Conclusion: The combined application of RNA sequencing, hierarchical cluster analysis, and gene network analysis identified Tnnt1 as the most important gene in the development of HFpEF., (© 2024 S. Karger AG, Basel.)

In this perspective, we discussed emerging data indicating a role for Notch signalling in inherited disorders of the heart failure with focus on hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM) linked to variants of genes encoding mutant proteins of the sarcomere. We recently reported an upregulation of elements in the Notch signalling cascade in cardiomyocytes derived from human inducible pluripotent stem cells expressing a TNNT2 variant encoding cardiac troponin T (cTnT-I79N +/- ), which induces hypertrophy, remodelling, abnormalities in excitation-contraction coupling and electrical instabilities (Shafaattalab S et al. 2021 Front . Cell Dev. Biol . 9 , 787581. (doi:10.3389/fcell.2021.787581)). Our search of the literature revealed the novelty of this finding and stimulated us to discuss potential connections between the Notch signalling pathway and familial cardiomyopathies. Our considerations focused on the potential role of these interactions in arrhythmias, microvascular ischaemia, and fibrosis. This finding underscored a need to consider the role of Notch signalling in familial cardiomyopathies which are trigged by sarcomere mutations engaging mechano-signalling pathways for which there is evidence of a role for Notch signalling with crosstalk with Hippo signalling. Our discussion included a role for both cardiac myocytes and non-cardiac myocytes in progression of HCM and DCM. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Obesity and hypertension, independently and combined, are associated with increased risk of heart failure and heart failure-related morbidity and mortality. Interest in circulating endothelial cell-derived microvesicles (EMVs) has intensified because of their involvement in the development and progression of endothelial dysfunction, atherosclerosis, and cardiomyopathy. The experimental aim of this study was to determine, in vitro, the effects of EMVs isolated from obese/hypertensive adults on key proteins regulating cardiomyocyte hypertrophy [cardiac troponin T (cTnT), α-actinin, nuclear factor-kB (NF-kB)] and fibrosis [transforming growth factor (TGF)-β, collagen1-α1], as well as endothelial nitric oxide synthase (eNOS) expression and nitric oxide (NO) production. EMVs (CD144 + microvesicles) were isolated from plasma by flow cytometry in 12 normal weight/normotensive [8 males/4 females; age: 56 ± 5 yr; body mass index (BMI): 23.3 ± 2.0 kg/m 2 ; blood pressure (BP): 117/74 ± 4/5 mmHg] and 12 obese/hypertensive (8 males/4 females; 57 ± 5 yr; 31.7 ± 1.8 kg/m 2 ; 138/83 ± 8/7 mmHg) adults. Human-induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) were cultured and treated with EMVs from either normal weight/normotensive or obese/hypertensive adults for 24 h. Expression of cTnT (64.1 ± 13.9 vs. 29.5 ± 7.8 AU), α-actinin (66.0 ± 14.7 vs. 36.2 ± 10.3 AU), NF-kB (166.3 ± 13.3 vs. 149.5 ± 8.8 AU), phosphorylated-NF-kB (226.1 ± 25.2 vs. 179.1 ± 25.5 AU), and TGF-β (62.1 ± 13.3 vs. 23.5 ± 8.8 AU) were significantly higher and eNOS activation (16.4 ± 4.3 vs. 24.8 ± 3.7 AU) and nitric oxide production (6.8 ± 1.2 vs. 9.6 ± 1.3 µmol/L) were significantly lower in iPSC-CMs treated with EMVs from obese/hypertensive compared with normal weight/normotensive adults. These data indicate that EMVs from obese/hypertensive adults induce a cardiomyocyte phenotype prone to hypertrophy, fibrosis, and reduced nitric oxide production, central factors associated with heart failure risk and development. NEW & NOTEWORTHY In the present study we determined the effect of endothelial microvesicles (EMVs) isolated from obese/hypertensive adults on mediators of cardiomyocyte hypertrophy [cardiac troponin T (cTnT), α-actinin, nuclear factor-kB (NF-kB)] and fibrosis [transforming growth factor (TGF-β), collagen1-α1] as well as endothelial nitric oxide synthase (eNOS) expression and NO production. EMVs from obese/hypertensive induced significantly higher expression of hypertrophic (cTnT, α-actinin, NF-kB) and fibrotic (TGF-β) proteins as well as significantly lower eNOS activation and NO production in cardiomyocytes than EMVs from normal weight/normotensive adults. EMVs are a potential mediating factor in the increased risk of cardiomyopathy and heart failure with obesity/hypertension.

In this study, the effects of different oxidation intensities on the degradation of myofibrillar protein by endogenous enzymes in iron-catalyzed oxidizing (IOS) and metmyoglobin oxidizing system (MOS) were compared. The results showed that carbonyl content and endogenous enzyme activities (caspase-3, caspase-6 and calpain-1) increased significantly and the total sulfhydryl content decreased significantly with H2O2 concentration in both oxidation systems. Meanwhile, the rate of carbonyl formation and the inhibition of endogenous enzymes activities of IOS were significantly lower than MOS for the same oxidation intensity. In addition, IOS and MOS mainly produced myosin light chains degradation products of 20-25 kDa and 20-17 kDa. At the same oxidation intensity, MOS of myofibrillar protein significantly enhanced the degradation of troponin-T and desmin by caspase-3/-6 compared with IOS, while inhibiting the degradation of troponin-T by calpain-1. In conclusion, MOS inhibited endogenous enzyme degradation in vitro more than IOS during post-slaughter maturation of yak meat., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The cardiac troponins (cTns), cardiac troponin C (cTnC), cTnT, and cTnI are key elements of myocardial apparatus, fixed as protein complex on the thin filament of sarcomere and are involved in the regulation of excitation-contraction coupling of cardiomyocytes in the presence of Ca 2+ . Circulating cTnT and cTnI (cTns) increase following cardiac tissue necrosis, and they are consolidated biomarkers of acute myocardial infarction (AMI). However, the use of high sensitivity (hs)-immunoassay tests for cTnT and cTnI has made it possible to identify a multitude of other clinical conditions associated with increased circulating levels of cTns. cTns can be measured also in the peripheral circulation of healthy subjects or athletes, suggesting that different mechanisms are involved in the release of cTns in the blood independently of cardiac cell necrosis. In this review, the molecular/cellular mechanisms involved in cTns release in blood and the exploitation of cTnI and cTnT as biomarkers of cardiac adverse events, in addition to cardiac necrosis, are discussed., (© 2022 International Union of Biochemistry and Molecular Biology.)

Cardiac muscle troponin T protein binds to tropomyosin and regulates the calcium-dependent actin-myosin interaction on thin filaments in cardiomyocytes. Recent genetic studies have revealed that TNNT2 mutations are strongly linked to dilated cardiomyopathy (DCM). In this study, we generated YCMi007-A, a human induced pluripotent stem cell (hiPSC) line from a DCM patient with a p. Arg205Trp mutation in the TNNT2 gene. The YCMi007-A cells show high expression of pluripotent markers, normal karyotype, and differentiation into three germ layers. Thus, YCMi007-A-an established iPSC-could be useful for the investigation of DCM., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Insects are the most widely distributed and successful animals on the planet. A large number of insects are capable of flight with functional wings. Wing expansion is an important process for insects to achieve functional wings after eclosion and healthy genital morphology is crucial for adult reproduction. Myofilaments are functional units that constitute sarcomeres and trigger muscle contraction. Here, we identified four myofilament proteins, including Myosin, Paramyosin, Tropomyosin and Troponin T, from the wing pads of nymphs in the American cockroach, Periplaneta americana. RNAi-mediated knockdown of Myosin, Paramyosin, Tropomyosin and Troponin T in the early stage of final instar nymphs caused a severely curly wing phenotype in the imaginal moult, especially in the Paramyosin and Troponin T knockdown groups, indicating that these myofilament proteins are involved in controlling wing expansion behaviours during the nymph-adult transition. In addition, the knockdown resulted in abnormal external genitalia, caused ovulation failure, and affected male accessory gland development. Interestingly, the expression of myofilament genes was induced by methoprene, a juvenile hormone (JH) analogue, and decreased by the depletion of the JH receptor gene Met. Altogether, we have determined that myofilament genes play an important role in promoting wing expansion and maintaining adult genitalia morphology, and their expression is induced by JH signalling. Our data reveal a novel mechanism by which wing expansion is regulated by myofilaments and the functions of myofilaments are involved in maintaining genitalia morphology., (© 2022 Royal Entomological Society.)

Introduction: N-terminal pro-B-type natriuretic peptide (NT-proBNP) and cardiac troponin T (cTnT) measurements are recommended in patients with acute dyspnea. We aimed to assess the prognostic merit of cTnT compared to NT-proBNP for 30-day readmission or death in patients hospitalized with acute dyspnea., Methods: We measured cTnT and NT-proBNP within 24 h in 314 patients hospitalized with acute dyspnea and adjudicated the cause of the index admission. Time to first event of readmission or death ≤30 days after hospital discharge was recorded, and cTnT and NT-proBNP measurements were compared head-to-head., Results: Patients who died (12/314) or were readmitted (71/314) within 30 days had higher cTnT concentrations (median: 32.6, Q1-Q3: 18.4-74.2 ng/L vs. median: 19.4, Q1-Q3: 8.4-36.1 ng/L; p for comparison &lt;0.001) and NT-proBNP concentrations (median: 1,753.6, Q1-Q3: 464.2-6,862.0 ng/L vs. median 984, Q1-Q3 201-3,600 ng/L; for comparison p = 0.027) compared to patients who survived and were not readmitted. cTnT concentrations were associated with readmission or death within 30 days after discharge both in the total cohort (adjusted hazard ratio [aHR]: 1.64, 95% confidence interval [CI]: 1.30-2.05) and in patients with heart failure (HF) (aHR: 1.58, 95% CI: 1.14-2.18). In contrast, NT-proBNP concentrations were not associated with short-term events, neither in the total cohort (aHR: 1.10, 95% CI: 0.94-1.30) nor in patients with adjudicated HF (aHR: 1.06, 95% CI: 0.80-1.40)., Conclusion: cTnT concentrations are associated with 30-day readmission or death in patients hospitalized with acute dyspnea, as well as in patients adjudicated HF., (© 2023 The Author(s). Published by S. Karger AG, Basel.)

Cardiomyopathies have unresolved genotype-phenotype relationships and lack disease-specific treatments. Here we provide a framework to identify genotype-specific pathomechanisms and therapeutic targets to accelerate the development of precision medicine. We use human cardiac electromechanical in-silico modelling and simulation which we validate with experimental hiPSC-CM data and modelling in combination with clinical biomarkers. We select hypertrophic cardiomyopathy as a challenge for this approach and study genetic variations that mutate proteins of the thick (MYH7 R403Q/+ ) and thin filaments (TNNT2 R92Q/+ , TNNI3 R21C/+ ) of the cardiac sarcomere. Using in-silico techniques we show that the destabilisation of myosin super relaxation observed in hiPSC-CMs drives disease in virtual cells and ventricles carrying the MYH7 R403Q/+ variant, and that secondary effects on thin filament activation are necessary to precipitate slowed relaxation of the cell and diastolic insufficiency in the chamber. In-silico modelling shows that Mavacamten corrects the MYH7 R403Q/+ phenotype in agreement with hiPSC-CM experiments. Our in-silico model predicts that the thin filament variants TNNT2 R92Q/+ and TNNI3 R21C/+ display altered calcium regulation as central pathomechanism, for which Mavacamten provides incomplete salvage, which we have corroborated in TNNT2 R92Q/+ and TNNI3 R21C/+ hiPSC-CMs. We define the ideal characteristics of a novel thin filament-targeting compound and show its efficacy in-silico. We demonstrate that hybrid human-based hiPSC-CM and in-silico studies accelerate pathomechanism discovery and classification testing, improving clinical interpretation of genetic variants, and directing rational therapeutic targeting and design., (© 2022. The Author(s).)

Tropomyosin (Tpm) mutations cause inherited cardiac diseases such as hypertrophic and dilated cardiomyopathies. We applied various approaches to investigate the role of cardiac troponin (Tn) and especially the troponin T (TnT) in the pathogenic effects of Tpm cardiomyopathy-associated mutations M8R, K15N, A277V, M281T, and I284V located in the overlap junction of neighboring Tpm dimers. Using co-sedimentation assay and viscosity measurements, we showed that TnT1 (fragment of TnT) stabilizes the overlap junction of Tpm WT and all Tpm mutants studied except Tpm M8R. However, isothermal titration calorimetry (ITC) indicated that TnT1 binds Tpm WT and all Tpm mutants similarly. By using ITC, we measured the direct K D of the Tpm overlap region, N-end, and C-end binding to TnT1. The ITC data revealed that the Tpm C-end binds to TnT1 independently from the N-end, while N-end does not bind. Therefore, we suppose that Tpm M8R binds to TnT1 without forming the overlap junction. We also demonstrated the possible role of Tn isoform composition in the cardiomyopathy development caused by M8R mutation. TnT1 dose-dependently reduced the velocity of F-actin-Tpm filaments containing Tpm WT, Tpm A277V, and Tpm M281T mutants in an in vitro motility assay. All mutations impaired the calcium regulation of the actin-myosin interaction. The M281T and I284V mutations increased the calcium sensitivity, while the K15N and A277V mutations reduced it. The Tpm M8R, M281T, and I284V mutations under-inhibited the velocity at low calcium concentrations. Our results demonstrate that Tpm mutations likely implement their pathogenic effects through Tpm interaction with Tn, cardiac myosin, or other protein partners.

Synthetic scaffolds are needed for generating organized neo-myocardium constructs to promote functional tissue repair. This study investigated the biocompatibility of an elastomeric electrospun degradable polar/hydrophobic/ionic polyurethane (D-PHI) composite scaffold with human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The composite material was electrospun to generate scaffolds, with nanofibres oriented in aligned or random directions. These features enabled the authors to evaluate the effect of characteristic elements which mimic that of the native extracellular matrix (alignment, chemical heterogeneity, and fiber topography) on hiPSC-CMs activity. The functional nature of the hiPSC-CM cultured on gelatin and Matrigel-coated scaffolds were assessed, investigating the influence of protein interactions with the synthetic substrate on subsequent cell phenotype. After 7 days of culture, high hiPSC-CM viability was observed on the scaffolds. The cells on the aligned scaffold were elongated and demonstrated aligned sarcomeres that oriented parallel to the direction of the fibers, while the cells on random scaffolds and a tissue culture polystyrene (TCPS) control did not exhibit such an organized morphology. The hiPSC-CMs cultured on the scaffolds and TCPS expressed similar levels of cardiac troponin-T, but there was a higher expression of ventricular myosin light chain-2 on the D-PHI composite scaffolds versus TCPS, indicating a higher proportion of hiPSC-CM exhibiting a ventricular cardiomyocyte like phenotype. Within 7 days, the hiPSC-CMs on aligned scaffolds and TCPS beat synchronously and had similar conductive velocities. These preliminary results show that aligned D-PHI elastomeric scaffolds allow hiPSC-CMs to demonstrate important cardiomyocytes characteristics, critical to enabling their future potential use for cardiac tissue regeneration., (© 2022 Wiley Periodicals LLC.)

Myocardial injury influenced by cisplatin (Cis) is a compelling reason to hunt out a treatment modality to overcome such a threat in cisplatin-treated patients. Breast Milk mesenchymal stem cells (Br-MSCs) are a non-invasive, highly reproducible source of stem cells. Herein, we investigate Br-MSCs' role in cardiotoxicity induced by cisplatin. Rats were divided into; control, Cis-treated (received 12 mg/kg single intraperitoneal injection), BrMSCs-treated (received single intraperitoneal injection of 0.5 ml sterilized phosphate-buffered saline containing 2 × 10 7 cells of Br-MSCs); metformin-treated (received 250 mg/kg/day orally) and BrMSCs + metformin + Cis treated groups. At the experiment end, serum creatine kinase (CK-MB) and cardiac troponin T (cTnT) activates were estimated, cardiac malondialdehyde (MDA), superoxide dismutase (SOD), interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α) levels were measured, cardiac expression of Bax and Bcl-2 and AMP-activated protein kinase (AMPK), as well as heart histopathology, were evaluated. Study results showed that Cis explored acute cardiotoxicity evidenced by deteriorated cardiac indices, induction of oxidative stress, and inflammation with myocardium histological alterations. Treatment with Br-MSCs restored heart function and structure deteriorated by Cis injection. The antioxidant/anti-inflammatory/anti-apoptotic results of Br-MSCs were supported by AMPK activation denoting their protective role against cisplatin-induced cardiac injury. These results were superior when metformin was added to the treatment protocol., (© 2022. The Author(s).)

Reprogramming of human dermal fibroblasts (HDFs) into induced cardiomyocyte-like cells (iCMs) represents a promising strategy for human cardiac regeneration. Different cocktails of cardiac transcription factors can convert HDFs into iCMs, although with low efficiency and immature phenotype. Here, GATA4, MEF2C, TBX5, MESP1, and MYOCD (GMTMeMy for short) were used to reprogram HDFs by retrovirus infection. We found that the exogenous expression stoichiometry of GATA4 (GATA4 stoichiometry) significantly affected reprogramming efficiency. When 1/8 dosage of GATA4 virus (GATA4 dosage) plus MTMeMy was used, the reprogramming efficiency was obviously improved compared with average pooled virus encoding each factor, which measured, by the expression level of cardiac genes, the percentage of cardiac troponin T and alpha-cardiac myosin heavy-chain immunopositive cells and the numbers of iCMs showing calcium oscillation or beating synchronously in co-culture with mouse CMs. In addition, we prepared conditioned maintenance medium (CMM) by CM differentiation of H9 human embryonic stem cell line. We found that compared with traditional maintenance medium (TMM), CMM made iCMs show well-organized sarcomere formation and characteristic calcium oscillation wave earlier. These findings demonstrated that appropriate GATA4 stoichiometry was essential for cardiac reprogramming and some components in CMM were important for maturation of iCMs.

Control on the moisture distribution, protein structure changes, and protein degradation of Antarctic krill meat during freeze-thaw (F-T) cycles by presoaking with antifreeze protein (AFP) was investigated. The results from the thawing loss rate and cooking loss rate indicated that 0.1% was the optimal AFP concentration. Magnetic resonance imaging and low-field nuclear magnetic resonance results showed that AFP inhibited the changes in moisture distribution and maintained the moisture in Antarctic krill meat. The contents of nonprotein nitrogen and trichloroacetic acid-soluble peptides indicated that AFP reduced protein degradation. Further, SDS-PAGE showed that AFP reduced the degradation of actin, troponin T, and myosin light chain. The results of fluorescence spectra, circular dichroism, and chemical bond contents indicated that AFP reduced the damage of the protein tertiary and secondary structures of Antarctic krill meat by holding it in a weak polar environment. This study supplied basic theory for the quality control of Antarctic krill meat. PRACTICAL APPLICATION: Protein degradation, moisture distribution, and protein structure changes occurred to Antarctic krill meat during freeze-thaw cycles due to ice crystal growth and recrystallization, which leads to the decrease in quality. Antifreeze protein has been proven to avoid ice crystals' growth and inhibit ice recrystallization. During freeze-thaw cycles, the moisture distribution of Antarctic krill meat treated with antifreeze protein was more uniform, the degree of protein degradation was lower, and the protein structure was protected. This study demonstrated the potential of antifreeze protein as a water and protein protectant of Antarctic krill meat during freeze-thaw cycles., (© 2022 Institute of Food Technologists®.)

The prevalence of cardiovascular complications in diabetes has become one of the major cause of diabetes related morbidity/mortality. The onset and progression of diabetic cardiomyopathy (DCM) has been majorly linked to lipid alterations, oxidative stress, inflammation and apoptosis. This present study investigated the cardioprotective role of Lycium chinense leaf extract (LCME) in fructose/streptozotocin induced diabetic rats. Diabetic animals were orally gavaged with LCME (100 and 400 mg/kg) for five weeks. The results indicated that diabetic rats showed increased blood glucose concentration, serum cardiac function markers (troponin T, creatine kinase-MB, aspartate aminotransferase and lactate dehydrogenase) and lipid profile (triglycerides and cholesterol). In addition, the cardiac tissues of diabetic rats showed increased levels of nuclear factor-κB (NF-κB), tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL 1β), interleukin 6 (IL-6), caspase-3 and malondialdehyde as well as significantly reduced activities of catalase, superoxide dismutase, reduced glutathione and glutathione peroxidase. LCME significantly ameliorated hyperglycemia and markedly decreased serum concentrations of troponin T, creatine kinase-MB, aspartate aminotransferase and lactate dehydrogenase, triglycerides and cholesterol. Furthermore, LCME notably suppressed cardiac oxido-inflammatory mediators and boosted cardiac antioxidant defense. Histopathologically, LCME restored cardiac structural alterations and also suppressed the immunohistochemical expression of collagen IV, smooth muscle alpha-actin (α-SMA) and p53, while Bcl2 expression was significantly increased. In conclusion, our result indicated that LCME protected against diabetic cardiomyopathy suppressing oxidative stress, inflammation, apoptosis and fibrosis., Competing Interests: Conflict of interest statement The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Aims: Genetic dilated cardiomyopathy (DCM) is a leading cause of heart failure. Despite significant progress in understanding the genetic aetiologies of DCM, the molecular mechanisms underlying the pathogenesis of familial DCM remain unknown, translating to a lack of disease-specific therapies. The discovery of novel targets for the treatment of DCM was sought using phenotypic sceening assays in induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) that recapitulate the disease phenotypes in vitro., Methods and Results: Using patient-specific iPSCs carrying a pathogenic TNNT2 gene mutation (p.R183W) and CRISPR-based genome editing, a faithful DCM model in vitro was developed. An unbiased phenotypic screening in TNNT2 mutant iPSC-derived cardiomyocytes (iPSC-CMs) with small molecule kinase inhibitors (SMKIs) was performed to identify novel therapeutic targets. Two SMKIs, Gö 6976 and SB 203580, were discovered whose combinatorial treatment rescued contractile dysfunction in DCM iPSC-CMs carrying gene mutations of various ontologies (TNNT2, TTN, LMNA, PLN, TPM1, LAMA2). The combinatorial SMKI treatment upregulated the expression of genes that encode serine, glycine, and one-carbon metabolism enzymes and significantly increased the intracellular levels of glucose-derived serine and glycine in DCM iPSC-CMs. Furthermore, the treatment rescued the mitochondrial respiration defects and increased the levels of the tricarboxylic acid cycle metabolites and ATP in DCM iPSC-CMs. Finally, the rescue of the DCM phenotypes was mediated by the activating transcription factor 4 (ATF4) and its downstream effector genes, phosphoglycerate dehydrogenase (PHGDH), which encodes a critical enzyme of the serine biosynthesis pathway, and Tribbles 3 (TRIB3), a pseudokinase with pleiotropic cellular functions., Conclusions: A phenotypic screening platform using DCM iPSC-CMs was established for therapeutic target discovery. A combination of SMKIs ameliorated contractile and metabolic dysfunction in DCM iPSC-CMs mediated via the ATF4-dependent serine biosynthesis pathway. Together, these findings suggest that modulation of serine biosynthesis signalling may represent a novel genotype-agnostic therapeutic strategy for genetic DCM., Competing Interests: Conflict of interest: M.M. is a shareholder and an advisory board member of Vala Sciences. R.S. is a consultant of Vala Sciences. The other authors declare no competing interests., (© The Author(s) 2022. Published by Oxford University Press on behalf of European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Cytokines such as tumor necrosis factor-α (TNFα) have been implicated in cardiac dysfunction and toxicity associated with doxorubicin (DOX). Although TNFα can elicit different cellular responses, including survival or death, the mechanisms underlying these divergent outcomes in the heart remain cryptic. The E3 ubiquitin ligase TRAF2 (TNF receptor associated factor 2) provides a critical signaling platform for K63-linked polyubiquitination of RIPK1 (receptor interacting protein 1), crucial for nuclear factor-κB (NF-κB) activation by TNFα and survival. Here, we investigate alterations in TNFα-TRAF2-NF-κB signaling in the pathogenesis of DOX cardiotoxicity., Methods: Using a combination of in vivo (4 weekly injections of DOX 5 mg·kg -1 ·wk -1 ) in C57/BL6J mice and in vitro approaches (rat, mouse, and human inducible pluripotent stem cell-derived cardiac myocytes), we monitored TNFα levels, lactate dehydrogenase, cardiac ultrastructure and function, mitochondrial bioenergetics, and cardiac cell viability., Results: In contrast to vehicle-treated mice, ultrastructural defects, including cytoplasmic swelling, mitochondrial perturbations, and elevated TNFα levels, were observed in the hearts of mice treated with DOX. While investigating the involvement of TNFα in DOX cardiotoxicity, we discovered that NF-κB was readily activated by TNFα. However, TNFα-mediated NF-κB activation was impaired in cardiac myocytes treated with DOX. This coincided with loss of K63- linked polyubiquitination of RIPK1 from the proteasomal degradation of TRAF2. Furthermore, TRAF2 protein abundance was markedly reduced in hearts of patients with cancer treated with DOX. We further established that the reciprocal actions of the ubiquitinating and deubiquitinating enzymes cellular inhibitors of apoptosis 1 and USP19 (ubiquitin-specific peptidase 19), respectively, regulated the proteasomal degradation of TRAF2 in DOX-treated cardiac myocytes. An E3-ligase mutant of cellular inhibitors of apoptosis 1 (H588A) or gain of function of USP19 prevented proteasomal degradation of TRAF2 and DOX-induced cell death. Furthermore, wild-type TRAF2, but not a RING finger mutant defective for K63-linked polyubiquitination of RIPK1, restored NF-κB signaling and suppressed DOX-induced cardiac cell death. Last, cardiomyocyte-restricted expression of TRAF2 (cardiac troponin T-adeno-associated virus 9-TRAF2) in vivo protected against mitochondrial defects and cardiac dysfunction induced by DOX., Conclusions: Our findings reveal a novel signaling axis that functionally connects the cardiotoxic effects of DOX to proteasomal degradation of TRAF2. Disruption of the critical TRAF2 survival pathway by DOX sensitizes cardiac myocytes to TNFα-mediated necrotic cell death and DOX cardiotoxicity.

Bergapten (BeG) is explored for its anti-inflammatory and antioxidant properties. Myocardial infarction (MI) is reported to be one of the leading cardiovascular diseases characterized by mitochondrial dysfunction and apoptosis. The main purpose of this study is to assess the cardiopreventive effects of BeG (50 mg/kg) in isoproterenol (ISO)-induced MI in Wistar rats. The increased infarct size after ISO induction was reduced simultaneously on treatment with BeG. Similarly, augmented levels of cardiac biomarkers, namely cardiac troponin T, creatine kinase (CK), cardiac troponin I, and CK-MB were also suppressed by BeG. The increased rate of lipid hydroperoxides and thiobarbituric acid reactive substances owing to the oxidative stress caused by free radical generation in ISO-induced rats were also inhibited by BeG. Antioxidants reduce oxidative stress by scavenging free radicals. ISO induction reduces these antioxidant enzymes glutathione peroxidase, catalase, superoxide dismutase, and glutathione, and levels causing oxidative cardiac damage to the heart tissue. BeG supplementation improved these enzymes synthesis preventing potential damage to the myocardium. Inflammation caused by ISO pretreatment increased the secretion of proinflammatory cytokines in ISO-induced rats. Pretreatment with BeG suppressed these inflammatory cytokines to a normal level in ISO + BeG-treated rats. The histopathological examination of the morphological characteristics showed that the intensity of cardiac damage caused by ISO induction was less in BeG pretreated rats with less inflammatory cells and no necrosis. BeG also showed promising results in the molecular alteration of AMP-activated protein kinase/endothelial nitric oxide synthase/protein kinase B signaling molecules. These observations emphasize the cardioprotective effects of BeG and its potential use as a drug in the near future., (© 2022 Wiley Periodicals LLC.)

Age-related muscle mass and strength decline (sarcopenia) impairs the performance of daily living activities and can lead to mobility disability/limitation in older adults. Biological pathways in muscle that lead to mobility problems have not been fully elucidated. Immunoglobulin G (IgG) infiltration in muscle is a known marker of increased fiber membrane permeability and damage vulnerability, but whether this translates to impaired function is unknown. Here, we report that IgG1 and IgG4 are abundantly present in the skeletal muscle (vastus lateralis) of ~ 50% (11 out of 23) of older adults (&gt; 65 years) examined. Skeletal muscle IgG1 was inversely correlated with physical performance (400 m walk time: r = 0.74, p = 0.005; SPPB score: r =  - 0.73, p = 0.006) and muscle strength (r =  - 0.6, p = 0.05). In a murine model, IgG was found to be higher in both muscle and blood of older, versus younger, C57BL/6 mice. Older mice with a higher level of muscle IgG had lower motor activity. IgG in mouse muscle co-localized with cardiac troponin T (cTnT) and markers of complement activation and apoptosis/necroptosis. Skeletal muscle-inducible cTnT knockin mice also showed elevated IgG in muscle and an accelerated muscle degeneration and motor activity decline with age. Most importantly, anti-cTnT autoantibodies were detected in the blood of cTnT knockin mice, old mice, and older humans. Our findings suggest a novel cTnT-mediated autoimmune response may be an indicator of sarcopenia., (© 2022. The Author(s), under exclusive licence to American Aging Association.)

Abstract: Hypoxia/reoxygenation (H/R) induces pyroptosis in the setting of acute myocardial infarction (AMI). Previous studies have shown that the expression of the miR-15 family is stimulated in myocardial ischemia-reperfusion injury or H/R-induced cardiomyocyte injury, and miR-15 is a promoter of cardiac ischemia-reperfusion or H/R injury. However, whether miR-15b-5p regulates H/R injury and cardiomyocyte pyroptosis and its mechanism still need to be further clarified. Bioinformatics analysis elicited that SIRT3 was the downstream regulatory target gene of miR-15b-5p. SIRT3 has been shown to participate in the regulation of pyroptosis by negatively regulating the NLRP3 inflammasome pathway. Therefore, we hypothesized that miR-15b-5p targets SIRT3 and activated the NLRP3 inflammasome pathway to promote H/R-induced cardiomyocyte pyroptosis. We first show that H/R increases miR-15b-5p in rat cardiomyocytes H9C2. Next, we tested the effects of inhibition of miR-15b-5p or overexpression of SIRT3. We found that miR-15b-5p downregulation or SIRT3 overexpression could reverse the H/R-induced pyroptosis. Furthermore, silencing SIRT3 antagonized the protective effect of miR-15b-5p downregulation on H9C2 cells. NLRP3 inhibitor MCC950 annulled the previously mentioned antagonistic effect of silencing SIRT3 on the protection of miR-15b-5p downregulation against pyroptosis. We then used a rat AMI model to analyze myocardial infarction area by triphenyl tetrazolium chloride staining and assess serum cardiac troponin T level by ELISA and found that miR-15b-5p silencing reduced AMI injury in rats. Collectively, these results suggest that miR-15b-5p increase H/R-induced pyroptosis in cardiomyocytes by targeting SIRT3 and activating the NLRP3 inflammasome., Competing Interests: The authors report no conflicts of interest., (Copyright © 2022 by the Shock Society.)

The structural analysis of large protein complexes has been greatly enhanced through the application of electron microscopy techniques. One such multiprotein complex, the cardiac thin filament (cTF), has cyclic interactions with thick filament proteins to drive contraction of the heart that has recently been the subject of such studies. As important as these studies are, they provide limited or no information on highly flexible regions that in isolation would be characterized as inherently disordered. One such region is the extended cardiac troponin T (cTnT) linker between the regions of cTnT which have been labeled TNT1 and TNT2. It comprises a hinge region (residues 158-166) and a highly flexible region (residues 167-203). Critically, this region modulates the troponin/tropomyosin complex's position across the actin filament. Thus, the cTnT linker structure and dynamics are central to the regulation of the function of cardiac muscles, but up to now, it was ill-understood. To establish the cTnT linker structure, we coupled an atomistic computational cTF model with time-resolved fluorescence resonance energy transfer measurements in both ±Ca 2+ conditions utilizing fully reconstituted cTFs. We mapped the cTnT linker's positioning across the actin filament, and by coupling the experimental results to computation, we found mean structures and ranges of motion of this part of the complex. With this new insight, we can now address cTnT linker structural dynamics in both myofilament activation and disease.

Background: Stem cell therapy has emerged as a novel treatment for heart failure after myocardial infarction (Ml). Bone marrow-derived mesenchymal stem cells (BM-MSCs) are commonly considered because of their accessibility and usability. However, their therapeutic potential remains controversial. In our previous in vitro study, chorion-derived mesenchymal stem cells (C-MSCs) and umbilical cord-derived mesenchymal stem cells (UC-MSCs) demonstrated an ability to differentiate into cardiomyocytes and neural cells, respectively. Thus, we examined whether C-MSCs had a better differentiation potential in an MI animal model., Methods: MI was induced by ligation of the left anterior descending artery, and DiI-labeled MSCs were injected into the border of the infarcted myocardium. The left ventricular ejection fraction (LVEF) and fractional shortening (FS) were measured using echocardiograms. Masson's Trichrome staining was performed to evaluate the viable myocardium. Alpha-sarcomeric actin (α-SA), cardiac troponin-T (cTnT), and isolectin were immunolabeled to evaluate differentiation and capillary formation., Results: After 8 weeks, the LVEF and FS significantly increased to a greater extent in the C-MSC-injected group with maintenance of viable myocardium, as compared to in the control, UC-MSC-, and BM-MSC-injected groups (p < 0.05). Compared to UC-MSCs and BM-MSCs, C-MSCs significantly increased the capillary density (p < 0.05) and demonstrated higher expressions of cTnT and α-SA., Conclusions: In conclusion, compared to UC-MSCs and BM-MSCs, C-MSCs showed a better therapeutic efficacy in a rat MI model., Competing Interests: Conflict of interest None., (Copyright © 2022 Hellenic Society of Cardiology. Published by Elsevier B.V. All rights reserved.)

Background: Cardiac troponin (cTn) T and cTnI are considered cardiac specific and equivalent in the diagnosis of acute myocardial infarction. Previous studies suggested rare skeletal myopathies as a noncardiac source of cTnT. We aimed to confirm the reliability/cardiac specificity of cTnT in patients with various skeletal muscle disorders (SMDs)., Methods: We prospectively enrolled patients presenting with muscular complaints (≥2 weeks) for elective evaluation in 4 hospitals in 2 countries. After a cardiac workup, patients were adjudicated into 3 predefined cardiac disease categories. Concentrations of cTnT/I and resulting cTnT/I mismatches were assessed with high-sensitivity (hs-) cTnT (hs-cTnT-Elecsys) and 3 hs-cTnI assays (hs-cTnI-Architect, hs-cTnI-Access, hs-cTnI-Vista) and compared with those of control subjects without SMD presenting with adjudicated noncardiac chest pain to the emergency department (n=3508; mean age, 55 years; 37% female). In patients with available skeletal muscle biopsies, TNNT/I1-3 mRNA differential gene expression was compared with biopsies obtained in control subjects without SMD., Results: Among 211 patients (mean age, 57 years; 42% female), 108 (51%) were adjudicated to having no cardiac disease, 44 (21%) to having mild disease, and 59 (28%) to having severe cardiac disease. hs-cTnT/I concentrations significantly increased from patients with no to those with mild and severe cardiac disease for all assays (all P <0.001). hs-cTnT-Elecsys concentrations were significantly higher in patients with SMD versus control subjects (median, 16 ng/L [interquartile range (IQR), 7-32.5 ng/L] versus 5 ng/L [IQR, 3-9 ng/L]; P <0.001), whereas hs-cTnI concentrations were mostly similar (hs-cTnI-Architect, 2.5 ng/L [IQR, 1.2-6.2 ng/L] versus 2.9 ng/L [IQR, 1.8-5.0 ng/L]; hs-cTnI-Access, 3.3 ng/L [IQR, 2.4-6.1 ng/L] versus 2.7 ng/L [IQR, 1.6-5.0 ng/L]; and hs-cTnI-Vista, 7.4 ng/L [IQR, 5.2-13.4 ng/L] versus 7.5 ng/L [IQR, 6-10 ng/L]). hs-cTnT-Elecsys concentrations were above the upper limit of normal in 55% of patients with SMD versus 13% of control subjects ( P <0.01). mRNA analyses in skeletal muscle biopsies (n=33), mostly (n=24) from individuals with noninflammatory myopathy and myositis, showed 8-fold upregulation of TNNT2 , encoding cTnT (but none for TNNI3 , encoding cTnI) versus control subjects (n=16, P Wald <0.001); the expression correlated with pathological disease activity ( R =0.59, P t-statistic <0.001) and circulating hs-cTnT concentrations ( R =0.26, P t-statistic =0.031)., Conclusions: In patients with active chronic SMD, elevations in cTnT concentrations are common and not attributable to cardiac disease in the majority. This was not observed for cTnI and may be explained in part by re-expression of cTnT in skeletal muscle., Registration: URL: https://www., Clinicaltrials: gov; Unique identifier: NCT03660969.

Purpose: Both immune checkpoint inhibitors (ICIs) and vascular endothelial growth factor receptor (VEGFR) inhibitors are approved for advanced renal cell carcinoma treatment and can cause cardiovascular events (CVs); thus, combination therapy could lead to major adverse CV events (MACE). Cardiac serum biomarker assessment and imaging, including left ventricular ejection fraction (LVEF) monitoring, can be used to evaluate MACE., Methods: To our knowledge, the JAVELIN Renal 101 trial, assessing avelumab plus axitinib versus sunitinib in patients with advanced renal cell carcinoma, is the first randomized study of ICI plus VEGFR inhibitor treatment to include prospective serial cardiac monitoring of LVEF and serum cardiac biomarkers., Results: MACE (defined as grade ≥ 3 CV AEs) occurred in 31 patients (7.1%) in the combination arm and 17 patients (3.9%) in the sunitinib arm. Patients in the combination arm who had high baseline troponin T values were at higher risk of MACE versus patients with low values (MACE in 6/35 v 7/135, respectively; relative risk, 3.31; 95% CI, 1.19 to 9.22). This association was not observed in patients treated with sunitinib. Other CV baseline risk factors and serum cardiac biomarkers were not significantly predictive for MACE, although a trend toward an association with dyslipidemia was seen in the combination arm. No clinical value of on-treatment routine monitoring of LVEF in relation to MACE was observed. Although LVEF decline was significantly more frequent in the combination arm, most patients recovered, and decline was not associated with other significant cardiac events or symptoms., Conclusion: Patients with high baseline troponin T levels receiving ICI and VEGFR combinations may need to be monitored more closely for MACE. Routine monitoring of LVEF in asymptomatic patients is not recommended., Competing Interests: Brian I. RiniLeadership: MJH Life SciencesStock and Other Ownership Interests: PTC TherapeuticsConsulting or Advisory Role: Pfizer, Merck & Co (Kenilworth, NJ), Synthorx, Bristol Myers Squibb, AVEO, Surface Oncology, 3D Medicines, Corvus Pharmaceuticals, Aravive, Arrowhead Pharmaceuticals, Shionogi, Eisai, GlaxoSmithKlineResearch Funding: Pfizer (Inst), Roche/Genentech (Inst), Bristol Myers Squibb (Inst), Merck & Co (Kenilworth, NJ) (Inst), AstraZeneca/MedImmune (Inst), Incyte (Inst), Arrowhead Pharmaceuticals (Inst), Taris (Inst), Seattle Genetics (Inst), Immunomedics (Inst), Surface Oncology (Inst), Dragonfly Therapeutics (Inst), Aravive (Inst), Exelixis (Inst)Travel, Accommodations, Expenses: Pfizer, Bristol Myers Squibb, Merck & Co (Kenilworth, NJ) Javid J. MoslehiConsulting or Advisory Role: Aerovate Therapeutics, AstraZeneca, Audentes Pharmaceuticals, Boehringer, Bristol Myers Squibb, Cytokinetics, Deciphera, GlaxoSmithKline, Kurome Therapeutics, Mallinckrodt Pharmaceuticals, Myokardia, Myovant, Novartis, Pfizer, Pharmacyclics, ProteinCure, Takeda, Silverback Therapeutics, Star Therapeutics Marc BonacaResearch Funding: ARCA BioPahrma (Inst), AstraZeneca/MedImmune (Inst), Bayer (Inst), Better Therapeutics (Inst), CellResearch (Inst), EverlyWell (Inst), Janssen (Inst), Novo Nordisk (Inst), Osiris Medical (Inst), Amgen (Inst), HDL Therapeutics (Inst) Manuela SchmidingerHonoraria: Pfizer, Bristol Myers Squibb, Ipsen, Roche/Genentech, Merck & Co (Kenilworth, NJ), EISAI, EUSA Pharma, ALKERMESConsulting or Advisory Role: Pfizer, Bristol Myers Squibb, Roche, Ipsen, Merck & Co (Kenilworth, NJ), EUSA Pharma, EisaiTravel, Accommodations, Expenses: Bristol Myers Squibb, Roche, Ipsen Laurence AlbigesConsulting or Advisory Role: Bristol Myers Squibb (Inst), Ipsen (Inst), Roche (Inst), Novartis (Inst), Pfizer (Inst), Astellas Pharma (Inst), Merck & Co (Kenilworth, NJ) (Inst), AstraZeneca (Inst), Janssen (Inst), Eisai (Inst), Corvus Pharmaceuticals (Inst), Bellerophon Theraeutics (Inst)Research Funding: Bristol Myers Squibb (Inst)Travel, Accommodations, Expenses: Bristol Myers Squibb, Merck & Co (Kenilworth, NJ) Toni K. ChoueiriEmployment: Dana Farber Cancer HospitalLeadership: Dana Farber Cancer Hospital, NCCN, KidneyCan, ASCOStock and Other Ownership Interests: Pionyr, Tempest TherapeuticsHonoraria: NCCN, UpToDate, Michael J. Hennessy Associates, ASCO, Harborside Press, Analysis Group, AstraZeneca, Alexion Pharmaceuticals, Sanofi/Aventis, Bayer, Bristol Myers Squibb, Genentech/Roche, GlaxoSmithKline, Merck & Co (Kenilworth, NJ), Novartis, Peloton Therapeutics, Pfizer, Corvus Pharmaceuticals, Ipsen, Foundation Medicine, Eisai, PlatformQ Health, Clinical Care Options, Navinata Health, Kidney Cancer Association, Exelixis, Prometheus, Lpath, The New England Journal of Medicine, Lancet Oncology, Cerulean Pharma, alligent, the healthcare business of Merck KGaA (Darmstadt, Germany), HERON, Lilly, Janssen Oncology, IQvia, Aveo, NCI Genitourinary Cancers Steering CommitteeConsulting or Advisory Role: Pfizer, Bayer, Novartis, GlaxoSmithKline, Merck & Co (Kenilworth, NJ), Bristol Myers Squibb, Roche/Genentech, Eisai, Foundation Medicine, Cerulean Pharma, AstraZeneca, Exelixis, Prometheus, alligent, Ipsen, Corvus Pharmaceuticals, Lpath, Alexion Pharmaceuticals, Sanofi/Aventis, Peloton Therapeutics, UpToDate, NCCN, Michael J. Hennessy Associates, Analysis Group, Kidney Cancer Association, Clinical Care Options, PlatformQ Health, Navinata Health, Harborside Press, ASCO, The New England Journal of Medicine, Lancet Oncology, the healthcare business of Merck KGaA (Darmstadt, Germany), HERON, Lilly, ESMO, NiKang Therapeutics, Kanaph Therapeutics, Infinity Pharmaceuticals, AraviveResearch Funding: Pfizer (Inst), Novartis (Inst), Merck & Co (Kenilworth, NJ) (Inst), Exelixis (Inst), TRACON Pharma (Inst), GlaxoSmithKline (Inst), Bristol Myers Squibb (Inst), AstraZeneca (Inst), Peloton Therapeutics (Inst), Roche/Genentech (Inst), Celldex (Inst), Agensys (Inst), Eisai (Inst), Takeda (Inst), Prometheus (Inst), Ipsen (Inst), Corvus Pharmaceuticals (Inst), Cerulean Pharma (Inst), Seattle Genetics/Astellas (Inst), Bayer (Inst), Foundation Medicine (Inst), Roche (Inst), Calithera Biosciences (Inst), Analysis Group (Inst), NCI (Inst), Gateway for Cancer Research (Inst), Congressionally Directed Medical Research Programs (DOD) (Inst)Patents, Royalties, Other Intellectual Property: International Patent Application No. PCT/US2018/058430, titled Biomarkers of Clinical Response and Benefit to Immune Checkpoint Inhibitor Therapy (Inst), International Patent Application No. PCT/US2018/12209, titled PBRM1 Biomarkers Predictive of Anti-Immune Checkpoint Response (Inst)Travel, Accommodations, Expenses: Pfizer, Bayer, Novartis, GlaxoSmithKline, Merck & Co (Kenilworth, NJ), Bristol Myers Squibb, Roche/Genentech, Eisai, Foundation Medicine, Cerulean Pharma, AstraZeneca, Exelixis, Prometheus, alligent, Ipsen, Corvus Pharmaceuticals, Lpath, Alexion Pharmaceuticals, Sanofi/Aventis, UpToDate, Peloton Therapeutics, NCCN, Michael J. Hennessy Associates, Analysis Group, Kidney Cancer Association, Clinical Care Options, PlatformQ Health, Harborside Press, Navinata Health, The New England Journal of Medicine, Lancet Oncology, the healthcare business of Merck KGaA (Darmstadt, Germany), HERON, Lilly, ESMOOther Relationship: Medical writing and editorial assistance support may have been funded by communications companies funded by pharmaceutical companies such as ClinicalThinking, Health Interactions, Envision Pharma Group, Fishawack Group of Companies, Parexel Robert J. MotzerConsulting or Advisory Role: Novartis, Eisai, Exelixis, Merck & Co (Kenilworth, NJ), Genentech/Roche, Incyte, Lilly, Pfizer, AstraZeneca, the healthcare business of Merck KGaA (Darmstadt, Germany), Calithera Biosciences, AveoResearch Funding: Pfizer (Inst), Bristol Myers Squibb (Inst), Eisai (Inst), Novartis (Inst), Genentech/Roche (Inst), Exelixis (Inst), Merck & Co (Kenilworth, NJ) (Inst), Aveo (Inst)Travel, Accommodations, Expenses: Bristol Myers Squibb Michael B. AtkinsStock and Other Ownership Interests: Werewolf Pharma, PyxisConsulting or Advisory Role: Genentech, Novartis, Bristol Myers Squibb, Merck & Co (Kenilworth, NJ), Exelixis, Eisai, Agenus, Arrowhead Pharmaceuticals, Werewolf Pharma, Surface Oncology, Iovance Biotherapeutics, Pyxis, Pneuma Respiratory, Leads Biolabs, Fathom Biotechnology, Aveo, Cota Healthcare, Neoleukin Therapeutics, Adagene, Idera, Ellipses Pharma, AstraZeneca, PACT Pharma, Seattle Genetics, Pfizer, Scholar Rock, Asher Biotherapeutics, Calithera Biosciences, Takeda, Sanofi, Simcha TherapeuticsResearch Funding: Bristol Myers Squibb (Inst) John HaanenStock and Other Ownership Interests: Neogene TherapeuticsConsulting or Advisory Role: Merck & Co (Kenilworth, NJ) (Inst), Pfizer (Inst), Bristol Myers Squibb (Inst), Novartis (Inst), Roche/Genentech (Inst), Ipsen (Inst), Achilles Therapeutics (Inst), Immunocore (Inst), Sanofi (Inst), Third Rock Ventures (Inst), Neogene Therapeutics, Molecular Partners (Inst), bioNTech (Inst), T-Knife (Inst), PokeAcel (Inst), Instil Bio (Inst), Iovance Biotherapeutics (Inst)Research Funding: Merck & Co (Kenilworth, NJ) (Inst), Bristol Myers Squibb (Inst), Novartis (Inst), Neon Therapeutics (Inst), Amgen (Inst), BioNTech (Inst), Asher Biotherapeutics (Inst) Mariangela MarianiStock and Other Ownership Interests: Pfizer Jing WangEmployment: PfizerStock and Other Ownership Interests: Pfizer Subramanian HariharanEmployment: PfizerStock and Other Ownership Interests: Pfizer James LarkinHonoraria: Bristol Myers Squibb, GlaxoSmithKline, Pfizer, Novartis, Roche/Genentech, Incyte, iOnctura, the healthcare business of Merck KGaA (Darmstadt, Germany), Eisai, Dynavax Technologies, Cancer Research UK, touchIME, touchEXPERTSConsulting or Advisory Role: Bristol Myers Squibb, GlaxoSmithKline, Pfizer, Novartis, Boston Biomedical, Incyte, iOnctura, Iovance Biotherapeutics, Immunocore, YKT Corporation, Apple Tree PartnersResearch Funding: Pfizer (Inst), Novartis (Inst), Merck & Co (Kenilworth, NJ) (Inst), Bristol Myers Squibb (Inst), Achilles Therapeutics (Inst), Roche (Inst), Nektar (Inst), Covance (Inst), Immunocore (Inst), AVEO (Inst), Pharmacyclics (Inst)Travel, Accommodations, Expenses: Bristol Myers Squibb, Pfizer, Novartis, Roche/Genentech, AstraZeneca, Incyte, GlaxoSmithKline, Pierre Fabre, the healthcare business of Merck KGaA (Darmstadt, Germany), iOnctura, British Uro-Oncology Group (BUG), ESMO, National Cancer Research Institute (NCRI), EUSA Pharma, Syneos Health, Kidney Cancer Association, Bioevents, MedConcept, RV MaisNo other potential conflicts of interest were reported.

Background Contemporary guidelines recommend using atherosclerotic cardiovascular disease screening tools to guide primary prevention. The performance of these scores is not well known in patients with moderate to advanced chronic kidney disease, particularly in combination with clinically available cardiac biomarkers including N-terminal pro-brain-type natriuretic peptide and high-sensitivity troponin T (hsTnT). Methods and Results We studied 1027 participants from the Chronic Renal Insufficiency Cohort without self-reported atherosclerotic cardiovascular disease who were not taking aspirin or statins at enrollment. Framingham Risk Score, Pooled Cohort Equation, N-terminal pro-brain-type natriuretic peptide, and hsTnT were measured at baseline. Outcomes included fatal and nonfatal myocardial infarction, stroke, and cardiac death. We calculated 10-fold cross-validated Harrell's C-indices for each risk score and cardiac biomarker alone and in combination. The C-index (95% CI) for discrimination of atherosclerotic cardiovascular disease was 0.72 (0.67, 0.77) for the Framingham Risk Score, and 0.72 (0.67, 0.76) for the Pooled Cohort Equation. HsTnT had comparable discrimination to each risk score, and improved the discrimination of each (change in Framingham 0.029, 95% CI 0.003, 0.055; change in Pooled Cohort Equation 0.027, 95% CI 0.002, 0.052). N-terminal pro-brain-type natriuretic peptide had poorer discrimination than the risk scores and did not significantly improve their discrimination (change in Framingham 0.009, 95% CI -0.001, 0.018; change in Pooled Cohort Equation 0.011, 95% CI -0.001, 0.024). Conclusions The Framingham Risk Score and Pooled Cohort Equation demonstrated moderate discrimination for atherosclerotic cardiovascular disease in patients with chronic kidney disease. HsTnT, but not N-terminal pro-brain-type natriuretic peptide, improved their discrimination overall. Until chronic kidney disease-specific atherosclerotic cardiovascular disease risk scores can be developed, it may be worth considering how to incorporate hsTnT into existing clinical risk scores.

Autosomal dominant mutations in sarcomere proteins such as the cardiac troponin T ( TNNT2 ) are the main genetic causes of human hypertrophic cardiomyopathy and dilated cardiomyopathy. Allele-specific silencing by RNA interference (ASP-RNAi) holds promise as a therapeutic strategy for downregulating a single mutant allele with minimal suppression of the corresponding wild-type allele. Here, we propose ASP-RNAi as a possible strategy to specifically knockdown mutant alleles coding for R92Q and R173W mutant TNNT2 proteins, identified in hypertrophic and dilated cardiomyopathy, respectively. Different siRNAs were designed and validated by luciferase reporter assay and following analysis in HEK293T cells expressing either the wild-type or mutant TNNT2 alleles. This study is the first exploration of ASP-RNAi on TNNT2 -R173W and TNNT2 -R92Q mutations in vitro and gives a base for further application of allele silencing as a therapeutic treatment for TNNT2 -mutation-associated cardiomyopathies.

Biomarkers of mineral bone disorders (MBD) including phosphorus, fibroblast growth factor (FGF)-23 and Klotho are strongly altered in patients with acute kidney injury (AKI) who have high cardiac outcomes and mortality rates. However, the crosslink between MBD and cardiac damage after an AKI episode still remains unclear. We tested MBD and cardiac biomarkers in an experimental AKI model after 24 or 72 hours of folic acid injection and we analyzed structural cardiac remodeling, intracellular calcium (Ca 2+ ) dynamics in cardiomyocytes and cardiac rhythm. AKI mice presented high levels of FGF-23, phosphorus and cardiac troponin T and exhibited a cardiac hypertrophy phenotype accompanied by an increase in systolic Ca 2+ release 24 hours after AKI. Ca 2+ transients and contractile dysfunction were reduced 72 hours after AKI while diastolic sarcoplasmic reticulum Ca 2+ leak, pro-arrhythmogenic Ca 2+ events and ventricular arrhythmias were increased. These cardiac events were linked to the activation of the calcium/calmodulin-dependent kinase II pathway through the increased phosphorylation of ryanodine receptors and phospholamban specific sites after AKI. Cardiac hypertrophy and the altered intracellular Ca 2+ dynamics were prevented in transgenic mice overexpressing Klotho after AKI induction. In a translational retrospective longitudinal clinical study, we determined that combining FGF-23 and phosphorus with cardiac troponin T levels achieved a better prediction of mortality in AKI patients at hospital admission. Thus, monitoring MBD and cardiac damage biomarkers could be crucial to prevent mortality in AKI patients. In this setting, Klotho might be considered as a new cardioprotective therapeutic tool to prevent deleterious cardiac events in AKI conditions., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Several studies have demonstrated the usefulness of cardiac troponin I (cTn) levels in predicting adverse clinical outcomes of patients with anerusmal subarachnoid hemorrhage (aSAH). However, it remains unclear whether cTn levels can be a useful factor in predicting adverse neurologic and cardiovascular outcomes regarding follow-up duration. The study aimed to evaluate the clinical value of cTn elevation among patients with aSAH. A systematic literature search was performed in PubMed and Cochrane to collect original studies that compared the adverse outcomes in patients with aSAH who had elevated cTn levels and those who did not have elevated cTn levels. Data on patient demographics and outcome measurements (mortality, major disability, delayed cerebral ischemia, cardiac dysfunction, and pulmonary edema) were extracted. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were computed by fitting a random effects model. A total of 4,117 patients with aSAH were included in the meta-analysis. Elevated cTn levels was associated with a higher all-cause mortality (OR 3.64; 95% CI 2.68-4.94; I 2  = 22.05%), poor major disability (OR 2.27; 95% CI 1.5-3.37; I 2  = 52.07%), delayed cerebral ischemia (OR 2.10; 95% CI 1.46-3.03; I 2  = 13.80%), cardiac dysfunction (OR 9.20; 95% CI 4.31-19.60; I 2  = 39.89), and pulmonary edema (OR 10.32; 95% CI 5.64-18.90; I 2  = 0.00%). Additionally, elevated cTn levels was associated with higher mortality in prospective studies (OR 3.66; 95% CI 2.61-5.14) as well as when compared with studies with short-term and long-term follow-up periods. Patients with aSAH who had elevated cTn levels also tended to experience poor short-term major disability (OR 2.36; 95% CI 1.48-3.76). Among patients with aSAH, elevated cTn levels was associated with higher mortality and adverse neurologic and cardiovascular outcomes. Given its clinical value, cardiac troponin levels may be included in the assessment of patients withs aSAH., (© 2021. Springer Science+Business Media, LLC, part of Springer Nature and Neurocritical Care Society.)