Single-Cell RNA Sequencing Reveals Cardiac Fibroblast-Specific Transcriptomic Changes in Dilated Cardiomyopathy.

Dilated cardiomyopathy (DCM) is the most common cause of heart failure, with a complex aetiology involving multiple cell types. We aimed to detect cell-specific transcriptomic alterations in DCM through analysis that leveraged recent advancements in single-cell analytical tools. Single-cell RNA sequencing (scRNA-seq) data from human DCM cardiac tissue were subjected to an updated bioinformatic workflow in which unsupervised clustering was paired with reference label transfer to more comprehensively annotate the dataset. Differential gene expression was detected primarily in the cardiac fibroblast population. Bulk RNA sequencing was performed on an independent cohort of human cardiac tissue and compared with scRNA-seq gene alterations to generate a stratified list of higher-confidence, fibroblast-specific expression candidates for further validation. Concordant gene dysregulation was confirmed in TGFß-induced fibroblasts. Functional assessment of gene candidates showed that AEBP1 may play a significant role in fibroblast activation. This unbiased approach enabled improved resolution of cardiac cell-type-specific transcriptomic alterations in DCM.

Dose-dependent changes in cardiac function, strain and remodelling in a preclinical model of heart base irradiation.

BACKGROUND AND PURPOSE: Radiation induced cardiotoxicity (RICT) is as an important sequela of radiotherapy to the thorax for patients. In this study, we aim to investigate the dose and fractionation response of RICT. We propose global longitudinal strain (GLS) as an early indicator of RICT and investigate myocardial deformation following irradiation. METHODS: RICT was investigated in female C57BL/6J mice in which the base of the heart was irradiated under image-guidance using a small animal radiation research platform (SARRP). Mice were randomly assigned to a treatment group: single-fraction dose of 16 Gy or 20 Gy, 3 consecutive fractions of 8.66 Gy, or sham irradiation; biological effective doses (BED) used were 101.3 Gy, 153.3 Gy and 101.3 Gy respectively. Longitudinal transthoracic echocardiography (TTE) was performed from baseline up to 50 weeks post-irradiation to detect structural and functional effects. RESULTS: Irradiation of the heart base leads to BED-dependent changes in systolic and diastolic function 50 weeks post-irradiation. GLS showed significant decreases in a BED-dependent manner for all irradiated animals, as early as 10 weeks after irradiation. Early changes in GLS indicate late changes in cardiac function. BED-independent increases were observed in the left ventricle (LV) mass and volume and myocardial fibrosis. CONCLUSIONS: Functional features of RICT displayed a BED dependence in this study. GLS showed an early change at 10 weeks post-irradiation. Cardiac remodelling was observed as increases in mass and volume of the LV, further supporting our hypothesis that dose to the base of the heart drives the global heart toxicity.

A multimodality assessment of the protective capacity of statin therapy in a mouse model of radiation cardiotoxicity.

PURPOSE: Despite technological advances in radiotherapy (RT), cardiotoxicity remains a common complication in patients with lung, oesophageal and breast cancers. Statin therapy has been shown to have pleiotropic properties beyond its lipid-lowering effects. Previous murine models have shown statin therapy can reduce short-term functional effects of whole-heart irradiation. In this study, we assessed the efficacy of atorvastatin in protecting against the late effects of radiation exposure on systolic function, cardiac conduction, and atrial natriuretic peptide (ANP) following a clinically relevant partial-heart radiation exposure. MATERIALS AND METHODS: Female, 12-week old, C57BL/6j mice received an image-guided 16 Gy X-ray field to the base of the heart using a small animal radiotherapy research platform (SARRP), with or without atorvastatin from 1 week prior to irradiation until the end of the experiment. The animals were followed for 50 weeks with longitudinal transthoracic echocardiography (TTE) and electrocardiography (ECG) every 10 weeks, and plasma ANP every 20 weeks. RESULTS: At 30-50 weeks, mild left ventricular systolic function impairment observed in the RT control group was less apparent in animals receiving atorvastatin. ECG analysis demonstrated prolongation of components of cardiac conduction related to the heart base at 10 and 30 weeks in the RT control group but not in animals treated with atorvastatin. In contrast to systolic function, conduction disturbances resolved at later time-points with radiation alone. ANP reductions were lower in irradiated animals receiving atorvastatin at 30 and 50 weeks. CONCLUSIONS: Atorvastatin prevents left ventricular systolic dysfunction, and the perturbation of cardiac conduction following partial heart irradiation. If confirmed in clinical studies, these data would support the use of statin therapy for cardioprotection during thoracic radiotherapy.

PLAC8-Mediated Activation of NOX4 Signalling Restores Angiogenic Function of Endothelial Colony-Forming Cells in Experimental Hypoxia.

Ischaemic cardiovascular disease is associated with tissue hypoxia as a significant determinant of angiogenic dysfunction and adverse remodelling. While cord blood-derived endothelial colony-forming cells (CB-ECFCs) hold clear therapeutic potential due to their enhanced angiogenic and proliferative capacity, their impaired functionality within the disease microenvironment represents a major barrier to clinical translation. The aim of this study was to define the specific contribution of NOX4 NADPH oxidase, which we previously reported as a key CB-ECFC regulator, to hypoxia-induced dysfunction and its potential as a therapeutic target. CB-ECFCs exposed to experimental hypoxia demonstrated downregulation of NOX4-mediated reactive oxygen species (ROS) signalling linked with a reduced tube formation, which was partially restored by NOX4 plasmid overexpression. siRNA knockdown of placenta-specific 8 (PLAC8), identified by microarray analysis as an upstream regulator of NOX4 in hypoxic versus normoxic CB-ECFCs, enhanced tube formation, NOX4 expression and hydrogen peroxide generation, and induced several key transcription factors associated with downstream Nrf2 signalling. Taken together, these findings indicated that activation of the PLAC8-NOX4 signalling axis improved CB-ECFC angiogenic functions in experimental hypoxia, highlighting this pathway as a potential target for protecting therapeutic cells against the ischaemic cardiovascular disease microenvironment.

The Potential Role of Salivary NT-proBNP in Heart Failure.

BACKGROUND: Serum natriuretic peptides (NPs) have an established role in heart failure (HF) diagnosis. Saliva NT-proBNP that may be easily acquired has been studied little. METHODS: Ninety-nine subjects were enrolled; thirty-six obese or hypertensive with dyspnoea but no echocardiographic HF findings or raised NPs served as controls, thirteen chronic HF (CHF) patients and fifty patients with acute decompensated HF (ADHF) requiring hospital admission. Electrocardiogram, echocardiogram, 6 min walking distance (6MWD), blood and saliva samples, were acquired in all participants. RESULTS: Serum NT-proBNP ranged from 60-9000 pg/mL and saliva NT-proBNP from 0.64-93.32 pg/mL. Serum NT-proBNP was significantly higher in ADHF compared to CHF (p = 0.007) and in CHF compared to controls (p < 0.05). There was no significant difference in saliva values between ADHF and CHF, or between CHF and controls. Saliva and serum levels were positively associated only in ADHF patients (R = 0.352, p = 0.012). Serum NT-proBNP was positively associated with NYHA class (R = 0.506, p < 0.001) and inversely with 6MWD (R = -0.401, p = 0.004) in ADHF. Saliva NT-proBNP only correlated with age in ADHF patients. CONCLUSIONS: In the current study, saliva NT-proBNP correlated with serum values in ADHF patients, but could not discriminate between HF and other causes of dyspnoea. Further research is needed to explore the value of saliva NT-proBNP.

FK506-Binding Protein like (FKBPL) Has an Important Role in Heart Failure with Preserved Ejection Fraction Pathogenesis with Potential Diagnostic Utility.

Heart failure (HF) is the leading cause of hospitalisations worldwide, with only 35% of patients surviving the first 5 years after diagnosis. The pathogenesis of HF with preserved ejection fraction (HFpEF) is still unclear, impeding the implementation of effective treatments. FK506-binding protein like (FKBPL) and its therapeutic peptide mimetic, AD-01, are critical mediators of angiogenesis and inflammation. Thus, in this study, we investigated-for the first time-FKBPL's role in the pathogenesis and as a biomarker of HFpEF. In vitro models of cardiac hypertrophy following exposure to a hypertensive stimulus, angiotensin-II (Ang-II, 100 nM), and/or AD-01 (100 nM), for 24 and 48 h were employed as well as human plasma samples from people with different forms of HFpEF and controls. Whilst the FKBPL peptide mimetic, AD-01, induced cardiomyocyte hypertrophy in a similar manner to Ang-II (p < 0.0001), when AD-01 and Ang-II were combined together, this process was abrogated (p < 0.01-0.0001). This mechanism appears to involve a negative feedback loop related to FKBPL (p < 0.05). In human plasma samples, FKBPL concentration was increased in HFpEF compared to controls (p < 0.01); however, similar to NT-proBNP and Gal-3, it was unable to stratify between different forms of HFpEF: acute HFpEF, chronic HFpEF and hypertrophic cardiomyopathy (HCM). FKBPL may be explored for its biomarker and therapeutic target potential in HFpEF.

Circulating biomarkers for management of cancer therapeutics-related cardiac dysfunction.

Cancer therapeutics-related cardiac dysfunction (CTRCD) has emerged as a major cause of morbidity and mortality in cancer survivors. Effective clinical management of CTRCD is impeded by a lack of sensitive diagnostic and prognostic strategies. Circulating molecular markers could potentially address this need as they are often indicative of cardiac stress before cardiac damage can be detected clinically. A growing understanding of the underlying physiological mechanisms for CTRCD has inspired research efforts to identify novel pathophysiologically relevant biomarkers that may also guide development of cardio-protective therapeutic approaches. The purpose of this review is to evaluate current circulating biomarkers of cardiac stress and their potential role in diagnosis and management of CTRCD. We also discuss some emerging avenues for CTRCD-focused biomarker investigations.

Families' expectations of an eHealth family-based cardiovascular disease-risk reduction programme.

AIM: Research has shown that families' participation in a cardiovascular disease (CVD) prevention programme could boost early adoption of healthy lifestyle behaviours in families. Behaviour-based, eHealth interventions are a potential means of achieving this. This study aimed to explore expectations of families-parents and children-at risk of CVD towards the design and functionality of an eHealth family-based CVD-risk reduction programme 'Health-e-Hearts'. METHODS AND RESULTS: Three online focus groups were conducted with six families comprising at least one parent at risk of CVD and at least one child aged 5-17 years. The focus groups were video and audio recorded and transcribed. Content analysis was used to synthesize and identify key categories and subcategories regarding development of and engagement with an eHealth programme. Three categories emerged: experiences of health apps and devices; eHealth application needs of family members; and motivators for using an eHealth programme. Experiences included using health apps individually and inconsistently. Needs included personalization, free and easy-to-use, time efficient, and multiple content formats. Motivators for engaging with the programme included goal setting, rewards, and competition. CONCLUSION: Families' expectations of an eHealth family-based CVD-risk reduction programme include the incorporation of personalized, easy-to-use design features and motivators for engaging with the programme. Family involvement in the development of an eHealth programme such as 'Health-e-Hearts' has the potential to boost early adoption of healthy lifestyle behaviours among all family members.

Spatial Gene Expression Changes in the Mouse Heart After Base-Targeted Irradiation.

PURPOSE: Radiation cardiotoxicity (RC) is a clinically significant adverse effect of treatment for patients with thoracic malignancies. Clinical studies in lung cancer have indicated that heart substructures are not uniformly radiosensitive, and that dose to the heart base drives RC. In this study, we aimed to characterize late changes in gene expression using spatial transcriptomics in a mouse model of base regional radiosensitivity. METHODS AND MATERIALS: An aged female C57BL/6 mouse was irradiated with 16 Gy delivered to the cranial third of the heart using a 6 × 9 mm parallel opposed beam geometry on a small animal radiation research platform, and a second mouse was sham-irradiated. After echocardiography, whole hearts were collected at 30 weeks for spatial transcriptomic analysis to map gene expression changes occurring in different regions of the partially irradiated heart. Cardiac regions were manually annotated on the capture slides and the gene expression profiles compared across different regions. RESULTS: Ejection fraction was reduced at 30 weeks after a 16 Gy irradiation to the heart base, compared with the sham-irradiated controls. There were markedly more significant gene expression changes within the irradiated regions compared with nonirradiated regions. Variation was observed in the transcriptomic effects of radiation on different cardiac base structures (eg, between the right atrium [n = 86 dysregulated genes], left atrium [n = 96 dysregulated genes], and the vasculature [n = 129 dysregulated genes]). Disrupted biological processes spanned extracellular matrix as well as circulatory, neuronal, and contractility activities. CONCLUSIONS: This is the first study to report spatially resolved gene expression changes in irradiated tissues. Examination of the regional radiation response in the heart can help to further our understanding of the cardiac base's radiosensitivity and support the development of actionable targets for pharmacologic intervention and biologically relevant dose constraints.

Pre-Eclampsia Biomarkers for Women With Type 1 Diabetes Mellitus: A Comprehensive Review of Recent Literature.

Background: Pre-eclampsia is a serious consideration for women with type 1 diabetes mellitus (T1DM) planning pregnancy. Risk stratification strategies, such as biomarkers measured in the first trimester of pregnancy, could help identify high-risk women. The literature on T1DM-specific pre-eclampsia biomarkers is expanding. We aimed to provide a narrative review of recently published evidence to identify the most promising biomarker candidates that could be targeted for clinical implementation in existing PE models. Methods: A search using MeSH terms was carried out of Medline, EMBASE, Maternity and Infant Care, Web of Science, and Scopus for relevant papers published since 2015 inclusive and in English. The time limit was applied from the publication of the preceding systematic review in this field. Included studies had pre-eclampsia as a primary outcome, measured one or more serum, plasma or urine biomarkers at any time during pregnancy, and had a distinct group of women with T1DM who developed pre-eclampsia. Studies with pre-eclampsia as a composite outcome were not considered. No restrictions on study types were applied. A narrative synthesis approach was adopted for analysis. Results: A total of 510 records were screened yielding 18 eligible studies relating to 32 different biomarkers. Higher first-trimester levels of HbA1c and urinary albumin were associated with an increased risk of pre-eclampsia development in women with T1DM. Urinary neutrophil gelatinase-associated lipocalin and adipokines were novel biomarkers showing moderate predictive ability before 15 gestational weeks. Two T1DM-specific pre-eclampsia prediction models were proposed, measuring adipokines or urinary neutrophil gelatinase-associated lipocalin together with easily attainable maternal clinical characteristics. Contradicting previous literature, pre-eclampsia risk in women with T1DM was correlated with vitamin D levels and atherogenic lipid profile in the context of haptoglobin phenotype 2-2. Pregnancy-associated plasma protein-A and soluble endoglin did not predict pre-eclampsia in women with T1DM, and soluble Fms-like tyrosine kinase 1 only predicted pre-eclampsia from the third trimester. Conclusion: Maternally derived biomarkers reflecting glycemic control, insulin resistance and renal dysfunction performed better as PE predictors among women with T1DM than those derived from the placenta. These biomarkers could be trialed in current PE prediction algorithms to tailor them for women with T1DM.

Clinical features and predictive biomarkers for bladder cancer in patients with type 2 diabetes presenting with haematuria.

AIMS: To identify clinical features and protein biomarkers associated with bladder cancer (BC) in individuals with type 2 diabetes mellitus presenting with haematuria. MATERIALS AND METHODS: Data collected from the Haematuria Biomarker (HaBio) study was used in this analysis. A matched sub-cohort of patients with type 2 diabetes and patients without diabetes was created based on age, sex, and BC diagnosis, using approximately a 1:2 fixed ratio. Randox Biochip Array Technology and ELISA were applied for measurement of 66 candidate serum and urine protein biomarkers. Hazard ratios and 95% confidence intervals were estimated by chi-squared and Wilcoxon rank sum test for clinical features and candidate protein biomarkers. Diagnostic protein biomarker models were identified using Lasso-based binominal regression analysis. RESULTS: There was no difference in BC grade, stage, and severity between individuals with type 2 diabetes and matched controls. Incidence of chronic kidney disease (CKD) was significantly higher in patients with type 2 diabetes (p = 0.008), and CKD was significantly associated with BC in patients with type 2 diabetes (p = 0.032). A biomarker model, incorporating two serum (monocyte chemoattractant protein 1 and vascular endothelial growth factor) and three urine (interleukin 6, cytokeratin 18, and cytokeratin 8) proteins, predicted incidence of BC with an Area Under the Curve (AUC) of 0.84 in individuals with type 2 diabetes. In people without diabetes, the AUC was 0.66. CONCLUSIONS: We demonstrate the potential clinical utility of a biomarker panel, which includes proteins related to BC pathogenesis and type 2 diabetes, for monitoring risk of BC in patients with type 2 diabetes. Earlier urology referral of patients with type 2 diabetes will improve outcomes for these patients. TRIAL REGISTRATION: http://www.isrctn.com/ISRCTN25823942.

A Systematic Review of Clinical Prediction Rules to Predict Hospitalisation in Children with Lower Respiratory Infection in Primary Care and their Validation in a New Cohort.

Background: Our goal was to identify existing clinical prediction rules for predicting hospitalisation due to lower respiratory tract infection (LRTI) in children in primary care, guiding antibiotic therapy. A validation of these rules was then performed in a novel cohort of children presenting to primary care in Malawi with World Health Organisation clinically defined pneumonia. Methods: MEDLINE & EMBASE databases were searched for studies on the development, validation and clinical impact of clinical prediction models for hospitalisation in children with lower respiratory tract infection between January 1st1946-June 30th 2021. Two reviewers screened all abstracts and titles independently. The study was conducted in accordance with the Preferred Reporting Items for Systematic Reviews & Meta-Analyses guidelines. The BIOTOPE cohort (BIOmarkers TO diagnose PnEumonia) recruited children aged 2-59 months with WHO-defined pneumonia from two primary care facilities in Mzuzu, Malawi. Validation of identified rules was undertaken in this cohort. Findings: 1023 abstracts were identified. Following the removal of duplicates, a review of 989 abstracts was conducted leading to the identification of one eligible model. The CHARMS checklist for prediction modelling studies was utilized for evaluation. The area under the curve (AUC) of the STARWAVe rule for hospitalisation in BIOTOPE was found to be 0.80 (95% C.I of 0.75-0.85). The AUC of STARWAVe for a confirmed diagnosis of bacterial pneumonia was 0.39 (95% C.I 0.25-0.54). Interpretation: This review highlights the lack of clinical prediction rules in this area. The STARWAVe rule identified was useful in predicting hospitalisation from bacterial infection as defined. However, in the absence of a gold standard indicator for bacterial LRTI, this is a reasonable surrogate and could lead to reductions in antibiotic prescription rates, should clinical impact studies prove its utility. Further work to determine the clinical impact of STARWAVe and to identify diagnostic tests for bacterial LRTI in primary care is required.

NT-proBNP/BNP ratio for prognostication in European Caucasian patients enrolled in a heart failure prevention programme.

AIMS: Guidelines support the role of B-type natriuretic peptide (BNP) and amino-terminal pro-BNP (NT-proBNP) for risk stratification of patients in programmes to prevent heart failure (HF). Although biologically formed in a 1:1 ratio, the ratio of NT-proBNP to BNP exhibits wide inter-individual variability. A report on an Asian population suggests that molar NT-proBNP/BNP ratio is associated with incident HF. This study aims to determine whether routine, simultaneous evaluation of both BNP and NT-proBNP is warranted in a European, Caucasian population. METHODS AND RESULTS: We determined BNP and NT-proBNP levels for 782 Stage A/B HF patients in the STOP-HF programme. The clinical, echocardiographic, and biochemical associates of molar NT-proBNP/BNP ratio were analysed. The primary endpoint was the adjusted association of baseline molar NT-proBNP/BNP ratio with new-onset HF and/or progression of left ventricular dysfunction (LVD). We estimated the C-statistic, integrated discrimination improvement, and the category-free net reclassification improvement metric for the addition of molar NT-proBNP/BNP ratio to adjusted models. The median age was 66.6 years [interquartile range (IQR) 59.5-73.1], 371 (47.4%) were female, and median molar NT-proBNP/BNP ratio was 1.91 (IQR 1.37-2.93). Estimated glomerular filtration rate, systolic blood pressure, left ventricular mass index, and heart rate were associated with NT-proBNP/BNP ratio in a linear regression model (all P < 0.05). Over a median follow-up period of 5 years (IQR 3.4-6.8), 247 (31.5%) patients developed HF or progression of LVD. Log-transformed NT-proBNP/BNP ratio is inversely associated with HF and LVD risk when adjusted for age, gender, diabetes, hypertension, vascular disease, obesity, heart rate, number of years of follow-up, estimated glomerular filtration rate, and baseline NT-proBNP (odds ratio 0.71, 95% confidence interval 0.55-0.91; P = 0.008). However, molar NT-proBNP/BNP ratio did not increase the C-statistic (Δ -0.01) and net reclassification improvement (0.0035) for prediction of HF and LVD compared with NT-proBNP or BNP alone. Substitution of NT-proBNP for BNP in the multivariable model eliminated the association with HF and LVD risk. CONCLUSIONS: This study characterized, for the first time in a Caucasian Stage A/B HF population, the relationship between NT-proBNP/BNP ratio and biological factors and demonstrated an inverse relationship with the future development of HF and LVD. However, this study does not support routine simultaneous BNP and NT-proBNP measurement in HF prevention programmes amongst European, Caucasian patients.

FKBPL and SIRT-1 Are Downregulated by Diabetes in Pregnancy Impacting on Angiogenesis and Endothelial Function.

Diabetes in pregnancy is associated with adverse pregnancy outcomes including preterm birth. Although the mechanisms leading to these pregnancy complications are still poorly understood, aberrant angiogenesis and endothelial dysfunction play a key role. FKBPL and SIRT-1 are critical regulators of angiogenesis, however, their roles in pregnancies affected by diabetes have not been examined before in detail. Hence, this study aimed to investigate the role of FKBPL and SIRT-1 in pre-gestational (type 1 diabetes mellitus, T1D) and gestational diabetes mellitus (GDM). Placental protein expression of important angiogenesis proteins, FKBPL, SIRT-1, PlGF and VEGF-R1, was determined from pregnant women with GDM or T1D, and in the first trimester trophoblast cells exposed to high glucose (25 mM) and varying oxygen concentrations [21%, 6.5%, 2.5% (ACH-3Ps)]. Endothelial cell function was assessed in high glucose conditions (30 mM) and following FKBPL overexpression. Placental FKBPL protein expression was downregulated in T1D (FKBPL; p<0.05) whereas PlGF/VEGF-R1 were upregulated (p<0.05); correlations adjusted for gestational age were also significant. In the presence of GDM, only SIRT-1 was significantly downregulated (p<0.05) even when adjusted for gestational age (r=-0.92, p=0.001). Both FKBPL and SIRT-1 protein expression was reduced in ACH-3P cells in high glucose conditions associated with 6.5%/2.5% oxygen concentrations compared to experimental normoxia (21%; p<0.05). FKBPL overexpression in endothelial cells (HUVECs) exacerbated reduction in tubule formation compared to empty vector control, in high glucose conditions (junctions; p<0.01, branches; p<0.05). In conclusion, FKBPL and/or SIRT-1 downregulation in response to diabetic pregnancies may have a key role in the development of vascular dysfunction and associated complications affected by impaired placental angiogenesis.

Salivary Biomarkers for Diagnosis and Therapy Monitoring in Patients with Heart Failure. A Systematic Review.

The aim of this study was to perform a systematic review on the potential value of saliva biomarkers in the diagnosis, management and prognosis of heart failure (HF). The correlation between saliva and plasma values of these biomarkers was also studied. PubMed was searched to collect relevant literature, i.e., case-control, cross-sectional studies that either compared the values of salivary biomarkers among healthy subjects and HF patients, or investigated their role in risk stratification and prognosis in HF patients. No randomized control trials were included. The search ended on 31st of December 2020. A total of 15 studies met the inclusion criteria. 18 salivary biomarkers were analyzed and the levels of all biomarkers studied were found to be higher in HF patients compared to controls, except for amylase, sodium, and chloride that had smaller saliva concentrations in HF patients. Natriuretic peptides are the most commonly used plasma biomarkers in the management of HF. Their saliva levels show promising results, although the correlation of saliva to plasma values is weakened in higher plasma values. In most of the publications, differences in biomarker levels between HF patients and controls were found to be statistically significant. Due to the small number of patients included, larger studies need to be conducted in order to facilitate the use of saliva biomarkers in clinical practice.

ECSIT is a critical limiting factor for cardiac function.

Evolutionarily conserved signaling intermediate in Toll pathways (ECSIT) is a protein with roles in early development, activation of the transcription factor NF-κB, and production of mitochondrial reactive oxygen species (mROS) that facilitates clearance of intracellular bacteria like Salmonella. ECSIT is also an important assembly factor for mitochondrial complex I. Unlike the murine form of Ecsit (mEcsit), we demonstrate here that human ECSIT (hECSIT) is highly labile. To explore whether the instability of hECSIT affects functions previously ascribed to its murine counterpart, we created a potentially novel transgenic mouse in which the murine Ecsit gene is replaced by the human ECSIT gene. The humanized mouse has low levels of hECSIT protein, in keeping with its intrinsic instability. Whereas low-level expression of hECSIT was capable of fully compensating for mEcsit in its roles in early development and activation of the NF-κB pathway, macrophages from humanized mice showed impaired clearance of Salmonella that was associated with reduced production of mROS. Notably, severe cardiac hypertrophy was manifested in aging humanized mice, leading to premature death. The cellular and molecular basis of this phenotype was delineated by showing that low levels of human ECSIT protein led to a marked reduction in assembly and activity of mitochondrial complex I with impaired oxidative phosphorylation and reduced production of ATP. Cardiac tissue from humanized hECSIT mice also showed reduced mitochondrial fusion and more fission but impaired clearance of fragmented mitochondria. A cardiomyocyte-intrinsic role for Ecsit in mitochondrial function and cardioprotection is also demonstrated. We also show that cardiac fibrosis and damage in humans correlated with low expression of human ECSIT. In summary, our findings identify a role for ECSIT in cardioprotection, while generating a valuable experimental model to study mitochondrial dysfunction and cardiac pathophysiology.

Effectiveness of family-based eHealth interventions in cardiovascular disease risk reduction: A systematic review.

Family-based eHealth interventions to reduce cardiovascular disease risk have potential as a primary prevention strategy to improve the health of parents and their children. This systematic review evaluated the effectiveness of such interventions in modifying parent and child/adolescent risk factors such as body mass index, physical activity, dietary intakes and alcohol use. Five electronic databases were searched up to April 2020. Of 2193 articles identified, seven randomised controlled trials met inclusion criteria and were reviewed. Data were extracted regarding study setting, design, methods, eHealth technology used, intervention and control group components, retention rates, outcome measures, incentives and limitations. Risk of bias and quality assessment were carried out using Cochrane methods. A qualitative narrative data synthesis of the studies was conducted. Our review found that three studies showed an improvement in alcohol use among parents and adolescents as a result of the eHealth intervention. Among children/adolescents, two studies showed an improvement in dietary intake, one study showed an improvement in physical activity, and one study showed an improvement in body mass index as a result of the eHealth intervention. Interventions appeared more likely to be effective if they were theory-based, had longer follow-up periods, were incentivised and included regular interaction. Our findings suggest that, despite a paucity of high-quality trials, there is some evidence that family-based eHealth interventions have potential to reduce cardiovascular disease risk. However, more sufficiently powered, higher-quality trials with theory driven, clearly described interventions and unambiguous outcomes are needed.

Multiplexed measurement of candidate blood protein biomarkers of heart failure.

AIMS: There is a critical need for better biomarkers so that heart failure can be diagnosed at an earlier stage and with greater accuracy. The purpose of this study was to design a robust mass spectrometry (MS)-based assay for the simultaneous measurement of a panel of 35 candidate protein biomarkers of heart failure, in blood. The overall aim was to evaluate the potential clinical utility of this biomarker panel for prediction of heart failure in a cohort of 500 patients. METHODS AND RESULTS: Multiple reaction monitoring (MRM) MS assays were designed with Skyline and Spectrum Mill PeptideSelector software and developed using nanoflow reverse phase C18 chromatographic Chip Cube-based separation, coupled to a 6460 triple quadrupole mass spectrometer. Optimized MRM assays were applied, in a sample-blinded manner, to serum samples from a cohort of 500 patients with heart failure and non-heart failure (non-HF) controls who had cardiovascular risk factors. Both heart failure with reduced ejection fraction (HFrEF) patients and heart failure with preserved ejection fraction (HFpEF) patients were included in the study. Peptides for the Apolipoprotein AI (APOA1) protein were the most significantly differentially expressed between non-HF and heart failure patients (P = 0.013 and P = 0.046). Four proteins were significantly differentially expressed between non-HF and the specific subtypes of HF (HFrEF and HFpEF); Leucine-rich-alpha-2-glycoprotein (LRG1, P < 0.001), zinc-alpha-2-glycoprotein (P = 0.005), serum paraoxanse/arylesterase (P = 0.013), and APOA1 (P = 0.038). A statistical model found that combined measurements of the candidate biomarkers in addition to BNP were capable of correctly predicting heart failure with 83.17% accuracy and an area under the curve (AUC) of 0.90. This was a notable improvement on predictive capacity of BNP measurements alone, which achieved 77.1% accuracy and an AUC of 0.86 (P = 0.005). The protein peptides for LRG1, which contributed most significantly to model performance, were significantly associated with future new onset HF in the non-HF cohort [Peptide 1: odds ratio (OR) 2.345 95% confidence interval (CI) (1.456-3.775) P = 0.000; peptide 2: OR 2.264 95% CI (1.422-3.605), P = 0.001]. CONCLUSIONS: This study has highlighted a number of promising candidate biomarkers for (i) diagnosis of heart failure and subtypes of heart failure and (ii) prediction of future new onset heart failure in patients with cardiovascular risk factors. Furthermore, this study demonstrates that multiplexed measurement of a combined biomarker signature that includes BNP is a more accurate predictor of heart failure than BNP alone.

Biomarker profiling for risk of future heart failure (HFpEF) development.

BACKGROUND: The purpose of this study was to investigate the utility of BNP, hsTroponin-I, interleukin-6, sST2, and galectin-3 in predicting the future development of new onset heart failure with preserved ejection fraction (HFpEF) in asymptomatic patients at-risk for HF. METHODS: This is a retrospective analysis of the longitudinal STOP-HF study of thirty patients who developed HFpEF matched to a cohort that did not develop HFpEF (n = 60) over a similar time period. Biomarker candidates were quantified at two time points prior to initial HFpEF diagnosis. RESULTS: HsTroponin-I and BNP at baseline and follow-up were statistically significant predictors of future new onset HFpEF, as was galectin-3 at follow-up and concentration change over time. Interleukin-6 and sST2 were not predictive of future development of new onset HFpEF in this study. Unadjusted biomarker combinations of hsTroponin-I, BNP, and galectin-3 could significantly predict future HFpEF using both baseline (AUC 0.82 [0.73,0.92]) and follow-up data (AUC 0.86 [0.79,0.94]). A relative-risk matrix was developed to categorize the relative-risk of new onset of HFpEF based on biomarker threshold levels. CONCLUSION: We provided evidence for the utility of BNP, hsTroponin-I, and Galectin-3 in the prediction of future HFpEF in asymptomatic event-free populations with cardiovascular disease risk factors.

Epigenetic Regulation of Endothelial Cell Function by Nucleic Acid Methylation in Cardiac Homeostasis and Disease.

Pathological remodelling of the myocardium, including inflammation, fibrosis and hypertrophy, in response to acute or chronic injury is central in the development and progression of heart failure (HF). While both resident and infiltrating cardiac cells are implicated in these pathophysiological processes, recent evidence has suggested that endothelial cells (ECs) may be the principal cell type responsible for orchestrating pathological changes in the failing heart. Epigenetic modification of nucleic acids, including DNA, and more recently RNA, by methylation is essential for physiological development due to their critical regulation of cellular gene expression. As accumulating evidence has highlighted altered patterns of DNA and RNA methylation in HF at both the global and individual gene levels, much effort has been directed towards defining the precise role of such cell-specific epigenetic changes in the context of HF. Considering the increasingly apparent crucial role that ECs play in cardiac homeostasis and disease, this article will specifically focus on nucleic acid methylation (both DNA and RNA) in the failing heart, emphasising the key influence of these epigenetic mechanisms in governing EC function. This review summarises current understanding of DNA and RNA methylation alterations in HF, along with their specific role in regulating EC function in response to stress (e.g. hyperglycaemia, hypoxia). Improved appreciation of this important research area will aid in further implicating dysfunctional ECs in HF pathogenesis, whilst informing development of EC-targeted strategies and advancing potential translation of epigenetic-based therapies for specific targeting of pathological cardiac remodelling in HF.