Development of a long noncoding RNA-based machine learning model to predict COVID-19 in-hospital mortality.

Tools for predicting COVID-19 outcomes enable personalized healthcare, potentially easing the disease burden. This collaborative study by 15 institutions across Europe aimed to develop a machine learning model for predicting the risk of in-hospital mortality post-SARS-CoV-2 infection. Blood samples and clinical data from 1286 COVID-19 patients collected from 2020 to 2023 across four cohorts in Europe and Canada were analyzed, with 2906 long non-coding RNAs profiled using targeted sequencing. From a discovery cohort combining three European cohorts and 804 patients, age and the long non-coding RNA LEF1-AS1 were identified as predictive features, yielding an AUC of 0.83 (95% CI 0.82-0.84) and a balanced accuracy of 0.78 (95% CI 0.77-0.79) with a feedforward neural network classifier. Validation in an independent Canadian cohort of 482 patients showed consistent performance. Cox regression analysis indicated that higher levels of LEF1-AS1 correlated with reduced mortality risk (age-adjusted hazard ratio 0.54, 95% CI 0.40-0.74). Quantitative PCR validated LEF1-AS1's adaptability to be measured in hospital settings. Here, we demonstrate a promising predictive model for enhancing COVID-19 patient management.

Persistent transcriptional changes in cardiac adaptive immune cells following myocardial infarction: New evidence from the re-analysis of publicly available single cell and nuclei RNA-sequencing data sets.

INTRODUCTION: Chronic immunopathology contributes to the development of heart failure after a myocardial infarction. Both T and B cells of the adaptive immune system are present in the myocardium and have been suggested to be involved in post-MI immunopathology. METHODS: We analyzed the B and T cell populations isolated from previously published single cell RNA-sequencing data sets (PMID: 32130914, PMID: 35948637, PMID: 32971526 and PMID: 35926050), of the mouse and human heart, using differential expression analysis, functional enrichment analysis, gene regulatory inferences, and integration with autoimmune and cardiovascular GWAS. RESULTS: Already at baseline, mature effector B and T cells are present in the human and mouse heart, having increased activity in transcription factors maintaining tolerance (e.g. DEAF1, JDP2, SPI-B). Following MI, T cells upregulate pro-inflammatory transcript levels (e.g. Cd11, Gzmk, Prf1), while B cells upregulate activation markers (e.g. Il6, Il1rn, Ccl6) and collagen (e.g. Col5a2, Col4a1, Col1a2). Importantly, pro-inflammatory and fibrotic transcription factors (e.g. NFKB1, CREM, REL) remain active in T cells, while B cells maintain elevated activity in transcription factors related to immunoglobulin production (e.g. ERG, REL) in both mouse and human post-MI hearts. Notably, genes differentially expressed in post-MI T and B cells are associated with cardiovascular and autoimmune disease. CONCLUSION: These findings highlight the varied and time-dependent dynamic roles of post-MI T and B cells. They appear ready-to-go and are activated immediately after MI, thus participate in the acute wound healing response. However, they subsequently remain in a state of pro-inflammatory activation contributing to persistent immunopathology.

Evolution of journal clubs: fostering collaborative learning in modern research.

Journal clubs have been a staple in scientific communities, facilitating discussions on recent publications. However, the overwhelming volume of biomedical information poses a challenge in literature selection. This article provides an overview of journal club types and their efficacy in training potential peer reviewers, enhancing communication skills, and critical thinking. Originating in the 19th century, journal clubs have evolved from traditional in-person meetings to virtual or hybrid formats, accelerated by the COVID-19 pandemic. Face-to-face interactions offer personal connections, while virtual events ensure wider participation and accessibility. Organizing journal clubs demands effort, but it has several benefits, including promoting new publications and providing a platform for meaningful discussions. The virtual CardioRNA J-club experience exemplifies successful multidisciplinary collaboration, fostering international connections and inspiring new research. Journal clubs remain a vital component of academic research, equipping senior researchers with the latest developments and nurturing the next generation of scientists. As millennial and Gen Z researchers join the scientific field, journal clubs continue to evolve as a fertile ground for education and collaborative learning in an ever-changing scientific landscape.

Non-coding RNAs as therapeutic targets and biomarkers in ischaemic heart disease.

The adult heart is a complex, multicellular organ that is subjected to a series of regulatory stimuli and circuits and has poor reparative potential. Despite progress in our understanding of disease mechanisms and in the quality of health care, ischaemic heart disease remains the leading cause of death globally, owing to adverse cardiac remodelling, leading to ischaemic cardiomyopathy and heart failure. Therapeutic targets are urgently required for the protection and repair of the ischaemic heart. Moreover, personalized clinical biomarkers are necessary for clinical diagnosis, medical management and to inform the individual response to treatment. Non-coding RNAs (ncRNAs) deeply influence cardiovascular functions and contribute to communication between cells in the cardiac microenvironment and between the heart and other organs. As such, ncRNAs are candidates for translation into clinical practice. However, ncRNA biology has not yet been completely deciphered, given that classes and modes of action have emerged only in the past 5 years. In this Review, we discuss the latest discoveries from basic research on ncRNAs and highlight both the clinical value and the challenges underscoring the translation of these molecules as biomarkers and therapeutic regulators of the processes contributing to the initiation, progression and potentially the prevention or resolution of ischaemic heart disease and heart failure.

HCG18, LEF1AS1 and lncCEACAM21 as biomarkers of disease severity in the peripheral blood mononuclear cells of COVID-19 patients.

BACKGROUND: Even after 3 years from SARS-CoV-2 identification, COVID-19 is still a persistent and dangerous global infectious disease. Significant improvements in our understanding of the disease pathophysiology have now been achieved. Nonetheless, reliable and accurate biomarkers for the early stratification of COVID-19 severity are still lacking. Long noncoding RNAs (LncRNAs) are ncRNAs longer than 200 nucleotides, regulating the transcription and translation of protein-coding genes and they can be found in the peripheral blood, thus holding a promising biomarker potential. Specifically, peripheral blood mononuclear cells (PBMCs) have emerged as a source of indirect biomarkers mirroring the conditions of tissues: they include monocytes, B and T lymphocytes, and natural killer T cells (NKT), being highly informative for immune-related events. METHODS: We profiled by RNA-Sequencing a panel of 2906 lncRNAs to investigate their modulation in PBMCs of a pilot group of COVID-19 patients, followed by qPCR validation in 111 hospitalized COVID-19 patients. RESULTS: The levels of four lncRNAs were found to be decreased in association with COVID-19 mortality and disease severity: HLA Complex Group 18-242 and -244 (HCG18-242 and HCG18-244), Lymphoid Enhancer Binding Factor 1-antisense 1 (LEF1-AS1) and lncCEACAM21 (i.e. ENST00000601116.5, a lncRNA in the CEACAM21 locus). Interestingly, these deregulations were confirmed in an independent patient group of hospitalized patients and by the re-analysis of publicly available single-cell transcriptome datasets. The identified lncRNAs were expressed in all of the PBMC cell types and inversely correlated with the neutrophil/lymphocyte ratio (NLR), an inflammatory marker. In vitro, the expression of LEF1-AS1 and lncCEACAM21 was decreased upon THP-1 monocytes exposure to a relevant stimulus, hypoxia. CONCLUSION: The identified COVID-19-lncRNAs are proposed as potential innovative biomarkers of COVID-19 severity and mortality.

Multi-response Mendelian randomization: Identification of shared and distinct exposures for multimorbidity and multiple related disease outcomes.

The existing framework of Mendelian randomization (MR) infers the causal effect of one or multiple exposures on one single outcome. It is not designed to jointly model multiple outcomes, as would be necessary to detect causes of more than one outcome and would be relevant to model multimorbidity or other related disease outcomes. Here, we introduce multi-response Mendelian randomization (MR2), an MR method specifically designed for multiple outcomes to identify exposures that cause more than one outcome or, conversely, exposures that exert their effect on distinct responses. MR2 uses a sparse Bayesian Gaussian copula regression framework to detect causal effects while estimating the residual correlation between summary-level outcomes, i.e., the correlation that cannot be explained by the exposures, and vice versa. We show both theoretically and in a comprehensive simulation study how unmeasured shared pleiotropy induces residual correlation between outcomes irrespective of sample overlap. We also reveal how non-genetic factors that affect more than one outcome contribute to their correlation. We demonstrate that by accounting for residual correlation, MR2 has higher power to detect shared exposures causing more than one outcome. It also provides more accurate causal effect estimates than existing methods that ignore the dependence between related responses. Finally, we illustrate how MR2 detects shared and distinct causal exposures for five cardiovascular diseases in two applications considering cardiometabolic and lipidomic exposures and uncovers residual correlation between summary-level outcomes reflecting known relationships between cardiovascular diseases.

Integration of epigenetic regulatory mechanisms in heart failure.

The number of "omics" approaches is continuously growing. Among others, epigenetics has appeared as an attractive area of investigation by the cardiovascular research community, notably considering its association with disease development. Complex diseases such as cardiovascular diseases have to be tackled using methods integrating different omics levels, so called "multi-omics" approaches. These approaches combine and co-analyze different levels of disease regulation. In this review, we present and discuss the role of epigenetic mechanisms in regulating gene expression and provide an integrated view of how these mechanisms are interlinked and regulate the development of cardiac disease, with a particular attention to heart failure. We focus on DNA, histone, and RNA modifications, and discuss the current methods and tools used for data integration and analysis. Enhancing the knowledge of these regulatory mechanisms may lead to novel therapeutic approaches and biomarkers for precision healthcare and improved clinical outcomes.

Cardiovascular complications of diabetes: role of non-coding RNAs in the crosstalk between immune and cardiovascular systems.

Diabetes mellitus, a group of metabolic disorders characterized by high levels of blood glucose caused by insulin defect or impairment, is a major risk factor for cardiovascular diseases and related mortality. Patients with diabetes experience a state of chronic or intermittent hyperglycemia resulting in damage to the vasculature, leading to micro- and macro-vascular diseases. These conditions are associated with low-grade chronic inflammation and accelerated atherosclerosis. Several classes of leukocytes have been implicated in diabetic cardiovascular impairment. Although the molecular pathways through which diabetes elicits an inflammatory response have attracted significant attention, how they contribute to altering cardiovascular homeostasis is still incompletely understood. In this respect, non-coding RNAs (ncRNAs) are a still largely under-investigated class of transcripts that may play a fundamental role. This review article gathers the current knowledge on the function of ncRNAs in the crosstalk between immune and cardiovascular cells in the context of diabetic complications, highlighting the influence of biological sex in such mechanisms and exploring the potential role of ncRNAs as biomarkers and targets for treatments. The discussion closes by offering an overview of the ncRNAs involved in the increased cardiovascular risk suffered by patients with diabetes facing Sars-CoV-2 infection.

Efficacy of propofol-supplemented cardioplegia on biomarkers of organ injury in patients having cardiac surgery using cardiopulmonary bypass: A protocol for a randomised controlled study (ProMPT2).

INTRODUCTION: Cardiac surgery with cardiopulmonary bypass and cardioplegic arrest is known to be responsible for ischaemia and reperfusion organ injury. In a previous study, ProMPT, in patients undergoing coronary artery bypass or aortic valve surgery we demonstrated improved cardiac protection when supplementing the cardioplegia solution with propofol (6 mcg/ml). The aim of the ProMPT2 study is to determine whether higher levels of propofol added to the cardioplegia could result in increased cardiac protection. METHODS AND ANALYSIS: The ProMPT2 study is a multi-centre, parallel, three-group, randomised controlled trial in adults undergoing non-emergency isolated coronary artery bypass graft surgery with cardiopulmonary bypass. A total of 240 patients will be randomised in a 1:1:1 ratio to receive either cardioplegia supplementation with high dose of propofol (12 mcg/ml), low dose of propofol (6 mcg/ml) or placebo (saline). The primary outcome is myocardial injury, assessed by serial measurements of myocardial troponin T up to 48 hours after surgery. Secondary outcomes include biomarkers of renal function (creatinine) and metabolism (lactate). ETHICS AND DISSEMINATION: The trial received research ethics approval from South Central - Berkshire B Research Ethics Committee and Medicines and Healthcare products Regulatory Agency in September 2018. Any findings will be shared though peer-reviewed publications and presented at international and national meetings. Participants will be informed of results through patient organisations and newsletters. TRIAL REGISTRATION: ISRCTN15255199. Registered in March 2019.

Consequences of the exposome to gestational diabetes mellitus.

The exposome is the cumulative measure of environmental influences and associated biological responses throughout the lifespan, including those from the environment, diet, behaviour, and endogenous processes. The exposome concept and the 2030 Agenda for the Sustainable Development Goals (SDGs) from the United Nations are the basis for understanding the aetiology and consequences of non-communicable diseases, including gestational diabetes mellitus (GDM). Pregnancy may be developed in an environment with adverse factors part of the immediate internal medium for fetus development and the external medium to which the pregnant woman is exposed. The placenta is the interface between maternal and fetal compartments and acts as a protective barrier or easing agent to transfer exposome from mother to fetus. Under and over-nutrition in utero, exposure to adverse environmental pollutants such as heavy metals, endocrine-disrupting chemicals, pesticides, drugs, pharmaceuticals, lifestyle, air pollutants, and tobacco smoke plays a determinant role in the development of GDM. This phenomenon is worsened by metabolic stress postnatally, such as obesity which increases the risk of GDM and other diseases. Clinical risk factors for GDM development include its aetiology. It is proposed that knowledge-based interventions to change the potential interdependent ecto-exposome and endo-exposome could avoid the occurrence and consequences of GDM.

Efficacy of treatments tested in COVID-19 patients with cardiovascular disease. A meta-analysis.

BACKGROUND: The COVID-19 pandemic has spread globally infecting and killing millions. Those with cardiovascular disease (CVD) are at higher risk of increased disease severity and mortality. We performed a systematic review and meta-analysis to estimate the rate of in-hospital mortality following different treatments on COVID-19 in patients with CVD. METHODS: Pertinent articles were identified from the PubMed, Google Scholar, Ovid MEDLINE, and Ovid EMBASE databases. This study protocol was registered under PROSPERO with the identifier CRD42020183057. RESULTS: Of the 1673 papers scrutinized, 46 were included in the review. Of the 2553 patients (mean age 63.9 ± 2.7 years/o; 57.2% male), the most frequent CVDs were coronary artery disease (9.09%) and peripheral arterial disease (5.4%) and the most frequent cardiovascular risk factors were hypertension (86.7%) and diabetes (23.7%). Most patients were on multiple treatments. 14 COVID-19 treatments were compared with controls. The pooled event rate for in-hospital mortality was 20% (95% confidence interval (CI): 11-33%); certain heterogeneity was observed across studies. CONCLUSIONS: COVID-19 is associated with a high in-hospital mortality rate in patients with CVD. This study shows that previous CVD determines mortality, regardless of the type of COVID-19 administered therapy. Treatments for at-risk patients should be administered carefully and monitored closely until further data are available.

Methods for the identification and characterization of extracellular vesicles in cardiovascular studies: from exosomes to microvesicles.

Extracellular vesicles (EVs) are nanosized vesicles with a lipid bilayer that are released from cells of the cardiovascular system, and are considered important mediators of intercellular and extracellular communications. Two types of EVs of particular interest are exosomes and microvesicles, which have been identified in all tissue and body fluids and carry a variety of molecules including RNAs, proteins, and lipids. EVs have potential for use in the diagnosis and prognosis of cardiovascular diseases and as new therapeutic agents, particularly in the setting of myocardial infarction and heart failure. Despite their promise, technical challenges related to their small size make it challenging to accurately identify and characterize them, and to study EV-mediated processes. Here, we aim to provide the reader with an overview of the techniques and technologies available for the separation and characterization of EVs from different sources. Methods for determining the protein, RNA, and lipid content of EVs are discussed. The aim of this document is to provide guidance on critical methodological issues and highlight key points for consideration for the investigation of EVs in cardiovascular studies.

The science behind soft skills: Do's and Don'ts for early career researchers and beyond. A review paper from the EU-CardioRNA COST Action CA17129.

Soft skills are the elementary management, personal, and interpersonal abilities that are vital for an individual to be efficient at workplace or in their personal life. Each work place requires different set of soft skills. Thus, in addition to scientific/technical skills that are easier to access within a short time frame, several key soft skills are essential for the success of a researcher in today's international work environment. In this paper, the trainees and trainers of the EU-CardioRNA COST Action CA17129 training school on soft skills present basic and advanced soft skills for early career researchers. Here, we particularly emphasize on the importance of transferable and presentation skills, ethics, literature reading and reviewing, research protocol and grant writing, networking, and career opportunities for researchers. All these skills are vital but are often overlooked by some scholars. We also provide tips to ace in aforementioned skills that are crucial in a day-to-day life of early and late career researchers in academia and industry.

A Segmental Approach from Molecular Profiling to Medical Imaging to Study Bicuspid Aortic Valve Aortopathy.

Bicuspid aortic valve (BAV) patients develop ascending aortic (AAo) dilation. The pathogenesis of BAV aortopathy (genetic vs. haemodynamic) remains unclear. This study aims to identify regional changes around the AAo wall in BAV patients with aortopathy, integrating molecular data and clinical imaging. BAV patients with aortopathy (n = 15) were prospectively recruited to surgically collect aortic tissue and measure molecular markers across the AAo circumference. Dilated (anterior/right) vs. non-dilated (posterior/left) circumferential segments were profiled for whole-genomic microRNAs (next-generation RNA sequencing, miRCURY LNA PCR), protein content (tandem mass spectrometry), and elastin fragmentation and degeneration (histomorphometric analysis). Integrated bioinformatic analyses of RNA sequencing and proteomic datasets identified five microRNAs (miR-128-3p, miR-210-3p, miR-150-5p, miR-199b-5p, and miR-21-5p) differentially expressed across the AAo circumference. Among them, three miRNAs (miR-128-3p, miR-150-5p, and miR-199b-5p) were predicted to have an effect on eight common target genes, whose expression was dysregulated, according to proteomic analyses, and involved in the vascular-endothelial growth-factor signalling, Hippo signalling, and arachidonic acid pathways. Decreased elastic fibre levels and elastic layer thickness were observed in the dilated segments. Additionally, in a subset of patients n = 6/15, a four-dimensional cardiac magnetic resonance (CMR) scan was performed. Interestingly, an increase in wall shear stress (WSS) was observed at the anterior/right wall segments, concomitantly with the differentially expressed miRNAs and decreased elastic fibres. This study identified new miRNAs involved in the BAV aortic wall and revealed the concomitant expressional dysregulation of miRNAs, proteins, and elastic fibres on the anterior/right wall in dilated BAV patients, corresponding to regions of elevated WSS.

PPMS: A framework to Profile Primary MicroRNAs from Single-cell RNA-sequencing datasets.

MOTIVATION: Single-cell/nuclei RNA-sequencing (scRNA-seq) technologies can simultaneously quantify gene expression in thousands of cells across the genome. However, the majority of the noncoding RNAs, such as microRNAs (miRNAs), cannot currently be profiled at the same scale. MiRNAs are a class of small noncoding RNAs and play an important role in gene regulation. MiRNAs originate from the processing of primary transcripts, known as primary-microRNAs (pri-miRNAs). The pri-miRNA transcripts, independent of their cognate miRNAs, can also function as long noncoding RNAs, code for micropeptides or even interact with DNA, acting like enhancers. Therefore, it is apparent that the significance of scRNA-seq pri-miRNA profiling expands beyond using pri-miRNA as proxies of mature miRNAs. However, there are no computational methods that allow profiling and quantification of pri-miRNAs at the single-cell-type resolution. RESULTS: We have developed a simple yet effective computational framework to profile pri-MiRNAs from single-cell RNA-sequencing datasets (PPMS). Based on user input, PPMS can profile pri-miRNAs at cell-type resolution. PPMS can be applied to both newly produced and publicly available datasets obtained via single cell or single-nuclei RNA-seq. It allows users to (i) investigate the distribution of pri-miRNAs across cell types and cell states and (ii) establish a relationship between the number of cells/reads sequenced and the detection of pri-miRNAs. Here, to demonstrate its efficacy, we have applied PPMS to publicly available scRNA-seq data generated from (i) individual chambers (ventricles and atria) of the human heart, (ii) human pluripotent stem cells during their differentiation into cardiomyocytes (the heart beating cells) and (iii) hiPSCs-derived cardiomyocytes infected with severe acute respiratory syndrome coronavirus 2.

Pathology-related changes in cardiac energy metabolites, inflammatory response and reperfusion injury following cardioplegic arrest in patients undergoing open-heart surgery.

Introduction: Changes in cardiac metabolites in adult patients undergoing open-heart surgery using ischemic cardioplegic arrest have largely been reported for non-ventricular tissue or diseased left ventricular tissue, with few studies attempting to assess such changes in both ventricular chambers. It is also unknown whether such changes are altered in different pathologies or linked to the degree of reperfusion injury and inflammatory response. The aim of the present work was to address these issues by monitoring myocardial metabolites in both ventricles and to establish whether these changes are linked to reperfusion injury and inflammatory/stress response in patients undergoing surgery using cold blood cardioplegia for either coronary artery bypass graft (CABG, n = 25) or aortic valve replacement (AVR, n = 16). Methods: Ventricular biopsies from both left (LV) and right (RV) ventricles were collected before ischemic cardioplegic arrest and 20 min after reperfusion. The biopsies were processed for measuring selected metabolites (adenine nucleotides, purines, and amino acids) using HPLC. Blood markers of cardiac injury (Troponin I, cTnI), inflammation (IL- 6, IL-8, Il-10, and TNFα, measured using Multiplex) and oxidative stress (Myeloperoxidase, MPO) were measured pre- and up to 72 hours post-operatively. Results: The CABG group had a significantly shorter ischemic cardioplegic arrest time (38.6 ± 2.3 min) compared to AVR group (63.0 ± 4.9 min, p = 2 x 10-6). Cardiac injury (cTnI release) was similar for both CABG and AVR groups. The inflammatory markers IL-6 and Il-8 were significantly higher in CABG patients compared to AVR patients. Metabolic markers of cardiac ischemic stress were relatively and significantly more altered in the LV of CABG patients. Comparing diabetic and non-diabetic CABG patients shows that only the RV of diabetic patients sustained major ischemic stress during reperfusion and that diabetic patients had a significantly higher inflammatory response. Discussion: CABG patients sustain relatively more ischemic stress, systemic inflammatory response and similar injury and oxidative stress compared to AVR patients despite having significantly shorter cross-clamp time. The higher inflammatory response in CABG patients appears to be at least partly driven by a higher incidence of diabetes amongst CABG patients. In addition to pathology, the use of cold blood cardioplegic arrest may underlie these differences.

Effect of cardioplegic arrest and reperfusion on left and right ventricular proteome/phosphoproteome in patients undergoing surgery for coronary or aortic valve disease.

Our earlier work has shown inter­disease and intra­disease differences in the cardiac proteome between right (RV) and left (LV) ventricles of patients with aortic valve stenosis (AVS) or coronary artery disease (CAD). Whether disease remodeling also affects acute changes occuring in the proteome during surgical intervention is unknown. This study investigated the effects of cardioplegic arrest on cardiac proteins/phosphoproteins in LV and RV of CAD (n=6) and AVS (n=6) patients undergoing cardiac surgery. LV and RV biopsies were collected during surgery before ischemic cold blood cardioplegic arrest (pre) and 20 min after reperfusion (post). Tissues were snap frozen, proteins extracted, and the extracts were used for proteomic and phosphoproteomic analysis using Tandem Mass Tag (TMT) analysis. The results were analysed using QuickGO and Ingenuity Pathway Analysis softwares. For each comparision, our proteomic analysis identified more than 3,000 proteins which could be detected in both the pre and Post samples. Cardioplegic arrest and reperfusion were associated with significant differential expression of 24 (LV) and 120 (RV) proteins in the CAD patients, which were linked to mitochondrial function, inflammation and cardiac contraction. By contrast, AVS patients showed differential expression of only 3 LV proteins and 2 RV proteins, despite a significantly longer duration of ischaemic cardioplegic arrest. The relative expression of 41 phosphoproteins was significantly altered in CAD patients, with 18 phosphoproteins showing altered expression in AVS patients. Inflammatory pathways were implicated in the changes in phosphoprotein expression in both groups. Inter­disease comparison for the same ventricular chamber at both timepoints revealed differences relating to inflammation and adrenergic and calcium signalling. In conclusion, the present study found that ischemic arrest and reperfusion trigger different changes in the proteomes and phosphoproteomes of LV and RV of CAD and AVS patients undergoing surgery, with markedly more changes in CAD patients despite a significantly shorter ischaemic period.