Optimizing RNA extraction methods for high-throughput transcriptome sequencing of formalin-fixed paraffin-embedded cardiac tissue specimens.

Archived FFPE cardiac tissue specimens are valuable for molecular studies aimed at identifying biomarkers linked to mortality in cardiovascular disease. Establishing a reliable and reproducible RNA extraction method is critical for generating high-quality transcriptome sequences for molecular assays. Here, the efficiency of four RNA extraction methods: Qiagen AllPrep DNA/RNA method (Method QP); Qiagen AllPrep DNA/RNA method with protocol modification on the ethanol wash step after deparaffinization (Method QE); CELLDATA RNA extraction (Method BP) and CELLDATA RNA extraction with protocol modifications on the lysis step (Method BL) was compared on 23 matching FFPE cardiac tissue specimens (n = 92).

The origins of unpredictability in life outcome prediction tasks.

Why are some life outcomes difficult to predict? We investigated this question through in-depth qualitative interviews with 40 families sampled from a multidecade longitudinal study. Our sampling and interviewing process was informed by the earlier efforts of hundreds of researchers to predict life outcomes for participants in this study. The qualitative evidence we uncovered in these interviews combined with a mathematical decomposition of prediction error led us to create a conceptual framework.

Transcriptome studies of congenital heart diseases: identifying current gaps and therapeutic frontiers.

Congenital heart disease (CHD) are genetically complex and comprise a wide range of structural defects that often predispose to - early heart failure, a common cause of neonatal morbidity and mortality. Transcriptome studies of CHD in human pediatric patients indicated a broad spectrum of diverse molecular signatures across various types of CHD. In order to advance research on congenital heart diseases (CHDs), we conducted a detailed review of transcriptome studies on this topic.

Mitochondrial MICOS complex genes, implicated in hypoplastic left heart syndrome, maintain cardiac contractility and actomyosin integrity.

Hypoplastic left heart syndrome (HLHS) is a severe congenital heart disease (CHD) with a likely oligogenic etiology, but our understanding of the genetic complexities and pathogenic mechanisms leading to HLHS is limited. We performed whole genome sequencing (WGS) on 183 HLHS patient-parent trios to identify candidate genes, which were functionally tested in the heart model. Bioinformatic analysis of WGS data from an index family of a HLHS proband born to consanguineous parents prioritized 9 candidate genes with rare, predicted damaging homozygous variants.

Autologous Umbilical Cord Blood-Derived Mononuclear Cell Therapy Promotes Cardiac Proliferation and Adaptation in a Porcine Model of Right Ventricle Pressure Overload.

Congenital heart diseases, including single ventricle circulations, are clinically challenging due to chronic pressure overload and the inability of the myocardium to compensate for lifelong physiological demands. To determine the clinical relevance of autologous umbilical cord blood-derived mononuclear cells (UCB-MNCs) as a therapy to augment cardiac adaptation following surgical management of congenital heart disease, a validated model system of right ventricular pressure overload due to pulmonary artery banding (PAB) in juvenile pigs has been employed. PAB in a juvenile porcine model and intramyocardial delivery of UCB-MNCs was evaluated in three distinct 12-week studies utilizing serial cardiac imaging and end-of-study pathology evaluations.

Uncoupling of Proliferative Capacity from Developmental Stage During Directed Cardiac Differentiation of Pluripotent Stem Cells.

Lineage-specific differentiation of human-induced pluripotent stem cells (hiPSCs) into cardiomyocytes (CMs) offers a patient-specific model to dissect development and disease pathogenesis in a dish. However, challenges exist with this model system, such as the relative immaturity of iPSC-derived CMs, which evoke the question of whether this model faithfully recapitulates in vivo cardiac development. As in vivo cardiac developmental stage is intimately linked with the proliferative capacity (or maturation is inversely correlated to proliferative capacity), we sought to understand how proliferation is regulated during hiPSC CM differentiation and how it compares with in vivo mouse cardiac development.

Shotgun Immunoproteomics for Identification of Nonhuman Leukocyte Antigens Associated With Cellular Dysfunction in Heart Transplant Rejection.

Background: The International Society for Heart and Lung Transplant consensus panel notes that too little data exist regarding the role of non-HLA in allograft rejection. We developed a novel shotgun immunoproteomic approach to determine the identities and potential roles non-HLA play in antibody-mediated rejection (AMR) in heart transplant recipients.

Methods: Serum was collected longitudinally from heart transplant recipients experiencing AMR in the absence of donor-specific anti-HLA antibodies (n = 6) and matched no rejection controls (n = 7).

Activation of P53 Via Nutlin-3a Reveals Role for P53 In ROS Signaling During Cardiac Differentiation of hiPSCs.

Activation of the transcription factor P53 within cancer cells is a well-characterized pathway, whereas the effects of P53 activation during development remain largely unexplored. Previous research has indicated that increased levels of P53 protein during key murine developmental stages cause defects in multiple embryonic tissues, including the heart. These findings were confirmed in several different mouse models of congenital heart defects, but P53 activation in a human system of cardiovascular development is not available.

Cone Reconstruction for Ebstein Anomaly: Ventricular Remodeling and Preliminary Impact of Stem Cell Therapy.

Objective: To define the impact of tricuspid valve cone reconstruction (CR) on ventricular performance in Ebstein anomaly, both independently and after stem cell therapy.

Patients And Methods: The control group included 257 patients who had CR between June 2007 and December 2019. Ten subjects of a phase I stem cell therapy trial (May 2017 - March 2019) were compared with the controls to assess the echocardiographic impact on ventricular remodeling.

Does Time with Dad in Childhood Pay Off in Adolescence?

Objective: We aim to understand the association between father involvement in middle childhood and adolescent behaviors and whether the relationship differs by father residence.

Background: Internalizing and externalizing behaviors in adolescence can trigger a cascade of negative outcomes later in life, including lower educational attainment, criminal justice involvement, and future psychological distress. Evidence, largely focusing on nonresidential fathers and older cohort, suggests that father involvement-particularly closeness and engagement-may reduce adolescents' internalizing and externalizing behaviors.

Point absorbers in Advanced LIGO.

Small, highly absorbing points are randomly present on the surfaces of the main interferometer optics in Advanced LIGO. The resulting nanometer scale thermo-elastic deformations and substrate lenses from these micron-scale absorbers significantly reduce the sensitivity of the interferometer directly though a reduction in the power-recycling gain and indirect interactions with the feedback control system. We review the expected surface deformation from point absorbers and provide a pedagogical description of the impact on power buildup in second generation gravitational wave detectors (dual-recycled Fabry-Perot Michelson interferometers).

Identifying gaps in parental support for families of children with hypoplastic left heart syndrome.

Purpose: The purpose of this study is to identify gaps in support for parents of children with Hypoplastic Left Heart Syndrome.

Design And Methods: Using a mixed-methods approach, the researchers first studied the parental and care team experience through interviews of Hypoplastic Left Heart Syndrome mothers and members of the inter-professional care team and then conducted an international survey of 690 Hypoplastic Left Heart Syndrome primary caregivers to validate the qualitative findings.

Results: Parental and care team interviews revealed three main gaps in parental support, including lack of open communication, unrealistic parental expectations, and unclear inter-professional team roles.

From Safety to Benefit in Cell Delivery During Surgical Repair of Ebstein Anomaly: Initial Results.

Background: The objective of this study is to assess the safety and early impact of intramyocardial delivery of autologous bone marrow-derived mononuclear cells (BM-MNC) at time of surgical Ebstein repair.

Methods: Patients with Ebstein anomaly (ages 6 months to 30 years) scheduled to undergo repair of the tricuspid valve were eligible to participate in this open-label, non-randomized phase I clinical trial. BM-MNC target dose was 1-3 million cells/kg.

Dysregulated ribonucleoprotein granules promote cardiomyopathy in RBM20 gene-edited pigs.

Ribonucleoprotein (RNP) granules are biomolecular condensates-liquid-liquid phase-separated droplets that organize and manage messenger RNA metabolism, cell signaling, biopolymer assembly, biochemical reactions and stress granule responses to cellular adversity. Dysregulated RNP granules drive neuromuscular degenerative disease but have not previously been linked to heart failure. By exploring the molecular basis of congenital dilated cardiomyopathy (DCM) in genome-edited pigs homozygous for an RBM20 allele encoding the pathogenic R636S variant of human RNA-binding motif protein-20 (RBM20), we discovered that RNP granules accumulated abnormally in the sarcoplasm, and we confirmed this finding in myocardium and reprogrammed cardiomyocytes from patients with DCM carrying the R636S allele.

Clinical Impact of Autologous Cell Therapy on Hypoplastic Left Heart Syndrome After Bidirectional Cavopulmonary Anastomosis.

Preservation of right ventricle function (RV) is a key to favorable outcome in Hypoplastic Left Heart Syndrome (HLHS), but methods to preserve or improve RV function are limited. Our goal was to assess the clinical and functional impact of autologous umbilical cord blood-derived mononuclear cells (UCB-MNC) therapy when given to patients with HLHS at Stage II surgery. UCB-MNC patients were enrolled prospectively in a phase I, FDA monitored trial as previously described (Burkhart et al.

Patient-specific genomics and cross-species functional analysis implicate LRP2 in hypoplastic left heart syndrome.

Congenital heart diseases (CHDs), including hypoplastic left heart syndrome (HLHS), are genetically complex and poorly understood. Here, a multidisciplinary platform was established to functionally evaluate novel CHD gene candidates, based on whole-genome and iPSC RNA sequencing of a HLHS family-trio. Filtering for rare variants and altered expression in proband iPSCs prioritized 10 candidates.

Intrinsic Endocardial Defects Contribute to Hypoplastic Left Heart Syndrome.

Hypoplastic left heart syndrome (HLHS) is a complex congenital heart disease characterized by abnormalities in the left ventricle, associated valves, and ascending aorta. Studies have shown intrinsic myocardial defects but do not sufficiently explain developmental defects in the endocardial-derived cardiac valve, septum, and vasculature. Here, we identify a developmentally impaired endocardial population in HLHS through single-cell RNA profiling of hiPSC-derived endocardium and human fetal heart tissue with an underdeveloped left ventricle.

Loss of Ventricular Function After Bidirectional Cavopulmonary Connection: Who Is at Risk?

Decline of single ventricle systolic function after bidirectional cavopulmonary connection (BDCPC) is thought to be a transient phenomenon. We analyzed ventricular function after BDCPC according to ventricular morphology and correlated this evolution to long-term prognosis. A review from Mayo Clinic databases was performed.

Mitochondrial DNA: Hotspot for Potential Gene Modifiers Regulating Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is a prevalent and untreatable cardiovascular disease with a highly complex clinical and genetic causation. HCM patients bearing similar sarcomeric mutations display variable clinical outcomes, implying the involvement of gene modifiers that regulate disease progression. As individuals exhibiting mutations in mitochondrial DNA (mtDNA) present cardiac phenotypes, the mitochondrial genome is a promising candidate to harbor gene modifiers of HCM.

m.3243A > G-Induced Mitochondrial Dysfunction Impairs Human Neuronal Development and Reduces Neuronal Network Activity and Synchronicity.

Epilepsy, intellectual and cortical sensory deficits, and psychiatric manifestations are the most frequent manifestations of mitochondrial diseases. How mitochondrial dysfunction affects neural structure and function remains elusive, mostly because of a lack of proper in vitro neuronal model systems with mitochondrial dysfunction. Leveraging induced pluripotent stem cell technology, we differentiated excitatory cortical neurons (iNeurons) with normal (low heteroplasmy) and impaired (high heteroplasmy) mitochondrial function on an isogenic nuclear DNA background from patients with the common pathogenic m.