An extensive ß1-adrenergic receptor gene signaling network regulates molecular remodeling in dilated cardiomyopathies.

We investigated the extent, biologic characterization, phenotypic specificity, and possible regulation of a ß1-adrenergic receptor-linked (ß1-AR-linked) gene signaling network (ß1-GSN) involved in left ventricular (LV) eccentric pathologic remodeling. A 430-member ß1-GSN was identified by mRNA expression in transgenic mice overexpressing human ß1-ARs or from literature curation, which exhibited opposite directional behavior in interventricular septum endomyocardial biopsies taken from patients with beta-blocker-treated, reverse remodeled dilated cardiomyopathies. With reverse remodeling, the major biologic categories and percentage of the dominant directional change were as follows: metabolic (19.3%, 81% upregulated); gene regulation (14.9%, 78% upregulated); extracellular matrix/fibrosis (9.1%, 92% downregulated); and cell homeostasis (13.3%, 60% upregulated). Regarding the comparison of ß1-GSN categories with expression from 19,243 nonnetwork genes, phenotypic selection for major ß1-GSN categories was exhibited for LV end systolic volume (contractility measure), ejection fraction (remodeling index), and pulmonary wedge pressure (wall tension surrogate), beginning at 3 months and persisting to study completion at 12 months. In addition, 121 lncRNAs were identified as possibly involved in cis-acting regulation of ß1-GSN members. We conclude that an extensive 430-member gene network downstream from the ß1-AR is involved in pathologic ventricular remodeling, with metabolic genes as the most prevalent category.

Impact of Neuroeffector Adrenergic Receptor Polymorphisms on Incident Ventricular Fibrillation During Acute Myocardial Ischemia.

Background Cardiac adrenergic receptor gene polymorphisms have the potential to influence risk of developing ventricular fibrillation (VF) during ST-segment-elevation myocardial infarction, but no previous study has comprehensively investigated those most likely to alter norepinephrine release, signal transduction, or biased signaling. Methods and Results In a case-control study, we recruited 953 patients with ST-segment-elevation myocardial infarction without previous cardiac history, 477 with primary VF, and 476 controls without VF, and genotyped them for ADRB1 Arg389Gly and Ser49Gly, ADRB2 Gln27Glu and Gly16Arg, and ADRA2C Ins322-325Del. Within each minor allele-containing genotype, haplotype, or 2-genotype combination, patients with incident VF were compared with non-VF controls by odds ratios (OR) of variant frequencies referenced against major allele homozygotes. Of 156 investigated genetic constructs, 19 (12.2%) exhibited significantly (P<0.05) reduced association with incident VF, and none was associated with increased VF risk except for ADRB1 Gly389 homozygotes in the subset of patients not receiving ß-blockers. ADRB1 Gly49 carriers (prevalence 23.0%) had an OR (95% CI) of 0.70 (0.49-0.98), and the ADRA2C 322-325 deletion (Del) carriers (prevalence 13.5%) had an OR of 0.61 (0.39-0.94). When present in genotype combinations (8 each), both ADRB1 Gly49 carriers (OR, 0.67 [0.56-0.80]) and ADRA2C Del carriers (OR, 0.57 [0.45- 0.71]) were associated with reduced VF risk. Conclusions In ST-segment-elevation myocardial infarction, the adrenergic receptor minor alleles ADRB1 Gly49, whose encoded receptor undergoes enhanced agonist-mediated internalization and ß-arrestin interactions leading to cardioprotective biased signaling, and ADRA2C Del322-325, whose receptor causes disinhibition of norepinephrine release, are associated with a lower incidence of VF. Registration URL: https://clinicaltrials.gov; Unique identifier: NCT00859300.

Myocardial Injury and Altered Gene Expression Associated With SARS-CoV-2 Infection or mRNA Vaccination.

SARS CoV-2 enters host cells via its Spike protein moiety binding to the essential cardiac enzyme angiotensin-converting enzyme (ACE) 2, followed by internalization. COVID-19 mRNA vaccines are RNA sequences that are translated into Spike protein, which follows the same ACE2-binding route as the intact virion. In model systems, isolated Spike protein can produce cell damage and altered gene expression, and myocardial injury or myocarditis can occur during COVID-19 or after mRNA vaccination. We investigated 7 COVID-19 and 6 post-mRNA vaccination patients with myocardial injury and found nearly identical alterations in gene expression that would predispose to inflammation, coagulopathy, and myocardial dysfunction.

Dynamic Regulation of SARS-Cov-2 Binding and Cell Entry Mechanisms in Remodeled Human Ventricular Myocardium.

Using serial analysis of myocardial gene expression employing endomyocardial biopsy starting material in a dilated cardiomyopathy cohort, we show that mRNA expression of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) cardiac myocyte receptor ACE2 is up-regulated with remodeling and with reverse remodeling down-regulates into the normal range. The proteases responsible for virus-cell membrane fusion were expressed but not regulated with remodeling. In addition, a new candidate for SARS-CoV-2 cell binding and entry was identified, the integrin encoded by ITGA5. Up-regulation in ACE2 in remodeled left ventricles may explain worse outcomes in patients with coronavirus disease 2019 who have underlying myocardial disorders, and counteracting ACE2 up-regulation is a possible therapeutic approach to minimizing cardiac damage.

Correction: Sequential analysis of myocardial gene expression with phenotypic change: Use of cross-platform concordance to strengthen biologic relevance.

[This corrects the article DOI: 10.1371/journal.pone.0221519.].

Sequential analysis of myocardial gene expression with phenotypic change: Use of cross-platform concordance to strengthen biologic relevance.

OBJECTIVES: To investigate the biologic relevance of cross-platform concordant changes in gene expression in intact human failing/hypertrophied ventricular myocardium undergoing reverse remodeling. BACKGROUND: Information is lacking on genes and networks involved in remodeled human LVs, and in the associated investigative best practices. METHODS: We measured mRNA expression in ventricular septal endomyocardial biopsies from 47 idiopathic dilated cardiomyopathy patients, at baseline and after 3-12 months of ß-blocker treatment to effect left ventricular (LV) reverse remodeling as measured by ejection fraction (LVEF). Cross-platform gene expression change concordance was investigated in reverse remodeling Responders (R) and Nonresponders (NR) using 3 platforms (RT-qPCR, microarray, and RNA-Seq) and two cohorts (All 47 subjects (A-S) and a 12 patient "Super-Responder" (S-R) subset of A-S). RESULTS: For 50 prespecified candidate genes, in A-S mRNA expression 2 platform concordance (CcpT), but not single platform change, was directly related to reverse remodeling, indicating CcpT has biologic significance. Candidate genes yielded a CcpT (PCR/microarray) of 62% for Responder vs. Nonresponder (R/NR) change from baseline analysis in A-S, and ranged from 38% to 100% in S-R for PCR/microarray/RNA-Seq 2 platform comparisons. Global gene CcpT measured by microarray/RNA-Seq was less than for candidate genes, in S-R R/NR 17.5% vs. 38% (P = 0.036). For S-R global gene expression changes, both cross-cohort concordance (CccT) and CcpT yielded markedly greater values for an R/NR vs. an R-only analysis (by 22 fold for CccT and 7 fold for CcpT). Pathway analysis of concordant global changes for R/NR in S-R revealed signals for downregulation of multiple phosphoinositide canonical pathways, plus expected evidence of a ß1-adrenergic receptor gene network including enhanced Ca2+ signaling. CONCLUSIONS: Two-platform concordant change in candidate gene expression is associated with LV biologic effects, and global expression concordant changes are best identified in an R/NR design that can yield novel information.

Serial gene expression profiling in the intact human heart.

BACKGROUND: In chronic heart failure due to a dilated cardiomyopathy phenotype, the molecular bases for contractile dysfunction and chamber remodeling remain largely unidentified. METHODS: To investigate the feasibility of measuring global gene expression serially in the intact failing human heart, we performed repeated messenger RNA (mRNA) expression profiling using RNA extracted from endomyocardial biopsy specimens and gene chip methodology in 8 subjects with idiopathic dilated cardiomyopathy. In patients treated with beta-blocking agents or placebo, myocardial gene expression was measured in endomyocardial biopsy material and radionuclide ejection fraction was measured at baseline and after 4 to 12 months of treatment. Gene expression was measured for 12,625 gene sequences by using Affymetrix U95 gene chips and commercially available software. For 6 mRNAs, gene chip results were compared with measurements made by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: In an unfiltered composite analysis of changes in expression detected in the patients with high-signal intensity chips, 241 genes showed an increase and 331 genes a decrease in mRNA abundance. There was good agreement between changes measured by quantitative RT-PCR and those determined by gene chips. There was less variance between differences in phenotype in patients sampled serially as compared between subjects with similar phenotypes sampled at baseline. CONCLUSIONS: Serial gene expression profiling with association to phenotypic change is feasible in the intact human heart and may offer advantages to cross-sectional expression profiling. This study suggests that the intact failing remodeled human heart is in an activated state of gene expression, with a large net reduction in gene expression occurring as phenotypic improvement occurs.

Altered expression of endothelin receptors in failing human left ventricles.

BACKGROUND: Endothelin signaling is activated in failing human hearts, and may contribute to progressive myocardial dysfunction and remodeling. However, the behavior of endothelin receptor systems (ET(A) and ET(B)) in failing human hearts is not well understood. METHODS AND RESULTS: (125)[I]-endothelin-1 binding assays conducted in the presence of a non-hydrolyzable guanine nucleotide to uncouple agonist binding demonstrated that membranes prepared from nonfailing left ventricles (LVs) exhibit a mixed pattern of ET(A) ( approximately 60%) and ET(B) ( approximately 40%) receptor protein expression. Chronic LV failure from either idiopathic dilated (IDC) or ischemic (ISC) cardiomyopathy was accompanied by a significant (P<0.001) increase in ET(A) receptor density, to approximately 80% of the total population, and a significant (P<0.02) decrease in ET(B) receptor density. Ribonuclease protection assays demonstrated an increase in ET(A) mRNA abundance in IDC and ISC LVs, and a significant (P<0.04) increase in ET(B) mRNA abundance in ISC LVs. Enzyme-linked immunoabsorbent assays demonstrated a significant increase in tissue immunoreactive endothelin-1 concentration in IDC (P=0.01) and in IDC+ISC LVs (P=0.02), but receptor subtype protein or mRNA level was not significantly correlated with tissue ET-1 across all LVs. In situ reverse-transcription polymerase chain reaction in LV sections demonstrated that in both failing and nonfailing LVs the ET(A) gene is expressed in cardiac myocytes, vascular smooth muscle and endothelium; the ET(B) gene is expressed in cardiac myocytes, fibroblasts and endothelium; and the prepro-endothelin-1 gene is expressed in myocytes and interstitial cells. CONCLUSIONS: In chronically failing human LVs, ET(A) receptor density is increased to become the dominant subtype while ET(B) receptor density is decreased. The ET(A), but not the ET(B) density change is accompanied by cognate regulation of mRNA abundance. Both receptor genes and prepro-endothelin-1 are expressed in cardiac myocytes. Finally, based on a lack of correlation with endothelin-1 tissue levels, it is unlikely that the failure-related changes in ET(A) and ET(B) receptor protein and mRNA expression result from homologous regulation by agonist exposure.

Myocardial gene expression in dilated cardiomyopathy treated with beta-blocking agents.

BACKGROUND: Beta-blocker therapy may improve cardiac function in patients with idiopathic dilated cardiomyopathy. We tested the hypothesis that beta-blocker therapy produces favorable functional effects in dilated cardiomyopathy by altering the expression of myocardial genes that regulate contractility and pathologic hypertrophy. METHODS: We randomly assigned 53 patients with idiopathic dilated cardiomyopathy to treatment with a beta-adrenergic-receptor blocking agent (metoprolol or carvedilol) or placebo. The amount of messenger RNA (mRNA) for contractility-regulating genes (those encoding beta1- and beta2-adrenergic receptors, calcium ATPase in the sarcoplasmic reticulum, and alpha- and beta-myosin heavy-chain isoforms) and of genes associated with pathologic hypertrophy (beta-myosin heavy chain and atrial natriuretic peptide) was measured with a quantitative reverse-transcription polymerase chain reaction in total RNA extracted from biopsy specimens of the right ventricular septal endomyocardium. Myocardial levels of beta-adrenergic receptors were also measured. Measurements were conducted at base line and after six months of treatment, and changes in gene expression were compared with changes in the left ventricular ejection fraction as measured by radionuclide ventriculography. RESULTS: Twenty-six of 32 beta-blocker-treated patients (those with complete mRNA measurements) had an improvement in left ventricular ejection fraction of at least 5 ejection-fraction (EF) units (mean [+/-SE] increase, 18.8+/-1.8). As compared with the six beta-blocker-treated patients who did not have a response (mean change, a decrease of 2.5+/-1.8 EF units), those who did have a response had an increase in sarcoplasmic-reticulum calcium ATPase mRNA and alpha-myosin heavy chain mRNA and a decrease in beta-myosin heavy chain mRNA. The change in sarcoplasmic-reticulum calcium ATPase was not present in the patients in the placebo group who had a spontaneous response. There were no differences between those who had a response and those who did not in terms of the change in mRNA or protein expression of beta-adrenergic receptors. CONCLUSIONS: In idiopathic dilated cardiomyopathy, functional improvement related to treatment with beta-blockers is associated with changes in myocardial gene expression.