Gut Microbiota-Generated Phenylacetylglutamine and Heart Failure.

BACKGROUND: The gut microbiota-dependent metabolite phenylacetylgutamine (PAGln) is both associated with atherothrombotic heart disease in humans, and mechanistically linked to cardiovascular disease pathogenesis in animal models via modulation of adrenergic receptor signaling. METHODS: Here we examined both clinical and mechanistic relationships between PAGln and heart failure (HF). First, we examined associations among plasma levels of PAGln and HF, left ventricular ejection fraction, and N-terminal pro-B-type natriuretic peptide in 2 independent clinical cohorts of subjects undergoing coronary angiography in tertiary referral centers (an initial discovery US Cohort, n=3256; and a validation European Cohort, n=829). Then, the impact of PAGln on cardiovascular phenotypes relevant to HF in cultured cardiomyoblasts, and in vivo were also examined. RESULTS: Circulating PAGln levels were dose-dependently associated with HF presence and indices of severity (reduced ventricular ejection fraction, elevated N-terminal pro-B-type natriuretic peptide) independent of traditional risk factors and renal function in both cohorts. Beyond these clinical associations, mechanistic studies showed both PAGln and its murine counterpart, phenylacetylglycine, directly fostered HF-relevant phenotypes, including decreased cardiomyocyte sarcomere contraction, and B-type natriuretic peptide gene expression in both cultured cardiomyoblasts and murine atrial tissue. CONCLUSIONS: The present study reveals the gut microbial metabolite PAGln is clinically and mechanistically linked to HF presence and severity. Modulating the gut microbiome, in general, and PAGln production, in particular, may represent a potential therapeutic target for modulating HF. REGISTRATION: URL: https://clinicaltrials.gov/; Unique identifier: NCT00590200 and URL: https://drks.de/drks_web/; Unique identifier: DRKS00020915.

Transcriptomics-based network medicine approach identifies metformin as a repurposable drug for atrial fibrillation.

Effective drugs for atrial fibrillation (AF) are lacking, resulting in significant morbidity and mortality. This study demonstrates that network proximity analysis of differentially expressed genes from atrial tissue to drug targets can help prioritize repurposed drugs for AF. Using enrichment analysis of drug-gene signatures and functional testing in human inducible pluripotent stem cell (iPSC)-derived atrial-like cardiomyocytes, we identify metformin as a top repurposed drug candidate for AF. Using the active compactor, a new design analysis of large-scale longitudinal electronic health record (EHR) data, we determine that metformin use is significantly associated with a reduced risk of AF (odds ratio = 0.48, 95%, confidence interval [CI] 0.36-0.64, p < 0.001) compared with standard treatments for diabetes. This study utilizes network medicine methodologies to identify repurposed drugs for AF treatment and identifies metformin as a candidate drug.

Atrial fibrillation rhythm is associated with marked changes in metabolic and myofibrillar protein expression in left atrial appendage.

Atrial fibrillation (AF) is strongly associated with risk of stroke and heart failure. AF promotes atrial remodeling that increases risk of stroke due to left atrial thrombogenesis, and increases energy demand to support high rate electrical activity and muscle contraction. While many transcriptomic studies have assessed AF-related changes in mRNA abundance, fewer studies have assessed proteomic changes. We performed a proteomic analysis on left atrial appendage (LAA) tissues from 12 patients with a history of AF undergoing elective surgery; atrial rhythm was documented at time of surgery. Proteomic analysis was performed using liquid chromatography with mass spectrometry (LC/MS-MS). Data-dependent analysis identified 3090 unique proteins, with 408 differentially expressed between sinus rhythm and AF. Ingenuity Pathway Analysis of differentially expressed proteins identified mitochondrial dysfunction, oxidative phosphorylation, and sirtuin signaling among the most affected pathways. Increased abundance of electron transport chain (ETC) proteins in AF was accompanied by decreased expression of ETC complex assembly factors, tricarboxylic acid cycle proteins, and other key metabolic modulators. Discordant changes were also evident in the contractile unit with both up and downregulation of key components. Similar pathways were affected in a comparison of patients with a history of persistent vs. paroxysmal AF, presenting for surgery in sinus rhythm. Together, these data suggest that while the LAA attempts to meet the energetic demands of AF, an uncoordinated response may reduce ATP availability, contribute to tissue contractile and electrophysiologic heterogeneity, and promote a progression of AF from paroxysmal episodes to development of a substrate amenable to persistent arrhythmia.

Genetic Control of Left Atrial Gene Expression Yields Insights into the Genetic Susceptibility for Atrial Fibrillation.

BACKGROUND: Genome-wide association studies have identified 23 loci for atrial fibrillation (AF), but the mechanisms responsible for these associations, as well as the causal genes and genetic variants, remain undefined. METHODS: To identify the effect of common genetic variants on gene expression that might explain the mechanisms linking genome-wide association loci with AF risk, we performed RNA sequencing of left atrial appendages from a biracial cohort of 265 subjects. RESULTS: Combining gene expression data with genome-wide single nucleotide polymorphism data, we found that approximately two-thirds of the expressed genes were regulated in cis by common genetic variants at a false discovery rate of <0.05, defined as cis-expression quantitative trait loci. Twelve of 23 reported AF genome-wide association loci displayed genome-wide significant cis-expression quantitative trait loci, at PRRX1 (chromosome 1q24), SNRNP27 (1q24), CEP68 (2p14), FKBP7 (2q31), KCNN2 (5q22), FAM13B (5q31), CAV1 (7q31), ASAH1 (8p22), MYOZ1 (10q22), C11ORF45 (11q24), TBX5 (12q24), and SYNE2 (14q23), suggesting that altered expression of these genes plays a role in AF susceptibility. Allelic expression imbalance was used as an independent method to characterize the cis-control of gene expression. One thousand two hundred forty-eight of 5153 queried genes had cis-single nucleotide polymorphisms that significantly regulated allelic expression at a false discovery rate of <0.05. CONCLUSIONS: We provide a genome-wide catalog of the genetic control of gene expression in human left atrial appendage. These data can be used to confirm the relevance of genome-wide association loci and to direct future functional studies to identify the genes and genetic variants responsible for complex diseases such as AF.

Rapid multislice T1 mapping of mouse myocardium: Application to quantification of manganese uptake in α-Dystrobrevin knockout mice.

PURPOSE: The aim of this study was to develop a rapid, multislice cardiac T1 mapping method in mice and to apply the method to quantify manganese (Mn(2+)) uptake in a mouse model with altered Ca(2+) channel activity. METHODS: An electrocardiography-triggered multislice saturation-recovery Look-Locker method was developed and validated both in vitro and in vivo. A two-dose study was performed to investigate the kinetics of T1 shortening, Mn(2+) relaxivity in myocardium, and the impact of Mn(2+) on cardiac function. The sensitivity of Mn(2+)-enhanced MRI in detecting subtle changes in altered Ca(2+) channel activity was evaluated in a mouse model with α-dystrobrevin knockout. RESULTS: Validation studies showed strong agreement between the current method and an established method. High Mn(2+) dose led to significantly accelerated T1 shortening. Heart rate decreased during Mn(2+) infusion, while ejection ratio increased slightly at the end of imaging protocol. No statistical difference in cardiac function was detected between the two dose groups. Mice with α-dystrobrevin knockout showed enhanced Mn(2+) uptake in vivo. In vitro patch-clamp study showed increased Ca(2+) channel activity. CONCLUSION: The saturation recovery method provides rapid T1 mapping in mouse hearts, which allowed sensitive detection of subtle changes in Mn(2+) uptake in α-dystrobrevin knockout mice.

Left atrial transcriptional changes associated with atrial fibrillation susceptibility and persistence.

BACKGROUND: Prior transcriptional studies of atrial fibrillation (AF) have been limited to specific transcripts, animal models, chronic AF, right atria, or small samples. We sought to characterize the left atrial transcriptome in human AF to distinguish changes related to AF susceptibility and persistence. METHODS AND RESULTS: Left atrial appendages from 239 patients stratified by coronary artery disease, valve disease, and AF history (no history of AF, AF history in sinus rhythm at surgery, and AF history in AF at surgery) were selected for genome-wide mRNA microarray profiling. Transcripts were examined for differential expression with AF phenotype group. Enrichment in differentially expressed genes was examined in 3 gene set collections: a transcription factor collection, defined by shared conserved cis-regulatory motifs, a miRNA collection, defined by shared 3' untranslated region motifs, and a molecular function collection, defined by shared Gene Ontology molecular function. AF susceptibility was associated with decreased expression of the targets of CREB/ATF family, heat-shock factor 1, ATF6, SRF, and E2F1 transcription factors. Persistent AF activity was associated with decreased expression in genes and gene sets related to ion channel function consistent with reported functional changes. CONCLUSIONS: AF susceptibility was associated with decreased expression of targets of several transcription factors related to inflammation, oxidation, and cellular stress responses. In contrast, changes in ion channel expression were associated with AF activity but were limited in AF susceptibility. Our results suggest that significant transcriptional remodeling marks susceptibility to AF, whereas remodeling of ion channel expression occurs later in the progression or as a consequence of AF.

Weighted gene coexpression network analysis of human left atrial tissue identifies gene modules associated with atrial fibrillation.

BACKGROUND: Genetic mechanisms of atrial fibrillation (AF) remain incompletely understood. Previous differential expression studies in AF were limited by small sample size and provided limited understanding of global gene networks, prompting the need for larger-scale, network-based analyses. METHODS AND RESULTS: Left atrial tissues from Cleveland Clinic patients who underwent cardiac surgery were assayed using Illumina Human HT-12 mRNA microarrays. The data set included 3 groups based on cardiovascular comorbidities: mitral valve (MV) disease without coronary artery disease (n=64), coronary artery disease without MV disease (n=57), and lone AF (n=35). Weighted gene coexpression network analysis was performed in the MV group to detect modules of correlated genes. Module preservation was assessed in the other 2 groups. Module eigengenes were regressed on AF severity or atrial rhythm at surgery. Modules whose eigengenes correlated with either AF phenotype were analyzed for gene content. A total of 14 modules were detected in the MV group; all were preserved in the other 2 groups. One module (124 genes) was associated with AF severity and atrial rhythm across all groups. Its top hub gene, RCAN1, is implicated in calcineurin-dependent signaling and cardiac hypertrophy. Another module (679 genes) was associated with atrial rhythm in the MV and coronary artery disease groups. It was enriched with cell signaling genes and contained cardiovascular developmental genes including TBX5. CONCLUSIONS: Our network-based approach found 2 modules strongly associated with AF. Further analysis of these modules may yield insight into AF pathogenesis by providing novel targets for functional studies.

Perioperative heart-type fatty acid binding protein levels in atrial fibrillation after cardiac surgery.

BACKGROUND: Postoperative atrial fibrillation (POAF) is common and associated with poor outcomes. Perioperative ischemia can alter arrhythmic substrate. OBJECTIVE: To demonstrate an association between perioperative measurements of heart-type fatty acid binding protein (HT-FABP), a sensitive marker of ischemic myocardial injury. METHODS: Blood samples from 63 inpatients undergoing coronary artery bypass surgery, valve surgery, or both were obtained before and up to 4 days after surgery. Continuous telemetry monitoring was used to detect POAF. Fifty-nine patients had at least 3 HT-FABP measurements. The relationship of enzyme-linked immunosorbent assay-measured HT-FABP with POAF was assessed by using joint logistic regression adjusted for age and surgery type. RESULTS: Thirty-five patients (55%) developed POAF; these were, on average, older (69.3±10 years vs 60±11 years; P = .0019), with a higher prevalence of heart failure (43% vs 17%; P = .034), chronic obstructive lung disease (26% vs 4%; P = .017), preoperative calcium channel blocker use (29% vs 7%; P = .031), and more likely to undergo combined surgery (21% vs 11%, P = .049). The joint age- and coronary artery bypass surgery-adjusted model revealed that postoperative but not preoperative HT-FABP levels predicted POAF (coefficient 1.9±0.87; P = .03). Longer bypass time, prior infarction, and worse renal function were all associated with higher postoperative HT-FABP. CONCLUSIONS: A greater rise of HT-FABP is associated with atrial fibrillation after cardiac surgery, suggesting that ischemic myocardial damage is a contributing underlying mechanism. Interventions that decrease perioperative ischemic injury may also decrease the occurrence of POAF.

Dietary ω3 fatty acids modulate the substrate for post-operative atrial fibrillation in a canine cardiac surgery model.

AIMS: Pre-treatment with dietary ω3 polyunsaturated fatty acids (ω3-PUFA) has been reported to reduce the incidence of new-onset atrial fibrillation (AF) following cardiac surgery. In a canine cardiac surgery model, we evaluated the impact of dietary ω3-PUFA on atrial electrophysiological properties, inflammatory markers, the atrial endothelin-1 (ET-1) system, and the expression and distribution of connexin 43. METHODS AND RESULTS: Adult mongrel dogs received either normal chow (NC, n = 11) or chow supplemented with fish oil (FO, 0.6 g ω3-PUFA/kg/day, n = 9) for 3 weeks before surgery. A left thoracotomy was performed, and the left atrial appendage (LAA) was excised. Atrial pacing/recording wires were placed, and the pericardium/chest was closed. The atrial ratio of ω6/ω3 lipids decreased from 15-20 in NC to 2-3 in FO. FO treatment lowered pre-surgical and stabilized post-surgical arachidonate levels. Peak neutrophil to lymphocyte ratio was lower and decayed faster in FO-treated animals. Extensive inflammatory cell infiltration was present in NC atria, but was reduced in FO-treated dogs. FO-treated animals had lower post-surgical atrial expression of inducible nitric oxide synthase (iNOS) and reduced plasma ET-1. Expression of ET-1 and inositol trisphosphate receptor type-2 proteins in the LAA was also reduced. FO treatment prolonged post-operative atrial effective refractory period, slowed heart rate, and enhanced heart rate variability. Importantly, AF (>30 s) was inducible in four of six NC dogs, but no FO dogs. CONCLUSION: Dietary FO attenuated AF inducibility following cardiac surgery by modulating autonomic tone and heart rate. FO also reduced atrial inflammation, iNOS, and ET-1 expression.

Kv1.5 is an important component of repolarizing K+ current in canine atrial myocytes.

Although the canine atrium has proven useful in several experimental models of atrial fibrillation and for studying the effects of rapid atrial pacing on atrial electrical remodeling, it may not fully represent the human condition because of reported differences in functional ionic currents and ion channel subunit expression. In this study, we reassessed the molecular components underlying one current, the ultrarapid delayed rectifier current in canine atrium [IKur(d)], by evaluating the mRNA, protein, immunofluorescence, and currents of the candidate channels. Using reverse transcriptase-polymerase chain reaction, we found that Kv1.5 mRNA was expressed in canine atrium whereas message for Kv3.1 was not detected. Western analysis on cytosolic and membrane fractions of canine tissues, using selective antibodies, showed that Kv3.1 was only detectable in the brain preparations, whereas Kv1.5 was expressed at high levels in both atrial and ventricular membrane fractions. Confocal imaging performed on isolated canine atrial myocytes clearly demonstrated the presence of Kv1.5 immunostaining, whereas that of Kv3.1 was equivocal. Voltage- and current-clamp studies showed that 0.5 mmol/L tetraethylammonium had variable effects on sustained K+ currents, whereas a compound with demonstrated selectivity for hKv1.5 versus Kv3.1, hERG or the sodium channel, fully suppressed canine atrial IKur tail currents and depressed sustained outward K+ current. This agent also increased action potential plateau potentials and action potential duration at 20% and 50% repolarization. These results suggest that in canine atria, as in other species including human, Kv1.5 protein is highly expressed and contributes to IKur.