General practitioners' perceptions on opportunistic single-time point screening for atrial fibrillation: A European quantitative survey.

Background: There is no clear guidance on how to implement opportunistic atrial fibrillation (AF) screening in daily clinical practice. Objectives: This study evaluated the perception of general practitioners (GPs) about value and practicalities of implementing screening for AF, focusing on opportunistic single-time point screening with a single-lead electrocardiogram (ECG) device. Methods: A descriptive cross-sectional study was conducted with a survey developed to assess overall perception concerning AF screening, feasibility of opportunistic single-lead ECG screening and implementation requirements and barriers. Results: A total of 659 responses were collected (36.1% Eastern, 33.4% Western, 12.1% Southern, 10.0% Northern Europe, 8.3% United Kingdom & Ireland). The perceived need for standardized AF screening was rated as 82.7 on a scale from 0 to 100. The vast majority (88.0%) indicated no AF screening program is established in their region. Three out of four GPs (72.1%, lowest in Eastern and Southern Europe) were equipped with a 12-lead ECG, while a single-lead ECG was less common (10.8%, highest in United Kingdom & Ireland). Three in five GPs (59.3%) feel confident ruling out AF on a single-lead ECG strip. Assistance through more education (28.7%) and a tele-healthcare service offering advice on ambiguous tracings (25.2%) would be helpful. Preferred strategies to overcome barriers like insufficient (qualified) staff, included integrating AF screening with other healthcare programs (24.9%) and algorithms to identify patients most suitable for AF screening (24.3%). Conclusion: GPs perceive a strong need for a standardized AF screening approach. Additional resources may be required to have it widely adopted into clinical practice.

Performance of an electronic health record-based predictive model to identify patients with atrial fibrillation across countries.

BACKGROUND: Atrial fibrillation (AF) burden on patients and healthcare systems warrants innovative strategies for screening asymptomatic individuals. OBJECTIVE: We sought to externally validate a predictive model originally developed in a German population to detect unidentified incident AF utilising real-world primary healthcare databases from countries in Europe and Australia. METHODS: This retrospective cohort study used anonymized, longitudinal patient data from 5 country-level primary care databases, including Australia, Belgium, France, Germany, and the UK. The study eligibility included adult patients (≥45 years) with either an AF diagnosis (cases) or no diagnosis (controls) who had continuous enrolment in the respective database prior to the study period. Logistic regression was fitted to a binary response (yes/no) for AF diagnosis using pre-determined risk factors. RESULTS: AF patients were from Germany (n = 63,562), the UK (n = 42,652), France (n = 7,213), Australia (n = 2,753), and Belgium (n = 1,371). Cases were more likely to have hypertension or other cardiac conditions than controls in all validation datasets compared to the model development data. The area under the receiver operating characteristic (ROC) curve in the validation datasets ranged from 0.79 (Belgium) to 0.84 (Germany), comparable to the German study model, which had an area under the curve of 0.83. Most validation sets reported similar specificity at approximately 80% sensitivity, ranging from 67% (France) to 71% (United Kingdom). The positive predictive value (PPV) ranged from 2% (Belgium) to 16% (Germany), and the number needed to be screened was 50 in Belgium and 6 in Germany. The prevalence of AF varied widely between these datasets, which may be related to different coding practices. Low prevalence affected PPV, but not sensitivity, specificity, and ROC curves. CONCLUSIONS: AF risk prediction algorithms offer targeted ways to identify patients using electronic health records, which could improve screening number and the cost-effectiveness of AF screening if implemented in clinical practice.

Preferences for and use of oral anticoagulants for stroke prevention in atrial fibrillation under real-world conditions in Germany: A survey among physicians.

In Germany, there is little real-world evidence on physicians' choice of oral anticoagulants (OACs). Our study aimed at assessing preferences for and prescribing patterns of treatment options for stroke prevention in atrial fibrillation in clinical practice in Germany. We conducted a nationwide quantitative online survey among office-based physicians in Germany. Physicians were asked about their preference for and use of treatment options as well as factors influencing their choice of a specific OAC. A total of n = 953 physicians was surveyed in September and October 2020 (general physicians: 36.0%; internists: 37.3%; cardiologists: 23.7%; neurologists: 10.5%; multiple specialties possible). Preference and use were highest for non-vitamin K oral anticoagulants (NOACs); followed by vitamin K antagonists (VKAs). Most preferred OACs were apixaban (39.3%), rivaroxaban (28.5%) and edoxaban (14.7%). Most used OACs were apixaban (24.3%), rivaroxaban (21.2%) and phenprocoumon (21.4%). NOACs were preferred more often than used (85.6% > 68.6%). VKAs were preferred less often than used (9.6% < 23.5%). OAC attributes and patient characteristics related to efficacy and safety, as well as patients' kidney function were most important when selecting a specific OAC. Federal and regional governance instruments likely influenced treatment decision-making. We found a high divergence between preferences for and use of available treatment options in clinical practice. Further exploration of the importance of OAC attributes, patient characteristics as well as federal and regional governance instruments for physicians' choice of a specific OAC may help to further optimize the healthcare of patients with atrial fibrillation in the long-term.

Feasible approaches and implementation challenges to atrial fibrillation screening: a qualitative study of stakeholder views in 11 European countries.

OBJECTIVES: Atrial fibrillation (AF) screening may increase early detection and reduce complications of AF. European, Australian and World Heart Federation guidelines recommend opportunistic screening, despite a current lack of clear evidence supporting a net benefit for systematic screening. Where screening is implemented, the most appropriate approaches are unknown. We explored the views of European stakeholders about opportunities and challenges of implementing four AF screening scenarios. DESIGN: Telephone-based semi-structured interviews with results reported using Consolidated criteria for Reporting Qualitative research guidelines. Data were thematically analysed using the framework approach. SETTING: AF screening stakeholders in 11 European countries. PARTICIPANTS: Healthcare professionals and regulators (n=24) potentially involved in AF screening implementation. INTERVENTION: Four AF screening scenarios: single time point opportunistic, opportunistic prolonged, systematic single time point/prolonged and patient-led screening. PRIMARY OUTCOME MEASURES: Stakeholder views about the challenges and feasibility of implementing the screening scenarios in the respective national/regional healthcare system. RESULTS: Three themes developed. (1) Current screening approaches: there are no national AF screening programmes, with most AF detected in symptomatic patients. Patient-led screening exists via personal devices, creating screening inequity. (2) Feasibility of screening: single time point opportunistic screening in primary care using single-lead ECG devices was considered the most feasible. Software algorithms may aid identification of suitable patients and telehealth services have potential to support diagnosis. (3) Implementation requirements: sufficient evidence of benefit is required. National screening processes are required due to different payment mechanisms and health service regulations. Concerns about data security, and inclusivity for those without primary care access or personal devices must be addressed. CONCLUSIONS: There is an overall awareness of AF screening. Opportunistic screening appears the most feasible across Europe. Challenges are health inequalities, identification of best target groups for screening, streamlined processes, the need for evidence of benefit and a tailored approach adapted to national realities.

Wearables, telemedicine, and artificial intelligence in arrhythmias and heart failure: Proceedings of the European Society of Cardiology Cardiovascular Round Table.

Digital technology is now an integral part of medicine. Tools for detecting, screening, diagnosis, and monitoring health-related parameters have improved patient care and enabled individuals to identify issues leading to better management of their own health. Wearable technologies have integrated sensors and can measure physical activity, heart rate and rhythm, and glucose and electrolytes. For individuals at risk, wearables or other devices may be useful for early detection of atrial fibrillation or sub-clinical states of cardiovascular disease, disease management of cardiovascular diseases such as hypertension and heart failure, and lifestyle modification. Health data are available from a multitude of sources, namely clinical, laboratory and imaging data, genetic profiles, wearables, implantable devices, patient-generated measurements, and social and environmental data. Artificial intelligence is needed to efficiently extract value from this constantly increasing volume and variety of data and to help in its interpretation. Indeed, it is not the acquisition of digital information, but rather the smart handling and analysis that is challenging. There are multiple stakeholder groups involved in the development and effective implementation of digital tools. While the needs of these groups may vary, they also have many commonalities, including the following: a desire for data privacy and security; the need for understandable, trustworthy, and transparent systems; standardized processes for regulatory and reimbursement assessments; and better ways of rapidly assessing value.

Machine learning-based identification of risk-factor signatures for undiagnosed atrial fibrillation in primary prevention and post-stroke in clinical practice.

AIMS: Atrial fibrillation (AF) carries a substantial risk of ischemic stroke and other complications, and estimates suggest that over a third of cases remain undiagnosed. AF detection is particularly pressing in stroke survivors. To tailor AF screening efforts, we explored German health claims data for routinely available predictors of incident AF in primary care and post-stroke using machine learning methods. METHODS AND RESULTS: We combined AF predictors in patients over 45 years of age using claims data in the InGef database (n = 1 476 391) for (i) incident AF and (ii) AF post-stroke, using machine learning techniques. Between 2013-2016, new-onset AF was diagnosed in 98 958 patients (6.7%). Published risk factors for AF including male sex, hypertension, heart failure, valvular heart disease, and chronic kidney disease were confirmed. Component-wise gradient boosting identified additional predictors for AF from ICD-codes available in ambulatory care. The area under the curve (AUC) of the final, condensed model consisting of 13 predictors, was 0.829 (95% confidence interval (CI) 0.826-0.833) in the internal validation, and 0.755 (95% CI 0.603-0.890) in a prospective validation cohort (n = 661). The AUC for post-stroke AF was of 0.67 (95% CI 0.651-0.689) in the internal validation data set, and 0.766 (95% CI 0.731-0.800) in the prospective clinical cohort. CONCLUSION: ICD-coded clinical variables selected by machine learning can improve the identification of patients at risk of newly diagnosed AF. Using this readily available, automatically coded information can target AF screening efforts to identify high-risk populations in primary care and stroke survivors.

Comprehensive plasma and tissue profiling reveals systemic metabolic alterations in cardiac hypertrophy and failure.

AIMS: Heart failure is characterized by structural and metabolic cardiac remodelling. The aim of the present study is to expand our understanding of the complex metabolic alterations in the transition from pathological hypertrophy to heart failure and exploit the results from a translational perspective. METHODS AND RESULTS: Mice were subjected to transverse aortic constriction (TAC) or sham surgery and sacrificed 2 weeks, 4 weeks, or 6 weeks after the procedure. Samples from plasma, liver, skeletal muscle, and heart were collected and analysed using metabolomics. Cardiac samples were also analysed by transcriptional profiling. Progressive alterations of key cardiac metabolic pathways and gene expression patterns indicated impaired mitochondrial function and a metabolic switch during transition to heart failure. Similar to the heart, liver, and skeletal muscle revealed significant metabolic alterations such as depletion of essential fatty acids and glycerolipids in late stages of heart failure. Circulating metabolites, particularly fatty acids, reflected cardiac metabolic defects, and deteriorating heart function. For example, inverse correlation was found between plasma and the heart levels of triacylglycerol (C18:1, C18:2, C18:3), and sphingomyelin (d18:1, C23:0) already at an early stage of heart failure. Interestingly, combining metabolic and transcriptional data from cardiac tissue revealed that decreased carnitine shuttling and transportation preceded mitochondrial dysfunction. We, thus, studied the therapeutic potential of OCTN2 (Organic Cation/Carnitine Transporter 2), an important factor for carnitine transportation. Cardiac overexpression of OCTN2 using an adeno-associated viral vector significantly improved ejection fraction and reduced interstitial fibrosis in mice subjected to TAC. CONCLUSION: Comprehensive plasma and tissue profiling reveals systemic metabolic alterations in heart failure, which can be used for identification of novel biomarkers and potential therapeutic targets.

Metabolic profiles in heart failure due to non-ischemic cardiomyopathy at rest and under exercise.

AIMS: Identification of metabolic signatures in heart failure (HF) patients and evaluation of their diagnostic potential to discriminate HF patients from healthy controls during baseline and exercise conditions. METHODS: Plasma samples were collected from 22 male HF patients with non-ischemic idiopathic cardiomyopathy and left ventricular systolic dysfunction and 19 healthy controls before (t0), at peak (t1) and 1 h after (t2) symptom-limited cardiopulmonary exercise testing. Two hundred fifty-two metabolites were quantified by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography (LC)-MS/MS-based metabolite profiling. RESULTS: Plasma metabolite profiles clearly differed between HF patients and controls at t0 (P < 0.05). The metabolic signature of HF was characterized by decreased levels of complex lipids and fatty acids, notably phosphatidylcholines, cholesterol, and sphingolipids. Moreover, reduced glutamine and increased glutamate plasma levels, significantly increased purine degradation products, as well as signs of impaired glucose metabolism were observed. The metabolic differences increased strongly according to New York Heart Association functional class and the addition of three metabolites further improved prediction of exercise capacity (Q2 = 0.24 to 0.35). Despite a high number of metabolites changing significantly with exercise (30.2% at t1/t0), the number of significant alterations between HF and controls was almost unchanged at t1 and t2 (30.7 and 29.0% vs. 31.3% at t0) with a similar predictive group separation (Q2 = 0.50 for t0, 0.52 for t1, and 0.56 for t2, respectively). CONCLUSIONS: Our study identified a metabolic signature of non-ischemic HF with prominent changes in complex lipids including phosphatidylcholines, cholesterol, and sphingolipids. The metabolic changes were already evident at rest and largely preserved under exercise.

A Novel Lipid Biomarker Panel for the Detection of Heart Failure with Reduced Ejection Fraction.

OBJECTIVES: In this study we aimed to identify novel metabolomic biomarkers suitable for improved diagnosis of heart failure with reduced ejection fraction (HFrEF). METHODS: We prospectively recruited 887 individuals consisting of HFrEF patients with either ischemic (ICMP, n = 257) or nonischemic cardiomyopathy (NICMP, n = 269), healthy controls (n = 327), and patients with pulmonary diseases (n = 34). A single-center identification (n = 238) was followed by a multicenter confirmation study (n = 649). Plasma samples from the single-center study were subjected to metabolite profiling analysis to identify metabolomic features with potential as HFrEF biomarkers. A dedicated analytical protocol was developed for the routine analysis of selected metabolic features in the multicenter cohort. RESULTS: In the single-center study, 92 of 181 metabolomic features with known chemical identity (51%) were significantly changed in HFrEF patients compared to healthy controls (P <0.05). Three specific metabolomic features belonging to the lipid classes of sphingomyelins, triglycerides, and phosphatidylcholines were selected as the cardiac lipid panel (CLP) and analyzed in the multicenter study using the dedicated analytical protocol. The combination of the CLP with N-terminal pro-B-type natriuretic peptide (NT-proBNP) distinguished HFrEF patients from healthy controls with an area under the curve (AUC) of 0.97 (sensitivity 80.2%, specificity 97.6%) and was significantly superior compared to NT-proBNP alone (AUC = 0.93, sensitivity 81.7%, specificity 88.1%, P <0.001), even in the subgroups with mildly reduced left ventricular EF (0.94 vs 0.87; P <0.001) and asymptomatic patients (0.95 vs 0.91; P <0.05). CONCLUSIONS: The new metabolomic biomarker panel has the potential to improve HFrEF detection, even in mild and asymptomatic stages. The observed changes further indicate lipid alterations in the setting of HFrEF.

Sex- and age-dependent effects of Gpr30 genetic deletion on the metabolic and cardiovascular profiles of diet-induced obese mice.

The G protein-coupled receptor 30 (GPR30) has been claimed as an estrogen receptor. However, the literature reports controversial findings and the physiological function of GPR30 is not fully understood yet. Consistent with studies assigning a role of GPR30 in the cardiovascular and metabolic systems, GPR30 expression has been reported in small arterial vessels, pancreas and chief gastric cells of the stomach. Therefore, we hypothesized a role of GPR30 in the onset and progression of cardiovascular and metabolic diseases. In order to test our hypothesis, we investigated the effects of a high-fat diet on the metabolic and cardiovascular profiles of Gpr30-deficient mice (GPR30-lacZ mice). We found that GPR30-lacZ female, rather than male, mice had significant lower levels of HDL along with an increase in fat liver accumulation as compared to control mice. However, two indicators of cardiac performance assessed by echocardiography, ejection fraction and fractional shortening were both decreased in an age-dependent manner only in Gpr30-lacZ male mice. Collectively our results point to a potential role of Gpr30 in preserving lipid metabolism and cardiac function in a sex- and age-dependent fashion.

A new metabolomic signature in type-2 diabetes mellitus and its pathophysiology.

OBJECTIVE: The objective of the current study was to find a metabolic signature associated with the early manifestations of type-2 diabetes mellitus. RESEARCH DESIGN AND METHOD: Modern metabolic profiling technology (MxP™ Broad Profiling) was applied to find early alterations in the plasma metabolome of type-2 diabetic patients. The results were validated in an independent study. Eicosanoid and single inon monitoring analysis (MxP™ Eicosanoid and MxP™ SIM analysis) were performed in subsets of samples. RESULTS: A metabolic signature including significantly increased levels of glyoxylate as a potential novel marker for early detection of type-2 diabetes mellitus was identified in an initial study (Study1). The signature was significantly altered in fasted diabetic and pre-diabetic subjects and in non-fasted subjects up to three years prior to the diagnosis of type-2 diabetes; most alterations were also consistently found in an independent patient group (Study 2). In Study 2 diabetic and most control subjects suffered from heart failure. In Study 1 a subgroup of diabetic subjects, with a history of use of anti-hypertensive medication further showed a more pronounced increase of glyoxylate levels, compared to a non-diabetic control group when tested in a hyperglycemic state. In the context of a prior history of anti-hypertensive medication, alterations in hexosamine and eicosanoid levels were also found. CONCLUSION: A metabolic signature including glyoxylate was associated with type-2 diabetes mellitus, independent of the fasting status and of occurrence of another major disease. The same signature was also found to be associated with pre-diabetic subjects. Glyoxylate levels further showed a specifically strong increase in a subgroup of diabetic subjects. It could represent a new marker for the detection of medical subgroups of diabetic subjects.

Doxorubicin and NRG-1/erbB4-Deficiency Affect Gene Expression Profile: Involving Protein Homeostasis in Mouse.

The accumulating evidence demonstrates the essential role of neuregulin-1 signaling in the adult heart, and, moreover, indicates that an impaired neuregulin signaling exacerbates the doxorubicin-mediated cardiac toxicity. Despite this strong data, the specific cardiomyocyte targets of the active erbB2/erbB4 heterodimer remain unknown. In this paper, we examined pathways involved in cardiomyocyte damage as a result of the cardiac sensitization to anthracycline toxicity in the ventricular muscle-specific erbB4 knockout mouse. We performed morphological analyses to evaluate the ventricular remodeling and employed a cDNA microarray to assess the characteristic gene expression profile, verified data by real-time RT-PCR, and then grouped into functional categories and pathways. We confirm the upregulation of genes related to the classical signature of a hypertrophic response, implicating an erbB2-dependent mechanism in doxorubicin-treated erbB4-KO hearts. Our results indicate the remarkable downregulation of IGF-I/PI-3' kinase pathway and extends our current knowledge by uncovering an altered ubiquitin-proteasome system leading to cardiomyocyte autophagic vacuolization.

High-mobility group A1 protein: a new coregulator of peroxisome proliferator-activated receptor-γ-mediated transrepression in the vasculature.

RATIONALE: The nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ) is an important regulator of gene transcription in vascular cells and mediates the vascular protection observed with antidiabetic glitazones. OBJECTIVE: To determine the molecular mechanism of ligand-dependent transrepression in vascular smooth muscle cells and their impact on the vascular protective actions of PPARγ. METHODS AND RESULTS: Here, we report a molecular pathway in vascular smooth muscle cells by which ligand-activated PPARγ represses transcriptional activation of the matrix-degrading matrix metalloproteinase-9 (MMP-9) gene, a crucial mediator of vascular injury. PPARγ-mediated transrepression of the MMP-9 gene was dependent on the presence of the high-mobility group A1 (HMGA1) protein, a gene highly expressed in vascular smooth muscle cells, newly identified by oligonucleotide array expression analysis. Transrepression of MMP-9 by PPARγ and regulation by HMGA1 required PPARγ SUMOylation at K367. This process was associated with formation of a complex between PPARγ, HMGA1, and the SUMO E2 ligase Ubc9 (ubiquitin-like protein SUMO-1 conjugating enzyme). After PPARγ ligand stimulation, HMGA1 and PPARγ were recruited to the MMP-9 promoter, which facilitated binding of SMRT (silencing mediator of retinoic acid and thyroid hormone receptor), a nuclear corepressor involved in transrepression. The relevance of HMGA1 for vascular PPARγ signaling was underlined by the complete absence of vascular protection through a PPARγ ligand in HMGA1(-/-) mice after arterial wire injury. CONCLUSIONS: The present data suggest that ligand-dependent formation of HMGA1-Ubc9-PPARγ complexes facilitates PPARγ SUMOylation, which results in the prevention of SMRT corepressor clearance and induction of MMP-9 transrepression. These data provide new information on PPARγ-dependent vascular transcriptional regulation and help us to understand the molecular consequences of therapeutic interventions with PPARγ ligands in the vasculature.

Load-reducing therapy prevents development of arrhythmogenic right ventricular cardiomyopathy in plakoglobin-deficient mice.

OBJECTIVES: We used a murine model of arrhythmogenic right ventricular cardiomyopathy (ARVC) to test whether reducing ventricular load prevents or slows development of this cardiomyopathy. BACKGROUND: At present, no therapy exists to slow progression of ARVC. Genetically conferred dysfunction of the mechanical cell-cell connections, often associated with reduced expression of plakoglobin, is thought to cause ARVC. METHODS: Littermate pairs of heterozygous plakoglobin-deficient mice (plako(+/-)) and wild-type (WT) littermates underwent 7 weeks of endurance training (daily swimming). Mice were randomized to blinded load-reducing therapy (furosemide and nitrates) or placebo. RESULTS: Therapy prevented training-induced right ventricular (RV) enlargement in plako(+/-) mice (RV volume: untreated plako(+/-) 136 ± 5 µl; treated plako(+/-) 78 ± 5 µl; WT 81 ± 5 µl; p < 0.01 for untreated vs. WT and untreated vs. treated; mean ± SEM). In isolated, Langendorff-perfused hearts, ventricular tachycardias (VTs) were more often induced in untreated plako(+/-) hearts (15 of 25), than in treated plako(+/-) hearts (5 of 19) or in WT hearts (6 of 21, both p < 0.05). Epicardial mapping of the RV identified macro-re-entry as the mechanism of ventricular tachycardia. The RV longitudinal conduction velocity was reduced in untreated but not in treated plako(+/-) mice (p < 0.01 for untreated vs. WT and untreated vs. treated). Myocardial concentration of phosphorylated connexin43 was lower in plako(+/-) hearts with VTs compared with hearts without VTs and was reduced in untreated plako(+/-) compared with WT (both p < 0.05). Plako(+/-) hearts showed reduced myocardial plakoglobin concentration, whereas ß-catenin and N-cadherin concentration was not changed. CONCLUSIONS: Load-reducing therapy prevents training-induced development of ARVC in plako(+/-) mice.

Female sex and estrogen receptor-beta attenuate cardiac remodeling and apoptosis in pressure overload.

We investigated sex differences and the role of estrogen receptor-beta (ERbeta) on myocardial hypertrophy in a mouse model of pressure overload. We performed transverse aortic constriction (TAC) or sham surgery in male and female wild-type (WT) and ERbeta knockout (ERbeta(-/-)) mice. All mice were characterized by echocardiography and hemodynamic measurements and were killed 9 wk after surgery. Left ventricular (LV) samples were analyzed by microarray profiling, real-time RT-PCR, and histology. After 9 wk, WT males showed more hypertrophy and heart failure signs than WT females. Notably, WT females developed a concentric form of hypertrophy, while males developed eccentric hypertrophy. ERbeta deletion augmented the TAC-induced increase in cardiomyocyte diameter in both sexes. Gene expression profiling revealed that WT male hearts had a stronger induction of matrix-related genes and a stronger repression of mitochondrial genes than WT female hearts. ERbeta(-/-) mice exhibited a different transcriptional response. ERbeta(-/-)/TAC mice of both sexes exhibited induction of proapoptotic genes with a stronger expression in ERbeta(-/-) males. Cardiac fibrosis was more pronounced in male WT/TAC than in female mice. This difference was abolished in ERbeta(-/-) mice. The number of apoptotic nuclei was increased in both sexes of ERbeta(-/-)/TAC mice, most prominent in males. Female sex offers protection against ventricular chamber dilation in the TAC model. Both female sex and ERbeta attenuate the development of fibrosis and apoptosis, thus slowing the progression to heart failure.

Expression pattern of G protein-coupled receptor 30 in LacZ reporter mice.

Multiple reports implicated the function of G protein-coupled receptor (GPR)-30 with nongenomic effects of estrogen, suggesting that GPR30 might be a G-protein coupled estrogen receptor. However, the findings are controversial and the expression pattern of GPR30 on a cell type level as well as its function in vivo remains unclear. Therefore, the objective of this study was to identify cell types that express Gpr30 in vivo by analyzing a mutant mouse model that harbors a lacZ reporter (Gpr30-lacZ) in the Gpr30 locus leading to a partial deletion of the Gpr30 coding sequence. Using this strategy, we identified the following cell types expressing Gpr30: 1) an endothelial cell subpopulation in small arterial vessels of multiple tissues, 2) smooth muscle cells and pericytes in the brain, 3) gastric chief cells in the stomach, 4) neuronal subpopulations in the cortex as well as the polymorph layer of the dentate gyrus, 5) cell populations in the intermediate and anterior lobe of the pituitary gland, and 6) in the medulla of the adrenal gland. In further experiments, we aimed to decipher the function of Gpr30 by analyzing the phenotype of Gpr30-lacZ mice. The body weight as well as fat mass was unchanged in Gpr30-lacZ mice, even if fed with a high-fat diet. Flow cytometric analysis revealed lower frequencies of T cells in both sexes of Gpr30-lacZ mice. Within the T-cell cluster, the amount of CD62L-expressing cells was clearly reduced, suggesting an impaired production of T cells in the thymus of Gpr30-lacZ mice.

Induction of cerebral arteriogenesis leads to early-phase expression of protease inhibitors in growing collaterals of the brain.

Cerebral arteriogenesis constitutes a promising therapeutic concept for cerebrovascular ischaemia; however, transcriptional profiles important for therapeutic target identification have not yet been investigated. This study aims at a comprehensive characterization of transcriptional and morphologic activation during early-phase collateral vessel growth in a rat model of adaptive cerebral arteriogenesis. Arteriogenesis was induced using a three-vessel occlusion (3-VO) rat model of nonischaemic cerebral hypoperfusion. Collateral tissue from growing posterior cerebral artery (PCA) and posterior communicating artery (Pcom) was selectively isolated avoiding contamination with adjacent tissue. We detected differential gene expression 24 h after 3-VO with 164 genes significantly deregulated. Expression patterns contained gene transcripts predominantly involved in proliferation, inflammation, and migration. By using scanning electron microscopy, morphologic activation of the PCA endothelium was detected. Furthermore, the PCA showed induced proliferation (PCNA staining) and CD68+ macrophage staining 24 h after 3-VO, resulting in a significant increase in diameter within 7 days after 3-VO, confirming the arteriogenic phenotype. Analysis of molecular annotations and networks associated with differentially expressed genes revealed that early-phase cerebral arteriogenesis is characterised by the expression of protease inhibitors. These results were confirmed by quantitative real-time reverse transcription-PCR, and in situ hybridisation localised the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) and kininogen to collateral arteries, showing that TIMP-1 and kininogen might be molecular markers for early-phase cerebral arteriogenesis.

Sex-specific pathways in early cardiac response to pressure overload in mice.

Pressure overload (PO) first causes cardiac hypertrophy and then heart failure (HF), which are associated with sex differences in cardiac morphology and function. We aimed to identify genes that may cause HF-related sex differences. We used a transverse aortic constriction (TAC) mouse model leading to hypertrophy without sex differences in cardiac function after 2 weeks, but with sex differences in hypertrophy 6 and 9 weeks after TAC. Cardiac gene expression was analyzed 2 weeks after surgery. Deregulated genes were classified into functional gene ontology (GO) categories and used for pathway analysis. Classical marker genes of hypertrophy were similarly upregulated in both sexes (alpha-actin, ANP, BNP, CTGF). Thirty-five genes controlling mitochondrial function (PGC-1, cytochrome oxidase, carnitine palmitoyl transferase, acyl-CoA dehydrogenase, pyruvate dehydrogenase kinase) had lower expression in males compared to females after TAC. Genes encoding ribosomal proteins and genes associated with extracellular matrix remodeling exhibited relative higher expression in males (collagen 3, matrix metalloproteinase 2, TIMP2, and TGFbeta2, all about twofold) after TAC. We confirmed 87% of the gene expression by real-time polymerase chain reaction. By GO classification, female-specific genes were related to mitochondria and metabolism and males to matrix and biosynthesis. Promoter studies confirmed the upregulation of PGC-1 by E2. Less downregulation of metabolic genes in female hearts and increased protein synthesis capacity and deregulation of matrix remodeling in male hearts characterize the sex-specific early response to PO. These differences could contribute to subsequent sex differences in cardiac function and HF.

Description of a local cardiac adiponectin system and its deregulation in dilated cardiomyopathy.

AIMS: Despite recent advances in medical therapy, heart failure remains a leading cause for cardiovascular mortality, and its complex pathogenesis is incompletely understood. This study was performed to identify possible new therapeutic targets in dilated cardiomyopathy (DCM). METHODS AND RESULTS: Oligonucleotide microarray analysis was performed on endomyocardial biopsies (EMBs) from patients with early DCM (LVEDD > or = 55 mm, LVEF < or = 55%, n = 5) and control subjects (LVEDD < 55 mm, LVEF > 60%, no cardiac pathology, n = 4). Adiponectin, an adipocytokine involved in cellular metabolism, survival, and immunmodulation, was six-fold downregulated in DCM patients. Microarray data for adiponectin were confirmed by TaqMan-PCR (9.2-fold downregulation, control n= 9 vs. DCM n= 9, respectively, P < 0.05). Immunohistological analysis of EMBs showed significant downregulation of cardiac adiponectin protein expression independent of serum adiponectin (P = 0.36, ns) or serum TNFalpha concentrations (P = 0.46, ns). Neither the adiponectin receptor 1 (adipo-R1) nor adipo-R2 was deregulated in early DCM. Adiponectin mRNA and protein downregulation were confirmed in explanted hearts of patients with advanced DCM (LVEF < 25%, n= 8). In vitro, adiponectin incubation of neonatal rat ventricular myocytes led to activation of the pro-survival kinase PKB/Akt, increased eNOS-phosphorylation, and prevented stress-induced apoptosis of cardiomyocytes in an Akt-dependent manner. Moreover, inhibition of adiponectin secretion was accompanied by an increase in the expression of the cytokine and its receptors. CONCLUSION: These data indicate the existence of a local cardiac adiponectin system regulated independent of adiponectin and TNFalpha serum levels and its disturbance in cardiac pathology. The study suggests a role for adiponectin in the pathogenesis of DCM and implicates the adipocytokine as a possible future therapeutic target in DCM.

Sexually dimorphic gene expression in the heart of mice and men.

The prevalence and clinical manifestation of several cardiovascular diseases vary considerably with sex and age. Thus, a better understanding of the molecular basis of these differences may represent a starting point for an improved gender-specific medicine. Despite the fact that sex-specific differences have been observed in the cardiovascular system of humans and animal models, systematic analyses of sexual dimorphisms at the transcriptional level in the healthy heart are missing. Therefore we performed gene expression profiling on mouse and human cardiac samples of both sexes and young as well as aged individuals and verified our results for a subset of genes using real-time polymerase chain reaction in independent left ventricular samples. To tackle the question whether sex differences are evolutionarily conserved, we also compared sexually dimorphic genes between both species. We found that genes located on sex chromosomes were the most abundant ones among the sexually dimorphic genes. Male-specific expression of Y-linked genes was observed in mouse hearts as well as in the human myocardium (e.g. Ddx3y, Eif2s3y and Jarid1d). Higher expression levels of X-linked genes were detected in female mice for Xist, Timp1 and Car5b and XIST, EIF2S3X and GPM6B in women. Furthermore, genes on autosomal chromosomes encoding cytochromes of the monoxygenase family (e.g. Cyp2b10), carbonic anhydrases (e.g. Car2 and Car3) and natriuretic peptides (e.g. Nppb) were identified with sex- and/or age-specific expression levels. This study underlines the relevance of sex and age as modifiers of cardiac gene expression.