A standardised FACS assay based on native, receptor transfected cells for the clinical diagnosis and monitoring of β1-adrenergic receptor autoantibodies in human heart disease

Bornholz, Beatrice; Benninghaus, Thomas; Reinke, Yvonne; Felix, Stephan B.; Roggenbuck, Dirk; Jahns-Boivin, Valerie; Boege, Fritz

doi:10.1515/cclm-2015-0603

Cited by 10 publications

(7 citation statements)

References 30 publications

(80 reference statements)

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“…To quantify anti-β 1 AR Ab, a commercially available CE-certified ELISA kit (CellTrend GmbH, Luckenwalde, Germany) was used. Native membrane extracts from cell lines overexpressing human ß 1 AdrR were used to measure IgG via 7-transmembrane-ß1-Receptors in its assumed physiological conformation, as previously described (21, 22).…”

Section: Methodsmentioning

confidence: 99%

Beta-1-Adrenergic Receptor Antibodies in Acute Coronary Syndrome: Is Less Sometimes More?

Ernst

Widera

Weiberg

et al. 2018

Front. Cardiovasc. Med.

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Background: Anti-beta-1-adrenergic receptor antibodies (anti-β1AR Ab) are associated with ischemic cardiomyopathies (ICM). Evidence continues to emerge supporting an autoimmune component to various cardiac diseases. This study compares anti-β1AR Ab concentrations in patients with different entities of acute coronary syndromes (ACS) to asymptomatic non-ACS patients with positron-emission computed tomography (PET/CT)-proven atherosclerosis, and healthy controls.Methods: Serum anti-β1AR Ab IgG concentrations were measured in 212 ACS patients, 100 atherosclerosis patients, and 62 controls using ELISA. All ACS patients underwent coronary angiography. All 374 patients participating completed a structured questionnaire regarding traditional cardiovascular risk factors. ACS patients were followed up for 6 months.Results: Patients with ACS exhibited lower anti-β1AR Ab levels compared to patients with atherosclerosis or healthy controls (both p < 0.001). No differences in the ab levels were evident between healthy controls and patients with atherosclerosis. In the ACS groups, lower concentrations were found in patients with ST-elevation myocardial infarction (STEMI) (0.67 μg/ml) compared to patients with angina pectoris (AP) and non-ST elevation myocardial infarction (NSTEMI) (both 0.76 μg/ml, p = 0.008). Anti-β1AR Ab levels ≤ 0.772 μg/ml were predictive for death and reinfarction (AUC 0.77, p = 0.006). No significant correlations between anti-β1AR Ab levels and atherosclerotic burden or traditional cardiovascular risk factors were identified.Conclusions: Lower anti-β1AR Ab concentrations appear to characterize ACS phenotypes and could serve as diagnostic and prognostic markers independent from traditional risk factors for atheroscle. The prognostic predictive value of anti-β1AR Ab in ACS remains to be confirmed in larger studies.

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Section: Methodsmentioning

confidence: 99%

Beta-1-Adrenergic Receptor Antibodies in Acute Coronary Syndrome: Is Less Sometimes More?

Ernst

Widera

Weiberg

et al. 2018

Front. Cardiovasc. Med.

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“…This classical bioassay has recently been transferred to a fully automated high throughput level by Joshi-Barr et al [13] . A FACS assay, based on native beta1-adrenoceptor conformation [14] , also proved to be appropriate and comparable to a high degree to the previously mentioned so-called “classical” bioassay of beta1-AAB measurement which is based on the spontaneously beating neonatal rat cardiomyocytes [15] . Other tests which are based on a biological read-out used receptor-mediated changes of contractility of small arterioles (in vitro) as successfully used and described by Li et al in 2014 [11] .…”

Section: Introductionmentioning

confidence: 74%

Performance and in-house validation of a bioassay for the determination of beta1-autoantibodies found in patients with cardiomyopathy

Wenzel

Schulze-Rothe

Haberland

et al. 2017

Heliyon

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BackgroundAutoantibodies specific for the adrenergic beta1-receptor were identified to be an essential factor for the pathogenesis of dilated cardiomyopathy. For the detection of these autoantibodies, a bioassay was developed and has been used, measuring the positive chronotropic effect on spontaneously beating neonatal rat cardiomyocytes. In order to use this bioassay as an analytical tool to monitor the effectiveness of autoantibody neutralizing therapy in a regulated field, there is a need to assess its analytical performance and validate it according to current guidelines.MethodsUsing standard autoantibody samples, the increased beat rate compared to the basal rate [delta beats/min] was recorded when investigating guideline required assay performance parameters.ResultsThe analytical specificity and sensitivity of the bioassay was demonstrated. The limit of detection and positivity cut-off level were determined to be 3.56 and 7.97 delta beats/min, respectively. The coefficient of variation (CV) of all tested single values (four technical replicates each) was ≤15.2%. The CV of precision within each measuring series did not exceed 20%. Furthermore, the sample stability under a variety of different storage conditions was assessed, as well as the robustness of the cardiomyocyte preparations, which were both given.ConclusionThis bioassay fulfilled guideline determined quality requirements and proved to be appropriate for its application in clinical trials.

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“… 46 , 47 , 48 , 49 , 50 The basic pathogenetic mechanism has been extensively demonstrated by ex vivo studies and animal models, 13 , 15 and we know of a human disease (Chagas' cardiomyopathy) based thereon. 51 However, published evidence regarding a putative role of humoral GPCR autoimmunity in human chronic HF other than Chagas' cardiomyopathy is inconsistent, and the measurements employed in most available clinical studies to determine potentially cardio‐pathogenic GPCR autoantibodies are either impractical, not certified, and/or poorly reproducible 52 , 53 or invalid. 35 , 36 , 37 …”

Section: Discussionmentioning

confidence: 99%

“…However, such measurements do not necessarily reflect the impact of the antibodies on receptor function. 35 , 36 , 37 , 53 Other previous studies have been exclusively based on functional readouts, which are only loosely related to circulating levels of the antibodies. 52 , 57 Here, we employed a type of assay that to the best of our knowledge combines both approaches as it measures IgG binding to the native receptor or a circular peptide faithfully reflecting the conformational autoepitope related to the active receptor conformation.…”

Section: Discussionmentioning

confidence: 99%

Receptor autoantibodies: Associations with cardiac markers, histology, and function in human non‐ischaemic heart failure

et al. 2023

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Aims A causal link between non-ischaemic heart failure (HF) and humoral autoimmunity against G-protein-coupled receptors (GPCR) remains unclear except for Chagas' cardiomyopathy. Uncertainty arises from ambiguous reports on incidences of GPCR autoantibodies, spurious correlations of autoantibody levels with disease activity, and lack of standardization and validation of measuring procedures for putatively cardio-pathogenic GPCR autoantibodies. Here, we use validated and certified immune assays presenting native receptors as binding targets. We compared candidate GPCR autoantibody species between HF patients and healthy controls and tested associations of serum autoantibody levels with serological, haemodynamic, metabolic, and functional parameters in HF. Methods Ninety-five non-ischaemic HF patients undergoing transcatheter endomyocardial biopsy and 60 healthy controls were included. GPCR autoantibodies were determined in serum by IgG binding to native receptors or a cyclic peptide (for β1AR autoantibodies). In patients, cardiac function, volumes, and myocardial structural properties were assessed by cardiac magnetic resonance imaging; right heart catheterization served for determination of cardiac haemodynamics; endomyocardial biopsies were used for histological assessment of cardiomyopathy and determination of cardiac mitochondrial oxidative function by high-resolution respirometry. Results Autoantibodies against β 1 adrenergic (β 1 AR ) , M5-muscarinic (M5AR), and angiotensin II type 2 receptors (AT2R) were increased in HF (all P < 0.001). Autoantibodies against α 1 -adrenergic (α 1 AR) and angiotensin II type 1 receptors (AT1R) were decreased in HF (all P < 0.001). Correlation of alterations of GPCR autoantibodies with markers of cardiac or systemic inflammation or cardiac damage, haemodynamics, myocardial histology, or left ventricular inflammation (judged by T2 mapping) were weak, even when corrected for total IgG. β 1 AR autoantibodies were related inversely to markers of left ventricular fibrosis indicated by T1 mapping (r = À0.362, P < 0.05) and global longitudinal strain (r = À0.323, P < 0.05). AT2R autoantibodies were associated with improved myocardial mitochondrial coupling as measured by high-resolution respirometry in myocardial biopsies (r = À0.352, P < 0.05). In insulin-resistant HF patients, AT2R autoantibodies were decreased (r = À.240, P < 0.05), and AT1R autoantibodies were increased (r = 0.212, P < 0.05). Conclusions GPCR autoantibodies are markedly altered in HF. However, they are correlated poorly or even inversely to haemodynamic, metabolic, and functional markers of disease severity, myocardial histology, and myocardial mitochondrial efficiency. These observations do not hint towards a specific cardio-pathogenic role of GPCR autoantibodies and suggest that further investigations are required before specific therapies directed at GPCR autoantibodies can be clinically tested in non-ischaemic HF.

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A standardised FACS assay based on native, receptor transfected cells for the clinical diagnosis and monitoring of β1-adrenergic receptor autoantibodies in human heart disease

Abstract: The novel assay possibly provides a tool to determine true prevalence and clinical impact of β1AR-autoantibodies. Furthermore, it may serve as companion diagnostic for therapies specifically directed at β1AR-autoantibodies.

Cited by 10 publications

References 30 publications

Beta-1-Adrenergic Receptor Antibodies in Acute Coronary Syndrome: Is Less Sometimes More?

Beta-1-Adrenergic Receptor Antibodies in Acute Coronary Syndrome: Is Less Sometimes More?

Performance and in-house validation of a bioassay for the determination of beta1-autoantibodies found in patients with cardiomyopathy

Receptor autoantibodies: Associations with cardiac markers, histology, and function in human non‐ischaemic heart failure

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