Indoxyl sulfate impairs electrophysiological function and triggers arrhythmias in cells isolated from human ventricular myocardium

Oliver Pfeuffer (Regensburg)1, T. Körtl (Gießen)2, D. Riedl (Gießen)2, N. Hankowitz (Regensburg)1, L. Stengel (Regensburg)3, L. S. Maier (Regensburg)4, K. Streckfuß-Bömeke (Würzburg)5, S. T. Sossalla (Gießen)2

1Universitätsklinikum Regensburg Klinik und Poliklinik für Innere Medizin II Regensburg, Deutschland; 2Universitätsklinikum Gießen und Marburg GmbH Medizinische Klinik I - Kardiologie und Angiologie Gießen, Deutschland; 3Universitätsklinikum Regensburg Innere Medizin II Regensburg, Deutschland; 4Universitätsklinikum Regensburg Klinik und Poliklinik für Innere Med. II, Kardiologie Regensburg, Deutschland; 5Universitätsklinikum Würzburg Institut für Pharmakologie und Toxikologie Würzburg, Deutschland


Background: Chronic kidney disease (CKD) affects more than 10% of the population worldwide and is associated with increased mortality and morbidity. In patients with CKD, cardiovascular diseases are the most common cause of death. The effects of CKD on cellular mechanisms in cardiomyocytes remain only partially understood. Here, we investigated the influence of the uremic toxin indoxyl sulfate (IS) on human ventricular myocardium.

Methods: Experiments were conducted using human left ventricular myocardium, which was acquired from patients with aortic stenosis undergoing surgical valve replacement. Additionally, human ventricular induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) from donors without any known cardiovascular disease were used. After digestion, single primary human cardiomyocytes were incubated with IS 120 mg/l (the highest level of IS found in dialysis patients) for 30 minutes before measurements were carried out. iPSC-CM were incubated for 7 days with IS 45 mg/l (the average level of IS in dialysis patients) or IS 120 mg/l before conducting measurements. Ca2+ homeostasis was assessed by epifluorescence microscopy, diastolic Ca2+ sparks were investigated by confocal microscopy, and cardiac actions potentials and the presence of arrhythmias were determined via patch clamp measurements.

Results: Human ventricular myocardium was obtained from 10 patients. The mean left ventricular ejection fraction was 51.5±12% and the mean glomerular filtration rate was 66.3±22.7 ml/min in these patients. After acute incubation with IS 120 mg/l, single primary human cardiomyocytes showed a higher systolic Ca2+ transient amplitude compared with the control group. These results were confirmed in iPSC-CM: after 7 days of incubation with IS 45 mg/l or IS 120 mg/l, Ca2+ transient amplitudes were higher (Figure 1). In addition, lower diastolic Ca2+ levels versus controls were observed under these conditions. Confocal microscopy revealed that in iPSC-CM treated with IS, diastolic Ca2+ sparks occurred more frequently. Furthermore, IS had no effect on cardiac action potential duration in iPSC-CM, but there was a higher number of delayed afterdepolarizations. 

Fig. 1: When primary human cardiomyocytes were treated with IS 120mg/l and iPSC-CM were treated with IS 45 mg/l or IS 120 mg/l, systolic Ca2+ transients were significantly increased. 0.5 Hz and 1 Hz were used as measurement frequencies. One-way ANOVA (Sidak) was carried out; *p<0.05, ****p<0.0001.


Conclusion: We demonstrate for the first time that IS alters cellular electrophysiological function in primary human myocardium and iPSC-CM. In addition, IS was identified as a proarrhythmogenic trigger. Thus, IS may contribute to the poor prognosis of patients suffering from end-stage renal disease. As IS is a protein-bound uremic toxin and is therefore not effectively removed by dialysis, new therapeutic concepts for CKD patients on dialysis are required to prevent them from experiencing deleterious effects.

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