Abstract
Thioridazine (THIO) is a phenothiazine derivative that is mainly used for the treatment of psychotic disorders. However, cardiac arrhythmias especially QT interval prolongation associated with the application of this compound have received serious attention after its introduction into clinical practice, and the mechanisms underlying the cardiotoxicity induced by THIO have not been well defined. The present study was aimed at exploring the long-term effects of THIO on the hERG and L-type calcium channels, both of which are relevant to the development of QT prolongation. The hERG current (IhERG) and the calcium current (ICa‐L) were measured by patch clamp techniques. Protein levels were analyzed by Western blot, and channel-chaperone interactions were determined by coimmunoprecipitation. Reactive oxygen species (ROS) were determined by flow cytometry and laser scanning confocal microscopy. Our results demonstrated that THIO induced hERG channel deficiency but did not alter channel kinetics. THIO promoted ROS production and stimulated endoplasmic reticulum (ER) stress and the related proteins. The ROS scavenger N-acetyl cysteine (NAC) significantly attenuated hERG reduction induced by THIO and abolished the upregulation of ER stress marker proteins. Meanwhile, THIO increased the degradation of hERG channels via disrupting hERG-Hsp70 interactions. The disordered hERG proteins were degraded in proteasomes after ubiquitin modification. On the other hand, THIO increased ICa‐L density and intracellular Ca2+ ([Ca2+]i) in neonatal rat ventricular cardiomyocytes (NRVMs). The specific CaMKII inhibitor KN-93 attenuated the intracellular Ca2+ overload, indicating that ROS-mediated CaMKII activation promoted calcium channel activation induced by THIO. Optical mapping analysis demonstrated the slowing effects of THIO on cardiac repolarization in mouse hearts. THIO significantly prolonged APD50 and APD90 and increased the incidence of early afterdepolarizations (EADs). In human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), THIO also resulted in APD prolongation. In conclusion, dysfunction of hERG channel proteins and activation of L-type calcium channels via ROS production might be the ionic mechanisms for QT prolongation induced by THIO.
Highlights
Thioridazine (THIO) is a phenothiazine derivative that has been used for the management of major psychotic disorders over the past few decades [1]
To determine the long-term effect of THIO on the hERG channels, hERG-HEK293 cells were treated with increasing concentrations of THIO (0.1, 1, and 3 μM) for 24 h. hERG expression was examined by Western blotting
To further explore whether the reduction of mature hERG protein causes a dysfunction in the hERG currents, we analyzed hERG currents recorded from hERG-HEK293 cells, which had been incubated with different concentrations of THIO for 24 h
Summary
Thioridazine (THIO) is a phenothiazine derivative that has been used for the management of major psychotic disorders over the past few decades [1]. THIO exhibits anticancer, antimicrobial, and antiviral activities, and its role in combinational drug application is being actively investigated [2,3,4]. The most common underlying mechanism for drug-induced QT prolongation is the inhibition of the hERG (human ether-a-go-go-related gene) potassium channel (including acute blockage and a decrease in the density of mature channels on the cell membrane), which plays a crucial role in phase 3 repolarization of cardiac action potential [8]. We demonstrate that THIO reduces hERG membrane expression by inhibiting forward trafficking from the ER to the cell surface and subsequently promoting mature channel degradation
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