Department of Cell and Molecular Physiology, Loyola University Chicago, Maywood, IL, 60153, USA.
Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, MN, 55455, USA.
Biochem Biophys Res Commun. 2023 Dec 10;685:149136. doi: 10.1016/j.bbrc.2023.149136. Epub 2023 Oct 19.
In cardiomyocytes, the sarco/endoplasmic reticulum Ca-ATPase (SERCA2a) is a central component of intracellular Ca regulation. Several heart diseases, including heart failure, are associated with reduced myocardial contraction due to SERCA2a downregulation. Therefore, the need for developing new drugs that could improve SERCA2a function is high. We have recently identified SERCA2a modulators (Compounds 6 and 8) from our screening campaigns and confirmed activation of biochemical SERCA2a ATPase activity and Ca uptake activity. In this study, confocal microscopy and in-cell Ca imaging were used to characterize the effects of these SERCA2a activators on Ca regulation in mouse ventricular myocytes and endoplasmic reticulum (ER) Ca uptake in a HEK293 cell expressing human SERCA2a. Analysis of cytosolic Ca dynamics in cardiomyocytes revealed that both Compounds (6 and 8) increase the action potential-induced Ca transients and sarcoplasmic reticulum (SR) Ca load. While Compound 6 induced a negligible effect on Ca transients invoked by the L-type Ca channel (LTCC) current, Compound 8 increased Ca transients during LTCC activation, suggesting an off-target protein interaction of Compound 8. Analysis of ER Ca transport by human SERCA2a in HEK cells showed that only Compound 6 increased both ER Ca uptake and ER Ca load significantly, whereas Compound 8 had no effect on SERCA2a Ca transport. This study revealed that Compound 6 exhibits promising characteristics that can improve intracellular Ca dynamics by selectively enhancing SERCA2a Ca uptake.
在心肌细胞中，肌浆网/内质网 Ca2+-ATP 酶（SERCA2a）是细胞内 Ca2+调节的核心组成部分。几种心脏病，包括心力衰竭，与 SERCA2a 下调导致心肌收缩力降低有关。因此，开发能够改善 SERCA2a 功能的新药的需求很高。我们最近从筛选活动中鉴定出 SERCA2a 调节剂（化合物 6 和 8），并证实它们能够激活生化 SERCA2a ATP 酶活性和 Ca2+摄取活性。在这项研究中，共聚焦显微镜和细胞内 Ca2+成像用于表征这些 SERCA2a 激活剂对小鼠心室肌细胞 Ca2+调节和表达人 SERCA2a 的 HEK293 细胞内质网（ER）Ca2+摄取的影响。对心肌细胞胞质 Ca2+动力学的分析表明，两种化合物（6 和 8）均增加了动作电位诱导的 Ca2+瞬变和肌浆网（SR）Ca2+负荷。虽然化合物 6 对 L 型 Ca2+通道（LTCC）电流引起的 Ca2+瞬变几乎没有影响，但化合物 8 在 LTCC 激活期间增加了 Ca2+瞬变，表明化合物 8 与非靶标蛋白相互作用。对 HEK 细胞中人 SERCA2a 的 ER Ca 转运的分析表明，只有化合物 6 显著增加了 ER Ca 摄取和 ER Ca 负荷，而化合物 8 对 SERCA2a Ca 转运没有影响。这项研究表明，化合物 6 具有通过选择性增强 SERCA2a Ca 摄取来改善细胞内 Ca2+动力学的有前景的特性。
