Western Medical Branch of PLA General Hospital, 100853 Beijing, China.
Senior Department of Cardiology, the Sixth Medical Center of PLA General Hospital, 100853 Beijing, China.
Front Biosci (Landmark Ed). 2024 Jul 23;29(7):263. doi: 10.31083/j.fbl2907263.
Telomere shortening is strongly associated with cardiovascular aging and disease, and patients with shorter telomeres in peripheral blood leukocytes are at higher risk of cardiovascular diseases such as heart failure and atrial fibrillation (AF). Telomerase reverse transcriptase (TERT) maintains telomere length, and overexpression of TERT has been shown to reduce cardiomyocyte apoptosis and myocardial infarct size, and extend the lifespan of aged mice. However, the specific impact of TERT on the electrophysiology of cardiomyocytes remains to be elucidated. The aims of this study were to evaluate the role of TERT in Ca2+ homeostasis and mitochondrial function in atrial myocytes as well as the underlying mechanisms.
TERT overexpressed and silenced HL-1 cells were constructed with lentiviruses, and the respective empty lentiviral vectors were used as negative controls. Then the patch clamp technique was used to record the electrophysiological characteristics such as cell action potential duration (APD) and L-type Ca2+ currents (ICa,L), flow cytometry was used to detect intracellular Ca2+ concentration and mitochondrial membrane potential (MMP), and the Seahorse assay was used to measure the oxygen consumption rate (OCR).
TERT silencing led to intracellular Ca2+ overload, shortened APD, decreased ICa,L current density, altered Ca2+ gating mechanism, decreased MMP and OCR, and increased reactive oxygen species (ROS), whereas TERT overexpression led to the reverse effects. Additionally, TERT silencing resulted in intracellular Ca2+ overload with decreased expression of the SERCA2a, CaV1.2, and NCX1.1, whereas TERT overexpression had opposing effects. Furthermore, we discovered that TERT could regulate the expression of p53 and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). The expression of PGC-1α was downregulated by the p53 agonist Tenovin-6 but upregulated by the p53 inhibitor PFTα. The effects of the PGC-1α inhibitor SR-18292 on intracellular Ca2+ and cell electrophysiology were similar to those of silencing TERT, whereas the PGC-1α agonist ZLN005 produced comparable outcomes to TERT overexpression.
TERT silencing-induced Ca2+ overload and mitochondrial dysfunction may be one mechanism of age-related AF. Overexpression of TERT reduced the basis for arrhythmia formation such as AF, suggesting a favorable safety profile for TERT therapy. TERT regulated intracellular Ca2+ homeostasis and mitochondrial function through the p53/PGC-1α pathway. In addition, PGC-1α might be a novel target for AF, suggesting that intervention for AF should be not limited to abnormal cation handling.
端粒缩短与心血管衰老和疾病密切相关，外周血白细胞中端粒较短的患者患心力衰竭和心房颤动（AF）等心血管疾病的风险较高。端粒酶逆转录酶（ TERT ）维持端粒长度，TERT 的过表达已被证明可减少心肌细胞凋亡和心肌梗死面积，并延长老年小鼠的寿命。然而，TERT 对心肌细胞电生理的确切影响仍有待阐明。本研究旨在评估 TERT 在心房肌细胞中钙稳态和线粒体功能中的作用及其潜在机制。
使用慢病毒构建 TERT 过表达和沉默的 HL-1 细胞，分别使用空慢病毒载体作为阴性对照。然后使用膜片钳技术记录细胞动作电位时程（ APD ）和 L 型钙电流（ ICa,L ）等电生理特征，使用流式细胞术检测细胞内钙浓度和线粒体膜电位（ MMP ），使用 Seahorse assay 测量耗氧量（ OCR ）。
TERT 沉默导致细胞内钙超载，缩短 APD ，降低 ICa,L 电流密度，改变钙门控机制，降低 MMP 和 OCR ，增加活性氧（ ROS ），而 TERT 过表达则产生相反的效果。此外，TERT 沉默导致细胞内钙超载， SERCA2a 、 CaV1.2 和 NCX1.1 的表达减少，而 TERT 过表达则有相反的作用。此外，我们发现 TERT 可以调节 p53 和过氧化物酶体增殖物激活受体γ共激活因子 1-α（ PGC-1α）的表达。 p53 激动剂 Tenovin-6 下调 PGC-1α 的表达，而 p53 抑制剂 PFTα上调 PGC-1α 的表达。PGC-1α 抑制剂 SR-18292 对细胞内钙和细胞电生理的影响与 TERT 沉默相似，而 PGC-1α 激动剂 ZLN005 则产生与 TERT 过表达相当的结果。
TERT 沉默诱导的钙超载和线粒体功能障碍可能是与年龄相关的 AF 的一种机制。TERT 的过表达减少了心律失常形成的基础，如 AF ，提示 TERT 治疗具有良好的安全性。TERT 通过 p53/PGC-1α 通路调节细胞内钙稳态和线粒体功能。此外，PGC-1α 可能是 AF 的一个新靶点，提示 AF 的干预不应仅限于异常阳离子处理。
