Cui Liu-Gen, Zhai Miao-Miao, Yin Jian-Jian, Wang Zhi-Mo, Wang Shu-Hui, Zhou Yue-Jiao, Li Pei-Pei, Wang Yang, Xia Li, Wang Pei, Cha Xue-Xiang, Zhang Li-Rong, Han Sheng-Na
Department of Pharmacology, School of Basic Medical Sciences, Zhengzhou University, Zhengzhou, 450001, China.
Department of Clinical Laboratory, Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, Zhengzhou, Henan, 450000, China.
Eur J Pharmacol. 2025 Feb 15;989:177247. doi: 10.1016/j.ejphar.2024.177247. Epub 2024 Dec 31.
Several studies have associated the epitranscriptomic RNA modification of N6-methyladenosine (mA) with cardiovascular diseases; however, how mA modification affects cardiomyocyte pyroptosis after myocardial infarction (MI) remains unknown. Here, we showed that AlkB homolog 5 (ALKBH5), an mA demethylase, is crucial in cardiomyocyte pyroptosis after MI. We used MI rat and mouse models, a cell hypoxia model of rat primary cardiomyocytes (RCMs), and rat embryonic ventricle cell line (H9c2) to explore the functional role of mA modification and ALKBH5 in the heart and cardiomyocytes. Using plasmids and small interfering RNAs, the expressions of ALKBH5 and NOD-like receptor family pyrin domain-containing 3 (NLRP3) were determined to study their functions in regulating cardiomyocyte mA and pyroptosis, respectively. We characterized the role of ALKBH5, which exhibited elevated expression in the ischemic heart tissue of rats and mice and hypoxic cardiomyocytes (RCMs and H9c2 cells). ALKBH5 knockdown alleviated hypoxia-induced H9c2 cell pyroptosis by inhibiting NLRP3 inflammasome activation, whereas ALKBH5 overexpression had the opposite effect. NLRP3 knockdown alleviated hypoxia-induced H9c2 cardiomyocyte pyroptosis by inhibiting ALKBH5 expression, whereas NLRP3 overexpression had the opposite effect. Mechanistically, ALKBH5 mediated mA modification of NLRP3 mRNA in an IGF2BP2-dependent manner, and NLRP3, as a nuclear transcription factor, regulated the ALKBH5 transcription process. Targeting the ALKBH5-NLRP3 loop with the small-molecule inhibitors alleviated cardiomyocyte pyroptosis. Our results highlight that ALKBH5-NLRP3 forms a positive feedback loop that promotes cardiomyocyte pyroptosis after MI. Therefore, inhibiting the ALKBH5-NLRP3 loop is a potential strategy for treating cardiovascular diseases.
多项研究已将N6-甲基腺苷（m⁶A）的表观转录组RNA修饰与心血管疾病联系起来；然而，m⁶A修饰如何影响心肌梗死后心肌细胞焦亡仍不清楚。在此，我们表明，作为一种m⁶A去甲基化酶的AlkB同源物5（ALKBH5）在心肌梗死后心肌细胞焦亡中起关键作用。我们使用心肌梗死大鼠和小鼠模型、大鼠原代心肌细胞（RCMs）的细胞缺氧模型以及大鼠胚胎心室细胞系（H9c2）来探究m⁶A修饰和ALKBH5在心脏和心肌细胞中的功能作用。使用质粒和小干扰RNA，分别测定ALKBH5和含NOD样受体家族吡啉结构域3（NLRP3）的表达，以研究它们在调节心肌细胞m⁶A和焦亡中的功能。我们对ALKBH5的作用进行了表征，其在大鼠和小鼠的缺血心脏组织以及缺氧心肌细胞（RCMs和H9c2细胞）中表达升高。敲低ALKBH5通过抑制NLRP3炎性小体激活减轻缺氧诱导的H9c2细胞焦亡，而ALKBH5过表达则产生相反的效果。敲低NLRP3通过抑制ALKBH5表达减轻缺氧诱导的H9c2心肌细胞焦亡，而NLRP3过表达则产生相反的效果。机制上，ALKBH5以依赖IGF2BP2的方式介导NLRP3 mRNA的m⁶A修饰，并且NLRP3作为一种核转录因子，调节ALKBH5的转录过程。用小分子抑制剂靶向ALKBH5-NLRP3环可减轻心肌细胞焦亡。我们的结果突出表明，ALKBH5-NLRP3形成一个正反馈环，促进心肌梗死后心肌细胞焦亡。因此，抑制ALKBH-5-NLRP3环是治疗心血管疾病的一种潜在策略。
