Wang Pi-Xiao, Zhu Ling, Xiang Mei, Zhang Rixin, Zheng Xiaolin, Zheng Zhi, Li Kai
Department of Hepatobiliary and Pancreatic Surgery, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Department of Cardiology, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Gastroenterol Res Pract. 2025 Jan 6;2025:5587859. doi: 10.1155/grp/5587859. eCollection 2025.
Despite N-methyladenosine (mA) being closely involved in various pathophysiological processes, its potential role in liver injury is largely unknown. We designed the current research to study the potential role of fat mass and obesity-associated protein (FTO), an mA demethylase, on hepatic ischemia-reperfusion injury (IRI). Wild-type mice injected with an adeno-associated virus carrying fat mass and obesity-associated protein (AAV-FTO) or adeno-associated virus carrying green fluorescent protein (GFP) (AAV-GFP) were subjected to a hepatic IRI model in vivo. Hematoxylin-eosin staining was performed to observe IRI. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was used to observe the cell apoptosis. Reverse transcription polymerase chain reaction (RT-PCR) was used to observe the expression of FTO. The protein levels of FTO, apoptosis, or autophagy-associated signaling proteins were detected by western blot. Reactive oxygen species (ROS) levels were determined by flow cytometry, and immunohistochemistry was used to detect the FTO and LC3-II expression. For in vitro experiments, cultured hepatocytes were subjected to hypoxia/reoxygenation (H/R) stimulation. Monodansylcadaverine (MDC) staining was used to visualize autophagic vesicles. In the present study, we showed that FTO was involved in hepatic IRI, apoptosis, and autophagy. Specifically, the expression level of FTO was significantly reduced in the hepatic IRI. Besides, increasing FTO expression (AAV-FTO) ameliorated the hepatic IRI in animal models, accompanied by decreased apoptosis and autophagy. Furthermore, the FTO inhibitor (FB23-2) aggravated autophagy in hepatocytes upon H/R-induced damage. FTO could act as a protective effector during hepatic IRI, associated with decreased apoptosis and autophagy. FTO-mediated mA demethylation modification may be an important therapeutic target for hepatic IRI.
尽管N-甲基腺苷（mA）密切参与各种病理生理过程，但其在肝损伤中的潜在作用在很大程度上仍不清楚。我们设计了本研究，以探讨mA去甲基化酶——脂肪量与肥胖相关蛋白（FTO）在肝脏缺血再灌注损伤（IRI）中的潜在作用。给野生型小鼠注射携带脂肪量与肥胖相关蛋白的腺相关病毒（AAV-FTO）或携带绿色荧光蛋白的腺相关病毒（AAV-GFP），然后在体内建立肝脏IRI模型。进行苏木精-伊红染色以观察IRI。采用末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）染色观察细胞凋亡。通过逆转录聚合酶链反应（RT-PCR）观察FTO的表达。采用蛋白质印迹法检测FTO、凋亡或自噬相关信号蛋白的水平。通过流式细胞术测定活性氧（ROS）水平，并用免疫组织化学法检测FTO和LC3-II的表达。对于体外实验，对培养的肝细胞进行缺氧/复氧（H/R）刺激。使用单丹磺酰尸胺（MDC）染色观察自噬小泡。在本研究中，我们发现FTO参与肝脏IRI、细胞凋亡和自噬。具体而言，肝脏IRI中FTO的表达水平显著降低。此外，增加FTO表达（AAV-FTO）可改善动物模型中的肝脏IRI，同时细胞凋亡和自噬减少。此外，FTO抑制剂（FB23-2）加重了H/R诱导损伤的肝细胞中的自噬。FTO在肝脏IRI期间可作为一种保护因子，与细胞凋亡和自噬减少有关。FTO介导的mA去甲基化修饰可能是肝脏IRI的一个重要治疗靶点。
