来自FNDC5工程化骨髓间充质干细胞的外泌体鸢尾素通过抑制YAP/EGR1/ACSL4介导的铁死亡改善缺血性中风。

Exosomal irisin from FNDC5-engineered BMSCs improves ischemic stroke via inhibiting YAP/EGR1/ACSL4-mediated ferroptosis.

作者信息

Fang Cuini, Huang Lei, Gu Jiayi, Song Tao

机构信息

Hunan Provincial People's Hospital (the First-Affiliated Hospital of Hunan Normal University), Changsha 410002, Hunan Province, China.

出版信息

Exp Neurol. 2025 May;387:115172. doi: 10.1016/j.expneurol.2025.115172. Epub 2025 Jan 29.

DOI:10.1016/j.expneurol.2025.115172

PMID:39889877

Abstract

BACKGROUND

BMSCs-derived exosomes play an important role in ischemic stroke. Irisin and its precursor fibronectin type III domain-containing protein 5 (FNDC5) are implicated in neuroprotective effect. We aimed to clarify the role of exosomal irisin from FNDC5-overexpressed BMSCs in ischemic stroke.

METHODS

Oxygen-glucose deprivation and reoxygenation (OGD/R) neuronal cells (HT-22 and Neuro-2a cells) model and middle cerebral artery occlusion (MCAO) mice model were established. Exosomes were isolated from FNDC5-overexpressed BMSCs (BMSCs-FNDC5-exos). Cell viability was estimated with CCK-8. Fe, lactate dehydrogenase (LDH), glutathione (GSH) and malondialdehyde (MDA) were determined with commercial kits. Moreover, lipid reactive oxygen species (ROS) was analyzed using flow cytometry. Neurological dysfunction, infarct volume and mitochondria injury were estimated with modified neurological severity score (mNSS), TTC staining and transmission electron microscopy (TEM). Chromatin immunoprecipitation and dual luciferase assay were applied to verify the molecular interactions. Western blot, RT-qPCR and ELISA were performed for the detection of related genes and proteins.

RESULTS

YAP, EGR1 and ACSL4 were increased in OGD/R-subjected cells. Irisin from BMSCs-FNDC5-exos elevated cell viability and suppressed ferroptosis. EGR1 transcriptionally upregulated ACSL4 and promoted OGD/R-induced ferroptosis. Additionally, YAP transcriptionally upregulated EGR1 and promoted OGD/R-induced ferroptosis. EGR1 or YAP overexpression could reverse the effects of BMSCs-FNDC5-exos. EGR1 silencing or BMSCs-FNDC5-exos overturned the facilitated ferroptosis induced by YAP overexpression, meanwhile, EGR1 silencing further enhanced the effect elicited by BMSCs-FNDC5-exos. BMSCs-FNDC5-exos reduced cerebral infarction, improved neurological impairment, inhibited ferroptosis, downregulated YAP, EGR1, ACSL4 and up-regulated irisin in MCAO mice.

CONCLUSION

Exosomal irisin from FNDC5-overexpressed BMSCs improves ischemic stroke via inhibiting YAP/EGR1/ACSL4-mediated ferroptosis, which shed light on discovering new strategy against ischemic stroke.

摘要

背景

骨髓间充质干细胞衍生的外泌体在缺血性卒中中发挥重要作用。鸢尾素及其前体含III型纤连蛋白结构域蛋白5（FNDC5）与神经保护作用有关。我们旨在阐明过表达FNDC5的骨髓间充质干细胞来源的外泌体鸢尾素在缺血性卒中中的作用。

方法

建立氧糖剥夺复氧（OGD/R）神经元细胞（HT-22和Neuro-2a细胞）模型和大脑中动脉闭塞（MCAO）小鼠模型。从过表达FNDC5的骨髓间充质干细胞（BMSCs-FNDC5-exos）中分离外泌体。用CCK-8评估细胞活力。用商业试剂盒测定铁、乳酸脱氢酶（LDH）、谷胱甘肽（GSH）和丙二醛（MDA）。此外，使用流式细胞术分析脂质活性氧（ROS）。用改良神经功能缺损评分（mNSS）、TTC染色和透射电子显微镜（TEM）评估神经功能障碍、梗死体积和线粒体损伤。应用染色质免疫沉淀和双荧光素酶测定来验证分子相互作用。进行蛋白质印迹、RT-qPCR和ELISA检测相关基因和蛋白质。

结果

在OGD/R处理的细胞中，YAP、EGR1和ACSL4增加。BMSCs-FNDC5-exos中的鸢尾素提高细胞活力并抑制铁死亡。EGR1转录上调ACSL4并促进OGD/R诱导的铁死亡。此外，YAP转录上调EGR1并促进OGD/R诱导的铁死亡。EGR1或YAP过表达可逆转BMSCs-FNDC5-exos的作用。EGR1沉默或BMSCs-FNDC5-exos可逆转YAP过表达诱导的铁死亡促进作用，同时，EGR1沉默进一步增强BMSCs-FNDC5-exos的作用。BMSCs-FNDC5-exos减少MCAO小鼠的脑梗死，改善神经功能缺损，抑制铁死亡，下调YAP、EGR1、ACSL4并上调鸢尾素。