Sheng Bin, Gao Sen, Chen XiangXin, Liu Yang, Lai Niansheng, Dong Jin, Sun Jiaqing, Zhou Yan, Wu Lingyun, Hang Chun-Hua, Li Wei
Department of Neurosurgery, Nanjing Drum Tower Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing, Jiangsu, China.
Department of Neurosurgery, Nanjing Drum Tower Hospital Clinical College of Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
Stroke Vasc Neurol. 2024 Oct 2. doi: 10.1136/svn-2024-003509.
Neuroinflammation participates in the pathogenesis of subarachnoid haemorrhage (SAH); however, no effective treatments exist. MicroRNAs regulate several aspects of neuronal dysfunction. In a previous study, we found that exosomal miR-486-3p is involved in the pathophysiology of SAH. Targeted delivery of miR-486-3p without blood-brain barrier (BBB) restriction to alleviate SAH is a promising neuroinflammation approach.
In this study, we modified exosomes (Exo) to form an RVG-miR-486-3p-Exo (Exo/miR) to achieve targeted delivery of miR-486-3p to the brain. Neurological scores, brain water content, BBB damage, flow cytometry and FJC staining were used to determine the effect of miR-486-3p on SAH. Western blot analysis, ELISA and RT-qPCR were used to measure relevant protein and mRNA levels. Immunofluorescence staining and laser confocal detection were used to measure the expression of mitochondria, lysosomes and autophagosomes, and transmission electron microscopy was used to observe the level of mitophagy in the brain tissue of mice after SAH.
Tail vein injection of Exo/miR improved targeting of miR-486-3p to the brains of SAH mice. The injection reduced levels of neuroinflammation-related factors by changing the phenotype switching of microglia, inhibiting the expression of sirtuin 2 (SIRT2) and enhancing mitophagy. miR-486-3p treatment alleviated neurobehavioral disorders, brain oedema, BBB damage and neurodegeneration. Further research found that the mechanism was achieved by regulating the acetylation level of peroxisome proliferator-activated receptor γ coactivator l alpha (PGC-1α) after SIRT2 enters the nucleus.
Exo/miR treatment attenuates neuroinflammation after SAH by inhibiting SIRT2 expression and stimulating mitophagy, suggesting potential clinical applications.
神经炎症参与蛛网膜下腔出血（SAH）的发病机制；然而，目前尚无有效的治疗方法。微小RNA调节神经元功能障碍的多个方面。在先前的一项研究中，我们发现外泌体miR-486-3p参与SAH的病理生理学过程。在无血脑屏障（BBB）限制的情况下靶向递送miR-486-3p以减轻SAH是一种有前景的神经炎症治疗方法。
在本研究中，我们对外泌体（Exo）进行修饰，形成RVG-miR-486-3p-Exo（Exo/miR），以实现miR-486-3p向脑内的靶向递送。采用神经功能评分、脑含水量、BBB损伤、流式细胞术和FJC染色来确定miR-486-3p对SAH的影响。采用蛋白质免疫印迹分析、酶联免疫吸附测定和逆转录定量聚合酶链反应来检测相关蛋白质和mRNA水平。采用免疫荧光染色和激光共聚焦检测来测定线粒体、溶酶体和自噬体的表达，并采用透射电子显微镜观察SAH后小鼠脑组织中的线粒体自噬水平。
尾静脉注射Exo/miR可改善miR-486-3p对SAH小鼠脑的靶向性。该注射通过改变小胶质细胞的表型转换、抑制沉默调节蛋白2（SIRT2）的表达和增强线粒体自噬，降低了神经炎症相关因子的水平。miR-486-3p治疗减轻了神经行为障碍、脑水肿、BBB损伤和神经退行性变。进一步研究发现，该机制是通过SIRT2进入细胞核后调节过氧化物酶体增殖物激活受体γ辅激活因子1α（PGC-1α）的乙酰化水平来实现的。
Exo/miR治疗通过抑制SIRT2表达和刺激线粒体自噬减轻SAH后的神经炎症，提示其具有潜在的临床应用价值。
