Department of Neurology, Shandong Provincial Qixia City People's Hospital, Yantai City, China.
Shock. 2024 Jul 1;62(1):85-94. doi: 10.1097/SHK.0000000000002348. Epub 2024 Mar 25.
Background: Cerebral ischemia-reperfusion (I/R) injury (CIRI) have severe consequences on brain function, and the exciting evidence has revealed protective role of acyl-CoA synthetase long chain family member 4 (Lin28a) against cerebral ischemia-reperfusion injury. The present work aims to reveal its molecular mechanism in regulating CIRI, with the hope of providing a therapeutic method for cerebral I/R injury. We hypothesized that the exosomal nuclear factor erythroid 2-related factor 2 (NRF2) derived from bone marrow mesenchymal stromal cells could transcriptionally activate Lin28a and thereby alleviate cerebral ischemia-reperfusion injury. This hypothesis was validated in the present work. Methods: Middle cerebral artery occlusion (MCAO) model was established using C57BL/6J mice, and the neurological deficit, infarct volume, and brain water content were assessed to evaluate neuron injury. Human glioblastoma cells (A172) were subjected to oxygen-glucose deprivation and reoxygenation (OGD/R) treatment to mimic a cerebral I/R injury cell model. Exosome isolation reagent was used to isolate exosomes from cell supernatant of bone marrow mesenchymal stromal cells through sequential centrifugation and filtration steps. mRNA expression level of Lin28a was detected by quantitative real-time polymerase chain reaction. Protein expression was analyzed by western blotting assay. TUNEL cell apoptosis detection kit was used to analyze cell apoptosis in brain tissues. Enzyme-linked immunosorbent assays and commercial kits were used to detect levels of inflammatory markers and oxidative stress markers. Ferrous Iron Colorimetric Assay Kit and Fe 2+ colorimetric assay kit were used to analyze Fe 2+ level. The association of Lin28a and NRF2 was identified by chromatin immunoprecipitation assay and dual-luciferase reporter assay. Results: The treatment of MCAO substantially augmented infarct volume in mice, impaired neurological function, and elevated brain water content. Lin28a was lowly expressed in brain tissues of mice with CIRI, and its overexpression protected against cerebral I/R injury of MCAO mice. Moreover, Lin28a overexpression protected A172 cells against OGD/R treatment-induced injury. Additionally, NRF2 transcriptionally activated Lin28a in A172 cells. Bone marrow mesenchymal stromal cell-derived exosomes increased Lin28a expression in a NRF2-dependent manner. Bone marrow mesenchymal stromal cell-derived exosomal NRF2 improved OGD/R-induced A172 cell injury by inducing Lin28a production. Conclusion: Bone marrow mesenchymal stromal cell-derived exosomal NRF2 improved CIRI by transcriptionally activating Lin28a.
背景：脑缺血再灌注（I/R）损伤（CIRI）对脑功能有严重后果，令人兴奋的证据表明酰基辅酶 A 合成酶长链家族成员 4（Lin28a）对脑缺血再灌注损伤具有保护作用。本研究旨在揭示其调节 CIRI 的分子机制，希望为脑 I/R 损伤提供一种治疗方法。我们假设骨髓间充质基质细胞衍生的核因子红细胞 2 相关因子 2（NRF2）外泌体可以转录激活 Lin28a，从而减轻脑缺血再灌注损伤。本研究验证了这一假设。方法：采用 C57BL/6J 小鼠建立大脑中动脉闭塞（MCAO）模型，通过神经功能缺损、梗死体积和脑水含量评估神经元损伤。用人神经胶质瘤细胞（A172）进行氧葡萄糖剥夺和再氧合（OGD/R）处理，模拟脑 I/R 损伤细胞模型。通过连续离心和过滤步骤，使用外泌体分离试剂从骨髓间充质基质细胞的细胞上清液中分离出外泌体。通过实时定量聚合酶链反应检测 Lin28a 的 mRNA 表达水平。通过 Western blot 分析检测蛋白表达。使用 TUNEL 细胞凋亡检测试剂盒分析脑组织中的细胞凋亡。酶联免疫吸附试验和商业试剂盒用于检测炎症标志物和氧化应激标志物的水平。使用亚铁比色法试剂盒和 Fe 2+比色法试剂盒分析 Fe 2+水平。通过染色质免疫沉淀分析和双荧光素酶报告基因分析鉴定 Lin28a 和 NRF2 的关联。结果：MCAO 处理显著增加了小鼠的梗死体积，损害了神经功能，并增加了脑含水量。Lin28a 在 CIRI 小鼠脑组织中低表达，过表达可保护 MCAO 小鼠免受脑 I/R 损伤。此外，Lin28a 过表达可保护 A172 细胞免受 OGD/R 处理诱导的损伤。此外，NRF2 在 A172 细胞中转录激活 Lin28a。骨髓间充质基质细胞衍生的外泌体通过 NRF2 依赖性方式增加 Lin28a 的表达。骨髓间充质基质细胞衍生的外泌体 NRF2 通过诱导 Lin28a 的产生来改善 OGD/R 诱导的 A172 细胞损伤。结论：骨髓间充质基质细胞衍生的外泌体 NRF2 通过转录激活 Lin28a 改善 CIRI。
J Cereb Blood Flow Metab. 2025-3-12
Zotero 插件