Department of Neurology, The First Affiliated Hospital, Hengyang Medical School, University of South China, Hengyang, China.
Department of Neurosurgery, Xiangya Hospital, Central South University, Changsha, China.
J Biochem Mol Toxicol. 2023 Jun;37(6):e23346. doi: 10.1002/jbt.23346. Epub 2023 Mar 29.
The protection of the blood-brain barrier (BBB) is the key direction to improving subarachnoid hemorrhage (SAH). Therefore, developing appropriate targeted drugs and therapies has become an urgent task for SAH patients. In this study, we investigated the role of dendritic cells (DCs) exosomal miR-3064-5p in repairing the BBB, providing a new basis for treating SAH. We detected the expression of miR-3064-5p in exosomes secreted by DCs (DCs-exo). An SAH rat model was constructed by intravascular perforation and characterized by HE and TUNEL-IF staining. We found that overexpression of miR-3064-5p in SAH rats suppressed iNOS expression and promoted the accumulation of tight junction proteins (Occludin, Claudin-3, ZO-1), whereas knockdown of miR-3064-5p exerted the opposite effect. Dual-LUC assay confirmed that miR-3064-5p could target and inhibit SIRT6. Knockdown of SIRT6 inhibited inflammatory cytokine (IL-6, IL-1β, IFN-γ, and TGF-β1) levels and apoptosis. The results of the co-IP assay showed that SIRT6 interacted with PCSK9, and knockdown of SIRT6 suppressed the expression of PCSK9. Moreover, DCs-exo reduced brain edema, upregulated miR-3064-5p and downregulated SIRT6 and PCSK9 in SAH rats. DCs-exo reduced inflammatory factors and increased tight junction proteins in SAH rats. Overexpression of miR-3064-5p enhanced the protective effect of DCs-exo, while overexpression of SIRT6 partially counteracted the effect. This study confirmed that DCs could secrete miR-3064-5p to ameliorate BBB damage after SAH. Mechanistically, miR-3064-5p alleviated BBB damage by targeting and inhibiting SIRT6/PCSk9 signaling pathway.
血脑屏障（BBB）的保护是改善蛛网膜下腔出血（SAH）的关键方向。因此，开发合适的靶向药物和治疗方法已成为 SAH 患者的迫切任务。在这项研究中，我们研究了树突状细胞（DC）来源的外泌体 miR-3064-5p 在修复 BBB 中的作用，为治疗 SAH 提供了新的依据。我们检测了 DC 分泌的外泌体（DC-exo）中 miR-3064-5p 的表达。通过血管穿孔构建 SAH 大鼠模型，通过 HE 和 TUNEL-IF 染色进行特征分析。我们发现，SAH 大鼠中 miR-3064-5p 的过表达抑制了 iNOS 的表达，促进了紧密连接蛋白（Occludin、Claudin-3、ZO-1）的积累，而 miR-3064-5p 的敲低则产生了相反的效果。双荧光素酶报告基因检测证实 miR-3064-5p 可靶向并抑制 SIRT6。SIRT6 的敲低抑制了炎症细胞因子（IL-6、IL-1β、IFN-γ 和 TGF-β1）水平和细胞凋亡。免疫共沉淀（co-IP）检测结果表明 SIRT6 与 PCSK9 相互作用，SIRT6 的敲低抑制了 PCSK9 的表达。此外，DC-exo 降低了脑水肿，上调了 miR-3064-5p，下调了 SIRT6 和 PCSK9 在 SAH 大鼠中的表达。DC-exo 降低了 SAH 大鼠的炎症因子，增加了紧密连接蛋白。miR-3064-5p 的过表达增强了 DC-exo 的保护作用，而过表达 SIRT6 则部分抵消了这种作用。本研究证实，DC 可以分泌 miR-3064-5p 来改善 SAH 后的 BBB 损伤。机制上，miR-3064-5p 通过靶向抑制 SIRT6/PCSk9 信号通路缓解 BBB 损伤。
