Department of Neurology, the First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001, Heilongjiang Province, PR China.
Department of Neurology, the First Affiliated Hospital of Harbin Medical University, 23 Youzheng Street, Harbin 150001, Heilongjiang Province, PR China; Department of Neurology, Heilongjiang Provincial Hospital, 405 Guogeli Street, Harbin 150036, Heilongjiang Province, PR China.
J Stroke Cerebrovasc Dis. 2023 Aug;32(8):107235. doi: 10.1016/j.jstrokecerebrovasdis.2023.107235. Epub 2023 Jun 30.
Ischemic stroke represents a major factor causing global morbidity and death. Bone marrow mesenchymal stem cell (BMSC)-derived exosomes (Exos) have important effects on treating ischemic stroke. Here, we investigated the therapeutic mechanism by which BMSC-derived exosomal miR-193b-5p affects ischemic stroke.
luciferase assay was performed to evaluate the regulatory relationship of miR-193b-5p with absent in melanoma 2 (AIM2). Additionally, an oxygen-glucose deprivation/reperfusion (OGD/R) model was constructed for the in vitro assay, while a middle cerebral artery occlusion (MCAO) model was developed for the in vivo assay. After exosome therapy, lactate dehydrogenase and MTT assays were conducted to detect cytotoxicity and cell viability, while PCR, ELISA, western blotting assay, and immunofluorescence staining were performed to detect changes in the levels of pyroptosis-related molecules. TTC staining and TUNEL assays were performed to assess cerebral ischemia/reperfusion (I/R) injury.
In the luciferase assay, miR-193b-5p showed direct binding to the 3'-untranslated region of AIM2. In both in vivo and in vitro assays, the injected exosomes could access the sites of ischemic injury and could be internalized. In the in vitro assay, compared to normal BMSC-Exos, miR-193b-5p-overexpressing BMSC-Exos showed greater effects on increasing cell viability and attenuating cytotoxicity; AIM2, GSDMD-N, and cleaved caspase-1 levels; and IL-1β/IL-18 generation. In the in vivo assay, compared to normal BMSC-Exos, miR-193b-5p-overexpressing BMSC-Exos showed greater effects on decreasing the levels of these pyroptosis-related molecules and infarct volume.
BMSC-Exos attenuate the cerebral I/R injury in vivo and in vitro by inhibiting AIM2 pathway-mediated pyroptosis through miR-193b-5p delivery.
缺血性脑卒中是导致全球发病率和死亡率的主要因素。骨髓间充质干细胞（BMSC）衍生的外泌体（Exos）对治疗缺血性脑卒中具有重要作用。在这里，我们研究了 BMSC 衍生的外泌体 miR-193b-5p 影响缺血性脑卒中的治疗机制。
采用荧光素酶报告基因实验评估 miR-193b-5p 与黑色素瘤缺失 2（AIM2）的调控关系。此外，构建了氧-葡萄糖剥夺/再灌注（OGD/R）模型进行体外检测，构建了大脑中动脉闭塞（MCAO）模型进行体内检测。外泌体治疗后，进行乳酸脱氢酶和 MTT 检测，检测细胞毒性和细胞活力，同时进行 PCR、ELISA、western blot 检测和免疫荧光染色，检测细胞焦亡相关分子水平的变化。TTC 染色和 TUNEL 检测评估脑缺血/再灌注（I/R）损伤。
在荧光素酶报告基因实验中，miR-193b-5p 与 AIM2 的 3'-非翻译区直接结合。在体内和体外实验中，注射的外泌体能够到达缺血损伤部位并被内化。在体外实验中，与正常 BMSC-Exos 相比，miR-193b-5p 过表达的 BMSC-Exos 对增加细胞活力和减轻细胞毒性的作用更大；AIM2、GSDMD-N 和裂解的 caspase-1 水平；和 IL-1β/IL-18 的产生。在体内实验中，与正常 BMSC-Exos 相比，miR-193b-5p 过表达的 BMSC-Exos 对降低这些细胞焦亡相关分子和梗死体积的水平的作用更大。
BMSC-Exos 通过 miR-193b-5p 递送来抑制 AIM2 通路介导的细胞焦亡，从而减轻体内和体外的脑 I/R 损伤。
