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脑震宁干预外周血来源外泌体对小胶质细胞诱导的神经元损伤的影响
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| 篇名： | 脑震宁干预外周血来源外泌体对小胶质细胞诱导的神经元损伤的影响 |
| TITLE： | Effects of peripheral blood-derived exosomes intervened by Naozhenning on injury of neuron induced by microglia |
| 摘要： | 目的 研究脑震宁干预外周血来源外泌体（Exo）对小胶质细胞BV-2诱导的神经元细胞HT22损伤的影响。方法选取Wistar大鼠制备脑震宁（66.83g/kg）干预外周血来源Exo（即NZN-Exo），同法制备生理盐水和吡拉西坦（1.62g/kg）干预外周血来源Exo（即KB-Exo、PLXT-Exo），并进行鉴定。以1μg/mL脂多糖（LPS）刺激BV-2细胞，制备经LPS刺激的上清液（同法制备未经LPS刺激的上清液）。将HT22细胞分为KB-Exo组（未经LPS刺激的上清液+KB-Exo）、模型组（即model组，经LPS刺激的上清液+KB-Exo）、PLXT-Exo组（经LPS刺激的上清液+PLXT-Exo）和NZN-Exo组（经LPS刺激的上清液+NZN-Exo），相应Exo的质量浓度均为50μg/mL。培养24h后，检测HT22细胞增殖情况（采用CCK-8法和EdU法），检测HT22细胞凋亡情况，观察HT22细胞超微结构，检测HT22细胞中白细胞介素1β（IL-1β）、IL-10、核因子κB（NF-κB）、肿瘤坏死因子α（TNF-α）含量，以及TNF-α、NOD样受体热蛋白结构域相关蛋白3（NLRP3）、胱天蛋白酶1（Caspase-1）、B细胞淋巴瘤2（Bcl-2）、Bcl-2相关X蛋白（Bax）的蛋白表达情况。结果本研究成功制备KB-Exo、PLXT-Exo、NZN-Exo，各Exo均呈现出较为典型的“杯状”轮廓和膜结构“包裹”特征。与KB-Exo组相比，model组的细胞增殖率（CCK-8法和EdU法）、细胞中IL-10含量和Bcl-2蛋白表达水平均显著降低（P＜0.05）；细胞凋亡率和细胞中IL-1β、TNF-α、NF-κB含量以及NLRP3、TNF-α、Caspase-1、Bax蛋白表达水平均显著升高（P＜0.05）；细胞体积膨胀，细胞膜不完整，核仁破裂，线粒体明显肿胀变形，空泡化严重。与model组相比，PLXT-Exo组、NZN-Exo组细胞上述定量指标均显著逆转（P＜0.05）；细胞核大而圆，核膜完整，线粒体空泡化减少。结论脑震宁干预外周血来源Exo可通过抑制炎症反应和细胞凋亡，减轻小胶质细胞诱导的神经元细胞HT22损伤。 |
| ABSTRACT： | OBJECTIVE To study the effects of peripheral blood-derived exosomes （Exo） intervened by Naozhenning （NZN） on injury of neuron cells HT22 induced by microglia BV-2 cells. METHODS Wistar rats were selected to prepare peripheral blood- derived Exo intervened by NZN （66.83 g/kg）， referred to as NZN-Exo； peripheral blood-derived Exo intervened by normal saline and piracetam （PLXT， 1.62 g/kg） were prepared using the same method， denoted as KB-Exo and PLXT-Exo respectively， and all Exo were subsequently identified. Meanwhile， BV-2 cells were stimulated with 1 μg/mL lipopolysaccharide （LPS） to prepare LPS- stimulated supernatant， and non-LPS-stimulated supernatant was prepared following the same protocol. HT22 cells were divided into four groups: KB-Exo group （treated with non-LPS-stimulated supernatant+KB-Exo）， model group （treated with LPS-stimulated supernatant+KB-Exo）， PLXT-Exo group （treated with LPS-stimulated supernatant+PLXT-Exo）， and NZN-Exo group （treated with LPS-stimulated supernatant+NZN-Exo）， with the concentration of the corresponding Exo in all groups being 50 μg/mL. After 24 hours of culture， the proliferation of HT22 cells was detected by the CCK-8 assay and EdU assay； the apoptosis of HT22 cells was detected； the microstructure of HT22 cells was observed； the contents of interleukin-1β （IL-1β）， IL-10， nuclear factor-κB （NF- κB）， and tumor necrosis factor-α （TNF-α） in HT22 cells were measured， as well as the expression levels of TNF-α， NOD-like receptor thermal protein domain associated protein 3 （NLRP3）， Caspase-1， B-cell lymphoma-2（ Bcl-2）， and Bcl-2-associated X protein （Bax）. RESULTS KB-Exo， PLXT-Exo and NZN-Exo were successfully prepared， and all Exo exhibited typical cup-shaped contours and membrane-enclosed characteristics. Compared with KB-Exo group， model group showed significantly decreased cell proliferation rates （detected by CCK-8 and EdU）， intracellular IL-10 levels， and Bcl-2 protein expression levels （P＜0.05）； while the cell apoptosis rate， intracellular levels of IL-1β， TNF-α， and NF-κB， as well as the expression levels of NLRP3， TNF-α， Caspase-1， and Bax proteins were significantly increased （P＜0.05）. Additionally， in the model group， the cells showed volume swelling， incomplete cell membrane， nucleolar rupture， significant swelling and deformation of mitochondria， and severe vacuolization. Compared with model group， the above quantitative indicators in the PLXT-Exo group and NZN-Exo group were significantly reversed （P＜0.05）， with large and round cell nuclei， intact nuclear membranes， and reduced mitochondrial vacuolization. CONCLUSIONS Peripheral blood-derived Exo intervened by naozhenning can alleviate the injury of neuronal cells HT22 by inhibiting inflammatory responses and cell apoptosis. |
| 期刊： | 2025年第36卷第19期 |
| 作者： | 高丽；赵乐；武立雅；张唯依；李难；魏楠楠；王永辉 |
| AUTHORS： | GAO Li，ZHAO Le，WU Liya，ZHANG Weiyi，LI Nan，WEI Nannan，WANG Yonghui |
| 关键字： | 脑震宁；小胶质细胞；神经元细胞；外泌体；凋亡；炎症 |
| KEYWORDS： | Naozhenning； microglia； neuronal cells； exosomes； apoptosis； inflammation |
| 阅读数： | 5 次 |
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