Department of Neurosurgery, Zhujiang Hospital, Southern Medical University, The National Key Clinical Specialty, The Engineering Technology Research Center of Education Ministry of China, Guangdong Provincial Key Laboratory on Brain Function Repair and Regeneration, Guangzhou, 510282, China.
J Neuroinflammation. 2019 Jan 15;16(1):8. doi: 10.1186/s12974-019-1396-5.
Activated microglia-mediated neuroinflammation has been regarded as an underlying key player in the pathogenesis of subarachnoid hemorrhage (SAH)-induced early brain injury (EBI). The therapeutic potential of bone marrow mesenchymal stem cells (BMSCs) transplantation has been demonstrated in several brain injury models and is thought to involve modulation of the inflammatory response. The present study investigated the salutary effects of BMSCs on EBI after SAH and the potential mechanism mediated by Notch1 signaling pathway inhibition.
The Sprague-Dawley rats SAH model was induced by endovascular perforation method. BMSCs (3 × 10 cells) were transplanted intravenously into rats, and N-[N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT), a Notch1 activation inhibitor, and Notch1 small interfering RNA (siRNA) were injected intracerebroventricularly. The effects of BMSCs on EBI were assayed by neurological score, brain water content (BWC), blood-brain barrier (BBB) permeability, magnetic resonance imaging, hematoxylin and eosin staining, and Fluoro-Jade C staining. Immunofluorescence and immunohistochemistry staining, Western blotting, and quantitative real-time polymerase chain reaction were used to analyze various proteins and transcript levels. Pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay.
BMSCs treatment mitigated the neurobehavioral dysfunction, BWC and BBB disruption associated with EBI after SAH, reduced ionized calcium binding adapter molecule 1 and cluster of differentiation 68 staining and interleukin (IL)-1 beta, IL-6 and tumor necrosis factor alpha expression in the left hemisphere but concurrently increased IL-10 expression. DAPT or Notch1 siRNA administration reduced Notch1 signaling pathway activation following SAH, ameliorated neurobehavioral impairments, and BBB disruption; increased BWC and neuronal degeneration; and inhibited activation of microglia and production of pro-inflammatory factors. The augmentation of Notch1 signal pathway agents and phosphorylation of nuclear factor-κB after SAH were suppressed by BMSCs but the levels of Botch were upregulated in the ipsilateral hemisphere. Botch knockdown in BMSCs abrogated the protective effects of BMSCs treatment on EBI and the suppressive effects of BMSCs on Notch1 expression.
BMSCs treatment alleviated neurobehavioral impairments and the inflammatory response in EBI after SAH; these effects may be attributed to Botch upregulation in brain tissue, which subsequently inhibited the Notch1 signaling pathway.
被激活的小胶质细胞介导的神经炎症被认为是蛛网膜下腔出血（SAH）诱导的早期脑损伤（EBI）发病机制中的关键因素。骨髓间充质干细胞（BMSCs）移植的治疗潜力已在几种脑损伤模型中得到证实，其被认为涉及炎症反应的调节。本研究旨在探讨 BMSCs 对 SAH 后 EBI 的有益作用，以及 Notch1 信号通路抑制介导的潜在机制。
采用血管内穿孔法建立 Sprague-Dawley 大鼠 SAH 模型。将 3×105个 BMSCs 经静脉注入大鼠体内，并用 Notch1 激活抑制剂 N-[N-(3,5-二氟苯乙酰基)-L-苯丙氨酸]-S-苯基甘氨酸叔丁酯（DAPT）和 Notch1 小干扰 RNA（siRNA）经侧脑室注射。通过神经功能评分、脑水含量（BWC）、血脑屏障（BBB）通透性、磁共振成像、苏木精和伊红染色以及氟来明 C 染色检测 BMSCs 对 EBI 的影响。免疫荧光和免疫组织化学染色、Western blot 和实时定量聚合酶链反应用于分析各种蛋白质和转录水平。通过酶联免疫吸附试验测量促炎细胞因子。
BMSCs 治疗减轻了 SAH 后 EBI 相关的神经行为功能障碍、BWC 和 BBB 破坏，减少了左半脑的离子钙结合接头蛋白 1 和分化簇 68 染色以及白细胞介素（IL）-1β、IL-6 和肿瘤坏死因子α的表达，但同时增加了 IL-10 的表达。SAH 后给予 DAPT 或 Notch1 siRNA 可减少 Notch1 信号通路的激活，改善神经行为障碍和 BBB 破坏；增加 BWC 和神经元变性；并抑制小胶质细胞的激活和促炎因子的产生。BMSCs 可抑制 SAH 后核因子-κB 的磷酸化和 Notch1 信号通路的激活，但同侧半球的 Botch 表达上调。BMSCs 中 Botch 的敲低消除了 BMSCs 治疗对 EBI 的保护作用，并抑制了 BMSCs 对 Notch1 表达的抑制作用。
BMSCs 治疗减轻了 SAH 后 EBI 的神经行为障碍和炎症反应；这些作用可能归因于脑组织中 Botch 的上调，随后抑制了 Notch1 信号通路。
