Department of Neurology, Hebei General Hospital, 348 West Heping Road, Shijiazhuang 050051, Hebei, China.
NYU School of Global Public Health, 708 Broadway, New York, NY, USA.
Brain Res Bull. 2022 Nov;190:244-255. doi: 10.1016/j.brainresbull.2022.10.011. Epub 2022 Oct 14.
Ligustrazine is a traditional Chinese herbal medicine that has long been used to treat cerebral ischemic disorders. However, the molecular mechanisms of ligustrazine in cerebral ischemia/reperfusion (I/R) damage have not been clear elucidated. The aim of this study was to examine the neuroprotective mechanisms of ligustrazine in cerebral I/R.
9 C57BL/6 mice were randomly divided to three groups: Sham group (n = 3), Middle cerebral artery occlusion (MCAO) group (n = 3), and MCAO + Ligustrazine group (n = 3). The neurological deficit score was evaluated, the cerebral infarct volume was measured by triphenylterazolium chloride (TTC) staining. Differentially expressed (DE) messenger RNAs (mRNAs), long noncoding RNAs (lncRNAs) and circular RNAs (circRNAs) were analyzed using the R package DEseq2 based on P-value < 0.05 and Log2 |fold change (FC)| ≥ 2 in sham group vs MCAO group and MCAO group vs ligustrazine group by high-throughput sequencing. Function enrichment analysis, the protein-protein interaction (PPI) of neurogenesis related genes were performed. The neurogenesis related competitive endogenous RNA (ceRNA) network was constructed.
The expression of circ_0008146 was considerably higher in the MCAO group than the Sham group, and ligustrazine treatment markedly decreased the expression of circ_0008146 in MCAO. Next, the circ_0008146 ceRNA network was established, including circ_0008146-miR-709-Cx3cr1 ceRNA network. Besides, real time quantitative polymerase chain reaction (RT-qPCR) assay identified that miR-709 expression was considerably lower and Cx3cr1 expression was higher in the MCAO group than Sham group, and ligustrazine treatment markedly increased the miR-709 expression and reduced Cx3cr1 expression in MCAO. Further, silencing of circ_0008146 inhibited the concentration of Interleukin 6 (IL-6), Tumor Necrosis Factor alpha (TNF-α) and reduced neuron cell death and up-regulated miR-709 expression and down-regulated Cx3cr1 expression in Lipopolysaccharide (LPS) induced BV-2 cells. Dual-Luciferase reporter gene assay verified that circ_0008146 targeted miR-709.
Ligustrazine targets circ_0008146/miR-709/Cx3cr1 axis to inhibit cell apoptosis and inflammation after cerebral ischemia/reperfusion injury.
川芎嗪是一种传统的中草药，长期以来一直用于治疗脑缺血性疾病。然而，川芎嗪在脑缺血/再灌注（I/R）损伤中的分子机制尚不清楚。本研究旨在探讨川芎嗪在脑 I/R 中的神经保护机制。
将 9 只 C57BL/6 小鼠随机分为三组：假手术组（n=3）、大脑中动脉闭塞组（MCAO 组，n=3）和 MCAO+川芎嗪组（n=3）。通过神经功能缺损评分、三苯基四唑氯（TTC）染色测量脑梗死体积。使用基于 P 值<0.05 和 sham 组与 MCAO 组、MCAO 组与川芎嗪组之间 Log2 |fold change (FC)|≥2 的高通量测序的 R 包 DEseq2 分析差异表达（DE）信使 RNA（mRNA）、长链非编码 RNA（lncRNA）和环状 RNA（circRNA）。进行功能富集分析，对神经发生相关基因的蛋白质-蛋白质相互作用（PPI）进行分析。构建神经发生相关竞争性内源性 RNA（ceRNA）网络。
MCAO 组 circ_0008146 的表达明显高于 Sham 组，而川芎嗪治疗显著降低了 MCAO 中 circ_0008146 的表达。接下来，建立了 circ_0008146 的 ceRNA 网络，包括 circ_0008146-miR-709-Cx3cr1 ceRNA 网络。此外，实时定量聚合酶链反应（RT-qPCR）检测发现，MCAO 组 miR-709 的表达明显低于 Sham 组，Cx3cr1 的表达明显高于 Sham 组，而川芎嗪治疗显著增加了 MCAO 中 miR-709 的表达并降低了 Cx3cr1 的表达。进一步，沉默 circ_0008146 抑制了脂多糖（LPS）诱导的 BV-2 细胞中白细胞介素 6（IL-6）和肿瘤坏死因子 alpha（TNF-α）的浓度，并上调了 miR-709 的表达和下调了 Cx3cr1 的表达。双荧光素酶报告基因实验证实 circ_0008146 靶向 miR-709。
川芎嗪通过靶向 circ_0008146/miR-709/Cx3cr1 轴抑制脑缺血再灌注损伤后的细胞凋亡和炎症。
