补阳还五汤干预脑缺血大鼠视束损伤的作用及关键药理途径的生物信息学分析

Effects of Buyang Huanwu Decoction on optic tract injury in cerebral ischemia rats and bioinformatics analysis of key pharmacological pathways involved

目的 观察补阳还五汤干预脑缺血大鼠视束损伤的作用,并探讨补阳还五汤干预脑缺血视束损伤的有效成分、核心靶点及调控机制。方法 采用线栓法制备右侧大脑中动脉栓塞致脑缺血的大鼠模型。将24只SD雄性大鼠随机分为假手术组、模型组和补阳还五汤组(16.1 g/kg),每组各8只,连续灌胃30 d。采用横向弛豫时间定量成像(T2 mapping)检测视束损伤,弥散张量成像(DTI)结合弥散张量纤维束成像(DTT)分析视束微结构的改变。采用TCMSP、PubMed和中国知网数据库获取补阳还五汤有效成分及对应靶点,基于GeneCards、OMIM、DisGeNET、DrugBank数据库筛选缺血性视束神经损伤相关靶点,导入Cytoscape软件绘制活性成分-治疗靶点网络,运用STRING数据库构建蛋白互作网络(PPI),并进行蛋白簇聚类分析,基于Metascape数据库对核心蛋白簇进行KEGG通路分析。结果 T2 mapping显示,与假手术组相比,模型组大鼠视束区域rT2值升高(P<0.01);与模型组相比,补阳还五汤组大鼠视束区域rT2值降低(P<0.01)。DTI结果显示,与假手术组比较,模型组大鼠及补阳还五汤组大鼠视束区域rFA值降低(P<0.01),rAD、rRD值增高(P<0.01);与模型组比较,补阳还五汤组大鼠视束区域rFA值升高(P<0.01),rAD、rRD值降低(P<0.01)。DTT结果显示,与假手术组比较,模型组与补阳还五汤组大鼠视束区域神经纤维的相对平均纤维束长度和密度均降低(P<0.05,P<0.01);与模型组相比较,补阳还五汤组大鼠视束区域神经纤维相对平均纤维束长度和密度均增加(P<0.05,P<0.01)。从补阳还五汤中筛选到与缺血性视束损伤相关的56个活性成分和400个潜在有效靶点,主要活性成分为黄芪甲苷Ⅳ、芍药苷和阿魏酸等,PPI核心基因为AKT1、TNF、TP53、CASP3、JUN等,PPI核心蛋白簇通过KEGG通路功能富集得到9条相关信号通路,在神经元、星型胶质细胞、小胶质细胞、少突胶质细胞中调控多种生物过程和分子功能,主要涉及磷脂酰肌醇3激酶(PI3K)-蛋白激酶B(Akt)信号通路、Toll样受体信号通路等,涉及细胞凋亡、能量代谢及炎性因子激活等重要生理生物过程。结论 补阳还五汤可减轻脑缺血大鼠视束神经损伤,促进视束超微结构损伤后的重塑,其干预缺血性视束损伤的作用可能是通过调控PI3K-Akt、丝裂原活化蛋白激酶、肿瘤坏死因子等多条信号通路影响神经元及胶质细胞功能来实现的。

Objective We aimed to observe the effects of Buyang Huanwu Decoction(BYHWD) on optic tract injury of cerebral ischemia rats and to explore the effective components, core targets, and regulatory mechanism of BYHWD on cerebral ischemia rats’ optic tract lesions.Methods A suture occlusion method was adopted to construct a rat model of cerebral ischemia induced by right middle cerebral artery embolization.In total, 24 male SD rats were randomly divided into the sham group, the model group, and the BYHWD group, eight rats in each group. The rats in BYHWD group were intragastrically injected with BYHWD for 30 days(16.1 g/kg).Optic tract damage was detected by T2 mapping, and changes in the optical tract microstructure were analyzed by diffusion tensor imaging(DTI) and diffusion tensor tractography(DTT).The active components and targets of BYHWD were collected from the TCMSP, PubMed, and CNKI databases, and the targets of ischemic optic tract injury were screened from the GeneCards, OMIM, DisGeNET, and DrugBank databases.Cytoscape was used to construct an active component-therapeutic target network.A protein-protein interaction(PPI) network was constructed, and protein cluster analysis was performed with the STRING platform.KEGG pathway analysis of core protein clusters was performed based on the Metascape database.Results T2 mapping showed that the rT2 value in the optic tract region was higher in the model group compared with the sham group(P<0.01).The rT2 value in the optic tract region of the BYHWD group was lower compared with the model group(P<0.01). The DTI result showed that the rFA value in the optic tract region of both the model group and the BYHWD group were lower(P<0.01) and the rAD and rRD were higher compared with the sham group(P<0.01).The rFA value was higher(P<0.01) and rAD and rRD were lower(P<0.01) in the BYHWD group compared with the model group.DTT analysis showed that the relative average length and density of nerve fibers in the optic tract region of the model group and BYHWD group were lower(P<0.05,P<0.01) compared with the sham group, while the relative average length and density of nerve fibers in the optic tract region of the BYHWD group were higher(P<0.05, P<0.01) compared with the model group.In total, 56 active components and 400 potential effective targets related to ischemic optic tract injury were screened from BYHWD.The main active components were Astragaloside Ⅳ, paeoniflorin, and ferulic acid.PPI core genes included AKT1, TNF, TP53, CASP3, and JUN.Nine related signaling pathways were obtained by KEGG pathway enrichment analysis of PPI core protein clusters.They regulate a variety of biological processes and molecular functions in neurons, astrocytes, microglia, and oligodendrocytes, mainly involving the PI3 K-Akt signaling pathway, Toll-like receptor signaling pathway the apoptosis pathway, energy metabolism, and inflammatory factor-related pathways.Conclusion BYHWD can alleviate optic tract nerve injury and promote remodeling of the optic tract ultrastructure after cerebral ischemia in rats.Its effects on ischemic optic tract injury may be mediated by multiple signaling pathways, including the PI3 K-Akt signaling pathway, the MAPK signaling pathway, and the TNF signaling pathway, affecting neuronal and glial cell functions.