[關鍵詞]
[摘要]
目的 旨在基于網絡藥理學篩選絞股藍皂苷XVII改善腦出血后神經損傷的潛在靶點與通路,并通過體內外實驗驗證其作用及對磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B1(Akt)信號通路的調控機制。方法 通過網絡藥理學平臺篩選絞股藍皂苷XVII與腦出血的交集靶點,構建蛋白質相互作用網絡(PPI)并進行關鍵通路的富集分析;通過分子對接驗證靶點結合能,利用細胞實驗、動物實驗探索絞股藍皂苷XVII治療腦出血的分子機制。通過氯化血紅素誘導SH-SY5Y細胞模擬腦出血模型后,使用CCK-8測細胞活性、熒光探針檢測細胞內活性氧(ROS)水平;實時熒光定量聚合酶鏈式反應(RT-qPCR)檢測腫瘤壞死因子(TNF)、白細胞介素(IL)-1β、胰島素樣生長因子1(IGF1)、表皮生長因子受體(EGFR)、磷酸肌醇3激酶調控亞基1(PIK3R1)的mRNA表達水平。將60只SD大鼠隨機分成假手術組、模型組、絞股藍皂苷XVII(20、40 mg/kg)組,采用自體血注射法構建大鼠腦出血模型,評估大鼠神經功能評分,干/濕質量法檢測腦組織含水量,HE染色及尼氏檢測血腫周圍腦組織病理變化,試劑盒檢測氧化應激指標超氧化物歧化酶(SOD)、丙二醛(MDA)及炎癥因子TNF-α、IL-1β水平,Western blotting檢測p-PI3K、PI3K、p-Akt、Akt蛋白表達水平。結果 網絡藥理學篩選出166個共同靶點,PPI分析揭示核心靶點(PIK3R1、Akt1等),KEGG富集提示PI3K/Akt為核心通路。體外實驗表明,絞股藍皂苷XVII顯著提高細胞存活率、抑制ROS生成,并下調TNF、IL-1β、IGF1、EGFR、PIK3R1 mRNA表達。體內實驗證實,絞股藍皂苷XVII能夠降低大鼠神經功能評分、減少腦水腫,改善血腫周圍腦組織病理損傷,提高SOD活性、降低MDA水平及炎癥因子TNF-α、IL-1β水平,顯著上調PI3K、Akt的磷酸化水平(P<0.05、0.01、0.001)。結論 絞股藍皂苷XVII可能通過調控PI3K/Akt通路改善腦出血神經損傷。
[Key word]
[Abstract]
Objective To screen potential targets and pathways of gypenoside XVII in ameliorating neurological impairment following intracerebral hemorrhage through network pharmacology approaches, and further validate its therapeutic effects along with the regulatory mechanisms on the PI3K/Akt signaling pathway via both in vitro and in vivo experiments. Methods Network pharmacology platforms were used to screen common targets of gypenoside XVII and intracerebral hemorrhage, followed by PPI network construction and pathway enrichment analysis. Molecular docking was performed to validate target binding activity, while cellular and animal experiments were conducted to explore the molecular mechanism of gypenoside XVII in intracerebral hemorrhage treatment. Hemin-induced SH-SY5Y cell model mimicking intracerebral hemorrhage was established, and cell viability was assessed using the CCK-8 assay. Intracellular reactive oxygen species (ROS) levels were measured via fluorescent probes. RT-qPCR was used to detect mRNA expression levels of TNF, IL-1β, IGF1, EGFR, and PIK3R1. Sixty Sprague-Dawley (SD) rats were randomly divided into sham operation group, model group, and gypenoside XVII (20, 40 mg/kg) group. Intracerebral hemorrhage model was constructed using autologous blood injection. Neurological function scores, cerebral edema (dry-wet weight method), histopathological changes in perihematomal brain tissue (HE and Nissl staining), oxidative stress markers (SOD, MDA), inflammatory cytokines (TNF-α, IL-1β), and protein expression levels of p-PI3K, PI3K, p-Akt, and Akt were evaluated. Results Network pharmacology identified 166 common targets, with PPI analysis revealing core targets (PIK3R1, Akt1), and KEGG enrichment highlighting the PI3K/Akt pathway as central. In vitro, gypenoside XVII significantly increased cell viability, inhibited ROS generation, and downregulated TNF, IL-1β, IGF1, EGFR, PIK3R1 mRNA expression. In vivo, gypenoside XVII reduced neurological deficits, decreased cerebral edema, alleviated perihematomal pathological damage, enhanced SOD activity, lowered MDA and inflammatory cytokine levels (TNF-α, IL-1β), and upregulated phosphorylation levels of PI3K and Akt (P < 0.05, 0.01, 0.001). Conclusion Gypenoside XVII may improve neurological injury after intracerebral hemorrhage by modulating the PI3K/Akt pathway.
[中圖分類號]
R286.1
[基金項目]
江蘇省衛(wèi)生健康委科研項目(K2023044);鹽城市科技項目基礎研究計劃(YCBK2024072)