目的 通過網(wǎng)絡藥理學及實驗驗證探討肺筋草對脂多糖（LPS）誘導大鼠急性肺損傷（ALI）的防治作用及機制。方法 查閱肺筋草相關文獻結(jié)合SwissADME數(shù)據(jù)庫對肺筋草的活性成分進行篩選，通過Swiss Target Prediction平臺預測肺筋草活性成分作用的潛在靶點，通過GeneCards數(shù)據(jù)庫、DrugBank數(shù)據(jù)庫、OMIM數(shù)據(jù)庫檢索與ALI相關的基因，與化合物的作用靶點進行比對，繪制韋恩圖得到交集靶點。STRING數(shù)據(jù)庫中輸入交集靶點信息，選擇綜合得分在0.9以上的靶點，利用Cytoscape 3.9.0軟件獲得蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）網(wǎng)絡，利用Metascape數(shù)據(jù)庫對關鍵靶點進行京都基因與基因組百科全書（KEGG）通路富集分析，篩選得到有關ALI的通路，通過Cytoscape 3.9.0軟件繪制“關鍵成分-靶點-通路”網(wǎng)絡圖。將SD大鼠隨機分為對照組、模型組、地塞米松（陽性藥，5 mg · kg^-1）組和肺筋草水提物（ ASWE ）高、中、低劑量（ 9.00、4.50、2.25 g · kg^-1）組，連續(xù)7 d對大鼠進行預防性ig給藥，7 d后ip脂多糖（LPS，5 mg· kg^-1）誘導ALI模型，對肺組織進行病理學觀察分析，試劑盒法檢測大鼠血清中的炎癥因子以及肺組織中氧化應激指標的表達水平，實時熒光定量PCR（qRT-PCR）檢測肺組織絲裂原活化蛋白激酶1（MAPK1）、Janus激酶2（JAK2）、核因子κB（NF-κB）、神經(jīng)細胞瘤鼠肉瘤癌基因（Nras）、信號傳導及轉(zhuǎn)錄激活蛋白（STAT）、內(nèi)磺肽α（Ensa）mRNA水平，Western blotting實驗檢測肺組織NF-κB、JAK2蛋白表達。結(jié)果 篩選出肺筋草的主要活性成分10個，包括阿魏酸甲酯、香豆酸、熊果酸等，得到肺筋草作用于ALI關鍵靶點189個，MAPK1、MAPK8、STAT3、AKT1、PIK3R1、TP53、ESR1、RELA等為核心靶點，KEGG富集分析結(jié)果顯示肺筋草可能通過MAPK、NF-κB、JAK-STAT等信號通路發(fā)揮藥效作用。與模型組比較，ASWE各劑量組炎癥細胞浸潤等肺組織病理表現(xiàn)明顯改善；中、高劑量組腫瘤壞死因子-α（TNF-α）水平顯著降低（P＜0.05），中劑量組白細胞介素（IL）-1β水平顯著降低（P＜0.05），各劑量組IL-6水平均有降低趨勢；高、中劑量組超氧化物歧化酶（SOD）、谷胱甘肽過氧化物酶（GSH-Px）水平明顯升高（P＜0.05）；低、中劑量組Nras、STAT3、MAPK1、JAK2、Ensa、NF-κB mRNA水平顯著降低（P＜0.05，0.01）；中劑量組NF-κB、JAK2蛋白的表達量顯著下降（P＜0.05、0.01）。結(jié)論 肺筋草可以抑制炎癥介質(zhì)的過度分泌，調(diào)節(jié)氧化應激平衡而對ALI有防治作用，其作用機制可能與抑制MAPK1、NF-κB、STAT3等基因表達，從而抑制NF-κB/JAK等炎癥信號通路相關。

Objective To explore the mechanism of the preventive and therapeutic effects of Aletris spicata on acute lung injury (ALI) induced by lipopolysaccharide (LPS) in rats through network pharmacology and experimental verification. Methods Review the relevant literature on A. spicata and use the SwissADME database to screen the active components of A. spicata. Predict the potential targets of the active components of A. spicata using the Swiss Target Prediction platform. Use the GeneCards database, DrugBank database, and OMIM database to search for genes related to ALI, compare them with the targets of the active compounds, and draw a Venn diagram to obtain the intersection targets. Input the intersection target information into the String database, select targets with a comprehensive score of 0.9 or above, and use Cytoscape 3.9.0 software to obtain a protein-protein interaction (PPI) network. Use the Metascape database to perform Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on key targets, and select pathways related to ALI. Draw a "key component-target-pathway" network diagram using Cytoscape 3.9.0 software. Randomly divide SD rats into a control group, a model group, a dexamethasone (positive drug, 5 mg· kg^-1) group, and A. spicata water extract (ASWE) high, medium, and low dose (9.00, 4.50, 2.25 g· kg^-1) groups. Continuously administer the rats with the drugs by ig for 7 days, ip LPS (5 mg· kg^-1) to induce the ALI model 7 days later, and observe and analyze the pathological changes in the lung tissue. Detect the expression levels of inflammatory factors in the serum and oxidative stress indicators in the lung tissue using kit assays, and detect the mRNA levels of β -activated protein kinase 2 (TAK2), Janus kinase 2 (JAK2), nuclear factor kappa B (NF-κB), Nras, signal transducer and activator of transcription (STAT), and Ensa in the lung tissue using real-time quantitative PCR (qRT-PCR), and detect the expression levels of NF-κB and JAK2 proteins in the lung tissue using Western blotting experiments. Results Ten main active compounds of A. spicata were screened out, including methyl salicylate, coumarin, ursolic acid, etc. A total of 189 key targets were identified for A. spicata acting on ALI, with MAPK1, MAPK8, STAT3, AKT1, PIK3R1, TP53, ESR1, and RELA as core targets. The KEGG enrichment analysis showed that A. spicata may exert its therapeutic effects through MAPK, NF-κB, and JAK-STAT signaling pathways. Compared with the model group, the pulmonary tissue pathological manifestations of inflammatory cell infiltration in the ASWE treatment groups were significantly improved. The TNF-α level was significantly lower in the middle and high-dose groups (P < 0.05), and the IL-1β level was significantly lower in the middle-dose group (P < 0.05). The IL-6 level showed a downward trend in all dosage groups; the SOD and GSH-Px levels were significantly higher in the high and middle-dose groups (P < 0.05); The Nras, STAT3, MAPK1, JAK2, Ensa, and NF-κB mRNA levels were significantly lower in the high and middle-dose groups (P < 0.05, 0.01). The expression levels of NF-κB and JAK2 proteins were significantly lower in the middle-dose group (P < 0.05, 0.01). Conclusions A. spicata can inhibit the excessive secretion of inflammatory mediators, regulate the balance of oxidative stress and have a preventive and therapeutic effect on ALI. The mechanism may be related to inhibiting the expression of MAPK1, NF-κB, STAT3 and other genes, thereby inhibiting the expression of NF-κB/JAK and other inflammatory signaling pathways.

R285.5

貴州省中醫(yī)藥管理局中醫(yī)藥、民族醫(yī)藥科學技術研究課題(QZYY-2022-015);貴州省科學技術基金(黔科合基礎-ZK[2021]一般515);貴州省教育廳2023年度自然科學研究項目(黔教技[2023]069號);貴州省特色食藥材高效綜合利用科技創(chuàng)新人才團隊建設(黔科合平臺人才-CXTD[2023]020)