目的 采用網(wǎng)絡(luò)藥理學(xué)、分子對(duì)接及實(shí)驗(yàn)驗(yàn)證的方法，探討大黃治療酒精性肝炎的作用機(jī)制，為臨床應(yīng)用提供依據(jù)。方法 運(yùn)用中藥系統(tǒng)藥理學(xué)數(shù)據(jù)庫(kù)與分析平臺(tái)（TCMSP）及查閱國(guó)內(nèi)外相關(guān)文獻(xiàn)收集大黃活性成分，通過(guò)GeneCards、OMIM、TTD數(shù)據(jù)庫(kù)收集酒精性肝炎靶點(diǎn)，將成分-疾病靶點(diǎn)取交集基因，運(yùn)用Cytoscape軟件構(gòu)建大黃-活性成分-酒精性肝炎-靶基因網(wǎng)絡(luò)圖，在STRING平臺(tái)制作蛋白質(zhì)-蛋白質(zhì)相互作用（PPI）網(wǎng)絡(luò)，通過(guò)Metascape平臺(tái)對(duì)交集靶點(diǎn)進(jìn)行富集分析，獲得靶點(diǎn)功能及通路。運(yùn)用Vina等軟件對(duì)大黃的關(guān)鍵活性成分與關(guān)鍵靶點(diǎn)進(jìn)行分子對(duì)接。建立酒精性肝炎小鼠模型，設(shè)置對(duì)照組、模型組、氫化可的松（20.8 mg·kg^-1）組和蘆薈大黃素低、中、高劑量（20、40、80 mg·kg^-1）組，觀察對(duì)各組小鼠肝臟指數(shù)影響；試劑盒法測(cè)定血清中天冬氨酸氨基轉(zhuǎn)移酶（AST）、丙氨酸氨基轉(zhuǎn)移酶（ALT）水平；蘇木精-伊紅（HE）染色和油紅O染色觀察小鼠肝組織病理學(xué)以及脂質(zhì)積累情況；Western blotting檢測(cè)小鼠肝組織中NOD樣受體熱蛋白結(jié)構(gòu)域相關(guān)蛋白3（NLRP3）、白細(xì)胞介素-18（IL-18）、IL-1β、剪切型半胱天冬酶-3（cleaved Caspase-3）、半胱天冬酶-3（Caspase-3）和半胱天冬酶-1（Caspase-1）的蛋白表達(dá)。結(jié)果 大黃16個(gè)活性成分對(duì)應(yīng)50個(gè)靶點(diǎn)，酒精性肝炎對(duì)應(yīng)6 888個(gè)靶點(diǎn)，二者取交集獲得45個(gè)靶點(diǎn)。其中IL-1β、CASP3、GSDMD、骨髓細(xì)胞瘤癌基因（MYC）、前列腺素內(nèi)過(guò)氧化物合酶2（PTGS2）以及蛋白激酶Cα（PRKCA）與核心成分蘆薈大黃素分子對(duì)接結(jié)果良好。基因本體（GO）功能富集分析和京都基因與基因組百科全書(shū)（KEGG）通路富集分析發(fā)現(xiàn)，細(xì)胞凋亡途徑可能是大黃治療酒精性肝炎的重要途徑。體內(nèi)實(shí)驗(yàn)結(jié)果顯示，蘆薈大黃素能夠緩解小鼠酒精性肝炎，并顯著降低肝臟指數(shù)（P＜0.001）和AST、ALT水平（P＜0.05）。HE染色與油紅O顯示，與模型組相比，蘆薈大黃素低、中、高劑量組恢復(fù)了肝組織結(jié)構(gòu)損傷，降低了炎癥細(xì)胞浸潤(rùn)，恢復(fù)了肝索排列紊亂狀態(tài)以及減少了脂滴堆積和脂肪變性。Western blotting結(jié)果顯示，與模型組相比，氫化可的松組以及蘆薈大黃素低、中、高劑量組NLRP3、IL-18、IL-1β、GSDMD、cleaved Caspase-3、Caspase-3和Caspase-1蛋白表達(dá)水平均有降低，其中蘆薈大黃素中劑量組IL-1β、GSDMD、Caspase-3和Caspase-1蛋白表達(dá)水平顯著降低（P＜0.05、0.01）。結(jié)論 大黃治療酒精性肝炎可通過(guò)多成分、多靶點(diǎn)、多通路途徑來(lái)發(fā)揮作用，其中蘆薈大黃素可減輕小鼠酒精性肝炎，其機(jī)制可能與抑制Caspase-1、Caspase-3介導(dǎo)的經(jīng)典細(xì)胞焦亡和凋亡通路有關(guān)。

Objective To explore the mechanism of Rhei Radix et Rhizoma in the treatment of alcoholic hepatitis by means of network pharmacology, molecular docking and experimental verification, and to provide evidence for the clinical application of Rhei Radix et Rhizoma in the treatment of alcoholic hepatitis.Methods The active components of Rhei Radix et Rhizoma were collected using TCMSP and relevant literature at home and abroad. The targets of alcoholic hepatitis were collected by GeneCards, OMIM and TTD databases, and the intersection genes of constituents and disease targets were selected. The Rhei Radix et Rhizoma-active ingredientalcoholic hepatitis-target gene network map was constructed by Cytoscape software. The protein interaction network was made on the STRING platform. The intersection targets were enriched and analyzed by Metascape platform to obtain the target functions and pathways. The key active components of Rhei Radix et Rhizoma were docked with key targets by Vina and other software. A mouse model of alcoholic hepatitis was established, and control group, model group, cortisone (20.8 mg·kg^-1) group and aloe emodin lowdose, medium-dose and high-dose (20, 40, 80 mg·kg^-1) groups were set up to observe the effects on liver index of mice in each group. Serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were determined by kit method. Liver histopathology and lipid accumulation were observed by HE staining and oil red O staining. Western blotting analysis of NLRP3, IL-18, IL-1β, cleaved Caspase-3, Caspase-3, and Caspase-1 protein expression in mouse liver tissues.Results A total of 16 active components of Rhei Radix et Rhizoma correspond to 50 targets, alcohol hepatitis correspond to 6 888 targets, and 45 targets were obtained by intersection of the two. IL-1β, CASP3, MYC, PTGS2 and PRKCA had good docking results with aloe emodin. GO and KEGG results showed that apoptosis pathway may be an important pathway for Rhei Radix et Rhizoma treatment of alcoholic hepatitis. The results of in vivo experiment showed that aloe emodin could alleviate alcoholic hepatitis in mice, and significantly reduce liver index (P<0.001) and AST and ALT levels (P<0.05). HE staining and oil red O showed that, compared with the model group, aloe emodin groups restored the damage of dry tissue structure, reduced inflammatory cell infiltration, restored the disorder of hepatic cord arrangement, and reduced lipid droplet accumulation and steatosis. Western blotting results showed that compared with the model group, protein expression levels of NLRP3, IL-18, IL-1β, GSDMD, cleaved Caspase-3, Caspase-3, and Caspase-1 were decreased in cortisone group and aloe albumin dose groups. The expression levels of IL-1β, GSDMD, Caspase-3 and Caspase-1 protein in aloe emodin medium dose group were significantly decreased (P<0.05).Conclusion Rhei Radix et Rhizoma can treat alcoholic hepatitis through multicomponent, multi-target and multi-pathway pathways, among which aloe emodin can alleviate alcoholic hepatitis in mice, and its mechanism may be related to the inhibition of classic cell pyrodeath and apoptosis pathways mediated by Caspase-1 and Caspase-3.

R285.5

國(guó)家自然科學(xué)基金聯(lián)合基金重點(diǎn)支持項(xiàng)目（U21A200211）；國(guó)家資助博士后研究人員計(jì)劃資助（GZC20231759）