Chaiqin chengqi decoction alleviates severity of acute pancreatitis via inhibition of TLR4 and NLRP3 inflammasome: Identification of bioactive ingredients via pharmacological sub-network analysis and experimental validation
Chaiqin chengqi decoction alleviates severity of acute pancreatitis via inhibition of TLR4 and NLRP3 inflammasome: Identification of bioactive ingredients via pharmacological sub-network analysis and experimental validation
复制标题
柴芩承气汤通过抑制TLR4和NLRP3炎性体减轻急性胰腺炎的严重程度：通过药理学子网络分析和实验验证鉴定生物活性成分
DOI：
10.1016/j.phymed.2020.153328
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发表时间：
2020-12-01
期刊：
影响因子：
7.9
通讯作者：
Xia, Qing
中科院分区：
文献类型：
--
作者：
Wen, Yongjian;Han, Chenxia;Xia, Qing
背景：柴芩承气汤是由大承气汤衍生而来的中药方剂。中西医结合治疗急性胰腺炎已有30多年的历史。虽然CQCQD具有良好的临床疗效，但对其生物活性成分，它们如何与不同的治疗靶点相互作用以及产生抗炎作用的途径知之甚少。目的：Toll样受体4（TLR 4）和含核甘酸结合寡聚化结构域样受体家族pyrin结构域3（NLRP 3）炎性小体介导的促炎信号传导途径，在AP中起决定胰腺损伤和全身炎症程度的核心作用。方法：采用UPLC-QTOF/MS法对CQCQD中的主要活性成分进行鉴定，并通过药理子网络分析确定CQCQD活性成分在AP中的作用靶点为TLR 4/NLRP 3。小鼠接受7次蛙皮素腹膜内注射（50 μ g/kg;每小时）以诱导AP（CER-AP），而在第3次注射蛙皮素时开始经口管饲CQCQD（5、10、15和20 μ g/kg; 3次剂量，2小时）。采用组织学和生化指标评估AP的严重程度，并采用聚合酶链反应、免疫印迹和免疫组化分析研究AP的发病机制。结果：CQCQD中主要成分为黄酮类化合物、环烯醚萜类化合物、酚类化合物、木脂素类化合物、蒽醌类化合物及相应的糖苷类化合物。子网络分析显示大黄素、大黄酸、黄芩苷和白杨素是直接调节TLR 4/NLRP 3相关蛋白TLR 4、RelA、NF-κ B和TNF-α最相关的化合物。在体内，CQCQD减轻了CER-AP的胰腺损伤和全身炎症，并与TLR 4/NLRP 3相关mRNA和蛋白的表达减少有关。大黄素、大黄酸、黄芩苷和白杨素显著减少胰腺腺泡细胞坏死，对TLR 4/NLRP 3相关mRNA的表达具有不同的抑制作用。大黄素，大黄酸和白杨素也减少了一氧化氮在巨噬细胞的生产和他们的组合有协同效应，减轻细胞死亡以及TLR 4/NLRP 3相关的蛋白的表达。结论：CQCQD衰减AP的严重程度至少部分通过抑制TLR 4/NLRP 3促炎通路。其有效成分大黄素、黄芩苷、大黄酸和白杨素有助于这些有益的效果。
Background: Chaiqin chengqi decoction (CQCQD) is a Chinese herbal formula derived from dachengqi decoction. CQCQD has been used for the management of acute pancreatitis (AP) in the West China Hospital for more than 30 years. Although CQCQD has a well-established clinical efficacy, little is known about its bioactive ingredients, how they interact with different therapeutic targets and the pathways to produce anti-inflammatory effects.Purpose: Toll-like receptor 4 (TLR4) and the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) inflammasome-mediated pro-inflammatory signaling pathways, play a central role in AP in determining the extent of pancreatic injury and systemic inflammation. In this study, we screened the bioactive ingredients using a pharmacological sub-network analysis based on the TLR4/NLRP3 signaling pathways followed by experimental validation.Methods: The main CQCQD bioactive compounds were identified by UPLC-QTOF/MS. The TLR4/NLRP3 targets in AP for CQCQD active ingredients were confirmed through a pharmacological sub-network analysis. Mice received 7 intraperitoneal injections of cerulein (50 mu g/kg; hourly) to induce AP (CER-AP), while oral gavage of CQCQD (5, 10, 15 and 20 g/kg; 3 doses, 2 hourly) was commenced at the 3rd injection of cerulein. Histopathology and biochemical indices were used for assessing AP severity, while polymerase chain reaction, Western blot and immunohistochemistry analyses were used to study the mechanisms. Identified active CQCQD compounds were further validated in freshly isolated mouse pancreatic acinar cells and cultured RAW264.7 macrophages.Results: The main compounds from CQCQD belonged to flavonoids, iridoids, phenols, lignans, anthraquinones and corresponding glycosides. The sub-network analysis revealed that emodin, rhein, baicalin and chrysin were the compounds most relevant for directly regulating the TLR4/NLRP3-related proteins TLR4, RelA, NF-kappa B and TNF-alpha. In vivo, CQCQD attenuated the pancreatic injury and systemic inflammation of CER-AP and was associated with reduced expression of TLR4/NLRP3-related mRNAs and proteins. Emodin, rhein, baicalin and chrysin significantly diminished pancreatic acinar cell necrosis with varied effects on suppressing the expression of TLR4/NLRP3-related mRNAs. Emodin, rhein and chrysin also decreased nitric oxide production in macrophages and their combination had synergistic effects on alleviating cell death as well as expression of TLR4/NLRP3-related proteins.Conclusions: CQCQD attenuated the severity of AP at least in part by inhibiting the TLR4/NLRP3 pro-inflammatory pathways. Its active ingredients, emodin, baicalin, rhein and chrysin contributed to these beneficial effects.