目的 基于网络药理学和动物实验验证探讨地黄环烯醚萜苷类（Rehmannia glutinosa iridoid glycosides，RIG）治疗2型糖尿病（type 2 diabetes，T2DM）的作用机制。方法 利用Swisstargetprediction和PharmMapper数据库预测筛选RIG相关作用靶点，OMMI、Genecard、DisGeNET数据库获取T2DM的相关靶标基因，将获得的共同靶点导入STRING数据库构建蛋白相互作用（protein-protein interaction，PPI）网络，Cytoscape软件构建“药物-成分-靶点”和核心靶点网络，通过DAVID数据库和微生信平台进行基因本体（gene ontology，GO）功能及京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）通路富集分析。采用高糖高脂喂养联合ip链脲佐菌素建立T2DM小鼠模型，将造模成功的小鼠随机分为模型组、二甲双胍（250 mg/kg）组和RIG低、高剂量（200、400 mg/kg）组，每组9只，另取10只正常小鼠作为对照组。药物干预8周，每周测定体质量、空腹血糖（fasting blood glucose，FBG）。给药结束后，分离血清，检测低密度脂蛋白胆固醇（low density lipoprotein cholesterol，LDL-C）、高密度脂蛋白胆固醇（high density lipoprotein cholesterol，HDL-C）、胰岛素（fasting insulin，FINS）水平，并计算胰岛素抵抗指数（homeostasis model assessment of insulin resistance，HOMA-IR）；采用苏木素-伊红（HE）、Masson和油红O染色观察小鼠肝脏病理变化；采用免疫组化法测定肝组织炎症因子白细胞介素-1（interleukin-1，IL-1）、IL-6、肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）的表达量；采用Western blotting法检测肝组织晚期糖基化终末产物（advanced glycation end products，AGEs）、晚期糖基化末端受体（receptor for advanced glycation end products，RAGE）蛋白表达及p-p38丝裂原活化蛋白激酶（mitogen activated protein kinase，MAPK）/p38 MAPK水平；采用qRT-PCR法检测肝组织RAGE、p38 MAPK mRNA表达。结果 共筛选得到RIG治疗T2DM潜在靶点175个，关键核心靶点有原癌基因酪氨酸蛋白激酶Src（proto-oncogene tyrosine-protein kinase Src，SRC）、表皮生长因子受体（epidermal growth factor receptor，EGFR）、信号转导和转录激活因子3（signal transducer and activator of transcription 3，STAT3）等。GO富集分析显示潜在作用靶点主要涉及炎症反应的调节等生物过程，KEGG通路分析筛选得到了277条信号通路，显示脂质和动脉粥样硬化、AGEs/RAGE信号通路和MAPK信号通路可能在治疗T2DM过程中发挥关键作用。动物实验结果显示，与模型组比较，RIG可以降低T2DM小鼠饮水量、FBG、FINS、HOMA-IR、LDL-C水平（P＜0.01），升高HDL-C水平（P＜0.01），改善肝细胞形态，减轻肝损伤，减少胶原沉积及脂质沉积，抑制肝组织IL-1、IL-6、TNF-α表达（P＜0.01），下调肝组织AGEs、RAGE、p-p38 MAPK/p38 MAPK蛋白表达及RAGE、p38 MAPK mRNA表达（P＜0.05、0.01）。结论 RIG能够有效降低T2DM小鼠FBG，改善胰岛素抵抗，减轻炎症反应，保护肝组织，其机制可能与调控AGEs/RAGE/MAPK信号通路有关。

Objective To reveal the mechanism of Rehmannia glutinosa iridoid glycosides (RIG) in treating type 2 diabetes mellitus (T2DM) based on network pharmacology and animal experimental verification.Methods The RIG related targets were predicted and screened by Swisstargetprediction and PharmMapper database, the target genes related to T2DM were selected using OMMI, Genecard and DisGeNET databases. The obtained common targets were imported into the STRING database to build the protein-protein interaction (PPI) network, and Cytoscape software was used to build the “drug-composition-target” and core target networks. Gene ontology (GO) function and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis were performed using the DAVID database and bioinformatics platform. T2DM mice model was established using high sugar and high-fat feeding combined with ip streptozotocin. The successfully modeled mice were randomly divided into model group, metformin (250 mg/kg) group, RIG low- and high-dose (200, 400 mg/kg) groups, with nine mice in each group, and 10 normal mice were selected as the control group. Drugs were given for intervention for eight weeks, body weight and fasting blood glucose (FBG) were measured weekly. After the last administration, serum was separated, the levels of low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) and fast insulin (FINS) were measured, the homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. The pathological changes of liver tissue in mice were observed using hematoxylin eosin (HE), Masson and oil red O staining. The expression levels of inflammatory factors interleukin-1 (IL-1), IL-6 and tumor necrosis factor-α (TNF-α) in liver tissue were measured using immunohistochemistry. Western blotting was used to detect the protein expressions of advanced glycation end products (AGEs), receptor for advanced glycation end products (RAGE), and p-p38 mitogen activated protein kinase (MAPK)/p38 MAPK levels in liver tissue. qRT-PCR was used to detect the expressions of RAGE and p38 MAPK mRNA in liver tissue.Results A total of 175 potential targets of RIG for T2DM were screened, and key core targets included proto-oncogene tyrosine-protein kinase (Src), epidermal growth factor receptor (EGFR), signal transducer and activator of transcription 3 (STAT3), etc. GO enrichment analysis showed that potential targets were mainly involved in biological processes such as the regulation of inflammatory response. KEGG pathway analysis screened 277 signal pathways, indicating that lipid and atherosclerosis, AGEs/RAGE signaling pathway and MAPK signaling pathway may play a key role in the treatment of T2DM. Animal experiment results showed that compared with model group, RIG could reduce the water intake, FBG, FINS, HOMA-IR, LDL-C levels (P < 0.01), increase HDL-C level (P < 0.01), improve liver cell morphology, alleviate liver injury, reduce collagen deposition and lipid deposition, inhibit the expressions of IL-1, IL-6, TNF-α in liver tissue (P < 0.01), and down-regulate the protein expressions of AGEs, RAGE, p-p38 MAPK/p38 MAPK and mRNA expressions of RAGE, p38 MAPK in liver tissue in T2DM mice (P < 0.05, 0.01).Conclusion RIG could effectively reduce FBG, improve insulin resistance, alleviate inflammatory response, and protect liver tissue in T2DM mice. Its mechanism may be related to the regulation of AGEs/RAGE/MAPK signaling pathway.

R285.5

河南省中医药科学研究专项重大课题（20-21ZYZD13）；河南省中医药科学研究专项重点课题（2024ZY1026）；河南省省属科研机构基本科研业务费项目（2404024）