| 引用本文: |
吴子政, 陈杏, 叶嘉豪, 王啸轶, 张君宇, 胡志希.基于网络药理学及实验验证探讨冠心宁注射液对冠心病大鼠的干预机制[J].湖南中医药大学学报,2025,45(6):1156-1168[点击复制] |
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| 基于网络药理学及实验验证探讨冠心宁注射液对冠心病大鼠的干预机制 |
| 吴子政,陈杏,叶嘉豪,王啸轶,张君宇,胡志希 |
| (湖南中医药大学中医诊断学湖南省重点实验室, 湖南 长沙 410208) |
| 摘要: |
| 目的 运用网络药理学方法探讨冠心宁注射液（GXNI）对冠心病（CHD）大鼠的作用机制，并通过体内实验进行验证。方法 运用UPLC-Q-TOF-MS技术、UNIFI数据库和文献检索，筛选并验证GXNI有效入血成分。运用网络药理学构建蛋白质-蛋白质相互作用（PPI）网络以识别潜在的治疗靶点，同时进行GO和KEGG分析以预测潜在的信号通路。将50只大鼠按照随机数字表法分为空白组，模型组，普萘洛尔组（10 mg/kg），冠心宁低、高剂量组（3.6、7.2 mL/kg），每组10只，分别连续预给药干预7 d。在干预第5、6天时，除空白组外，其他组均采用背部皮下注射异丙肾上腺素（85 mg/kg）的方式构建CHD模型。检测各组大鼠心电图情况；采用HE染色、Masson染色观察心肌病理学改变；ELISA检测血清白细胞介素（IL）-6、乳酸脱氢酶（LDH）、肌酸激酶同工酶（CK-MB）、IL-1β、肿瘤坏死因子（TNF）-α以及丙二醛（MDA）和超氧化物歧化酶（SOD）水平；Western blot检测心肌组织自噬相关蛋白（Atg）3和Atg5蛋白表达水平。结果 UPLC-Q-TOF-MS 分析显示，从GXNI中筛选出有效入血成分11种化合物，GXNI-有效成分-共同靶点网络图得到核心有效成分为丁烯基苯酞、开环异落叶松树脂酚、正丁基苯酞、丹酚酸A等。PPI网络筛选得到Degree前5的靶点为TNF、白蛋白、类固醇受体辅激活因子、表皮生长因子受体（EGFR）、胱天蛋白酶-3。通过KEGG富集分析预测出GXNI治疗CHD可能与环磷酸腺苷信号通路、神经活性配体-受体相互作用、钙信号通路等疾病通路相关。心肌组织病理检测显示，与空白组比较，模型组心肌细胞肥大，心肌纤维排列紊乱，心肌组织粥样化、胶原化；冠心宁注射液低、高剂量组心肌细胞排列紊乱减轻，心肌细胞间隙轻度增宽，心肌组织粥样化，胶原化减轻。ELISA结果显示，与空白组相比，模型组LDH、CK-MB、IL-6、IL-1β、TNF-α、MAD水平显著升高（P<0.05，P<0.01），SOD活性降低（P<0.01）；与模型组相比，普萘洛尔组和冠心宁高剂量组CK-MB、IL-6、IL-1β、TNF-α、LDH、MDA水平显著降低（P<0.05），SOD活性升高（P<0.01）。与空白组比较，模型组大鼠心肌组织中Atg3、Atg5蛋白相对表达显著升高（P<0.01）；与模型组相比，普萘洛尔组和冠心宁低、高剂量组中的Atg3、Atg5蛋白表达下降（P<0.05，P<0.01）。结论 GXNI可能通过抑制炎症反应、调控自噬，使心肌细胞得到改善，从而治疗CHD。 |
| 关键词: 冠心病 冠心宁注射液 心肌缺血 网络药理学 细胞自噬 |
| DOI：10.3969/j.issn.1674-070X.2025.06.027 |
| 投稿时间：2025-03-03 |
| 基金项目:国家自然科学基金项目（82274412）。 |
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| Intervention mechanism of Guanxinning Injection on rats with coronary heart disease based on network pharmacology and experimental verification |
| WU Zizheng, CHEN Xing, YE Jiahao, WANG Xiaoyi, ZHANG Junyu, HU Zhixi |
| (Hunan Key Laboratory of Chinese Medicine Diagnostics, Hunan University of Chinese Medicine, Changsha, Hunan 410208, China) |
| Abstract: |
| Objective To investigate the intervention mechanism of Guanxinning Injection (GXNI) on rats with coronary heart disease (CHD) using network pharmacology and molecular docking, and to validate it through in vivo experiments.Methods UPLC-Q-TOF-MS technology, UNIFI database, and literature retrieval were employed to screen and verify the effective blood-entry components of GXNI. A protein-protein interaction (PPI) network was constructed using network pharmacology to identify potential therapeutic targets, while GO and KEGG analyses were performed to predict potential signaling pathways. Molecular docking was conducted between key effective blood-entry components and core targets. Fifty rats were randomly divided into control, model, propranolol (10 mg/kg), low-dose GXNI (3.6 mL/kg), and high-dose GXNI (7.2 mL/kg) groups, with ten rats in each group. All groups received pre-administration interventions for seven consecutive days. Except for the blank group, the other groups were administered isoprenaline (85 mg/kg) via subcutaneous injection on the back to establish the CHD model on the 5th and 6th days of intervention. Electrocardiograms (ECG) were monitored. Myocardial pathological changes were observed by HE and Masson staining. Serum levels of interleukin (IL)-6, lactate dehydrogenase (LDH), creatine kinase-MB isoenzyme (CK-MB), IL-1β, tumor necrosis factor-α (TNF-α), and plasma malondialdehyde (MDA), and superoxide dismutase (SOD) were measured by ELISA. Western blot was used to measure the expression levels of autophagy-related protein (Atg) 3 and Atg5 in myocardial tissue.Results UPLC-Q-TOF-MS analysis identified 11 effective blood-entry compounds in GXNI, including butylidenephthalide, secoisolariciresinol, n-butylphthalide, and salvianolic acid A as core active components based on the "GXNI-active components-common targets" network. The PPI network highlighted the top five targets by degree: tumor necrosis factor (TNF), albumin, steroid receptor coactivator, epidermal growth factor receptor (EGFR), and caspase-3. KEGG enrichment analysis predicted that GXNI's therapeutic effects on CHD may involve disease pathways such as cyclic adenosine monophosphate signaling pathway, neuroactive ligand-receptor interactions, and calcium signaling pathway. Molecular docking showed binding affinities (absolute values) in descending order: EGFR-salvianolic acid A (-9.6 kcal/mol), EGFR-secoisolariciresinol (-9.1 kcal/mol), and EGFR-butylidenephthalide (-8.4 kcal/mol). Myocardial histopathology revealed that compared to the blank group, the model group exhibited myocardial hypertrophy, disordered myocardial fiber arrangement, atherosclerosis, and collagen deposition. The low- and high-dose GXNI groups showed reduced disarray of myocardial cells, mildly widened intercellular gaps, and attenuated atherosclerosis and collagen deposition compared with the model group. ELISA indicated that compared with the blank group, the model group had significantly elevated levels of LDH, CK-MB, IL-6, IL-1β, TNF-α, and MDA (P<0.05, P<0.01), along with decreased SOD activity (P<0.01). Compared with the model group, the propranolol and high-dose GXNI groups showed significantly reduced levels of CK-MB, IL-6, IL-1β, TNF-α, LDH, and MDA (P<0.05), along with increased SOD activity (P<0.01). Compared with the blank group, the model group had significantly elevated relative protein expression levels of Atg3 and Atg5 in myocardial tissue (P<0.01). Compared with the model group, the propranolol, low-and high-dose GXNI groups showed decreased protein expression levels of Atg3 and Atg5 (P<0.05, P<0.01).Conclusion GXNI may improve myocardial cell function and treat CHD by inhibiting inflammatory responses and regulating autophagy. |
| Key words: coronary heart disease Guanxinning Injection myocardial ischemia network pharmacology autophagy |
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