中国组织工程研究 ›› 2025, Vol. 29 ›› Issue (24): 5140-5147.doi: 10.12307/2025.728

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

水苏碱激活自噬改善高脂喂养小鼠动脉粥样硬化的作用机制

杨  珺,殷  鹏,郑中华   

  1. 浙江省人民医院毕节医院血管外科,贵州省毕节市  551700


  • 收稿日期:2024-06-17 接受日期:2024-08-24 出版日期:2025-08-28 发布日期:2025-01-24
  • 作者简介:杨珺,男,1977年生,贵州省毕节市人,汉族,副主任医师,主要从事血管外科方面的研究。
  • 基金资助:
    贵州省自然科学基础研究计划项目(2022JM-580),项目负责人:杨珺

Mechanism of stachydrine-induced autophagy in improving atherosclerosis in high-fat-fed mice

Yang Jun, Yin Peng, Zheng Zhonghua   

  1. Department of Vascular Surgery, Bijie Hospital, Zhejiang Provincial People’s Hospital, Bijie 551700, Guizhou Province, China
  • Received:2024-06-17 Accepted:2024-08-24 Online:2025-08-28 Published:2025-01-24
  • About author:Yang Jun, Associate chief physician, Department of Vascular Surgery, Bijie Hospital, Zhejiang Provincial People’s Hospital, Bijie 551700, Guizhou Province, China
  • Supported by:
    Guizhou Province Natural Science Basic Research Program, No. 2022JM-580 (to YJ)

摘要:




文题释义:
动脉粥样硬化:是一种由脂质代谢异常和炎症引起的慢性炎症性疾病,以内皮细胞损伤及功能障碍为起始,导致低密度脂蛋白沉积到血管壁并被氧化,受损的内皮细胞分泌趋化因子和黏附分子,进而招募循环中的单核细胞迁移并黏附至内皮下血管间隙,吞噬脂蛋白转化为泡沫细胞,形成早期病变。
水苏碱:是中国草本植物益母草的主要成分,具有抗炎、抗氧化、抗血小板和促血管生成的特性,对心血管系统、中枢神经系统有潜在益处。最近研究发现,水苏碱可调节内皮细胞、诱导血管舒张,从而改善血管内环境稳态。

背景:水苏碱具有抗炎、抗氧化、抗血小板和促血管生成的特性,对心血管系统和中枢神经系统有潜在益处。最近研究发现,水苏碱通过增加鸟苷三磷酸环化水解酶和二氢叶酸还原酶的表达有效逆转同型半胱氨酸诱导的内皮功能障碍,改善内皮功能紊乱,但水苏碱在动脉粥样硬化中的作用尚不清楚。
目的:探讨水苏碱对高脂喂养诱导ApoE-/-基因敲除小鼠动脉粥样硬化的影响及分子机制。
方法:采用随机数字表法将48只ApoE-/-基因敲除小鼠随机分为空白对照组、模型组、水苏碱组、阿托伐他汀组,每组12只,模型组、水苏碱组、阿托伐他汀组给予高脂饲料喂养12周(建立动脉粥样硬化模型),空白对照组给予基础饲料喂养12周。造模成功后,水苏碱组给予30 mg/kg水苏碱灌胃,阿托伐他汀组给予2.6 mg/kg阿托伐他汀灌胃,空白对照组和模型组给予等体积羧甲基纤维素钠灌胃,1次/d,连续给药30 d。给药结束后,苏木精-伊红染色观察主动脉根部病理变化,油红O染色观察主动脉斑块及主动脉根部脂质沉积情况,RT-qPCR检测主动脉中黏附分子(细胞间黏附分子1、血管细胞黏附分子1、选择素E)及趋化因子(CXCL1、CXCL4、单核细胞趋化蛋白1)mRNA表达,RNA测序分析各组主动脉组织基因表达差异并富集上调显著的信号通路,免疫印迹检测主动脉中自噬标志蛋白LC3BⅡ/LC3BⅠ、SQSTM1、p-腺苷酸活化蛋白激酶α、沉默信息调节因子1表达,透射电子显微镜下观察自噬溶酶体变化。
结果与结论:①与空白对照组相比,模型组小鼠主动脉粥样硬化斑块增多、脂质沉积增多,黏附分子及趋化因子mRNA表达升高(P < 0.05);与模型组相比,水苏碱组、阿托伐他汀组小鼠主动脉粥样硬化斑块减少、脂质沉积减少,黏附分子及趋化因子mRNA表达降低(P < 0.05);②RNA组学分析显示,与模型组相比,水苏碱组基因表达上调972个、表达下调781个,对表达上调的基因进行KEGG富集分析,自噬信号通路及腺苷酸活化蛋白激酶信号通路上调显著;③免疫印迹检测结果显示,与模型组相比,水苏碱组LC3BⅡ/LC3BⅠ比值、p-腺苷酸活化蛋白激酶α和沉默信息调节因子蛋白表达均升高(P < 0.05),SQSTM1蛋白表达下降(P < 0.05);④透射电子显微镜下可见水苏碱组自噬溶酶体数量多于模型组;⑤结果表明,水苏碱可能通过上调腺苷酸活化蛋白激酶/沉默信息调节因子1信号通路激活细胞自噬,进而缓解动脉粥样硬化小鼠的血管内皮炎症和斑块沉积。
https://orcid.org/0009-0001-1957-5264(杨珺)

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程

关键词: 水苏碱, 动脉粥样硬化, 自噬, AMPK/SIRT1信号通路, 炎性反应, 工程化组织构建

Abstract: BACKGROUND: Stachydrine has anti-inflammatory, antioxidant, and antiplatelet properties that promote angiogenesis and has potential benefits on the cardiovascular system and central nervous system. Recently, it has been found that stachydrine effectively reverses homocysteine-induced endothelial dysfunction and ameliorates endothelial dysfunction by increasing the expression of guanosine triphosphate cyclase hydrolase and dihydrofolate reductase, but the role of stachydrine in atherosclerosis is yet unclear.
OBJECTIVE: To explore the effect and molecular mechanism of stachydrine on atherosclerosis induced by a high-fat diet in ApoE-/- mice.
METHODS: A total of 48 ApoE-/- mice were randomly divided into blank control group, model group, stachydrine group and atorvastatin group, with 12 mice in each group. Mice in the latter three groups were fed with high-fat diet for 12 weeks to establish animal models of atherosclerosis. After successful modeling, the stachydrine group was treated with stachydrine (30 mg/kg) by gavage, the atorvastatin group was treated with atorvastatin (2.6 mg/kg) by gavage, and the blank control group and the model group were treated with the same volume of sodium carboxymethyl cellulose by gavage once a day for 30 days. After administration, hematoxylin-eosin staining was used to observe the pathological changes of the aortic root. Oil red O staining was used to detect lipid deposition in aortic plaques and the aortic root. Real-time fluorescent quantitative PCR was used to detect mRNA expression of adhesion molecules (intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and selectin E) and chemokines (CXCL1, CXCL4, and monocyte chemotactic protein 1) in the aorta. RNA sequencing was used to analyze differential expression of genes between groups of aortic tissues and enrich for significantly upregulated signaling pathways. Western blot was used to detect the expression levels of autophagy marker proteins, autophagy microtubule-associated protein light chain β3 antibody (LC3BII/LC3BI), SQSTM1, phosphorylated AMp-activated protein kinase α and silent information regulator. Autophagy-lysosome changes were observed under transmission electron microscope.
RESULTS AND CONCLUSION: Compared with the blank control group, the model group had increased aortic plaques and lipid deposition, and increased mRNA expression of adhesion molecules and chemokines (P < 0.05). Compared with the model group, the stachydrine group or atorvastatin group had reduced aortic plaques and lipid deposition, and decreased mRNA expression of adhesion molecules and chemokines (P < 0.05). RNA sequencing analysis showed that 972 genes were up-regulated and 781 genes were down-regulated in the stachydrine group compared with the model group. KEGG enrichment analysis of the up-regulated genes showed that autophagy signaling pathway and AMPK signaling pathway were significantly up-regulated. Western blot results showed that compared with the model group, the stachydrine group had a significantly increased LC3BII/LC3BI ratio and protein expression of phosphorylated AMp-activated protein kinase α and silent information regulator (P < 0.05), and a significantly decreased protein level of SQSTM1. Transmission electron microscope analysis of mouse aorta showed that the stachydrine group had a significantly increased number of autophagolysosomes compared with the model group. To conclude, stachydrine may activate autophagy by up-regulating AMp-activated protein kinase/silent information regulator signaling pathway, thereby alleviating vascular endothelial inflammation and plaque deposition in atherosclerosis mice. 

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程

Key words: stachydrine, atherosclerosis, autophagy, AMPK/SIRT1, inflammatory response, engineered tissue construction

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