中国组织工程研究

中国组织工程研究 ›› 2025, Vol. 29 ›› Issue (30): 6398-6408.doi: 10.12307/2025.766

• 软骨组织构建 cartilage tissue construction • 上一篇    下一篇

二甲双胍抑制铁死亡改善骨关节炎模型大鼠的软骨损伤

樊佳欣1，2，贾  祥1，2，徐田杰1，2，刘凯楠3，郭小玲1，2，张  辉4，王  茜1，2   

  1. 1华北理工大学基础医学院，河北省唐山市  063210；2河北省慢性疾病基础医学重点实验室，河北省唐山市  063210；3邢台医学院基础医学部，河北省邢台市  054000；4唐山市第二医院关节一科，河北省唐山市  063000


  • 收稿日期:2024-08-10 接受日期:2024-09-29 出版日期:2025-10-28 发布日期:2025-03-27
  • 通讯作者: 王茜，博士，硕士生导师，华北理工大学基础医学院，河北省唐山市 063210；河北省慢性疾病基础医学重点实验室，河北省唐山市063210
  • 作者简介:樊佳欣，女，1997年生，华北理工大学在读硕士，主要从事骨组织工程方面的研究。
  • 基金资助:
    三三人才工程(C20221117)，项目负责人：张辉；河北省2023年度医学科学研究课题(20230215)，项目负责人：王茜

Metformin inhibits ferroptosis and improves cartilage damage in osteoarthritis model rats

Fan Jiaxin1, 2, Jia Xiang1, 2, Xu Tianjie1, 2, Liu Kainan3, Guo Xiaoling1, 2, Zhang Hui4, Wang Qian1, 2    

  1. 1School of Basic Medical Sciences, North China University of Science and Technology, Tangshan 063210, Hebei Province, China; 2Key Laboratory of Basic Medicine for Chronic Diseases, Tangshan 063210, Hebei Province, China; 3College of Basic Medicine, Xingtai Medical College, Xingtai 054000, Hebei Province, China; 4First Department of Joint Surgery, Tangshan Second Hospital, Tangshan 063000, Hebei Province, China
  • Received:2024-08-10 Accepted:2024-09-29 Online:2025-10-28 Published:2025-03-27
  • Contact: Wang Qian, MD, Master’s supervisor, School of Basic Medical Sciences, North China University of Science and Technology, Tangshan 063210, Hebei Province, China; Key Laboratory of Basic Medicine for Chronic Diseases, Tangshan 063210, Hebei Province, China
  • About author:Fan Jiaxin, Master candidate, School of Basic Medical Sciences, North China University of Science and Technology, Tangshan 063210, Hebei Province, China; Key Laboratory of Basic Medicine for Chronic Diseases, Tangshan 063210, Hebei Province, China
  • Supported by:
    Three-Three Talent Project, No. C20221117 (to ZH); Hebei Province 2023 Medical Science Research Project, No. 20230215 (to WQ) 

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摘要：


文题释义：
铁死亡：是一种新型的细胞死亡形式，与传统的细胞死亡方式(如凋亡、坏死和自噬)有所不同，它由铁依赖的脂质过氧化物积累引起，通常与细胞内铁代谢失调密切相关。
P53/SLC7A11/谷胱甘肽过氧化物酶4信号通路：是铁死亡的经典信号通路，SLC7A11是P53的关键靶点之一，P53通过抑制胱氨酸的摄取来促进铁死亡，而SLC7A11是System-Xc(胱氨酸/谷氨酸反转运蛋白)的重要组成部分，抑制SLC7A11可减少胱氨酸摄取，导致胱氨酸依赖性谷胱甘肽过氧化物酶4失活，增强细胞内脂质过氧化和铁死亡。

背景：二甲双胍目前被视为治疗2型糖尿病的首选药物。研究发现，二甲双胍可延缓骨关节炎进展，但具体的作用机制仍不明确。
目的：评估二甲双胍对骨关节炎大鼠的治疗效果及相关作用机制。
方法：①网络药理学：使用CTD、SwissTargetPrediction、GeneCards 和OMIM数据库筛选二甲双胍、骨关节炎、铁死亡的潜在共同靶标，导入STRING数据库进行蛋白质-蛋白质相互作用分析得到二甲双胍、骨关节炎和铁死亡的关键靶点。②分子对接：从PDB数据库下载P53及其下游因子SLC7A11蛋白质的PDB格式，将二甲双胍的2D结构式转换为3D结构式，使用Discovery Studio 2019 Client进行二甲双胍和蛋白质的分子对接。③体内实验：取30只雄性SD大鼠，随机分为3组(n=10)：空白组不进行任何手术，模型组、二甲双胍组采用改良Hulth法建立骨关节炎模型；术后1 d，二甲双胍组大鼠灌胃二甲双胍200 mg/(kg·d)，空白组、模型组灌胃生理盐水，连续给药4周。苏木精-伊红染色、番红O-固绿染色观察大鼠膝关节软骨形态结构并进行Mankin评分；ELISA检测血清中肿瘤坏死因子α、白细胞介素6水平；微板法检测血清中铁死亡相关指标谷胱甘肽、丙二醛、Fe2+水平；免疫荧光染色、Western blot、Real-time qPCR检测大鼠软骨组织中P53、SLC7A11、谷胱甘肽过氧化物酶4、软骨蛋白聚糖、基质金属蛋白酶13的蛋白及mRNA表达。
结果与结论：①二甲双胍、骨关节炎与铁死亡交集靶点共96个，经蛋白质-蛋白质相互作用分析发现潜在靶点共有77个，进一步筛选得到核心靶点依次为TP53、AKT1、JUN、白细胞介素6、MYC、白细胞介素1β、肿瘤坏死因子α等。②对接分析结果显示，二甲双胍与P53及其下游因子SLC7A11结合牢固稳定。③模型组大鼠膝关节软骨表面不规则，软骨组织出现缺损，软骨细胞数量减少；相较于模型组，二甲双胍组大鼠膝关节软骨结构损伤明显改善，软骨表面趋于平整，软骨细胞数量增加。模型组Mankin评分较空白组明显升高，二甲双胍组Mankin评分较模型组明显降低。④与模型组比较，二甲双胍组大鼠血清中肿瘤坏死因子α、白细胞介素6、丙二醛、Fe2+水平明显降低，谷胱甘肽水平明显升高。⑤与模型组比较，二甲双胍组大鼠软骨组织中SLC7A11、谷胱甘肽过氧化物酶4、软骨蛋白聚糖的蛋白和mRNA表达明显升高，P53、基质金属蛋白酶13的蛋白和mRNA表达明显降低。⑥结果表明：二甲双胍可有效改善骨关节炎大鼠关节软骨损伤，并通过抑制异常激活的P53/SLC7A11/谷胱甘肽过氧化物酶4信号通路改善软骨细胞铁代谢和脂质过氧化反应，从而缓解软骨细胞铁死亡，减少软骨基质降解，防止软骨损伤和炎症反应进一步发展。
https://orcid.org/0009-0008-7941-1051(樊佳欣)

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: 骨关节炎, 关节软骨, 铁死亡, 铁代谢, 二甲双胍, P53, SLC7A11, 谷胱甘肽过氧化物酶4, 工程化组织构建

Abstract: BACKGROUND: Metformin is currently considered the first-line medication for the treatment of type 2 diabetes. Metformin may delay the progression of osteoarthritis, but its specific mechanism of action remains unclear.
OBJECTIVE: To evaluate the therapeutic effects and the related action mechanisms of metformin on osteoarthritis in rats.
METHODS: (1) Network pharmacology: Potential common targets for metformin, osteoarthritis, and ferroptosis were screened using the CTD, SwissTargetPrediction, GeneCards, and OMIM databases. After importing the targets into the STRING database, protein-protein interaction analysis was conducted to identify the key targets for metformin, osteoarthritis, and ferroptosis. (2) Molecular docking: P53 and its downstream factor SLC7A11 protein structures in PDB format were downloaded from the PDB database. The 2D structure of metformin was converted to a 3D structure, and molecular docking of metformin with the proteins was performed using Discovery Studio 2019 Client. (3) In vivo experiments: Thirty male SD rats were randomly divided into three groups (n=10). The blank group did not receive surgery. The osteoarthritis model was established using the modified Hulth method for the model and metformin groups. One day after the surgery, rats in the metformin group were gavaged with metformin 200 mg/kg per day, while the blank and model groups were gavaged with physiological saline. Treatment continued for 4 weeks. Hematoxylin-eosin staining and Safranin O-fast green staining were used to observe the pathological morphology and structure of the knee cartilage, and Mankin scoring was performed. ELISA was used to measure the levels of tumor necrosis factor-α and interleukin-6 in the serum. The microplate method was used to measure serum ferroptosis-related indicators, including glutathione, malondialdehyde, and Fe2+. Immunofluorescence staining, western blot assay, and real-time qPCR were used to detect the protein and mRNA expression of P53, SLC7A11, glutathione peroxidase 4, proteoglycans, and matrix metalloproteinase 13 in the cartilage tissue of the rats.
RESULTS AND CONCLUSION: (1) A total of 96 intersecting targets among metformin, osteoarthritis, and ferroptosis were identified. After protein-protein interaction analysis, 77 potential targets were found. Further screening identified the core targets as TP53, AKT1, JUN, interleukin-6, MYC, interleukin-1β, and tumor necrosis factor-α, among others. (2) Docking analysis results showed that metformin bound strongly and stably with P53 and its downstream factor SLC7A11. (3) In the model group, the knee cartilage surface was irregular, with cartilage tissue defects and reduced chondrocyte numbers. Compared to the model group, the knee cartilage structure damage in the metformin group was significantly improved, with a smoother cartilage surface and increased chondrocyte numbers. The Mankin score in the model group was significantly higher than that in the blank group, while the Mankin score in the intervention group was significantly lower than that in the model group. (4) Compared with the model group, the metformin group had significantly lower levels of tumor necrosis factor-α, interleukin-6, malondialdehyde, and Fe2+, and significantly higher glutathione levels. (5) Compared to the model group, the metformin group had significantly increased protein and mRNA expression of SLC7A11, glutathione peroxidase 4, and proteoglycans, and significantly decreased protein and mRNA expression of P53 and matrix metalloproteinase 13 in their cartilage tissue. (6) The results indicate that metformin can effectively improve cartilage damage in osteoarthritis rats and alleviate chondrocyte ferroptosis by inhibiting the aberrantly activated P53/SLC7A11/glutathione peroxidase 4 signaling pathway. This improvement in chondrocyte iron metabolism and lipid peroxidation response further reduces cartilage matrix degradation and prevents further cartilage damage and inflammatory response.

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: osteoarthritis, articular cartilage, ferroptosis, iron metabolism, metformin, P53, SLC7A11, glutathione peroxidase 4, engineered tissue construction

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