口腔医学研究

口腔医学研究 ›› 2025, Vol. 41 ›› Issue (3): 243-249.DOI: 10.13701/j.cnki.kqyxyj.2025.03.012

• 口腔生物学研究 • 上一篇    下一篇

ANXA2调控YAP和p38 MAPK信号通路改善ATDC5软骨细胞炎症损伤

王心茹, 王心怡, 杨畅, 董伟, 王家伟*   

  1. 口颌系统重建与再生全国重点实验室,口腔生物医学教育部重点实验室,口腔医学湖北省重点实验室,武汉大学口腔医(学)院 湖北 武汉 430079
  • 出版日期:2025-03-28 发布日期:2025-03-25
  • 通讯作者: *王家伟,E-mail:wb000238@whu.edu.cn
  • 作者简介:王心茹(1999~ ),女,山东人,硕士在读,主要从事软骨内成骨相关的分子机制研究。
  • 基金资助:
    国家自然科学基金(编号:82170930、82201025、81870744);中央高校基本科研业务费

ANXA2 Improves Inflammatory Injury in ATDC5 Cells by Regulating YAP and p38 MAPK Signaling Pathways

WANG Xinru, WANG Xinyi, YANG Chang, DONG Wei, WANG Jiawei*   

  1. State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration, Key Laboratory of Oral Biomedicine Ministry of Education, Hubei Key Laboratory of Stomatology, School & Hospital of Stomatology, Wuhan University, Wuhan 430079, China
  • Online:2025-03-28 Published:2025-03-25

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摘要： 目的: 探究膜联蛋白A2 (Annexin A2, ANXA2) 对白细胞介素-1β(interleukin-1β,IL-1β)诱导的ATDC5软骨细胞炎症损伤的作用和分子机制。方法: 用碘乙酸钠(monosodium iodoacetate,MIA)和生理盐水分别注射小鼠颞下颌关节部位2周,取髁突切片进行番红/固绿和免疫组织化学染色,分析ANXA2在关节炎软骨中的表达水平。用不同浓度的IL-1β作用ATDC5细胞,荧光定量聚合酶链反应(real-time fluorescence quantitative PCR,qRT-PCR)检测ANXA2和软骨合成代谢标志物Y染色体性别决定区-盒转录因子9(SRY-box transcription factor 9, Sox9)、Ⅱ型胶原蛋白(collagentypeⅡalphalchain, Col2a1),软骨分解代谢标志物基质金属蛋白酶13(matrix metalloproteinase 13, MMP-13)、血小板反应蛋白解整合素金属肽酶5(a disintegrin and metalloproteinase with thrombospondin 5, Adamts5),炎症反应相关因子环氧合酶2(cyclooxygenase 2, Cox2)、诱导型一氧化氮合酶(inducible nitric oxide sythase, iNOS)的表达。用敲低和过表达ANXA2的慢病毒转染ATDC5细胞后,Western blot检测转染效率,qRT-PCR和Western blot分别检测生理和炎症条件下ANXA2对ATDC5细胞中软骨合成、分解以及炎症反应相关标志物的mRNA和蛋白质表达量的影响。Western blot检测Yes相关蛋白(Yes-associated protein, YAP)和p38丝裂原活化蛋白激酶 (p38 mitogen-activated protein kinase, p38 MAPK)的磷酸化水平,以及添加YAP抑制剂维替泊芬(Verteporfin)后SOX9、MMP-13和iNOS的蛋白表达水平。结果: MIA诱导的颞下颌关节炎小鼠的髁突软骨明显退化,ANXA2在关节炎软骨中的表达明显升高。随着作用ATDC5细胞的IL-1β浓度增加,软骨合成代谢标志物表达降低,软骨分解代谢和炎症反应标志物表达升高,ANXA2表达也明显升高。ANXA2敲低加重炎症刺激抑制ATDC5细胞的软骨合成,促进软骨分解和炎症相关因子的表达;ANXA2过表达则得到相反的结果。ANXA2过表达促进炎症刺激下的YAP磷酸化,抑制p38磷酸化。抑制YAP可以部分逆转ANXA2对ATDC5细胞炎症损伤的保护作用。结论: ANXA2部分通过调控YAP和p38 MAPK通路来改善IL-1β诱导的ATDC5软骨细胞炎症损伤。

关键词: 软骨细胞, 炎症损伤, YAP信号通路, p38 MAPK信号通路

Abstract: Objective: To investigate the role and molecular mechanism of Annexin A2 (ANXA2) in inflammatory injury of ATDC5 cells induced by IL-1β. Methods: After injecting the temporomandibular joint of mice with monosodium iodoacetate (MIA) or saline for 2 weeks,the condylar sections were analyzed the expression level of ANXA2 in arthritis cartilage. qRT-PCR was used to detect the effects of different concentrations of IL-1β on ANXA2 and cartilage anabolism markers SRY-box transcription factor 9 (Sox9) and collagen type Ⅱ alphal chain (Col2a1), cartilage catabolism markers matrix metalloproteinase 13 (MMP-13) and a disintegrin and metalloproteinase with thrombospondin 5 (Adamts5), and inflammatory response-related factors cyclooxygenase 2 (Cox2) and inducible nitric oxide synthase (iNOS) in ATDC5 cells. After ATDC5 cells were transfected with lentiviruses that knocked down and overexpressed ANXA2, transfection efficiency was detected by Western blotting. qRT-PCR and Western blot were used to detect the effects of ANXA2 on the mRNA and protein expressions of cartilage catabolism, anabolism, and inflammation-related factors in ATDC5 cells under physiological and inflammatory conditions, respectively. Western blot was used to detect the phosphorylation levels of Yes-associated protein (YAP) and p38 mitogen-activated protein kinase (p38 MAPK), as well as the protein expression levels of SOX9, MMP-13, and iNOS after the addition of the YAP inhibitor Verteporfin. Results: The condylar cartilage of MIA-induced temporomandibular joint osteoarthritis mice significantly degenerated, and the expression of ANXA2 in arthritic cartilage increased.As the concentration of IL-1β increased, the expression of cartilage anabolism markers decreased, while the expression of cartilage catabolism and inflammatory response markers and ANXA2 increased significantly.Knockdown of ANXA2 inhibited cartilage anabolism under inflammatory conditions and promoted cartilage catabolism and the expression of inflammatory-related factors; overexpression of ANXA2 showed the opposite results.Overexpression of ANXA2 promoted YAP phosphorylation under inflammatory conditions and inhibited p38 phosphorylation. Inhibition of YAP could partially reverse the protective effect of ANXA2 on inflammatory injury in ATDC5 cells. Conclusion: ANXA2 improves IL-1β-induced inflammatory injury of ATDC5 cells partially by regulating YAP and p38 MAPK pathways.

Key words: chondrocytes, inflammatory injury, YAP signaling pathway, p38 MAPK signaling pathway