欢迎访问《中国实验方剂学杂志》编辑部网站!
苍膝通痹胶囊调控p38 MAPK/NLRP3/Caspase-1通路介导膝关节骨性关节炎软骨细胞焦亡的机制
作者:
作者单位:

1.山东中医药大学 第一临床医学院,济南 250014;2.山东体育学院,济南 250102;3.山东中医药大学 附属医院,济南 250014

作者简介:

张志猛,在读硕士,从事中医骨科学基础与临床研究,E-mail:15610145105@163.com

通讯作者:

谢文鹏,博士,副主任医师,从事中医骨科学基础与临床研究,E-mail:xiewenpeng0925@163.com

中图分类号:

R2-0;R22;R285.5;R289;R33

基金项目:

国家自然科学基金青年科学基金项目(82104897);山东省自然科学基金青年项目(ZR2020QH312);美罗青年医师创新发展项目(GSKQNJJ-2023-006);山东省中医药科技青年项目(Q-2022063);山东省医药卫生科技项目(202304071516)


Mechanism of Cangxitongbi Capsules in Mediating Chondrocyte Pyroptosis in Knee Osteoarthritis via Regulating p38 MAPK/NLRP3/Caspase-1 Pathway
Author:
Affiliation:

1.The First Clinical Medical College of Shandong University of Traditional Chinese Medicine(TCM),Jinan 250014,China;2.Shandong Sport University,Jinan 250102,China;3.Affiliated Hospital of Shandong University of TCM,Jinan 250014,China

Fund Project:

  • 摘要
  • |
  • 图/表
  • |
  • 访问统计
  • |
  • 参考文献
  • |
  • 相似文献
  • |
  • 引证文献
  • |
  • 资源附件
  • |
  • 文章评论
    摘要:

    目的 探究苍膝通痹胶囊(CXTB)调控p38丝裂原活化蛋白激酶(p38 MAPK)/NOD样受体蛋白3(NLRP3)/胱天蛋白酶-1(Caspase-1)信号通路抑制膝关节骨性关节炎(KOA)软骨细胞焦亡的机制。方法 60只雄性SD大鼠随机分为假手术组、模型组、CXTB低、中、高剂量组及阳性药组,每组10只。使用改良Hulth法构建膝关节骨性关节炎大鼠模型,根据分组给予苍膝通痹胶囊(0.25、0.5、1.0 g·kg-1)及塞来昔布(24 mg·kg-1)灌胃,假手术组与模型组给予等体积生理盐水,连续灌胃28 d,1次/d。微计算机断层扫描(Micro-CT)检测骨体积分数(BV/TV)、骨小梁分离度(Tb.Sp),苏木素-伊红(HE)染色、番红固绿(SO)染色及国际骨关节炎研究协会(OARSI)评分观察膝关节退变水平,蛋白免疫印迹法(Western blot)检测p38 MAPK、磷酸化p38 MAPK(p-p38 MAPK)、NLRP3、Caspase-1、消皮素D(GSDMD)蛋白表达,实时荧光定量聚合酶链式反应(Real-time PCR)检测p38 MAPK、NLRP3、Caspase-1、GSDMD mRNA表达,酶联免疫吸附测定法(ELISA)检测肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-18(IL-18)炎性因子的含量。膝关节置换术后的软骨组织以Western blot检测p38 MAPK、p-p38 MAPK、NLRP3、Caspase-1、GSDMD蛋白表达,Real-time PCR检测p38 MAPK、NLRP3、Caspase-1、GSDMD mRNA表达。结果 Micro-CT显示,与假手术组比较,KOA大鼠关节间隙明显狭窄并见骨赘增生,BV/TV值减少、Tb.Sp值增加(P<0.01),血清中TNF-α、IL-1β、IL-18含量升高(P<0.01),软骨中p-p38 MAPK、NLRP3、Caspase-1、GSDMD蛋白表达升高(P<0.01),p38 MAPK、NLRP3、Caspase-1、GSDMD mRNA表达增强(P<0.01),与膝关节置换术后的正常软骨组织相比,病变软骨中的p-p38 MAPK、NLRP3、Caspase-1、GSDMD蛋白表达升高(P<0.05),p38 MAPK、NLRP3、Caspase-1、GSDMD mRNA表达增强(P<0.01)。HE染色及SO染色中可见KOA大鼠关节面粗糙,软骨厚度变薄,细胞排列无序杂乱,同时OARSI评分增加(P<0.01);与模型组比较,CXTB低、中、高浓度组大鼠膝关节的BV/TV值增加、Tb.Sp值减少(P<0.01),HE染色及SO染色可见关节面趋于平滑,OARSI评分减少(P<0.01),p-p38 MAPK、NLRP3、Caspase-1、GSDMD蛋白表达降低(P<0.05),p38 MAPK、NLRP3、Caspase-1、GSDMD mRNA的表达下降(P<0.01),血清中TNF-α、IL-1β、IL-18含量降低(P<0.01)。结论 苍膝通痹胶囊干预后可减缓KOA大鼠的膝关节退变并且抑制炎性因子的表达及软骨细胞焦亡从而保护软骨,而机制可能是通过p38 MAPK/NLRP3/Caspase-1信号通路来调控的。

    Abstract:

    Objective To explore the mechanism of Cangxi Tongbi capsules (CXTB) in regulating the p38 mitogen-activated protein kinase (p38 MAPK)/NOD-like receptor protein 3 (NLRP3)/cysteine protease-1 (Caspase-1) signaling pathway to inhibit pyroptosis of cartilage cells in knee osteoarthritis (KOA).Method Sixty male SD rats were randomly divided into a sham operation group, a model group, low, medium, and high dose CXTB groups, and a positive control group, with 10 rats per group. The modified Hulth method was employed to establish a rat model of KOA. According to their respective assignments, rats were administered CXTB (0.25, 0.5, 1.0 g·kg-1) and Celecoxib (24 mg·kg-1) by gavage. The sham operation and model groups were given an equivalent volume of physiological saline. Treatment was performed once daily for 28 days. Micro-computed tomography (Micro-CT) was used to assess bone volume/total volume (BV/TV) and trabecular separation (Tb.Sp). Joint degeneration was evaluated through hematoxylin-eosin (HE) staining, safranin-fast green (SO) staining, and Osteoarthritis Research Society International (OARSI) scoring. Western blot analysis was conducted to measure the expression levels of p38 MAPK, phosphorylated p38 MAPK (p-p38 MAPK), NLRP3, Caspase-1, and gasdermin D (GSDMD) proteins. Real-time PCR was used to assess mRNA expression levels of p38 MAPK, NLRP3, Caspase-1, and GSDMD genes. Enzyme-linked immunosorbent assay (ELISA) was used to measure serum concentrations of inflammatory cytokines including tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-18 (IL-18). After knee replacement surgery, cartilage tissue was analyzed using Western blot to assess the protein expression levels of p38 MAPK, p-p38 MAPK, NLRP3, Caspase-1, and GSDMD, and Real-time PCR was used to evaluate gene expression levels of p38 MAPK, NLRP3, Caspase-1, and GSDMD.Result Micro-CT analysis revealed significant narrowing of the joint space and increased bone spur formation in KOA rats compared with the sham operation group, with a decrease in BV/TV ratio and an increase in Tb.Sp value (P<0.01). Serum levels of TNF-α, IL-1β, and IL-18 were elevated (P<0.01). The protein expression levels of p-p38 MAPK, NLRP3, Caspase-1, and GSDMD in cartilage were significantly increased (P<0.01), and the mRNA expression levels of p38 MAPK, NLRP3, Caspase-1, and GSDMD were also enhanced (P<0.01). Significant differences in protein expression of p-p38 MAPK, NLRP3, Caspase-1, and GSDMD were observed between normal and diseased cartilage tissues after knee replacement surgery (P<0.05), and the gene expression of p38 MAPK, NLRP3, Caspase-1, and GSDMD were also significantly different (P<0.01). HE and SO staining showed roughened joint surfaces, reduced cartilage thickness, and disordered cellular arrangement in KOA rats. OARSI scores were significantly higher (P<0.01). Compared with the model group, treatment with low, medium, and high concentrations of CXTB resulted in increased BV/TV ratios and decreased Tb.Sp values in the knee joints of rats (P<0.01). HE and SO staining indicated a trend towards smoother joint surfaces and reduced OARSI scores (P<0.01). The protein expression levels of p-p38 MAPK, NLRP3, Caspase-1, and GSDMD were notably decreased (P<0.05), as were the mRNA expression levels of p38 MAPK, NLRP3, Caspase-1, and GSDMD (P<0.01). Additionally, serum concentrations of TNF-α, IL-1β, and IL-18 were significantly reduced (P<0.01).Conclusion CXTB intervention may alleviate knee joint degeneration in KOA rats and inhibit the expression of inflammatory factors and pyroptosis of cartilage cells, thereby protecting cartilage. The underlying mechanism may involve modulation of the p38 MAPK/NLRP3/Caspase-1 signaling pathway.

    参考文献
    相似文献
    引证文献
引用本文

张志猛,袁道通,靳希民,巩瑞,宋振龙,张永奎,王晓乐,毕荣修,谢文鹏.苍膝通痹胶囊调控p38 MAPK/NLRP3/Caspase-1通路介导膝关节骨性关节炎软骨细胞焦亡的机制[J].中国实验方剂学杂志,2024,30(22):61~68

复制
分享
文章指标
  • 点击次数:
  • 下载次数:
  • HTML阅读次数:
  • 引用次数:
历史
  • 收稿日期:2024-07-16
  • 最后修改日期:
  • 录用日期:
  • 在线发布日期: 2024-10-17
  • 出版日期:

地址:北京东直门内南小街16号

邮编:100700

电话:010-84076882

E-mail:syfjx_2010@188.com

中国实验方剂学杂志 ® 2024 版权所有

技术支持:北京勤云科技发展有限公司