| 摘要: |
| 目的 基于p38 MAPK信号通路，探究电针刺激足三里、三阴交激活腺苷A2A受体（A2AR）减轻胶原诱导性关节炎（CIA）大鼠关节炎症及骨侵蚀的作用机制。方法 将32只SD大鼠随机分为对照组、模型组、药物组和电针组，每组8只。模型组、药物组和电针组大鼠均通过胎牛Ⅱ型胶原与弗氏佐剂混合液2次注射免疫复制CIA大鼠模型，药物组于造模成功后每隔7 d腹腔注射甲氨蝶呤2 mg/kg、连续14 d，电针组于造模成功后予大鼠双侧足三里、三阴交每天电针（2/100 Hz疏密波，输出电流2 mA）刺激20 min、连续14 d。通过CT观察大鼠后足骨质破坏情况，H-E、番红O-固绿及抗酒石酸酸性磷酸酶（TRAP）染色分别观察膝关节滑膜、软骨及破骨细胞病理组织学变化，蛋白质印迹法测定膝关节滑膜组织中A2AR、p38、磷酸化p38（P-p38）、NF-κB、T细胞核因子c1（NFATc1）蛋白表达，ELISA法测定外周血清IL-1β含量。结果 与对照组相比，模型组大鼠后足骨密度下降，空间结构紊乱；膝关节软骨明显受损，滑膜组织过度增生；TRAP阳性的破骨细胞数目增加（P<0.01）；膝关节滑膜组织中A2AR、p38、P-p38、NF-κB、NFATc1表达均上调（P均<0.01）；外周血清中IL-1β含量升高（P<0.01）。与模型组比较，电针组和药物组大鼠后足整体结构、膝关节滑膜及软骨破坏明显改善；TRAP阳性破骨细胞数目减少（P<0.01）；A2AR表达上调（P<0.01），p38、P-p38、NF-κB、NFATc1表达下调（P均<0.01）；外周血清中IL-1β含量降低（P<0.01）。与药物组比较，电针组的TRAP阳性破骨细胞数目增加（P<0.05）；A2AR蛋白表达下调（P<0.01），NF-κB、NFATc1蛋白表达上调（P均<0.05），p38表达变化不明显（P>0.05），但P-p38表达上调（P<0.05）。结论 电针刺激足三里、三阴交可减轻CIA大鼠的关节骨侵蚀损害，其关节保护作用机制可能与电针激活A2AR后抑制p38 MAPK通路进而减少破骨细胞生成有关。 |
| 关键词: 电针 甲氨蝶呤 胶原诱导性关节炎 腺苷A2A受体 p38丝裂原活化蛋白激酶信号通路 |
| DOI：10.16781/j.0258-879x.2021.07.0740 |
| 投稿时间：2021-04-26修订日期：2021-06-30 |
| 基金项目:国家自然科学基金面上项目（81674053），浙江省自然科学基金探索项目（LY20H270015）. |
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| Inhibiting p38 MAPK signaling pathway to reduce joint bone erosion in rats with collagen-induced arthritis by activating A2A adenosine receptor after treating with electroacupuncture |
| TANG Ke-jing1,ZHENG Qi-qi1,SONG Zhi-wei1,DU Zhong-heng2,YE Tian-shen2* |
(1. School of First Clinical Medicine, School of Information and Engineering, Wenzhou Medical University, Wenzhou 325035, Zhejiang, China;
2. Department of Acupuncture, Tuina and Physiotherapy, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, Zhejiang, China
*Corresponding author) |
| Abstract: |
| Objective To explore the mechanism of electroacupuncture at "Zusanli" (ST36) and "Sanyinjiao" (SP06) to activate A2A adenosine receptor (A2AR) for reducing joint inflammation and bone erosion in rats with collagen-induced arthritis (CIA) based on p38 mitogen-activated protein kinase (MAPK) signaling pathway. Methods A total of 32 male SD rats were evenly randomized into control, CIA model, methotrexate (MTX)-treated and electroacupuncture-treated groups. The rats in model, MTX-treated and electroacupuncture-treated groups were immunized to replicate the CIA model by twice injection of fetal bovine typeⅡ collagen and Freund's adjuvant mixture. After the model was successfully established, the rats in the MTX-treated group were injected with MTX 2 mg/kg every 7 days for 14 days, and those in the electroacupuncture-treated group were treated with electroacupuncture (2/100 Hz density wave and output current of 2 mA) bilaterally at ST36 and SP06 for 20 min every day for 14 days. The pathological changes of hind foot bone were observed by computed tomography (CT) imaging; the histopathological changes of synovium, cartilage and osteoclasts of the knee joint were observed by hematoxylin-eosin (H-E), safranin O-fast green and tartrate-resistant acid phosphatase (TRAP) staining; the expression of A2AR, p38, phosphorylated p38 (P-p38), nuclear factor κB (NF-κB) and nuclear factor-activated T cell 1 (NFATc1) proteins in the knee synovial tissue was determined by Western blotting; and the peripheral serum interleukin 1β (IL-1β) content was measured by enzyme-linked immunosorbent assay (ELISA). Results Compared with the control group, the bone density of hind foot in the model group was decreased and the spatial structure was disordered; the cartilage of knee joint was obviously damaged and the synovial tissue was hyperplastic; the number of TRAP positive osteoclasts was increased significantly (P<0.01), while the expression of A2AR, p38, P-p38, NF-κB and NFATc1 in the knee synovial tissue was significantly up-regulated (P<0.01); and the peripheral serum IL-1β content was significantly increased (P<0.01). Compared with the model group, the overall structure of hind foot, synovium and cartilage damage of knee joint in the electroacupuncture-treated group and MTX-treated group were significantly improved; the number of TRAP positive osteoclasts was decreased (P<0.01); the expression of A2AR was significantly up-regulated (P<0.01) and expression of p38, P-p38, NF-κB and NFATc1 was significantly down-regulated (all P<0.01); and the serum IL-1β content was greatly reduced (P<0.01). Compared with the MTX-treated group, the number of TRAP positive osteoclasts was increased in the electroacupuncture-treated group (P<0.05); the expression of A2AR protein was down-regulated (P<0.01), the expression of NF-κB and NFATc1 proteins was up-regulated (P<0.05); and there was no significant difference in the expression of total p38 (P>0.05), but the expression of P-p38 was up-regulated (P<0.05). Conclusion Electroacupuncture at ST36 and SP06 can alleviate the joint bone erosion in CIA rats by inhibiting p38 MAPK pathway to reduce the production of osteoclasts after activating A2AR. |
| Key words: electroacupuncture methotrexate collagen-induced arthritis A2A adenosine receptor p38 mitogen-activated protein kinase pathway |
