Objectives: To observe the effect of electroacupuncture (EA) of "Jiaji" (EX-B2) on expression of high mobility group box 1 (HMGB1)/Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) related proteins and inflammation-related factors in spinal cord injury (SCI) rats, so as to explore the dynamic process of inhibiting inflammatory response in rats with SCI, and to provide a new idea and theoretical basis of molecular biology for the treatment of SCI.
Methods: Fifty-four SD rats were randomly divided into sham operation, model, and EA groups (n=18 in each group), which were further divided into 3 d, 7 d and 14 d subgroups, with 6 rats at each time point. The SCI model was established according to the Allen's method. Rats in EA group received EA (1 mA, 100 Hz) intervention at EX-B2 of T9 and T11 24 hours after modeling for 30 min, once a day for 3 d, 7 d, or 14 d, respectively. The motor function of rats' hind limbs was evaluated using BBB scale, the morphological structure of rats' spinal cord tissue was observed by H.E. staining. The contents of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in spinal cord tissue were detected by ELISA. Western blot and immunofluorescence staining were used to detect the relative protein expression and fluorescence positive expression of HMGB1, TLR4 and NF-κB p65 in spinal cord tissue, respectively.
Results: Compared with the sham operation group, the BBB scores after modeling and on day 3, 7 and 14 were all decreased (P<0.05), while the contents of iNOS and COX-2 in spinal cord tissue, the protein expression and fluorescence positive expression of HMGB1, TLR4, and NF-κB p65 were increased (P<0.05) in the model group. In comparison with the model group, the BBB scores on day 3, 7 and 14 were obviously increased (P<0.05), while the contents of iNOS and COX-2, the protein expression and fluorescence positive expression of HMGB1, TLR4, and NF-κB p65 were significantly down-regulated in the EA group (P<0.05).
Conclusions: EA of "Jiaji" can promote the recovery of SCI neurological function, which may be related to its function in regulating the HMGB1/TLR4/NF-κB signaling pathway, down-regulating the expression of iNOS and COX-2, and inhibiting the neuroinflammatory response after SCI.
目的: 观察“夹脊”电针对脊髓损伤（SCI）大鼠高迁移率族蛋白1（HMGB1）/Toll样受体4（TLR4）/核转录因子-κB（NF-κB）通路相关蛋白及炎性因子表达的影响，探讨“夹脊”电针抑制SCI大鼠炎性反应促进神经功能恢复的可能机制及时间动态规律。方法: 54只SD大鼠随机分为假手术组、模型组、电针组，每组分为3、7、14 d 3个亚组，每个亚组6只。采用Allen’s法制备SCI大鼠模型。电针组于造模后1 d取双侧胸（T）9、T11“夹脊”行电针干预，密波（100 Hz），电流强度1 mA，1次/d，每次30 min，分别治疗3、7、14 d。采用运动功能（BBB）评分量表观察各组大鼠后肢运动功能；HE染色观察脊髓组织病理形态；ELISA法检测脊髓组织中诱导型一氧化氮合酶（iNOS）、环氧合酶-2（COX-2）的含量；Western blot及免疫荧光染色法分别检测脊髓组织HMGB1、TLR4、NF-κB p65的蛋白相对表达量和荧光阳性表达。结果: 与假手术组比较，模型组大鼠造模后和治疗后各时间点BBB评分降低（P<0.05），神经元数量大幅减少、炎性浸润严重，脊髓组织iNOS、COX-2含量升高（P<0.05），HMGB1、TLR4、NF-κB p65的蛋白表达及荧光阳性表达升高（P<0.05）。与模型组比较，分别治疗3、7、14 d后电针组大鼠的BBB评分升高（P<0.05），神经元丢失及炎性浸润情况得到改善，脊髓组织iNOS、COX-2含量降低（P<0.05），HMGB1、TLR4、NF-κB p65的蛋白表达及荧光阳性表达减少（P<0.05）。结论: “夹脊”电针通过调节HMGB1/TLR4/NF-κB信号通路，下调iNOS和COX-2的表达，从而抑制SCI后神经炎性反应，促进SCI神经功能恢复。.
Keywords: Electroacupuncture; High mobility group box 1; Nuclear factor-κB; Spinal cord injury; Toll like receptor 4; “Jiaji”(EX-B2).