艾灸环跳穴对坐骨神经损伤大鼠神经再生的影响

Effects of moxibustion at Huantiao (GB30) acupoint on nerve regeneration in rats with sciatic nerve injury

  • 摘要:
    目的 观察艾灸环跳穴对坐骨神经损伤(SNI)大鼠神经修复再生和功能的影响,探讨艾灸改善SNI的可能机制。
    方法 将70只无特定病原体(SPF)级雄性SD大鼠随机分为对照组(n = 10)和造模组(n = 60)。造模组复制SNI模型大鼠,造模后将60只SNI大鼠随机分为SNI 1 d、3 d、7 d组,艾灸1 d、3 d、7 d组,每组10只。艾灸组在异氟烷呼吸麻醉下在距离皮肤5 cm处予艾灸患侧环跳穴,每日1次,每次20 min,分别治疗1 d、3 d、7 d。对照组和SNI组每日固定呼吸麻醉20 min,每日1次。在造模前1天、治疗第1、3、7天分别对各组大鼠进行旷场试验、热痛阈值测试并观察大鼠一般情况。治疗结束后,免疫荧光法检测大鼠神经轴突生长速度情况、压伤段中间1 mm横截面神经轴突生长密度和施万细胞增殖情况;取大鼠健患两侧腓肠肌,称重后计算大鼠健患两侧腓肠肌湿重比,观察大鼠肌肉萎缩情况,苏木素-伊红(HE)染色观察患侧腓肠肌病理形态学改变;定量实时聚合酶链式反应(qPCR)检测大鼠坐骨神经组织内神经生长因子(NGF)、干扰素(IFN)-γ、巨噬细胞迁移抑制因子(MIF)、白介素(IL)-4、转化生长因子(TGF)-β表达水平。
    结果 造模后,与对照组相比,艾灸组和SNI组大鼠均出现显著的疼痛行为(患侧后脚掌弯曲蜷缩、舔爪、跛行)及活动量下降。艾灸干预的主要效应自第3天起呈现:与SNI组相比,艾灸组大鼠疼痛行为显著改善,活动量和运动轨迹回升,热痛阈值降低。同时,艾灸显著促进了SNI修复,表现为压伤段轴突生长速度、生长密度及施万细胞增殖密度显著优于对照组(P < 0.01)。艾灸还调节了损伤局部微环境,上调促神经修复因子NGF、IL-4、TGF-β(P < 0.05),下调促炎因子IFN-γ(P < 0.01)和MIF(P < 0.05)。治疗第7天,艾灸组的腓肠肌组织形态得到改善,可见肌纤维增大、肌细胞形态更规则,患健侧湿重比增加(P < 0.05),同时NGF、IL-4、TGF-β持续高表达(分别为P < 0.05、P < 0.05、P < 0.01),IFN-γ维持低水平(P < 0.01),MIF水平较SNI组无统计学意义(P > 0.05)。
    结论 艾灸环跳穴可有效改善SNI大鼠运动、感觉功能恢复和神经再生,可能与调控局部炎症反应,促进神经生长因子生长,改善再生微环境,加快受损神经内施万细胞增殖和轴突生长速度有关。

     

    Abstract:
    Objective To observe the effects of moxibustion at Huantiao (GB30) acupoint on nerve repair, regeneration, and function in rats with sciatic nerve injury (SNI), and explore the possible mechanism of SNI improvement via moxibustion.
    Methods A total of 70 specific pathogen-free (SPF) grade male Sprague-Dawley (SD) rats were randomly assigned to control group (n = 10) and model group (n = 60). Following replication of SNI to model group rats, 60 SNI model rats were randomly allocated to SNI groups of 1 d, 3 d, and 7 d and moxibustion groups of 1 d, 3 d, and 7 d with 10 rats in each group. Moxibustion groups were given moxibustion at the Huantiao (GB30) acupoint on the affected side with a 5 cm distance from the skin under isoflurane respiratory anesthesia and treated once a day for 20 min for 1 d, 3 d, and 7 d, respectively. Control and SNI groups were anesthetized with isoflurane daily for 20 min. Open field tests and thermal pain threshold tests were conducted, and the general condition of rats was observed in each group pre-modeling and on treatment day 1, 3, and 7. At the end of the treatment, immunofluorescence was used to detect the axonal growth rate, axonal growth density, and Schwann cells (SCs) proliferation in the middle 1-mm cross-section of the crush injury segment in rats. The gastrocnemius muscles on both sides of the rats were taken and weighed to calculate the wet weight ratio of the gastrocnemius muscles on both sides to observe the muscle atrophy of the rats, and hematoxylin-eosin (HE) staining was used to observe the pathomorphological changes of the gastrocnemius muscles on the affected side. Quantitative real-time polymerase chain reaction (qPCR) was used to detect the expression levels of nerve growth factor (NGF), interferon (IFN), macrophage migration inhibitory factor (MIF), interleukin (IL)-4, and transforming growth factor (TGF)-β in the sciatic nerve tissue of the rats.
    Results After modeling, rats in both moxibustion and SNI groups showed typical signs of pain behaviors (bending and curling of the hind soles of the affected side, licking claws, and lameness) and decreased activity compared with control group. The main benefits of moxibustion were evident from day 3: compared with SNI group, rats in moxibustion group had marked relief of pain behavior, increased activity levels and movement, and a lower response to thermal pain. At the same time, moxibustion significantly promoted the repair of SNI, as evidenced by the significantly better axonal growth rate, growth density, and SCs proliferation density in the crush injury segment compared with SNI group (P < 0.01). Moxibustion also regulated the local microenvironment of the injury, up-regulated the pro-nerve repair factors NGF, IL-4, and TGF-β (P < 0.05), and down-regulated the pro-inflammatory factors IFN-γ (P < 0.01) and MIF (P < 0.05). By day 7, the histomorphology of the gastrocnemius muscle in moxibustion group was improved, as indicated by enlarged muscle fibers, elevated regular myocyte morphology and wet weight ratio of the affected and unaffected sides (P < 0.05), as well as a sustained high expression levels of NGF, IL-4, and TGF-β (P < 0.05, P < 0.05, and P < 0.01, respectively), and a maintenance of low level of IFN-γ (P < 0.01). Concurrently, the MIF level was not significantly different from SNI group (P > 0.05).
    Conclusion Moxibustion at the Huantiao (GB30) acupoint effectively improves motor function and promotes recovery of sensory function and nerve regeneration in SNI rats, which may be related to the regulation of local inflammatory response, the promotion of nerve growth factor expression, the improvement of regenerative microenvironment, and the acceleration of SCs proliferation and axonal growth rate in damaged nerves.

     

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