基于BDNF- TrkB -p38-JNK信号通路研究电针治疗筋膜疼痛综合征的机制*

Study on the mechanism of electroacupuncture intervention in treating Myofascial Pain Syndrome based on BDNF- TrkB -p38/JNK signal pathway*

摘要:

基于“BDNF-TrkB-p38/JNK”信号通路，通过体外、体内实验研究针刺激痛点治疗肌筋膜疼痛综合征（MPS）的机制。方法：采用打击结合离心运动方式建立MPS大鼠模型，电针针刺激痛点和塞来昔布灌胃给药干预后，制备电针、塞来昔布大鼠血清。通过脂多糖（LPS）诱导BV-2细胞在MPS的炎症状态，分别用电针大鼠血清、塞来昔布含药血清和BDNF抑制剂(K-252a)进行干预。同时将SD大鼠分为正常组、模型组、电针组和西药组，MPS大鼠模型复制和干预方式同上。实验结束后，通过倒置显微镜观察各组细胞形态，CCK8法检测各组细胞的活性，Elisa法检测各组细胞上清液中IL-1β、IL-6、TNF-α、IL-10水平，免疫细胞化学和免疫荧光检测各组BV-2细胞OX-42的平均光密度值和平均荧光强度，Masson染色观察各组大鼠肌肉组织的形态，RT-PCR检各组细胞、肌肉组织的BDNF、TrkB、P38、JNKmRNA表达水平，Wes检测各组细胞和肌肉组织BDNF、TrkB、p-TrkB、p38、p-P38、JNK、p-JNK的蛋白水平。结果：体外实验各干预方式均能降低各组经LPS诱导BV-2细胞上清液炎性中IL-1β、IL-6、TNF-α水平，升高IL-10水平，同时能降低细胞的活化程度和凋亡水平；电针和西药干预能通过降低各组BV-2细胞和肌肉组织的BDNF、TrkB、P38、JNK的mRNA表达水平和BDNF、TrkB、p-TrkB、p38、p-P38、JNK、p-JNK的蛋白表达水平以降低中枢敏化进而治疗MPS。体外实验BDNF抑制剂和西药组好于电针组，体内实验电针组好于西药组。结论：体外、体内实验结果共同证实针刺激痛点可通过降低中枢敏化治疗MPS，具体机制与抑制BDNF- TrkB-p38/JNK信号通路有关。

Abstract:

Abstract: Objective: This study investigates the mechanism of acupuncture stimulation of myofascial trigger points in the treatment of Myofascial Pain Syndrome (MPS) Based on the BDNF TrkB p38/JNK signaling pathway. Method: Establishing an MPS rat model using a combination of percussion and centrifugal exercise, after intervention with electroacupuncture stimulation of pain points and oral administration of celecoxib, preparing electroacupuncture rat serum. Lipopolysaccharide (LPS) induced BV-2 cell inflammatory state in MPS, and intervene with electroacupuncture rat serum, celecoxib containing serum, and BDNF inhibitor (K-252a), respectively. At the same time, SD rats were divided into normal group, model group, electroacupuncture group, and Western medicine group. The replication method of MPS rat model and intervene method is the same as above. After the experiment, the morphology of each group of cells was observed under an inverted microscope, the activity of each group of cells was detected by CCK8 method, and the level of IL-1β, IL-6, TNF-α, IL-10 in cell supernatant of each group as detected by Elisa. The average optical density and average fluorescence intensity in OX-42 of the cells in each group were detected by immunocytochemistry and immunofluorescence. Masson staining was used to observe the morphology of muscle tissue in each group of rats. RT-PCR was used to detect the expression levels of BDNF, TrkB, P38, and JNK mRNA in each group of cells and muscle tissue. Wes was used to detect the protein levels of BDNF, TrkB, p-TrkB, p38, p-P38, JNK, and p-JNK in each group of cells and muscle tissue. Result: In vitro experiments showed that all intervention methods could reduce the level of IL-1β, IL-6, TNF-α and increase the level of IL-10, while reducing cell activation and apoptosis levels; Electroacupuncture and Western medicine intervention can reduce the mRNA expression levels of BDNF, TrkB, P38, and JNK in each group of BV-2 cells and muscle tissue, as well as the protein expression levels of BDNF, TrkB, p-TrkB, p38, p-P38, JNK, and p-JNK, in order to reduce central sensitization and then treat MPS. In vitro experiments showed that the BDNF inhibitor and Western medicine group performed better than the electroacupuncture group, while in vivo experiments showed that the electroacupuncture group performed better than the Western medicine group. Conclusion: The experimental results in vitro and in vivo confirm that acupuncture stimulation can treat MPS by reducing central sensitization, and the specific mechanism is related to the inhibition the BDNF TrkB p38/JNK signaling pathway.