电针对多囊卵巢综合征小鼠机械痛阈及TRPV 1和CGRP表达的影响

Effects of electroacupuncture on mechanical pain threshold and expression of TRPV 1 and CGRP in mice with polycystic ovary syndrome

摘要:

目的：观察电针关元及敏化点对多囊卵巢综合征 (PCOS) 小鼠关元穴、敏化点机械痛阈值及T13-L2节段脊髓、背根神经节 (DRG) 中瞬时感受器电位香草酸受体1 (TRPV 1)、降钙素基因相关肽 (CGRP) 表达的影响，探讨PCOS小鼠模型体表敏化的部分机制。方法：32只SPF级成年雌性ICR小鼠，随机分为对照组、模型组、电针关元组、电针敏化点组，各8只。采用双酚A灌胃的方式制备PCOS小鼠模型，对照组以同等条件进行玉米油灌胃；电针关元及敏化点，每天20 min，1次/天，连续5天，共治疗4周。电针结束后第3天，用电子Von Frey测量小鼠体表敏化点及关元穴的机械痛阈值；HE染色法观察小鼠卵巢组织病理形态；蛋白印迹法、免疫荧光法检测脊髓、DRG组织中TRPV 1、CGRP表达水平。结果：与对照组比较，模型组卵巢组织囊性扩张、闭锁卵泡增加，关元穴、敏化点痛阈值显著下降（P < 0.01），脊髓组织中TRPV 1及CGRP蛋白表达显著升高（P < 0.05、P < 0.01）、DRG组织中TRPV 1及CGRP蛋白表达显著升高（P < 0.05），脊髓中TRPV 1/CGRP共表达光密度显著升高（P < 0.01）、DRG组织中TRPV 1/CGRP共表达神经元显著增加（P < 0.05）；与模型组比较，电针关元组及电针敏化点组卵泡及黄体发育良好，电针关元组关元穴及敏化点痛阈值显著升高（P < 0.05）、电针敏化点组关元穴及敏化点痛阈值显著升高（P < 0.05、P < 0.01），电针关元组和电针敏化点组脊髓、DRG组织中TRPV 1及CGRP蛋白表达显著降低（P < 0.05）、脊髓中TRPV 1/CGRP共表达光密度显著降低（P < 0.01）、DRG组织中共表达神经元显著减少（P < 0.01）；电针关元组与电针敏化点组比较，差异均无统计学意义（P > 0.05）。结论：电针敏化点与关元可提高PCOS小鼠关元穴及敏化点机械痛阈值，调节PCOS小鼠体表敏化现象，其作用机制可能与下调TRPV 1、CGRP的异常高表达有关。

Abstract:

Objective: observe the effects of electroacupuncture(EA) at Guan Yuan (CV 4) and sensitization point on the mechanical pain threshold of CV 4 point and sensitization point and the expression of transient receptor potential vanilloid 1 (TRPV 1) and calcitonin gene related peptide (CGRP) in T13-L2 segment spinal cord and dorsal root ganglion (DRG) in mice with polycystic ovary syndrome (PCOS), to explore partial mechanisms of PCOS mice model body surface sensitization. Methods: 32 SPF adult female ICR mice were randomly divided into control group，model group, electroacupuncture (EA) Guanyuan (CV 4) group and EA-sensitization points group, with 8 mice in each group. PCOS mice models were intragastrically administered with bisphenol A, and the control group were intragastrically administered with corn oil. EA-CV 4 and EA-sensitization point, 20 min a day, once a day, for 5 consecutive days, a total of 4 weeks . On the third day after the EA, the mechanical pain threshold of the sensitized point on the body surface and CV 4 were measured by electron Von Frey . HE staining was used to observe the pathological morphology of mice ovarian tissue. The expression levels of TRPV 1 and CGRP in spinal cord and DRG tissues were detected by Western blot and Immunofluorescence. Results Compared with the control group, the cystic dilatation and atresia of ovarian tissue increased in the model group, and the pressure pain threshold of CV 4 acupoint and sensitized point were significantly decreased (P < 0.01), and the protein expressions of TRPV 1 and CGRP in spinal cord tissue were significantly increased(P < 0.05，P < 0.01)，the expression of TRPV 1 and CGRP protein in DRG tissue were significantly increased （P < 0.05），the optical density of TRPV 1/CGRP co-expression in spinal cord was significantly increased （P < 0.01），and the TRPV 1/CGRP co-expression neurons in the DRG was significantly increased（P < 0.05）.Compared with model group， the follicles and luteum of the EA-CV 4 group and the EA-sensitization points group were well developed，the pressure pain threshold of CV 4 acupoint and sensitized point in the EA-CV 4 group were significantly increased (P < 0.05), the pressure pain threshold of CV 4 acupoint and sensitized point in the EA-sensitization points group were significantly increased (P < 0.05，P < 0.01), the protein expression of TRPV 1 and CGRP in spinal cord and DRG tissues were significantly decreased (P < 0.05), and the optical density of TRPV 1/CGRP co-expression were significantly decreased (P < 0.01, P < 0.05), and the TRPV 1/CGRP co-expression neurons in the DRG was significantly decreased（P < 0.05） . There was no significant difference between EA-CV 4 and EA-sensitization points group (P > 0.05). Conclusion EA-sensitization points and EA-CV 4 can increase the mechanical pain threshold of CV 4 point and sensitization point of PCOS mice, and regulate the body surface sensitization of PCOS mice. The mechanism of which may be related to the down-regulation of the abnormally high expression of TRPV 1 and CGRP.