基于GnRH神经元kisspeptin-GPR54通路探究振腹法治疗寒凝证类痛经大鼠的机理

Nowadays, cold coagulating and blood stasis syndrome dysmenorrhea has attracted the attention of the medical community. The patients troubled by feelings of cold and painful, which seriously affects their physical and mental health. Early research has shown that abdominal vibration method can warm the channel and alleviating pain.The abdominal vibration method(1) is preliminarily established as an effective treatment of cold coagulating and blood stasis syndrome dysmenorrhea patients based on improved VAS scores;(2)has kinematic patterns, and the optimal vibrational parameters have ascertained. However, the mechanism of abdominal vibration method has not been elucidated yet. This study aims to give cold coagulating and blood stasis syndrome model rats treatment using an abdominal vibration method simulator to maintain quantitative and qualitative controls. The parameters of time, frequency and amplitude will be assessed. Ethology, morphology and functional science will be used as the primary index of effectiveness, cytological histology and molecular biology techniques will be used to detect changes in the expression of hypothalamic CNS regulating target molecules (key indicators of the kisspeptin - GPR54 signaling pathway), HPOA endocrine regulating hormone and receptor (FSH, LH, E2, P and related receptors), and uterine local analgesic target molecules (PGF2α and β-EP). In order to explore the mechanism of applying abdominal vibration method on cold coagulating syndrome dysmenorrhea model rats from the perspective of GnRH neuron kisspeptin-GPR54 signal pathway regulating HPOA and then regulating changes of uterine local analgesic indexes.

临床中以“寒”、“痛”两症状为主的寒凝证痛经严重影响患者的身心健康和生活质量，倍受医学界关注。课题组前期研究表明：具有温经通络、缓急止痛作用的振腹法①可改善寒凝血瘀证痛经患者VAS评分，初步确立振腹法治疗寒凝证痛经有效性；②有一定运动学规律，已确定优效振腹参数。但振腹法治疗寒凝证痛经机理尚未阐明。本课题建立寒凝证类痛经模型大鼠，以振动时间、频率、幅度为参数，采用振腹模拟仪定时定量模拟手法，将行为学、形态学、功能学作为评价标准，以组织细胞学及分子生物技术检测寒凝证类痛经大鼠下丘脑中枢调控指标（kisspeptin-GPR54信号通路关键指标）、HPOA中关键激素及受体（FSH、LH、E2、P及相关受体）、子宫局部镇痛指标（PGF2α及β-EP）的变化，以期从GnRH神经元kisspeptin-GPR54信号通路调控HPOA，进而调控子宫局部镇痛指标变化的层面，探究振腹法治疗寒凝证痛经的机理。

前期研究中围绕振腹推拿疗法申报了一系列的理论和临床课题，进一步证实了振腹疗法在治疗临床内、妇科疾病的优势。虽然振腹推拿疗法治疗寒凝血瘀型痛经有一定的临床基础，但其机理研究尚不充足。振腹推拿手法可以调节神经内分泌系统，从而达到治疗痛经的目的，本研究设想振腹法是否能够通过调控HPOA，进而影响子宫局部相关指标，从而达到温经止痛的治疗效果？因此本实验研究以寒凝血瘀型痛经模型大鼠为研究对象，通过观察各组大鼠扭体反应及子宫体表投影温度，并以组织细胞学及分子生物技术检测寒凝血瘀型痛经大鼠下丘脑中GnRH，HPOA中关键激素及子宫局镇痛指标的表达变化，以期通过调控中枢通路影响HPOA，进而调控子宫局部镇痛指标的表达变化探究振腹法治疗寒凝血瘀型痛经的温经止痛作用机理。.研究表明，模型组大鼠血清性激素FSH、LH水平与对照组比较明显升高，P水平降低，差异具有统计学意义（P<0.05）；振腹组血清性激素FSH、LH水平与模型组比较明显降低，P水平升高，差异具有统计学意义（P<0.05）。FSH与LH通过血液循环，与卵巢表面的FSHR及LHR特异性结合，刺激卵巢合成释放E2及P。E2及P对于子宫局部PGF2α及β-EP含量的作用是相互制约，进而维持子宫平滑肌的正常生理功能。振腹法干预机体HPOA的信号传导途径，使大鼠血清性激素FSH、LH水平降低，P水平升高，抑制卵巢合成释放E2，进而降低子宫局部PGF2α含量，提高β-EP水平，缓解痛经症状。而且提取大鼠下丘脑及垂体组织检测发现，与模型组比较，振腹组下丘脑GnRH与垂体前叶GnRH-R表达降低，差异具有统计学意义（P<0.05），从而进一步减少垂体前叶分泌促性腺激素FSH及LH。因此，振腹法可以通过降低下丘脑GnRH与GnRH-R表达，进一步减少促性腺激素FSH及LH释放，增加P值水平，进而降低子宫局部PGF2α含量，提高β-EP水平，最终缓解痛经症状。因此，振腹法可通过下调下丘脑GnRH及相关受体的表达，调节HPOA中相关性激素含量，进而在子宫局部抑制PGF2α的生成并上调β-EP含量，达到“温经止痛”的作用。