基于RS-fMRI及ERK-CREB-BDNF信号通路杜仲腰痛丸干预腰椎间盘突出症慢性疼痛的脑机制研究

Because the mechanism for lumbar disc herniation (LDH) is not clear, which results in noneffective reaction for management, and gives serious burden for patients and society. Currently, it is found that patients’ structure and function of brain have reconstructed by cerebral function imaging, because of that could leads to long painful pathogenesis and correlated with severity , effectiveness and prognosis. Some study demonstrated that signal molecules, such as ERK, CREB and BDNF, are important for happening and persistence of chronic pain. Our study certificated that Duzhongyaotong Pills can relive pain of LDH. So, we hypothesis that LDH activated signal pathway of ERK-CREB-BDNF and gave rise to transformation of structure and function of brain, as well as led to persistence for pain in waist and lower extremities. Duzhongyaotong Pills would improve structure and function of brain and play a role of relieve pain via regulate signal pathway of ERK-CREB-BDNF. This study will product pain model of lower extremities, employ magnetic resonance morphology technology based on voxel, Resting state functional magnetic resonance technology, pain behavior and molecular biology technique to verify this hypothesis. This research will explore brain mechanism of chronic pain in lower extremities result from LDH and supply experimental and theoretical evidence for novel target spot and improve effectiveness. So, this project will has importantly value.

由于致痛机制不明，腰椎间盘突出症（LDH）慢性腰腿痛缺乏有效的治疗响应，疼痛迁延难愈，给患者和社会造成沉重负担。近年脑功能成像技术发现慢性痛者的大脑会发生结构和功能的重塑，由此可能导致慢性疼痛的长期维持，并与严重程度、治疗效果及愈后密切相关。研究表明ERK、CREB、BDNF等信号分子对慢性疼痛的发生和维持起着重要作用。前期研究表明，杜仲腰痛丸能有效缓解LDH腰腿痛。因此我们假设：腰椎间盘突出激活了大脑的ERK-CREB-BDNF信号通路，进而引起大脑结构和功能的重塑，导致慢性腰腿痛的长期维持。杜仲腰痛丸通过调控ERK-CREB-BDNF信号通路，改善大脑结构和功能的重塑，而发挥止痛效应。本项目拟通过建立LDH慢性下肢痛大鼠模型，采用基于体素的磁共振形态学、静息态功能磁共振、疼痛行为学及分子生物学等技术验证此假说。本项目可望能初步揭示LDH慢性痛的脑机制，发现治疗新靶点，具有重大意义。

腰椎间盘突出症（LDH）慢性腰腿痛缺乏有效的治疗响应；前期研究表明，杜仲腰痛丸能有效缓解LDH慢性腰腿痛，但作用机制不明，本课题研究目的在于寻找杜仲腰痛丸治疗LDH慢性腰腿痛的效应环节和作用靶点，结合运用医学物理学、生物信息学、蛋白质生物化学等方法与技术，对杜仲腰痛丸治疗LDH慢性腰腿痛进行了影像学和分子机制的初步研究。首先通过自体髄核移植方法建立LDH慢性下肢痛大鼠模型，利用静息态功能磁共振（RS-fMRI）及脑功能分化低频振幅率分析（ALFF）、比率低频振幅率分析（fALFF）、RS-fMRI局部一致性分析（ReHo）的方法分析了杜仲腰痛丸干预前后LDH慢性下肢痛模型大鼠脑功能连接的变化差异，除此，利用基于体素的磁共振形态学（VBM）方法分析了药物干预前后LDH 慢性下肢痛大鼠脑区灰质密度间的差异，然后取海马切片，利用免疫荧光（IF）、实时荧光定量PCR(RT-qPCR)和蛋白免疫印迹（WB），观察分析杜仲腰痛丸干预前后ERK-CREB-BDNF信号通路关键信号分子 Ras、Raf、ERK、CREB、BDNF、TrkB蛋白及mRNA的表达水平。结果显示杜仲腰痛丸干预后LDH慢性下肢痛大鼠模型的ReHo值显著增高的脑区有左初级躯体运动皮层，ALFF值存在显著差异的脑区是左内嗅皮层，fALFF值显著增高的脑区有左外侧内嗅皮层、右海马；VBM技术分析后，模型大鼠左外侧内嗅皮层、左初级躯体感觉皮层、右嗅球、右视皮层、右海马、胼胝体等脑区的灰质体积显著增加；IF、RT-PCR和WB技术检测大鼠海马切片后，ERK-CREB-BDNF 信号通路关键信号分子 Ras、Raf、ERK、CREB 、BDNF、TrkB蛋白及mRNA 及的表达量明显升高。综合上述结果，本研究为探讨中药复方杜仲腰痛丸治疗LDH慢性下肢痛发挥效应的靶点和作用机制奠定了坚实的工作基础，同时为中医腰痛与脑的关系提供了影像学和分子学依据，进一步丰富了腰痛的脏象理论，为中医关于腰痛的先进理论的推广做了科学铺垫。. 培养硕士生4名，均已取得硕士学位。发表高质量研究论文6篇，项目投入经费34万元，支出33.9002万元，各项支出基本与预算相符。剩余经费0.0998万元，剩余经费计划用于本项目研究后续支出。