MicroRNA29家族在帕金森病中的作用与机制研究

Parkinson's disease is the second most common neurodegenerative disease, characterized by the progressive loss of dopaminergic neurons in the substantia nigra of the midbrain. There is still a lack of effective diagnosis and treatments for PD at yet. With research advances in PD, MicroRNA has been proved to play an important role in PD pathology. MicroRNA29 family (miR29s) is including two gene clusters: miR-29a/b1 and miR-29b2/c, which is widely expressed in the central nervous system (CNS). It has been recognized that there exists a closely relationship between miR29s and neurodegenerative diseases, but the specific mechanisms remain to be established. In our previous studies, we found that the levels of miR29s markedly decreased in the serum of PD patients; In a cellular PD model, MPP+ significantly reduced the expression of miR29s; However, miR29s increased dramatically in LPS-treated BV2 cells, indicating that miR29s might be involved in PD by the regulation of neuronal apoptosis and the activation of glial cells. In addition, the miR-29a/b1 and miR-29b2/c gene deficient mice and cell lines have been generated. In this project, we will decipher the physiological functions of miR29s in the brain and demonstrate the functions and mechanisms of miR29s in PD. Additionally, the possibility of miR29s in serum as a biomarker of PD and PD HY scales will be determined. Our work might provide new diagnostic and therapeutic alterations for PD.

帕金森病(Parkinson's disease, PD)是中枢神经系统第二大退行性病，以中脑黑质多巴胺能神经元进行性丢失为特征，目前仍缺乏非常有效的诊治手段。MicroRNA在PD病理机制中具有重要作用。MicroRNA29家族包括两个基因簇miR-29a/b1与b2/c，其在脑内的功能尚未明确。前期我们发现在PD病人的血清中miR-29s含量显著降低；MPP+处理的多巴胺能细胞中miR-29s表达亦明显下降；而在LPS处理的小胶质细胞内miR-29s表达则显著上升，提示miR-29s可能通过调节神经元凋亡和胶质细胞激活参与了PD；我们进而构建了miR-29a/b1与b2/c基因缺失突变的小鼠和细胞系。本申请拟利用这些模型并结合临床样本进一步阐明miR-29s在脑内的功能及在PD发病中的作用和机制，探讨血液中miR-29s含量作为PD发病的生物标志物的可行性，为PD的诊治提供新的思路。

miR-29家族包括miR-29a和miR-29b-1以及miR-29b-2和miR-29c。miR-29家族在帕金森病(PD)发病中的作用与机制是本课题所关注的。我们发现，与正常人群相比，miR-29s表达水平在PD患者的血清中明显下调；而在PD患者的脑脊液中，特别是女性PD患者的脑脊液中miR-29a表达水平升高，提示miR-29s与PD发病相关。进一步利用CRISPR-Cas9技术构建miR-29a/b-1和miR-29b-2/c基因敲除小鼠(以下称为29aKO和29cKO)，研究了miR-29s在衰老和PD中的作用。与野生型(WT)小鼠相比，29aKO小鼠和29cKO小鼠均表现出显着的体重减轻、后凸畸形、真皮明显增厚以及皱纹增加和加深。此外，29aKO和29cKO小鼠的腹部和棕色脂肪组织均显著减少。然而在MPTP诱导的PD模型中，miR-29s缺失降低了多巴胺能系统的损伤程度，减轻了黑质-纹状体通路中神经胶质细胞的激活，从而缓解了小鼠的运动障碍。从机制上讲，MPP+处理的原代混合胶质细胞内，miR-29a/b-1缺失促进了神经营养因子的表达，而miR-29b-2/c缺失则抑制了炎性因子的表达；在MPP+处理的原代星形胶质细胞中，miR-29s缺失下调了炎症因子的表达；在LPS处理的原代小胶质细胞中，敲除miR-29a/b-1和miR-29b-2/c均抑制了炎症因子的表达，敲除miR-29a/b-1还促进了抗炎因子和神经营养因子的表达。在信号通路方面，敲除miRNA-29s可增加神经胶质细胞内AMPK活性，抑制NF-κB/p65信号传导。我们的结果表明29aKO和29cKO小鼠表现出外周早衰；而在MPTP诱导的小鼠PD模型中，miR-29s缺失的神经胶质细胞激活减弱，炎症反应减轻，进而起到多巴胺能神经保护作用。