基于微流控芯片的发育性、变形性大脑疾患的表观遗传学调控机制研究

The study on neuroprotection and neuroregeneration has been a hotspot and difficulty of biomedical research. In the central nervous system, the epigenetic regulation of gene expression is critical for developmental processes and for complex brain functions. Dysregulation of the epigenetic profile of nerve cells can be detrimental, and has been associated with neurodevelopmental and neurodegenerative disorders. The dynamics and reversibility of epigenetic regulation may provide new options for potential diagnostic and for the development of epigenetic drug treatment.For finding the target proteins associated with brain disorders, this project plans to use ELISA to analyze specific target protein expression differences of endothelial cells (E), astrocytes(A) and neuron(N).Furthermore, the molecular biology and biochemical techniques such as RT-PCR, Western Blot, Immunofluorescence cytochemistry staining, and methylation specific PCR would be used to observe the influences of epigenetic regulation on the target proteins expression in vitro.Finally, high throughput DNA methylation chip will be used to detect clinical samples. Using Transwell coculture system, we has established a novel four-dimensional model in primary culture of tricellular elements, E, A, and N, from same-origin of brain or spinal cord. This four-dimensional model provides a accurate and reliable method for in vitro study of epigenetic regulation mechanism on neurodevelopmental and neurodegeneration disorders. This study will explore new ideas of deeply understanding pathogenesis of neurodevelopmental and neurodegeneration disorders, and screening, investigating and eveluating drugs with effects of neuroprotection and/or neuroregeneration.

脑保护与脑损伤修复研究一直是生物医药研究领域的热点和难点。表观遗传学调控机制在发育性、变性性大脑疾患发展中起着至关重要的作用，且具有可逆性，为疾病治疗提供了乐观的前景。本研究拟采用ELISA分析神经细胞，如毛细管内皮细胞（E）、星形胶质细胞（A）、神经元（N）特异性靶蛋白的表达差异，寻找与大脑疾患相关的蛋白靶分子；采用RT-PCR、Western Blot、免疫荧光细胞化学染色以及甲基化特异性PCR方法体外研究影响各蛋白靶分子表达的表观遗传学调控机制；利用高通量的DNA甲基化微流控芯片分析检测临床样品。为进行体外研究，我们首次利用Transwell将来源于同一种属动物同一部位的E、A、N三种原代细胞进行立体共培养，构建原代同源EAN三细胞四维模型，为机制的体外研究提供更准确可靠的研究载体。本研究将为深入了解发育性、变性性大脑疾患的发病机制和研发新型治疗药物拓展新思路。

大脑疾患是最重大、最复杂的健康负担和医学难题之一。本研究通过对临床样本检测提出UCH-L1可能是反映脑白质病变程度的一个生物标记物；通过甲基化芯片分析MS和NMO病人临床样本中表观遗传学差异，为疾病诊断治疗提供指导意义；构建MS动物模型及LPS诱导的神经炎症模型，进一步研究PGE2通路在神经退行性疾病神经炎症中的调控作用。本项目的顺利实施为深入了解神经退行性疾病的发病机制和研发新型治疗药物拓展新思路。项目资助发表SCI论文2篇，接收SCI论文1篇，待发表1篇。辅助培养硕士研究生3名，其中2名已取得硕士学位，1名在读。国际会议上口头汇报1次。项目投入经费26万，支出22.77万元，各项支出基本与预算相符。剩余经费3.23万元，剩余经费计划用于本项目研究后续支出。