脑靶向输送新型神经营养因子MANF的磁性纳米载体构建及其在帕金森病治疗中的应用

Novel neurotrophic factors CDNF/MANF have received considerable attentions for their specific protection of dopaminergic neurons. However, they hardly enter the brain due to the presence of blood brain barrier (BBB), which decreases the therapeutic effects of Parkinson's disease (PD) significantly. Therefore, it has great importance to construct a targeted nanocarrier for brain delivery of NTFs. To construct such targeted delivery system, the self-made magnetic mesoporous Fe3O4@mSiO2 nanoparticles are employed as the carrier, the common ligand lactoferrin (Lf) of the BBB and PD lesion site is selected as the targeting molecule and NTFs are encapsulated in the Lf-modified nanoparticles. In vitro toxicity of the nanocarrier will be carried on the b.End3 cell lines by using MTT assay and the intracellular uptake will be examined by using CLSM and FACS analysis. Before the in vivo experiment, NTFs are loaded into the nanopores of the mesoporous layer on the surface of the nanocarrier. Then MANF-His protein delivered to the brain is verified by double immunofluorescence staining method. Finaly, the therapeutic effects of the nano carrier system are investigated on the built PD rat model via the behavioral test, the neuron surviral/apoptosis analysis, and striatal transmitter content detection.

新型神经营养因子家族的CDNF/MANF因能特异性保护多巴胺能神经元而倍受关注，但由于血脑屏障（BBB）的存在难以通过外周给药到达脑内，从而影响了其在帕金森病中的应用。因此，构建具有靶向功能的纳米载体，对实现MANF在脑内的成功转输具有重要意义。本项目拟以磁性/介孔Fe3O4@mSiO2纳米粒子作为载体，利用纳米粒子表面的介孔SiO2层实现对重组人MANF-His的包载；选择BBB和病灶组织高表达的乳铁蛋白受体（LfR）的配体Lf作为靶向分子，使其穿越BBB后在病灶部位富集；通过检测小鼠脑毛细管内皮细胞b.End3的活性和增殖情况来评价纳米载体的毒性；通过流式细胞仪和激光共聚焦显微镜等研究细胞摄取及胞内定位；用免疫荧光双标法鉴定被输送到脑内的MANF-His；在大鼠帕金森模型上，通过检测行为学、神经元存活/凋亡、神经递质等来考察该载药系统递送的MANF对帕金森病的治疗作用。