Collins Will, Rouleau Nicolas, Bonzanni Mattia, Kapner Kevin, Jeremiah Alex, Du Chuang, Pothos Emmanuel N, Kaplan David L
Department of Pharmacology & Experimental Therapeutics, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Tufts University, Boston, Massachusetts 02111, United States.
Department of Biomedical Engineering, Tufts University, 4 Colby Street, Medford, Massachusetts 02155, United States.
ACS Biomater Sci Eng. 2019 Jan 14;5(1):308-317. doi: 10.1021/acsbiomaterials.8b00976. Epub 2018 Dec 17.
The substantia nigra pars compacta (SNpc) is a discrete region of the brain that exhibits a dark pigment, neuromelanin (NM), a biomaterial with unique properties and the subject of ongoing research pertaining to neurodegenerative conditions like Parkinson's disease (PD). Obtaining human tissue to isolate this pigment is costly and labor intensive, making it necessary to find alternatives to model the biochemical interaction of NM and its implications on PD. To address this limitation, we modified our established silk 3D brain tissue model to emulate key characteristics of the SNpc by using a structural analogue of NM to examine the effects of the material on dopaminergic neurons using Lund's human mesencephalon (LUHMES) cells. We utilized a sepia-melanin, squid ink, derived NM analogue (NM-sim) to chelate ferric iron, and this iron-neuromelanin precipitate (Fe-NM) was purified and characterized. We then exposed LUHMES dopaminergic cells to the NM-sim, Fe-NM-sim, and control vehicle within 3D silk protein scaffolds. The presence of both NM-sim and Fe-NM-sim in the scaffolds negatively impacted spontaneous electrical activity from the LUMES networks, as evidenced by changes in local field potential (LFP) electrophysiological recordings. Furthermore, the Fe-NM-sim precipitate generated peroxides, depleted nutrients/antioxidants, and increased protein oxidation by carbonylation in sustained (>2 weeks) 3D cultures, thereby contributing to cell dysfunction. The results suggest that this 3D tissue engineered brain-like model may provide useful readouts related to PD neuro-toxicology research.
黑质致密部（SNpc）是大脑中的一个离散区域，其呈现出一种深色色素，即神经黑色素（NM），这是一种具有独特性质的生物材料，也是目前关于帕金森病（PD）等神经退行性疾病研究的主题。获取人体组织来分离这种色素成本高昂且劳动强度大，因此有必要寻找替代方法来模拟NM的生化相互作用及其对PD的影响。为了解决这一局限性，我们对已建立的丝绸三维脑组织模型进行了改进，通过使用NM的结构类似物来模拟SNpc的关键特征，利用隆德人脑中脑（LUHMES）细胞研究该材料对多巴胺能神经元的影响。我们使用了一种乌贼墨衍生的NM类似物（NM-sim）来螯合三价铁，然后对这种铁 - 神经黑色素沉淀物（Fe-NM）进行了纯化和表征。接着，我们将LUHMES多巴胺能细胞暴露于三维丝绸蛋白支架中的NM-sim、Fe-NM-sim和对照载体中。支架中NM-sim和Fe-NM-sim的存在均对LUMES网络的自发电活动产生负面影响，局部场电位（LFP）电生理记录的变化证明了这一点。此外，在持续（>2周）的三维培养中，Fe-NM-sim沉淀物产生过氧化物，消耗营养物质/抗氧化剂，并通过羰基化增加蛋白质氧化，从而导致细胞功能障碍。结果表明，这种三维组织工程化的类脑模型可能为与PD神经毒理学研究相关的有用读数提供依据。
