Duan Zhibing, Sesti Federico
Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, USA.
J Vis Exp. 2013 May 17(75):e50435. doi: 10.3791/50435.
Amylopathy is a term that describes abnormal synthesis and accumulation of amyloid beta (Aβ) in tissues with time. Aβ is a hallmark of Alzheimer's disease (AD) and is found in Lewy body dementia, inclusion body myositis and cerebral amyloid angiopathy (1-4). Amylopathies progressively develop with time. For this reason simple organisms with short lifespans may help to elucidate molecular aspects of these conditions. Here, we describe experimental protocols to study Aβ-mediated neurodegeneration using the worm Caenorhabditis elegans. Thus, we construct transgenic worms by injecting DNA encoding human Aβ42 into the syncytial gonads of adult hermaphrodites. Transformant lines are stabilized by a mutagenesis-induced integration. Nematodes are age synchronized by collecting and seeding their eggs. The function of neurons expressing Aβ42 is tested in opportune behavioral assays (chemotaxis assays). Primary neuronal cultures obtained from embryos are used to complement behavioral data and to test the neuroprotective effects of anti-apoptotic compounds.
淀粉样病变是一个描述β淀粉样蛋白(Aβ)随着时间在组织中异常合成和积累的术语。Aβ是阿尔茨海默病(AD)的标志,在路易体痴呆、包涵体肌炎和脑淀粉样血管病中也有发现(1 - 4)。淀粉样病变会随着时间逐渐发展。因此,寿命较短的简单生物体可能有助于阐明这些病症的分子机制。在此,我们描述了使用秀丽隐杆线虫研究Aβ介导的神经退行性变的实验方案。于是,我们通过将编码人Aβ42的DNA注射到成年雌雄同体线虫的合胞性腺中来构建转基因线虫。通过诱变诱导整合使转化株系稳定。通过收集和播种线虫卵使其年龄同步。在适当的行为测定(趋化性测定)中测试表达Aβ42的神经元的功能。从胚胎中获得的原代神经元培养物用于补充行为数据并测试抗凋亡化合物的神经保护作用。