Breid Sara, Bernis Maria E, Tachu Julius B, Garza Maria C, Wille Holger, Tamgüney Gültekin
German Center for Neurodegenerative Diseases (DZNE).
Centre for Prions and Protein Folding Diseases & Department of Biochemistry, University of Alberta.
J Vis Exp. 2017 Apr 13(122):55503. doi: 10.3791/55503.
To study the prion-like behavior of misfolded alpha-synuclein, mouse models are needed that allow fast and simple transmission of alpha-synuclein prionoids, which cause neuropathology within the central nervous system (CNS). Here we describe that intraglossal or intraperitoneal injection of alpha-synuclein fibrils into bigenic Tg(M83:Gfap-luc) mice, which overexpress human alpha-synuclein with the A53T mutation from the prion protein promoter and firefly luciferase from the promoter for glial fibrillary acidic protein (Gfap), is sufficient to induce neuropathologic disease. In comparison to homozygous Tg(M83) mice that develop severe neurologic symptoms beginning at an age of 8 months, heterozygous Tg(M83:Gfap-luc) animals remain free of spontaneous disease until they reach an age of 22 months. Interestingly, injection of alpha-synuclein fibrils via the intraperitoneal route induced neurologic disease with paralysis in four of five Tg(M83:Gfap-luc) mice with a median incubation time of 229 ±17 days. Diseased animals showed severe deposits of phosphorylated alpha-synuclein in their brains and spinal cords. Accumulations of alpha-synuclein were sarkosyl-insoluble and colocalized with ubiquitin and p62, and were accompanied by an inflammatory response resulting in astrocytic gliosis and microgliosis. Surprisingly, inoculation of alpha-synuclein fibrils into the tongue was less effective in causing disease with only one of five injected animals showing alpha-synuclein pathology after 285 days. Our findings show that inoculation via the intraglossal route and more so via the intraperitoneal route is suitable to induce neurologic illness with relevant hallmarks of synucleinopathies in Tg(M83:Gfap-luc) mice. This provides a new model for studying prion-like pathogenesis induced by alpha-synuclein prionoids in greater detail.
为了研究错误折叠的α-突触核蛋白的朊病毒样行为，需要能够快速简单地传播α-突触核蛋白类朊病毒的小鼠模型，这些类朊病毒会在中枢神经系统（CNS）内引起神经病理学变化。在此，我们描述了向双转基因Tg(M83:Gfap-luc)小鼠舌内或腹腔注射α-突触核蛋白纤维，该小鼠从朊病毒蛋白启动子过表达具有A53T突变的人α-突触核蛋白，并从胶质纤维酸性蛋白（Gfap）启动子过表达萤火虫荧光素酶，足以诱发神经病理学疾病。与从8个月大开始出现严重神经症状的纯合Tg(M83)小鼠相比，杂合Tg(M83:Gfap-luc)动物在22个月龄之前没有自发性疾病。有趣的是，通过腹腔途径注射α-突触核蛋白纤维在五只Tg(M83:Gfap-luc)小鼠中的四只诱发了伴有麻痹的神经疾病，中位潜伏期为229±17天。患病动物的大脑和脊髓中出现了严重的磷酸化α-突触核蛋白沉积。α-突触核蛋白的聚集物对 Sarkosyl 不溶，与泛素和p62共定位，并伴有炎症反应，导致星形胶质细胞增生和小胶质细胞增生。令人惊讶的是，将α-突触核蛋白纤维接种到舌头中诱发疾病的效果较差，五只注射动物中只有一只在285天后出现α-突触核蛋白病理学变化。我们的研究结果表明，通过舌内途径接种，更重要的是通过腹腔途径接种，适合在Tg(M83:Gfap-luc)小鼠中诱发具有突触核蛋白病相关特征的神经疾病。这为更详细地研究由α-突触核蛋白类朊病毒诱导的朊病毒样发病机制提供了一个新模型。
