Karttunen Marja J, Lyons David A
Centre for Discovery Brain Sciences, The University of Edinburgh, Edinburgh, UK.
Methods Mol Biol. 2019;1936:227-238. doi: 10.1007/978-1-4939-9072-6_13.
To study cellular and molecular mechanisms of demyelination and remyelination in vivo, we developed a transgenic zebrafish line, Tg(mbp:mCherry-NTR), in which expression of the bacterial enzyme nitroreductase (NTR) is driven under the myelin basic protein promoter (mbp) and thus is expressed in myelinating glia. When NTR-expressing larvae are treated with the prodrug metronidazole, the reaction between NTR and Mtz results in a toxic metabolite which selectively kills NTR-expressing cells. Using the Tg(mbp:mCherry-NTR) line, we can ablate two-thirds of oligodendrocytes following a 2-day MTZ treatment. Demyelination is evident seven days later, and remyelination is observed 16 days after Mtz treatment. The Tg(mbp:mCherry-NTR) model can be used to image cell behavior during, and to test how genetic manipulations or chemical compounds regulate, demyelination and remyelination. In this chapter, we describe the methods we used to characterize the oligodendrocyte loss, demyelination and remyelination in the Tg(mbp:mCherry-NTR) model.
为了研究体内脱髓鞘和再髓鞘化的细胞和分子机制，我们构建了一种转基因斑马鱼品系Tg(mbp:mCherry-NTR)，其中细菌酶硝基还原酶（NTR）的表达由髓鞘碱性蛋白启动子（mbp）驱动，因此在髓鞘形成胶质细胞中表达。当用前药甲硝唑处理表达NTR的幼虫时，NTR与甲硝唑之间的反应会产生一种有毒代谢物，该代谢物会选择性地杀死表达NTR的细胞。使用Tg(mbp:mCherry-NTR)品系，经过2天的甲硝唑处理后，我们可以消除三分之二的少突胶质细胞。7天后脱髓鞘明显，甲硝唑处理16天后观察到再髓鞘化。Tg(mbp:mCherry-NTR)模型可用于在脱髓鞘和再髓鞘化过程中对细胞行为进行成像，并测试基因操作或化合物如何调节脱髓鞘和再髓鞘化。在本章中，我们描述了用于表征Tg(mbp:mCherry-NTR)模型中少突胶质细胞损失、脱髓鞘和再髓鞘化的方法。
