Farzanehfar Parisa, Horne Malcolm K, Aumann Tim D
Florey Institute for Neuroscience & Mental Health, The University of Melbourne, Parkville, Victoria 3010, Australia.
Florey Institute for Neuroscience & Mental Health, The University of Melbourne, Parkville, Victoria 3010, Australia.
Neurosci Lett. 2017 May 1;648:34-40. doi: 10.1016/j.neulet.2017.03.028. Epub 2017 Mar 29.
Generation of new dopamine (DA) neurons in the adult midbrain is a controversial issue in development of better treatments for Parkinson's disease (PD). Previous research suggests Nestin-expressing neural precursor cells (NPCs) have a propensity to differentiate into neurons here, including DA neurons. In the present study we sought confirmation of this by studying gene expression in single Nestin-expressing cells and their progeny/ontogeny within the adult mouse midbrain. Cells were identified by administering a pulse of Tamoxifen to adult Nestin-CreER×R26eYFP transgenic mice. Samples of cytoplasm were harvested 4 days to 8 months later from individual eYFP+ cells in acutely prepared midbrain slices and analysed by RT-qPCR for gene expression. Remarkably, most eYFP+ cells co-expressed genes associated with mature (including DA) neurons (i.e. NeuN, Gad1, Gad2, vGlut2, TH and/or D2R) and neurogenesis (i.e. Ki67, Dcx, Ncam, Pax6, Ngn2 and/or Msx1), and this was true at all time-points following Tamoxifen. Indeed, cell proliferation genes (Nestin, Ki67) were exclusively expressed by eYFP+ cells with mature neuronal morphology and gene expression, and only at early time-points after Tamoxifen. Expression of proneuronal genes (Pax6, Msx1, Ngn2) was, however, higher in eYFP+ cells with immature morphology compared with mature morphology. Gene expression bore no relationship to cell location indicating that, in contrast to development, Nestin-expressing cells arise throughout the midbrain parenchyma and do not migrate long distances. On the other hand, gene expression did change with time after Tamoxifen, although not in a way consistent with neurogenesis. Overall, our results suggest that Nestin expression in the adult midbrain occurs in mature neurons, casting doubt on the premise of neurogenesis from Nestin+ NPCs here.
在成体中脑生成新的多巴胺（DA）神经元是帕金森病（PD）更好治疗方法开发中的一个有争议的问题。先前的研究表明，表达巢蛋白的神经前体细胞（NPCs）有在此处分化为神经元的倾向，包括DA神经元。在本研究中，我们通过研究成年小鼠中脑单个表达巢蛋白的细胞及其后代/个体发育中的基因表达来寻求对此的证实。通过向成年Nestin-CreER×R26eYFP转基因小鼠给予他莫昔芬脉冲来鉴定细胞。在4天至8个月后，从急性制备的中脑切片中的单个eYFP+细胞中收集细胞质样本，并通过RT-qPCR分析基因表达。值得注意的是，大多数eYFP+细胞共表达与成熟（包括DA）神经元相关的基因（即NeuN、Gad1、Gad2、vGlut2、TH和/或D2R）和神经发生相关的基因（即Ki67、Dcx、Ncam、Pax6、Ngn2和/或Msx1），并且在给予他莫昔芬后的所有时间点都是如此。事实上，细胞增殖基因（Nestin、Ki67）仅由具有成熟神经元形态和基因表达的eYFP+细胞表达，并且仅在给予他莫昔芬后的早期时间点表达。然而，与成熟形态相比，具有未成熟形态的eYFP+细胞中神经前体基因（Pax6、Msx1、Ngn2）的表达更高。基因表达与细胞位置无关，这表明与发育过程不同，表达巢蛋白的细胞出现在整个中脑实质中，并不进行长距离迁移。另一方面，给予他莫昔芬后的基因表达确实随时间变化，尽管其变化方式与神经发生不一致。总体而言，我们的结果表明，成年中脑中巢蛋白的表达发生在成熟神经元中，这对在此处由巢蛋白+ NPCs进行神经发生的前提提出了质疑。
