Machado Lucas Simões, Recchia Kaiana, Pieri Naira Caroline Godoy, Botigelli Ramon Cesar, de Castro Raquel Vasconcelos Guimarães, Brunhara Cruz Jéssica, Pessôa Laís Vicari de Figueiredo, Bressan Fabiana Fernandes
Department of Surgery, Faculty of Veterinary Medicine and Animal Sciences, University of São Paulo.
Department of Surgery, Faculty of Veterinary Medicine and Animal Sciences, University of São Paulo; Department of Veterinary Medicine, Faculty of Animal Sciences and Food Engineering, University of São Paulo.
J Vis Exp. 2021 Jun 11(172). doi: 10.3791/62209.
iPSC-derived neurons are attractive in vitro models to study neurogenesis and early phenotypic changes in mental illness, mainly when most animal models used in pre-clinical research, such as rodents, are not able to meet the criteria to translate the findings to the clinic. Non-human primates, canines, and porcine are considered more adequate models for biomedical research and drug development purposes, mainly due to their physiological, genetic, and anatomical similarities to humans. The swine model has gained particular interest in translational neuroscience, enabling safety and allotransplantation testing. Herein the generation of porcine iPSCs is described along with its further differentiation into neural progenitor cells (NPCs). The generated cells expressed NPC markers Nestin and GFAP, confirmed by RT-qPCR, and were positive for Nestin, b-Tubulin III, and Vimentin by immunofluorescence. These results show the evidence for the generation of NPC-like cells after in vitro induction with chemical inhibitors from a large animal model, an interesting and adequate model for regenerative and translational medicine research.
诱导多能干细胞衍生的神经元是研究神经发生和精神疾病早期表型变化的有吸引力的体外模型，主要是因为临床前研究中使用的大多数动物模型，如啮齿动物，无法满足将研究结果转化到临床的标准。非人灵长类动物、犬类和猪被认为是生物医学研究和药物开发目的更合适的模型，主要是由于它们在生理、遗传和解剖学上与人类相似。猪模型在转化神经科学中引起了特别的兴趣，可用于安全性和同种异体移植测试。本文描述了猪诱导多能干细胞的生成及其进一步分化为神经祖细胞（NPC）的过程。通过RT-qPCR证实，所生成的细胞表达NPC标志物巢蛋白（Nestin）和胶质纤维酸性蛋白（GFAP），并且通过免疫荧光检测，这些细胞对巢蛋白、β-微管蛋白III（b-Tubulin III）和波形蛋白（Vimentin）呈阳性。这些结果表明，在体外使用化学抑制剂诱导后，从大型动物模型中生成了类NPC细胞，这是再生医学和转化医学研究中一个有趣且合适的模型。
