Department of Neurology, The Second Xiangya Hospital, Central South University, Changsha, 410011, Hunan, China; Department of Pharmacology & Experimental Neuroscience, University of Nebraska Medical Center, Omaha, Nebraska, 68198, United States.
Department of Pharmacology & Experimental Neuroscience, University of Nebraska Medical Center, Omaha, Nebraska, 68198, United States; Center for Translational Neurodegeneration and Regenerative Therapy, Shanghai Tenth People's Hospital affiliated to Tongji University School of Medicine, Shanghai, China.
Neurosci Lett. 2019 Apr 23;699:16-23. doi: 10.1016/j.neulet.2019.01.033. Epub 2019 Jan 18.
Transplantation of dopaminergic precursors (DPs) is a promising therapeutic strategy of Parkinson's disease (PD). However, limited cell source for dopaminergic precursors has become a major obstacle for transplantation therapy. Our group demonstrated previously that mouse fibroblasts can be reprogrammed into induced dopaminergic precursors (iDPs) with high differentiation efficiency. In the current study, we hypothesized that a similar strategy can be applied to generate human iDPs for future cell therapy of PD. We overexpressed transcription factors Brn2, Sox2, and Foxa2 in human fibroblasts and observed formation of neurospheres. Subsequent characterization of the precursor colonies confirmed the generation of human induced dopaminergic precursors (hiDPs). These hiDPs were capable of self-renewal, proliferation, and differentiation. The hiDPs demonstrated high immunoreactivity for neural progenitor markers and high levels of gene expression for ventral mesencephalon-related neural progenitor markers such as Lmx1a, NIKX6.1, Corin, Otx2 and Mash1. Furthermore, the hiDPs could be differentiated into dopaminergic neurons with ˜80% efficiency, which significantly increased major functionally relevant proteins such as TH, DAT, AADC, Lmx1B, and VMAT2 compared to hiDPs. Additionally, hiDPs are more dopaminergic progenitor-restricted compare to those hiDP-like cells reprogrammed only by Brn2 and Sox2. Together, these results suggest that hiDPs with high differentiation efficiency can be generated by direct lineage reprogramming of fibroblasts with transcription factors Brn2, Sox2, and Foxa2. These hiDPs may serve as a safe and effective cell source for transplantation treatment of PD.
移植多巴胺能前体细胞（DPs）是治疗帕金森病（PD）的一种很有前途的治疗策略。然而，多巴胺能前体细胞的有限细胞来源已成为移植治疗的主要障碍。我们的研究小组之前已经证明，小鼠成纤维细胞可以通过高分化效率重编程为诱导性多巴胺能前体细胞（iDPs）。在本研究中，我们假设可以应用类似的策略来产生人类 iDPs，用于未来 PD 的细胞治疗。我们在人成纤维细胞中转染转录因子 Brn2、Sox2 和 Foxa2，并观察到神经球的形成。随后对前体细胞集落进行鉴定，证实了人类诱导性多巴胺能前体细胞（hiDPs）的产生。这些 hiDPs 能够自我更新、增殖和分化。hiDPs 对神经祖细胞标记物具有高免疫反应性，并且高水平表达与中脑神经祖细胞相关的标记物，如 Lmx1a、NIKX6.1、Corin、Otx2 和 Mash1。此外，hiDPs 可以分化为多巴胺能神经元，效率约为 80%，与 hiDPs 相比，显著增加了主要功能相关蛋白，如 TH、DAT、AADC、Lmx1B 和 VMAT2。此外，与仅由 Brn2 和 Sox2 重编程的 hiDP 样细胞相比，hiDPs 更受多巴胺能祖细胞限制。总之，这些结果表明，通过 Brn2、Sox2 和 Foxa2 等转录因子直接谱系重编程，可以产生具有高分化效率的 hiDPs。这些 hiDPs 可能成为 PD 移植治疗的安全有效细胞来源。
