The Florey Institute of Neuroscience and Mental Health, The University of Melbourne, Parkville, VIC, Australia.
Australian Regenerative Medicine Institute, Monash University, Melbourne, VIC, Australia.
Nat Commun. 2021 May 27;12(1):3275. doi: 10.1038/s41467-021-23125-9.
Despite advancements in human pluripotent stem cells (hPSCs) differentiation protocols to generate appropriate neuronal progenitors suitable for transplantation in Parkinson's disease, resultant grafts contain low proportions of dopamine neurons. Added to this is the tumorigenic risk associated with the potential presence of incompletely patterned, proliferative cells within grafts. Here, we utilised a hPSC line carrying a FailSafe suicide gene (thymidine kinase linked to cyclinD1) to selectively ablate proliferative cells in order to improve safety and purity of neural transplantation in a Parkinsonian model. The engineered FailSafe hPSCs demonstrated robust ventral midbrain specification in vitro, capable of forming neural grafts upon transplantation. Activation of the suicide gene within weeks after transplantation, by ganciclovir administration, resulted in significantly smaller grafts without affecting the total yield of dopamine neurons, their capacity to innervate the host brain or reverse motor deficits at six months in a rat Parkinsonian model. Within ganciclovir-treated grafts, other neuronal, glial and non-neural populations (including proliferative cells), were significantly reduced-cell types that may pose adverse or unknown influences on graft and host function. These findings demonstrate the capacity of a suicide gene-based system to improve both the standardisation and safety of hPSC-derived grafts in a rat model of Parkinsonism.
尽管人类多能干细胞（hPSCs）分化方案在生成适合帕金森病移植的适当神经元祖细胞方面取得了进展，但产生的移植物中多巴胺神经元的比例仍然较低。此外，移植物中存在未完全定型、增殖细胞的致瘤风险也增加了这一问题。在这里，我们利用携带失效安全自杀基因（与细胞周期蛋白 D1 相连的胸苷激酶）的 hPSC 系，选择性地消除增殖细胞，以提高帕金森病模型中神经移植的安全性和纯度。工程化的失效安全 hPSC 在体外表现出强大的中脑神经节特异性，能够在移植后形成神经移植物。在移植后数周内通过给予更昔洛韦激活自杀基因，导致移植物明显缩小，而不影响多巴胺神经元的总产量、它们对宿主大脑的支配能力或在帕金森病大鼠模型中六个月时的运动缺陷的逆转。在更昔洛韦处理的移植物中，其他神经元、神经胶质和非神经元群体（包括增殖细胞）明显减少——这些细胞类型可能对移植物和宿主功能产生不良或未知的影响。这些发现证明了基于自杀基因的系统在帕金森病大鼠模型中改善 hPSC 衍生移植物的标准化和安全性的能力。
