背景：睡眠病由细胞外寄生物Trypanosoma brucei引起，与神经炎症和神经精神障碍有关，包括打乱睡眠/清醒模式，现在被认为是一种昼夜节律失调。由于缺乏全面重现人脑细胞多样性与功能的替代体外系统，睡眠病传统上使用小鼠模型进行研究。本研究旨在开发一种急需的体外系统，以减少和取代对中枢神经系统感染的各种动物进行研究，使用睡眠病作为模型感染。方法：我们使用干细胞诱导形成的皮质人脑器官样体和人体病原体T. b. gambiense开发了一种共培养系统，以模拟体外的宿主-病原体相互作用。在共培养下，我们对脑器官样体在两个时间点上对T. b. gambiense的转录反应进行了分析。结果：与未经处理的器官样体相比，我们检测到暴露于T. b. gambiense的脑器官样体中出现了广泛的转录变化，主要与先天免疫反应、趋化性和血管分化相关。结论：我们的共培养系统为研究大脑中的宿主-病原体相互作用提供了新的、更符合伦理要求的途径，作为对小鼠实验感染模型的替代品。尽管我们的数据支持使用器官样体模拟T. brucei感染期间宿主-病原体相互作用，作为替代体内模型，但未来的工作需要增加器官样体的复杂性（如添加小胶质细胞和血管系统）。我们预计器官样体系统的采用对于研究原虫寄生物感染大脑机制的研究人员具有益处。此外，器官样体系统有潜力用于研究其他严重影响大脑的寄生虫，从而大大减少进行与神经炎症和脑感染研究相关的中等和/或严重的动物实验的数量。版权：© 2023 Chandrasegaran P et al.

Background: Sleeping sickness is caused by the extracellular parasite Trypanosoma brucei and is associated with neuroinflammation and neuropsychiatric disorders, including disruption of sleep/wake patterns, and is now recognised as a circadian disorder. Sleeping sickness is traditionally studied using murine models of infection due to the lack of alternative in vitro systems that fully recapitulate the cellular diversity and functionality of the human brain. The aim of this study is to develop a much-needed in vitro system that reduces and replaces live animals for the study of infections in the central nervous system, using sleeping sickness as a model infection. Methods: We developed a co-culture system using induced pluripotent stem cell (iPSC)-derived cortical human brain organoids and the human pathogen T. b. gambiense to model host-pathogen interactions in vitro. Upon co-culture, we analysed the transcriptional responses of the brain organoids to T. b. gambiense over two time points. Results: We detected broad transcriptional changes in brain organoids exposed to T. b. gambiense, mainly associated with innate immune responses, chemotaxis, and blood vessel differentiation compared to untreated organoids. Conclusions: Our co-culture system provides novel, more ethical avenues to study host-pathogen interactions in the brain as alternative models to experimental infections in mice. Although our data support the use of brain organoids to model host-pathogen interactions during T. brucei infection as an alternative to in vivo models, future work is required to increase the complexity of the organoids ( e.g., addition of microglia and vasculature). We envision that the adoption of organoid systems is beneficial to researchers studying mechanisms of brain infection by protozoan parasites. Furthermore, organoid systems have the potential to be used to study other parasites that affect the brain significantly reducing the number of animals undergoing moderate and/or severe protocols associated with the study of neuroinflammation and brain infections.Copyright: © 2023 Chandrasegaran P et al.