Department of Bioengineering, Faculty of Engineering, Ege University, 35100, Izmir, Turkey.
Izmir Biomedicine and Genome Center (IBG), Dokuz Eylul University Health Campus, 35340, Izmir, Turkey.
Commun Biol. 2023 Feb 14;6(1):173. doi: 10.1038/s42003-023-04547-1.
The bioengineerined and whole matured human brain organoids stand as highly valuable three-dimensional in vitro brain-mimetic models to recapitulate in vivo brain development, neurodevelopmental and neurodegenerative diseases. Various instructive signals affecting multiple biological processes including morphogenesis, developmental stages, cell fate transitions, cell migration, stem cell function and immune responses have been employed for generation of physiologically functional cerebral organoids. However, the current approaches for maturation require improvement for highly harvestable and functional cerebral organoids with reduced batch-to-batch variabilities. Here, we demonstrate two different engineering approaches, the rotating cell culture system (RCCS) microgravity bioreactor and a newly designed microfluidic platform (µ-platform) to improve harvestability, reproducibility and the survival of high-quality cerebral organoids and compare with those of traditional spinner and shaker systems. RCCS and µ-platform organoids have reached ideal sizes, approximately 95% harvestability, prolonged culture time with Ki-67 + /CD31 + /β-catenin+ proliferative, adhesive and endothelial-like cells and exhibited enriched cellular diversity (abundant neural/glial/ endothelial cell population), structural brain morphogenesis, further functional neuronal identities (glutamate secreting glutamatergic, GABAergic and hippocampal neurons) and synaptogenesis (presynaptic-postsynaptic interaction) during whole human brain development. Both organoids expressed CD11b + /IBA1 + microglia and MBP + /OLIG2 + oligodendrocytes at high levels as of day 60. RCCS and µ-platform organoids showing high levels of physiological fidelity a high level of physiological fidelity can serve as functional preclinical models to test new therapeutic regimens for neurological diseases and benefit from multiplexing.
经过生物工程设计并完全成熟的人类大脑类器官是高度有价值的三维体外脑模拟模型，可重现体内大脑发育、神经发育和神经退行性疾病。各种影响包括形态发生、发育阶段、细胞命运转变、细胞迁移、干细胞功能和免疫反应在内的多个生物过程的指导信号已被用于生成具有生理功能的大脑类器官。然而，目前的成熟方法需要改进，以获得具有高收获率和功能的大脑类器官，并减少批次间的变异性。在这里，我们展示了两种不同的工程方法，即旋转细胞培养系统（RCCS）微重力生物反应器和新设计的微流控平台（µ 平台），以提高收获率、重现性和高质量大脑类器官的存活率，并将其与传统的旋转瓶和摇床系统进行比较。RCCS 和 µ 平台类器官达到了理想的大小，约有 95%的可收获性，延长了培养时间，Ki-67+/CD31+/β-连环蛋白+增殖、黏附和内皮样细胞的存活率提高，表现出丰富的细胞多样性（丰富的神经/胶质/内皮细胞群体）、结构脑形态发生、进一步的功能神经元特征（谷氨酸能谷氨酸能、GABA 能和海马神经元）和突触发生（突触前-突触后相互作用），这是整个人类大脑发育的特征。两种类器官在第 60 天均表达高水平的 CD11b+/IBA1+/小胶质细胞和 MBP+/OLIG2+/少突胶质细胞。RCCS 和 µ 平台类器官表现出高水平的生理保真度，可作为功能临床前模型，用于测试神经疾病的新治疗方案，并受益于多重分析。
