Shaji Maneesha, Tamada Atsushi, Fujimoto Kazuya, Muguruma Keiko, Karsten Stanislav L, Yokokawa Ryuji
Department of Micro Engineering, Graduate School of Engineering, Kyoto University, Kyoto daigaku-Katsura, Nishikyo-ku, Kyoto - 615-8540, Japan.
Department of iPS Cell Applied Medicine, Kansai Medical University, 2-5-1 Shin-machi, Hirakata City, Osaka - 573-1010, Japan.
Lab Chip. 2024 Feb 13;24(4):680-696. doi: 10.1039/d3lc00930k.
The lack of functional vascular system in stem cell-derived cerebral organoids (COs) limits their utility in modeling developmental processes and disease pathologies. Unlike other organs, brain vascularization is poorly understood, which makes it particularly difficult to mimic . Although several attempts have been made to vascularize COs, complete vascularization leading to functional capillary network development has only been achieved transplantation into a mouse brain. Understanding the cues governing neurovascular communication is therefore imperative for establishing an efficient system for vascularized cerebral organoids that can emulate human brain development. Here, we used a multidisciplinary approach combining microfluidics, organoids, and transcriptomics to identify molecular changes in angiogenic programs that impede the successful vascularization of human induced pluripotent stem cell (iPSC)-derived COs. First, we established a microfluidic cerebral organoid (CO)-vascular bed (VB) co-culture system and conducted transcriptome analysis on the outermost cell layer of COs cultured on the preformed VB. Results revealed coordinated regulation of multiple pro-angiogenic factors and their downstream targets. The VEGF-HIF1A-AKT network was identified as a central pathway involved in the angiogenic response of cerebral organoids to the preformed VB. Among the 324 regulated genes associated with angiogenesis, six transcripts represented significantly regulated growth factors with the capacity to influence angiogenic activity during co-culture. Subsequent on-chip experiments demonstrated the angiogenic and vasculogenic potential of cysteine-rich angiogenic inducer 61 (CYR61) and hepatoma-derived growth factor (HDGF) as potential enhancers of organoid vascularization. Our study provides the first global analysis of cerebral organoid response to three-dimensional microvasculature for vascularization.
干细胞衍生的脑类器官（COs）中缺乏功能性血管系统，限制了它们在模拟发育过程和疾病病理方面的应用。与其他器官不同，脑血管生成的机制尚不清楚，这使得模拟脑血管生成尤为困难。尽管已经多次尝试使COs血管化，但只有通过移植到小鼠大脑中才能实现导致功能性毛细血管网络发育的完全血管化。因此，了解控制神经血管通讯的线索对于建立一个能够模拟人类大脑发育的血管化脑类器官的有效系统至关重要。在这里，我们采用了一种多学科方法，结合微流体技术、类器官技术和转录组学，来识别血管生成程序中的分子变化，这些变化阻碍了人类诱导多能干细胞（iPSC）衍生的COs成功血管化。首先，我们建立了一个微流体脑类器官（CO）-血管床（VB）共培养系统，并对在预制VB上培养的COs最外层细胞层进行了转录组分析。结果揭示了多种促血管生成因子及其下游靶点的协同调控。VEGF-HIF1A-AKT网络被确定为参与脑类器官对预制VB血管生成反应的核心途径。在与血管生成相关的324个受调控基因中，有6个转录本代表了在共培养过程中具有影响血管生成活性能力的显著受调控生长因子。随后的芯片实验证明了富含半胱氨酸的血管生成诱导因子61（CYR61）和肝癌衍生生长因子（HDGF）作为类器官血管化潜在增强剂的血管生成和血管发生潜力。我们的研究首次对脑类器官对三维微脉管系统进行血管化的反应进行了全面分析。
