Evaluation of the Effect of Perfusion on Human Ips Cell-Derived Brain Microvascular Endothelial Cells by Using a Closed Two-Channel Microfluidic Device

The blood-brain barrier (BBB), which tightly regulates the exchange of substances between the blood vessel and the brain, maintains central nervous system (CNS). Therefore, the development of BBB in vitro model is expected to significantly contribute to the CNS-targeted drug development. Organ-on-a-chip systems using microfluidic devices under flow conditions have recently attracted attention for enabling cell culture environment that closely resemble in vivo. In particular, organ-on-a-chip systems utilizing closed, dual-channel microfluidic devices with a porous membrane are valuable tools for analyzing inter-organ interactions, such as those at the BBB. In this study, we aimed to investigate the influence of shear stress, induced by perfusion, on the gene expression in human induced pluripotent stem cell-derived brain microvascular endothelial cells (iBMECs) cultured in such a device for the development of a BBB-on-a-chip.iBMECs were seeded into the upper channel of a closed, two-channel microfluidic device and cultured under static conditions for 12 h. Following this, the culture medium was perfused for 48 h, and the cells were then for gene expression analysis. As a control, iBMECs cultured under static conditions using conventional cell culture inserts were similarly analyzed.We first optimized the perfusion parameters to prevent cell detachment. As a result, a trend toward increased expression of genes, including those encoding drug efflux transporters, was observed in the perfused group compared to the static culture. Notably, gene expression of Breast Cancer Resistance Protein markedly increased to levels comparable to those observed in vivo. In addition, gene expression of a glycosyltransferase C1GALT1, which has been reported to contribute to the barrier function of BBB, also showed a tendency to increase in the perfused group.In this study, we successfully established perfusion culture condition for iBMECs within a microfluidic device and demonstrated that flow-induced mechanical stimuli can significantly enhance the expression of key BBB-related genes. These findings suggest that the perfusion culture of iBMECs is a valuable approach for developing a functional BBB-on-a-chip model.