Closed, pumpless microphysiological system with unidirectional flow for co-culture and focal irradiation

Intercellular communication is important for biological phenomena such as radiation-induced bystander effects (RIBEs). Microphysiological systems (MPSs), in which multiple groups of cells are co-cultured in a circulatory system, have been used to study complex intercellular communication. Because they can provide models that closely resemble the human body, human cells have been used in MPSs to replicate it. For recapitulating the intercellular communication in human bodies, experiments using cells differentiated from human induced pluripotent stem cells or human primary cells should be performed; however, they are expensive for a limited cell number. Therefore, MPSs should be scaled down as much as possible to analyze secreted substances with a minimum number of cells. In this study, we propose a closed, pumpless MPS (CPMS) that allows focal irradiation of X-rays to a part of minimized culture space. The CPMS was designed to operate with a minimum of approximately 200 µL of medium by isolating the culture space from the external space to prevent its evaporation. For the efficient circulation of the substances secreted from cells, a gravity-driven passive unidirectional flow was generated using water head pressure with a maximum flow rate of approximately 15.7 µL min^-1. In the CPMS, cultured neurons survived for 21 d in a static condition and formed synapses. Even under unidirectional flow for 7 d, cultured neurons extended neurites and formed branches. Furthermore, focal irradiation of X-rays induced apoptosis of the hippocampal cells in the irradiated chamber. These results suggest that the CPMS will be useful for analyzing intercellular communication, such as secreted substance-mediated RIBEs. The CPMS is suitable for analyzing small quantities of rare and/or expensive cells, such as commercially available human cells, because of its compactness.