Graduate Student Institute of Science Tokyo, Tokyo, Japan
Abstract: Enterovirus A71 (EV-A71), one of the viruses responsible for hand, foot, and mouth disease, infects and replicates in the human intestine. While EV-A71 infection is known to cause the damage of the central nervous system (CNS), the pathophysiology of intestine including the enteric nervous system (ENS) remains unknown. Additionally, there are no therapeutic drugs to treat EV-A71 infection. In this study, we developed intestinal models with enteric neurons and infected them with EV-A71 to elucidate the intestinal pathophysiology of EV-A71 infection and to evaluate whether these models can be used for pharmaceutical research. We previously generated an intestinal model by differentiating human ES/iPS cells in microfluidic devices (micro-intestine system). The single-cell RNA-sequencing (scRNA-seq) analysis revealed that this model contained not only intestinal epithelial cells but also enteric neurons. Thus, we used the micro-intestine system to study EV-A71 infection. EV-A71 genome was detected in the culture supernatant of virus-infected systems, suggesting that EV-A71 can efficiently infect the micro-intestine system. RNA-seq analysis showed that the expressions of neuron and epithelial markers in the micro-intestine system were downregulated by EV-A71 infection. Histological analysis also indicated that Tubulin Beta 3 (TUBB3)-positive neurons were decreased, and the epithelial layer was disrupted by EV-A71 infection, indicating that EV-A71 infection caused the neural and epithelial damage. We evaluated the anti-viral effect of rupintirivir, which is one of the 3C protease inhibitors, using micro-intestine system. The viral genome copy number in the cell culture supernatant of the infected micro-intestine system was decreased by rupintrivir treatment. This result suggested that this model is useful for evaluating the effectiveness of drugs. In conclusion, the micro-intestine system with enteric neurons is a valuable tool for elucidating the detailed pathophysiology of EV-A71-infected intestine and evaluating the effect of therapeutic drugs.
