Dual effects of indoxyl sulfate on modulation of human hepatic CYP3A activity, with individual differences

by Masao Togao, Naoyuki Asakawa, Gaku Wagai, Yuki Ohta-Takada, Jun Otsuka, Minoru Ando, Akinobu Kurita, Koji Kawakami

This study aimed to identify gut microbiota-derived metabolites governing the activity of hepatic CYP3A in blood level. Indole propionic acid (IPA) and lithocholic acid, ligands of the pregnane X receptor, a transcriptional regulator of CYP3A, and various gut microbiota-derived metabolites in blood level were analyzed. Results revealed that IPA and lithocholic acid did not affect CYP3A activity, while indoxyl sulfate (IS), a uremic toxin, affected CYP3A across different cell lines. The effects of IS on primary hepatocytes from three donors were analyzed, and a concentration-dependent impact was observed, as the CYP3A activity decreased in one donor and increased in another. These findings offer initial insights into blood-level gut microbiota-derived metabolites influencing hepatic CYP3A. Furthermore, the study demonstrates that the response to IS, beyond its concentration, can cause variations in hepatic CYP3A activity among individuals. This study advocates accounting for the dual effects of IS and the benefits of personalized medicine.

Potential consequences of phototoxicity on cell function during live imaging of intestinal organoids

by Yuki Yokoi, Ryu Nakamura, Shuya Ohira, Shota Takemi, Tokiyoshi Ayabe, Kiminori Nakamura

Live imaging visualizes the structure, dynamics, and function of cells and tissues to reveal the molecular mechanisms, and has contributed to the advancement of life science. In live imaging, it has been well known that there is a trade-off between higher-resolution analysis and cell damage caused by light illumination, i.e., phototoxicity. However, despite the risk of unknowingly distorting experimental results, phototoxicity is an unresolved issue in live imaging because overall consequences occurring inside cells due to phototoxicity remains unknown. Here, we determined the molecular process of phototoxicity-induced cell damage systematically under low- and high-dose light illumination conditions by analyzing differential gene expression using RNA-sequencing in a three-dimensional organoid of small intestinal epithelial cells, enteroid. The low-dose light illumination already induced various abnormalities in functional molecules involved in the response to reactive oxygen species generated by the excitation of fluorescent dyes, intracellular metabolism, mitosis, immune responses, etc., at mRNA expression level. Together with the behavior toward apoptosis caused by high-dose light illumination, the light dose-dependent progression of intracellular damage was revealed. About visible impairment of intestinal epithelial function, failures in both the structure-forming ability of enteroids and Paneth cell granule secretion were observed under high-dose light illumination, while the drug efflux was not disturbed despite abnormal drug efflux transporter mRNA expression. Based on the gene expression profiles, we comprehensively clarified phenomena in the cells at mRNA level that cannot be recognized both morphologically and functionally during live imaging, further providing a new insight into the risk of phototoxicity. This study warns from the aspect of mRNA expression that awareness of phototoxic artifacts is needed when analyzing cellular function and the mechanism in live imaging.