Multi-lineage hepatic organoids reveal toxic exosome mediated indirect hepatotoxicity

Indirect hepatotoxicity remains a major challenge in drug development because of limited evaluation tools and mechanistic insight. Here, we construct three-dimensional multi-lineage hepatic organoids comprising five liver cell types derived from human embryonic stem cells, and in a screen of 58 hepatotoxic drugs, identify imipramine as an inducer of indirect hepatotoxicity. Imipramine specifically engages tyrosine kinase receptor B expressed by non-parenchymal hepatic stellate cells, activating the p53/hnRNPA2B1/DGCR8 pathway, which drives selective enrichment of microRNA-34a-3p in hepatic stellate cell–derived exosomes, thereby converting them into toxic exosomes. These toxic exosomes transfer microRNA-34a-3p to hepatocytes, where it disrupts cellular homeostasis by downregulating the anti-apoptotic protein XIAP, thereby activating caspase3 and inducing apoptosis. Consistently, imipramine long-term gavage in vivo triggers hepatic stellate cell apoptosis and toxic exosome–mediated hepatocyte injury without direct hepatocyte toxicity. Our findings establish a biomimetic organoid platform for precision drug testing and highlight the central role of intercellular communication in drug-induced liver injury. Assessing the hepatotoxicity of a drug is critical in drug development. Here, the authors develop human multi-lineage liver organoids and identify imipramine as an inducer of indirect hepatotoxicity from a drug screen. The study offers insight into indirect hepatotoxicity and an in vitro platform for drug safety testing.