A high-throughput liver-kidney metabolic interaction chip for insights into the nephrotoxicity mechanisms of triptolide

Abstract

The kidney organ-on-a-chip (OoC) is a powerful tool for studying drug-induced nephrotoxicity, but its application is limited by the absence of liver metabolism and low throughput. Here, we developed a high-throughput liver-kidney OoC system (HLKOC) featuring microfluidics, plug-in biomimetic cups (MIMICups), scalable flow channel plates, precision-cut liver slices (PCLS), and 3D HK-2 cell spheroids. We first established a functional endothelial barrier by optimizing cell types, biomimetic blood flow rate, serum content, and membrane pore size. The structural and functional integrity of the PCLS and HK-2 spheroids within the MIMICups was then confirmed through histological staining, metabolic assays, and functional tests for viability, polarization, and transport. To evaluate the system's utility, we integrated the HLKOC with a single kidney OoC control and multidisciplinary techniques—including biochemical analysis, computational toxicology, molecular docking, metabolomics, and transcriptomics—to investigate the nephrotoxicity of triptolide (TPL) and its underlying mechanisms. Results showed that, compared to the single kidney OoC, the HLKOC exhibited higher levels of urea, total protein, and albumin in the biomimetic blood, confirming the robust biosynthetic capacity of the PCLS-based liver chip and its ability to better simulate in vivo conditions. Notably, a TPL-induced elevation in urea was observed only in the HLKOC, demonstrating the superior sensitivity of the liver-kidney co-culture. Multi-omics analysis revealed that TPL induced distinct metabolic and transcriptional responses in the HLKOC, involving pathways related to linoleic acid metabolism and vesicle-mediated processes, and led to the significant downregulation of transport proteins cubilin and GLUT1. These findings highlight the advantages of the HLKOC over single-organ systems for drug toxicity assessment and provide new insights into the mechanisms of TPL-induced nephrotoxicity.