A Microfibrous Extracellular Matrix Platform for <i>In Vitro</i> Hepatotoxicity Screening: A Next-Generation Model for Humanized Liver Research

Drug-induced hepatotoxicity is a significant concern in drug development and patient safety. The liver, responsible for drug metabolism, can suffer from damage caused by certain medications, leading to inflammation, cellular damage, and even failure. Traditional two-dimensional (2D) cell cultures fail to replicate the complex <i>in vivo</i> environment, making it difficult to assess drug-induced liver toxicity accurately. Here, we develop a more advanced model using three-dimensional (3D) cultured human and mice hepatocytes in a solid microfibrous extracellular matrix (smfECM). This 3D system showed improved hepatocyte growth and long-term viability compared with 2D cultures. Functional assays, such as albumin secretion and urea production, demonstrated the superior functionality of hepatocytes cultured in 3D. Additionally, immunocytochemical analysis confirmed CYP3A4 expression in immortalized human liver cells (THLE-2) 3D spheroids, indicating that these hepatic cells retain their normal metabolic functions. smfECM also enhanced the accuracy of high-throughput drug screening, with hepatotoxin screening results aligning closely with clinical data. Furthermore, when implanted <i>in vivo</i>, smfECM supported human cell survival, promoted angiogenesis, and showed no signs of immunogenicity. Overall, smfECM provides a reliable, humanized model for accurate hepatotoxicity assessment in drug development, making it a promising tool for drug screening and toxicity evaluation.