Monocrotaline toxicity in ExVive 3D bioprinted human liver tissue
Monocrotaline (MCT), a pyrrolizidine alkaloid causes liver injury in animals similar to that of hepatic venoocclusive disorder in humans. MCT induced liver injury occurs through a complex set of cellular insults involving multiple cell types which can ultimately lead to fibrotic changes. In the study, we evaluated the effects of MCT in 3D-bioprinted human liver tissue comprising of primary hepatocytes, hepatic stellate cells, and endothelial cells (ExVive™ Human Liver Tissue). The bioprinted tissues were treated with MCT for fourteen days. MCT treatment led to time- and dose-dependent decreases in tissue health as measured by LDH leakage and albumin synthesis and by histopathologic changes in the tissues, as well as increases in the production of the pro-inflammatory cytokines IL-1β, IL-4, IL-8 and IL-10. Histologic assessment of formalin-fixed, paraffin-embedded tissue revealed signs of tissue damage, including dissociation of the network of hepatocytes and reduced cellularity within the tissues. Immunohistochemical analyses revealed a dose-dependent increase in CD31+ cells and a marked increase in the appearance of large, CD31+ bright cells that co-expressed smooth muscle actin (α-SMA), often forming clusters or complex multicellular structures. Changes in organization of CD31 expressing endothelial cells and appearance of α-SMA expressing cells are indicative of remodeling and initiation of fibrotic events. Observations which emerged from this study capture the spectrum of changes induced by MCT ranging from reduced hepatocellular function and vascular remodeling, which may involve endothelial cell migration, organization, proliferation, apoptosis, and endothelial-to-mesenchymal transformation to early fibrotic events.