Postdoctoral Researcher Federal University of Rio de Janeiro Rio de Janeiro , Rio de Janeiro, Brazil
Abstract: The large number of discarded marginal livers offers a promising source of organs to expand the donor pool and reduce global organ shortages. However, the post-transplantation use of a marginal liver derived - extracellular matrix (ECM) has not been demonstrated. In this context, we performed in vitro and in vivo analyses to determine whether the Metabolic dysfunction-associated steatohepatitis (MASH)- derived ECM from rat and human livers could be used to reduce organ shortage. For in vivo assays, Wistar rats (n=24) were submitted to MASH induction model based on a high-fat diet (45% fat, 20% fructose, 2% cholesterol) followed by 6% sucrose in water for 16 weeks and intraperitoneal injections of CCl4 (1mL/Kg), 3x a week for the last 3 weeks (Animal Care approval-UFRJ 69/24). Biochemical and histological analyses were performed before, and after the MASH induction and post-transplantation. MASH rat livers (n=5) were submitted to perfusion-based decellularization to obtain MASH-derived acellular liver scaffold (MASH-ALS). Macroscopic and microscopic analyses revealed cell-free, dense, opaque, and rigid ECM. After characterization, MASH-AHA were partially and orthotopically transplanted into MASH receptor rats (n=8). Macroscopic analysis revealed a well-distributed and connected MASH-AHA 30 days after transplantation. Inflammation, absence of steatosis and intense recellularization confirming the presence of liver cells, including liver stem/progenitor cells were detected into the MASH-ALS after transplantation. No significant differences were observed in body weight, glucose levels, total cholesterol, triglycerides, very high-, high- and low-density lipoproteins 30 days after transplantation. For in vitro assays, MASH-derived human livers cubes (n=20) were submitted to a decellularization process to generate human-derived MASH ALS (hMASH-ALS). HepG2 cells (500k) were cultured in hMASH-ALS for 14 days and then, submitted to histological analysis and an intracellular Ca2+ signalization assay (n=5). Sequential confocal images identified an increase in cytosolic Ca2+ after ATP stimulation and revealed that HepG2 cells acquired a synchronized signal in hMASH-ALS. Taken together, these data show that MASH-derived ECM can be transplanted, allowing for cell survival both in vivo and in vitro.
