Treatment of liver failure utilizing multi-cell lineage liver microtissue

The shortage of liver donors remains a critical challenge in the treatment of end-stage liver failure. Previous studies have explored ectopic transplantation of hepatocyte organoids or spheroids to support unstable liver function. We hypothesized that the addition of non-parenchymal liver cells to hepatocyte organoids would improve overall survival in acute liver failure. Here, we aimed to engineer DNA origami-mediated multi-lineage liver microtissue (NAC-liver microtissue) and evaluate its therapeutic efficacy in a mouse model of acute liver failure.

We employed self-assembling nucleic acid nanostructures (NAC) to construct two types of liver microtissues: one composed exclusively of hepatocytes (NAC-hepatocyte microtissue) and another incorporating both hepatocytes and non-parenchymal cells (NAC-liver microtissue). We then investigated their hepatic functional properties in vitro and evaluated therapeutic efficacy in vivo through intraperitoneal transplantation into mice undergoing extended hepatectomy. To explore the underlying mechanisms, we performed histological analysis and RNA sequencing of the remnant liver.

NAC-liver microtissue exhibited superior scalability, uniformity, and stable, enhanced hepatic functional activity compared to NAC-hepatocyte microtissue in vitro (n = 4, p <0.001). In vivo, transplantation of NAC-liver microtissue markedly improved survival in mice undergoing extended hepatectomy compared with untreated controls (70% [n = 10] vs. 8.3% [n = 12], p <0.001), whereas NAC-hepatocyte microtissue demonstrated moderate efficacy (25%, n = 8). Furthermore, NAC-liver microtissue transplantation improved liver function, lipid oxidation, and sinusoidal vascular network formation in the remnant liver of mice following extended hepatectomy.

These findings provide therapeutic evidence supporting NAC-liver microtissue transplantation as a potential treatment for acute liver failure and highlight a novel strategy for extrahepatic cellular support in severe liver injury.

Although hepatocyte organoid or spheroid transplantation has long been explored as a potential therapy for liver disease, these approaches fail to replicate the cellular complexity of the liver, resulting in suboptimal functionality and limited therapeutic benefit. The therapeutic potential of multi-lineage liver microtissue for liver failure has not previously been reported. Here, we demonstrate that NAC-liver microtissue, composed of hepatocytes and non-parenchymal cells, provides superior functional outcomes both in vitro and in vivo compared with NAC-hepatocyte microtissue. These findings highlight the indispensable role of non-parenchymal cells in maintaining hepatic functionality and support the therapeutic potential of multi-lineage liver microtissue for patients with severe liver disease.