Preconditioned umbilical-cord mesenchymal stem-cell exosomes preserve functional hepatocytes in bile duct-ligated rats through alternate miRNA signaling rather than HGF signaling.

Abstract

Biliary atresia (BA) is a prominent cause of liver cirrhosis in pediatric patients with a high pediatric end-stage liver disease (PELD) score and a Laennec score of 4 at the Cipto Mangunkusumo National General Hospital, Jakarta, Indonesia. Bile duct-ligated rat models have been used to mimic the clinical condition of BA partially. This study aims to evaluate the potential miRNAs of preconditioned umbilical-cord mesenchymal stem-cell exosomes (UC-MSC-exosomes) in comparison with the combined exosomes-HGF effect on liver histology, liver function, and hepatocyte-signaling in bile duct-ligated rats. Characterization of preconditioned UC-MSC-exosomes fulfilled the ISEV 2018 criteria. Liver histology showed trends of hepatocytes, inflammatory cells, and oval cell numbers returning to slightly below normal levels post-preconditioned UC-MSC-exosomes or the exosomes-HGF combination, with no statistically significant difference. A significant difference in bile duct proliferation post-preconditioned UC-MSC-exosomes or the exosomes-HGF combination was observed when compared with the control. Trends towards lower deposition of collagen around the porta area and total area fractions post-preconditioned UC-MSC-exosomes or the exosomes-HGF combination showed no statistically significant difference; however, there was a slightly lower total collagen area fraction in the exosomes-HGF combination group. Functional hepatocyte preservation was indicated by increased albumin expression and partially improved CYP3A4 levels post-preconditioned UC-MSC-exosomes or the exosomes-HGF combination. There was also a different trend in functional hepatocyte biochemical markers, SGOT (AST) and SGPT (ALT), between preconditioned UC-MSC-exosomes and the exosomes-HGF combination. NanoString microarray analysis of liver tissue and bioinformatics analysis were done to investigate in-depth hepatocyte signaling. miRNA profiling of the liver from preconditioned UC-MSC-exosomes identified the upregulation of hsa-miR-1-3p and hsa-miR-372-3p, which inhibit key signaling pathways, e.g., MAPK, PI3K-AKT, and NF-κB, and the downregulation of hsa-miR-520a-3p, which promotes hepatocyte proliferation and survival. miRNA profiles of the liver from the exosomes-HGF combination identified five different miRNA expressions (upregulated hsa-miR-144-3p, hsa-let7-5p, hsa-let7a-5p, hsa-let7e-5p, and hsa-miR-32-5p) and shared only one similar miR with the exosomes-treated liver (downregulated hsa-520a-3p). These miR differences contribute to the hepatocyte signaling in the exosomes-HGF combination group. Preconditioned UC-MSC-exosomes preserve functional hepatocytes in bile duct-ligated rats through alternate miR-mediated signaling rather than HGF signaling.