Vectors Trgeting Hepatocyte: Reconstructive Umbilical Cord Mesenchymal Stem Cell-Derived Exosomes.

BACKGROUND

Specific binding of the hepatitis B virus (HBV) pre-S1 protein to Na+/taurocholate Cotransporting Polypeptide (NTCP) on the surface of hepatocytes mediates viral entry into host cells. Exosomes, which serve as natural delivery vehicles, can protect their cargo from degradation in the extracellular environment.

METHODS

Umbilical cord mesenchymal stem cell (UC-MSC)-derived exosomes were isolated and characterized. The gene sequence encoding the CD90 C-terminal signal peptide was fused with the pre-S1 gene and cloned into a lentiviral vector (VP045-U6-PGK-preS1-CD90-hygro), enabling membrane anchoring of pre-S1 on UC-MSC-derived exosomes. PKH26 probe, Phalloidin-AF488 probe, and DiR dye were used to explore the targeting of reconstituted UC-MSC-derived exosomes on hepatocytes and liver.

RESULTS

Isolated UC-MSCs exhibited fibroblast-like morphology and retained the capacity to differentiate into osteoblasts, adipocytes, and chondrocyte. The immunophenotype of UC-MSCs was characterized as CD34-CD45-CD44+CD73+CD90+CD105+Oct-4+. UC-MSC-derived exosomes exhibited a cup-shaped bilayer structure, with a diameter of 78.47 ± 13.46 nm and a concentration of 1.28 × 1010 particles/mL. The VP045-U6-PGK-preS1-CD90-hygro lentivirus successfully mediated pre-S1 expression on the membranes of UC-MSC-derived exosomes. In vitro, hepatocytes (HepG2.2.15 and HepaRG cells) internalized significantly greater amounts of reconstituted UC-MSC-derived exosomes than native exosomes, with the effect being particularly pronounced in HepG2.2.15 cells. No significant difference was observed in lung cells. In vivo, reconstituted UC-MSC-derived exosomes were predominantly accumulated in the liver of BALB/c mice, whereas minimal signals were detected in the lung, spleen and kidney.

CONCLUSION

UC-MSC-derived exosomes expressing pre-S1 exhibit specific tropism for hepatocytes and liver tissue. These engineered exosomes represent a promising delivery platform for therapeutic drugs and nucleic acid cargos in HBV-targeted therapy.