Nonintegrating Direct Conversion Using mRNA into Hepatocyte-Like Cells

Abstract EN

Recently, several researchers have reported that direct reprogramming techniques can be used to differentiate fibroblasts into hepatocyte-like cells without a pluripotent intermediate step.

However, the use of viral vectors for conversion continues to pose important challenges in terms of genome integration.

Herein, we propose a new method of direct conversion without genome integration with potential clinical applications.

To generate hepatocyte-like cells, mRNA coding for the hepatic transcription factors Foxa3 and HNF4α was transfected into mouse embryonic fibroblasts.

After 10-12 days, the fibroblasts converted to an epithelial morphology and generated colonies of hepatocyte-like cells (R-iHeps).

The generated R-iHeps expressed hepatocyte-specific marker genes and proteins, including albumin, alpha-fetoprotein, HNF4α, CK18, and CYP1A2.

To evaluate hepatic function, indocyanine green uptake, periodic acid-Schiff staining, and albumin secretion were assessed.

Furthermore, mCherry-positive R-iHeps were engrafted in the liver of Alb-TRECK/SCID mice, and we confirmed FAH enzyme expression in F a h 1 R T y r c /RJ models.

In conclusion, our data suggest that the nonintegrating method using mRNA has potential for cell therapy.