Hepatic Differentiation of Marmoset Embryonic Stem Cells and Functional Characterization of ESC-Derived Hepatocyte-Like Cells

Rajagopal N Aravalli; Daniel P Collins; Joel H Hapke; Andrew T Crane; Clifford J Steer

doi:10.2147/HMER.S243277

Hepatic Differentiation of Marmoset Embryonic Stem Cells and Functional Characterization of ESC-Derived Hepatocyte-Like Cells

Hepat Med. 2020 Feb 13:12:15-27. doi: 10.2147/HMER.S243277. eCollection 2020.

Authors

Rajagopal N Aravalli¹, Daniel P Collins², Joel H Hapke², Andrew T Crane³, Clifford J Steer^{4

5}

Affiliations

¹ Department of Electrical and Computer Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
² Cytomedical Design Group LLC, St. Paul, MN 55127, USA.
³ Department of Neurosurgery, University of Minnesota Medical School, Minneapolis, MN 55455, USA.
⁴ Department of Medicine, University of Minnesota Medical School, Minneapolis, MN 55455, USA.
⁵ Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455, USA.

Abstract

Background: Primary human hepatocytes (PHHs) are the ideal candidates for studying critical liver functions such as drug metabolism and toxicity. However, as they are isolated from discarded livers that are unsuitable for transplantation, they possess limited expansion ability in vitro and their enzymatic functions deteriorate rapidly because they are often of poor quality. Therefore, there is a compelling reason to find reliable alternative sources of hepatocytes.

Methods: In this study, we report on efficient and robust differentiation of embryonic stem cells (ESC) from the common marmoset Callithrix jacchus into functional hepatocyte-like cells (HLC) using a simple, and reproducible three-step procedure. ESC-derived HLCs were examined by morphological analysis and tested for their expression of hepatocyte-specific markers using a combination of immunohistochemistry, RT-PCR, and biochemical assays. Primary human hepatocytes were used as controls.

Results: ESC-derived HLCs expressed each of the hepatocyte-specific markers tested, including albumin; α-fetoprotein; asialoglycoprotein receptor 1; α-1 antitrypsin; hepatocyte nuclear factors 1α and 4; cytokeratin 18; hepatocyte growth factor receptor; transferrin; tyrosine aminotransferase; alkaline phosphatase; c-reactive protein; cytochrome P450 enzymes CYP1A2, CYP2E1 and CYP3A4; and coagulation factors FVII and FIX. They were functionally competent as demonstrated by biochemical assays in addition to producing urea.

Conclusion: Our data strongly suggest that marmoset HLCs possess characteristics similar to those of PHHs. They could, therefore, be invaluable for studies on drug metabolism and cell transplantation therapy for a variety of liver disorders. Because of the similarities in the anatomical and physiological features of the common marmoset to that of humans, Callithrix jacchus is an appropriate animal model to study human disease conditions and cellular functions.

Keywords: Callithrix jacchus; differentiation; embryonic stem cell; hepatocyte; liver; marmoset.