Reprogramming of Human Hepatic Non-Parenchymal Cells: Step-by-Step Protocol

Varvara A. Kirchner, Kirk Twaroski, Kelli Wysoglad, Jenna Algoo, Edward L. LeCluyse, Gi Won Song, Eunyoung Tak, Weili Chen, Sung Gyu Lee, Timothy L. Pruett, Jakub Tolar

Research output: Contribution to journalArticlepeer-review

Abstract

Human induced pluripotent stem cells (h-iPSCs) represent a potentially unlimited source for the generation of human hepatocyte-like cells (h-iHLCs) for the establishment of platforms to study drug-induced hepatotoxicity, liver disease modeling, and ultimately the application of h-iHLCs in treatment of patients with end-stage liver disease. To understand the impact of donor-specific factors on the generation of h-iHLCs, the model for the direct comparison of h-iHLCs and primary human hepatocytes (PHHs) from the same human donor is needed. This study proposes a step-by-step protocol for plating, expansion, and characterization of primary human hepatic non-parenchymal cells (h-NPCs) isolated from the human liver, the reprogramming of generated h-NPCs into h-iPSCs and subsequent differentiation of reprogrammed h-iPSCs into h-iHLCs. The ultimate goal is to compare the gene expression involved in hepatocyte metabolism between h-iHLCs and PHHs from the same human donor thus eliminating interdonor variability. This newly developed protocol of h-NPC culture, expansion, and reprogramming into h-iPSCs allows: (1) utilization of a single organ source (i.e., liver) for isolation of PHHs and h-NPCs and (2) the in-depth study of donor factors involved in the generation of h-iHLCs with direct comparison to PHHs from the same donor.

Original languageEnglish (US)
Article numbere112
JournalCurrent Protocols in Stem Cell Biology
Volume53
Issue number1
DOIs
StatePublished - Jun 1 2020

Bibliographical note

Funding Information:
Authors would like to thank Courtney Popp, Christopher Lees, Cindy Eide, and Ellen Florek for their assistance in this project and manuscript. The research was supported by Asan Minnesota Institute for Innovating Transplantation (AMIT) and research funds from the National Research Foundation of Korea (NRF‐2015K1A4A3046807).

Publisher Copyright:
© 2020 Wiley Periodicals LLC

Keywords

  • h-NPCs
  • h-iPSCs
  • human hepatic non-parenchymal cells
  • human liver
  • reprogramming

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