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Generation of In Vivo Traceable Hepatocyte-Like Cells from Human iPSCs

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

Original languageEnglish
Title of host publicationHepatocytes
Pages15-49
Number of pages35
Volume2544
ISBN (Electronic)978-1-0716-2557-6
DOIs
Accepted/In press5 Oct 2021
Published21 Sep 2022

Publication series

NameMethods in molecular biology (Clifton, N.J.)

Bibliographical note

Funding Information: The authors were supported by Guy’s and St Thomas’ Charity (PhD studentship to CAH), the United Kingdom Regenerative Medicine Platform (CAH), and Cancer Research UK via a Multidisciplinary Project Award (C48390/A21153) to G.O.F. Furthermore, the authors are supported by the National Institute for Health Research (NIHR) Biomedical Research Centre based at Guy’s and St Thomas’ NHS Foundation Trust and King’s College London. The views expressed are those of the authors and not necessarily those of the NHS, the NIHR, or the DoH. Publisher Copyright: © 2022, The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.

King's Authors

Abstract

In this chapter, we describe a protocol for differentiation of human-induced pluripotent stem cells (iPSCs) into hepatocyte-like cells (HLCs) and their transduction with a lentivirus for gene transfer. Here, we engineer them to express the human sodium iodide symporter, which can be exploited as a radionuclide reporter gene, thereby enabling these cells to be tracked in vivo by single-photon emission computed tomography (SPECT) or positron emission tomography (PET) imaging. Differentiation of HLCs from iPSCs involves three steps: induction of iPSCs to definitive endoderm, differentiation to a hepatic progenitor cell population, and maturation of immature HLCs. Once proliferation of hepatic progenitors has ceased and an immature HLC population is generated, lentiviral transduction can be performed. The immature hepatic gene expression profile/morphology at the stage of transduction will be compatible with further maturation following transgene expression either in vitro or in vivo, with expression of the transgene retained. We detail how transgenic cells can be imaged in vivo. While we provide a protocol for the NIS reporter gene, the cell engineering aspects of this protocol are transferable for use with other (reporter) genes if desired.

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