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Cited by 9Liver physiology and pathologyTissue Engineering and Regenerative MedicineDrug-Induced Hepatotoxicity and Protection

A two-step strategy to expand primary human hepatocytes in vitro with efficient metabolic and regenerative capacities

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Xie, Huangfan, Li, Guangya, Fu, Yunxi, Jiang, Nan, Yi, Simeng, Kong, Xi, Shi, Jihang, Yin, Shigang +5 more·2024·Stem Cell Research & Therapy
DOI 10.1186/s13287-024-03911-0PMC PMC11373096
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Abstract

BACKGROUND: Primary human hepatocytes (PHHs) are highly valuable for drug-metabolism evaluation, liver disease modeling and hepatocyte transplantation. However, their availability is significantly restricted due to limited donor sources, alongside their constrained proliferation capabilities and reduced functionality when cultured in vitro. To address this challenge, we aimed to develop a novel method to efficiently expand PHHs in vitro without a loss of function. METHODS: By mimicking the in vivo liver regeneration route, we developed a two-step strategy involving the de-differentiation/expansion and subsequent maturation of PHHs to generate abundant functional hepatocytes in vitro. Initially, we applied SiPer, a prediction algorithm, to identify candidate small molecules capable of activating liver regenerative transcription factors, thereby formulating a novel hepatic expansion medium to de-differentiate PHHs into proliferative human hepatic progenitor-like cells (ProHPLCs). These ProHPLCs were then re-differentiated into functionally mature hepatocytes using a new hepatocyte maturation condition. Additionally, we investigated the underlying mechanism of PHHs expansion under our new conditions. RESULTS: The novel hepatic expansion medium containing hydrocortisone facilitated the de-differentiation of PHHs into ProHPLCs, which exhibited key hepatic progenitor characteristics and demonstrated a marked increase in proliferation capacity compared to cells cultivated in previously established expansion conditions. Remarkably, these subsequent matured hepatocytes rivaled PHHs in terms of transcriptome profiles, drug metabolizing activities and in vivo engraftment capabilities. Importantly, our findings suggest that the enhanced expansion of PHHs by hydrocortisone may be mediated through the PPARα signaling pathway and regenerative transcription factors. CONCLUSIONS: This study presents a two-step strategy that initially induces PHHs into a proliferative state (ProHPLCs) to ensure sufficient cell quantity, followed by the maturation of ProHPLCs into fully functional hepatocytes to guarantee optimal cell quality. This approach offers a promising means of producing large numbers of seeding cells for hepatocyte-based applications.

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  • Primary hepatocytes exhibit considerable inter-donor variation in proliferative capacity, drug metabolism and stress tolerance, influenced by donor age, genetic background, clinical status and drug exposure history.

    “PHs exhibit considerable inter-donor variation in proliferative capacity, drug metabolism and stress tolerance, which are influenced by donor age, genetic background, clinical status and drug exposure history [15].”

    From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [15]

    The Results section, “Hydrocortisone promotes PHHs expansion in vitro” (page unavailable), explicitly considers donor-related variability in hepatocyte expansion; the Discussion also refers to variability between PHH donor batches. The paper does not establish inter-donor variation in drug metabolism or stress tolerance, or attribute such variation to donor age, genetics, clinical status, or drug exposure history. (LLM, conf=0.86)

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  • Human primary hepatocytes show substantial donor-to-donor variability not typically observed to the same degree in inbred mouse models, and this variability can persist even within engineered expansion or maturation protocols derived from the same donor.

    “Human primary hepatocytes also show substantial donor-to-donor variability that is not typically observed to the same degree in inbred mouse models, and this variability can persist even within engineered expansion or maturation protocols derived from the same donor [4,15].”

    From The Hepatocyte Expansion Paradox: A Review of In Vitro Challenges and Advances (2026) · cites this paper as [15]

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Cited by

Sorted by citations

Authors
Xie, H., Li, G., Fu, Y., Jiang, N., Yi, S., Kong, X., Shi, J., Yin, S., Peng, J., Jiang, Y., Lu, S., Deng, H., & Xie, B.
Affiliations
  • Southwest Medical University (CN)
  • Peking University (CN)
  • Chinese PLA General Hospital (CN)
Corresponding author
  • Lu, Shichun
  • Deng, Hongkui
  • Xie, Bingqing
ORCID
Venue
Stem Cell Research & Therapy
Publisher
Springer Science and Business Media LLC
Published
2024-09-04
Volume
15
Issue
1
Pages
281-281
ISSN
1757-6512 (ISSN-L)
DOI
10.1186/s13287-024-03911-0
PMC
PMC11373096
OpenAlex
W4402252655
Cited by
9
References
3
Citations by year
2026: 7 · 2025: 2
FWCI
2.36
Type
journal article
Language
en
Open access
Licence
Locations
  • Stem Cell Research & Therapy (publishedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • National Natural Science Foundation of China (32300675, 32288102)
  • Sichuan Science and Technology Program (2022YFS0615)
  • Luzhou Science and Technology Bureau (2023SYF136)
  • Luzhou Science and Technology Program of China (2023JYJ019)
  • Doctoral Research Initiation Fund of Affiliated Hospital of Southwest Medical University (22069, 22070)
Topics
  • Health Sciences › Medicine › Hepatology › Liver physiology and pathology
  • Health Sciences › Medicine › Surgery › Tissue Engineering and Regenerative Medicine
  • Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Drug-Induced Hepatotoxicity and Protection
Keywords
primary human hepatocytes, hepatocyte expansion, hepatocyte maturation, cortisol, PPARα signaling, drug metabolism, hepatocyte transplantation, liver regeneration
MeSH terms
Animals, Cell Differentiation, Cells, Cultured, Humans, Liver Regeneration, Cell Culture Techniques, Hepatocytes, Cell Proliferation

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