The Effect of FOXA3 Overexpression on Hepatocyte Differentiation and Liver Regeneration in a Fah cKO Mouse Model

IF 7.1 1区医学 Q1 GASTROENTEROLOGY & HEPATOLOGY

Cellular and Molecular Gastroenterology and Hepatology Pub Date : 2025-01-01 DOI:10.1016/j.jcmgh.2024.101438

Shao Li , Chupeng Ou , Jiajun Zhang , Min Zeng , Kangyan Liang , Qing Peng , Yi Gao

{"title":"The Effect of FOXA3 Overexpression on Hepatocyte Differentiation and Liver Regeneration in a Fah cKO Mouse Model","authors":"Shao Li , Chupeng Ou , Jiajun Zhang , Min Zeng , Kangyan Liang , Qing Peng , Yi Gao","doi":"10.1016/j.jcmgh.2024.101438","DOIUrl":null,"url":null,"abstract":"<div><h3>Background & Aims</h3><div>Stimulated by injury or disease, hepatocytes can regenerate and repair liver tissues through proliferation and differentiation. Partial hepatectomy and liver transplantation are effective treatments for liver diseases. This study investigated the effect of FOXA3 on cell differentiation in HepaRG cell lines under 2- and 3-dimensional culture conditions.</div></div><div><h3>Methods</h3><div>Experiments were performed using a HepaRG cell line that stably overexpressed FOXA3 (RF3) and hepatocyte-specific functions. Moreover, a Fah conditional knockout mouse model (Fah cKO mice) was constructed using the CRISPR-Cas9 method and treated with RF3 spheroids for transplantation. Various molecular biology and immunostaining experiments were performed to assess liver function, hepatocyte structure, and expression levels of cell cycle–related proteins.</div></div><div><h3>Results</h3><div>HepaRG cells that overexpressed FOXA3 had hepatocyte-specific functions. RF3 spheroids expressed liver markers following gene and protein expression analysis. After RF3 spheroid transplantation, Fah cKO mice exhibited increased survival, reduced weight loss, normalization of liver function and hepatocyte structure, and enhanced expression of hepatocyte differentiation factors. However, the expression of cell cycle–related proteins, including p53 and p21, was decreased in vivo. Injection of an HNF4α antagonist revealed that inhibition of HNF4α effectively suppressed the regenerative capacity of the liver after RF3 spheroid transplantation, resulting in an increase in the number of p53- and p21-positive cells and a decrease in the expression levels of liver function–related genes.</div></div><div><h3>Conclusions</h3><div>FOXA3 plays an important role in hepatocyte function. RF3 spheroid transplantation had a therapeutic effect in the Fah cKO mouse model, improving liver function and promoting liver regeneration.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 3","pages":"Article 101438"},"PeriodicalIF":7.1000,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11786892/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Cellular and Molecular Gastroenterology and Hepatology","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2352345X24001930","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"GASTROENTEROLOGY & HEPATOLOGY","Score":null,"Total":0}

引用次数: 0

Abstract

Background & Aims

Stimulated by injury or disease, hepatocytes can regenerate and repair liver tissues through proliferation and differentiation. Partial hepatectomy and liver transplantation are effective treatments for liver diseases. This study investigated the effect of FOXA3 on cell differentiation in HepaRG cell lines under 2- and 3-dimensional culture conditions.

Methods

Experiments were performed using a HepaRG cell line that stably overexpressed FOXA3 (RF3) and hepatocyte-specific functions. Moreover, a Fah conditional knockout mouse model (Fah cKO mice) was constructed using the CRISPR-Cas9 method and treated with RF3 spheroids for transplantation. Various molecular biology and immunostaining experiments were performed to assess liver function, hepatocyte structure, and expression levels of cell cycle–related proteins.

Results

HepaRG cells that overexpressed FOXA3 had hepatocyte-specific functions. RF3 spheroids expressed liver markers following gene and protein expression analysis. After RF3 spheroid transplantation, Fah cKO mice exhibited increased survival, reduced weight loss, normalization of liver function and hepatocyte structure, and enhanced expression of hepatocyte differentiation factors. However, the expression of cell cycle–related proteins, including p53 and p21, was decreased in vivo. Injection of an HNF4α antagonist revealed that inhibition of HNF4α effectively suppressed the regenerative capacity of the liver after RF3 spheroid transplantation, resulting in an increase in the number of p53- and p21-positive cells and a decrease in the expression levels of liver function–related genes.

Conclusions

FOXA3 plays an important role in hepatocyte function. RF3 spheroid transplantation had a therapeutic effect in the Fah cKO mouse model, improving liver function and promoting liver regeneration.

查看原文本刊更多论文

FOXA3过表达对Fah cKO小鼠肝细胞分化和肝再生的影响。

背景与目的：在损伤或疾病的刺激下，肝细胞可以通过增殖和分化来再生和修复肝组织。肝部分切除和肝移植是治疗肝脏疾病的有效方法。本研究考察了FOXA3在二维和三维培养条件下对HepaRG细胞系细胞分化的影响。方法：采用稳定过表达FOXA3 （RF3）和肝细胞特异性功能的HepaRG细胞系进行实验。此外，利用CRISPR-Cas9方法构建Fah条件敲除小鼠模型（Fah cKO小鼠），并用RF3球体处理用于移植。通过各种分子生物学和免疫染色实验来评估肝功能、肝细胞结构和细胞周期相关蛋白的表达水平。结果：过表达FOXA3的HepaRG细胞具有肝细胞特异性功能。通过基因和蛋白表达分析，RF3球体表达肝脏标志物。RF3球体移植后，Fah cKO小鼠的存活率提高，体重减轻，肝功能和肝细胞结构正常化，肝细胞分化因子表达增强。然而，细胞周期相关蛋白，包括p53和p21的表达在体内下降。注射HNF4α拮抗剂发现，抑制HNF4α可有效抑制RF3球体移植后肝脏的再生能力，导致p53-和p21阳性细胞数量增加，肝功能相关基因表达水平降低。结论：FOXA3在肝细胞功能中起重要作用。RF3球体移植在Fah cKO小鼠模型中具有改善肝功能、促进肝脏再生的治疗作用。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

Cellular and Molecular Gastroenterology and Hepatology Medicine-Gastroenterology

CiteScore

13.00

自引率

2.80%

发文量

246

审稿时长

42 days

期刊介绍： "Cell and Molecular Gastroenterology and Hepatology (CMGH)" is a journal dedicated to advancing the understanding of digestive biology through impactful research that spans the spectrum of normal gastrointestinal, hepatic, and pancreatic functions, as well as their pathologies. The journal's mission is to publish high-quality, hypothesis-driven studies that offer mechanistic novelty and are methodologically robust, covering a wide range of themes in gastroenterology, hepatology, and pancreatology. CMGH reports on the latest scientific advances in cell biology, immunology, physiology, microbiology, genetics, and neurobiology related to gastrointestinal, hepatobiliary, and pancreatic health and disease. The research published in CMGH is designed to address significant questions in the field, utilizing a variety of experimental approaches, including in vitro models, patient-derived tissues or cells, and animal models. This multifaceted approach enables the journal to contribute to both fundamental discoveries and their translation into clinical applications, ultimately aiming to improve patient care and treatment outcomes in digestive health.