title: Engineering EMT using 3D micro-scaffold to promote hepatic functions for drug hepatotoxicity evaluation.
abstract: Accompanied by decreased hepatic functions, epithelial-mesenchymal transition (EMT) was observed in two dimensional (2D) cultured hepatocytes with elongated morphology, loss of polarity and weakened cell-cell interaction, while upgrading to 3D culture has been considered as significant improvement of its 2D counterpart for hepatocyte maintenance. Here we hypothesize that 3D culture enhances hepatic functions through regulating the EMT status. Biomaterial-engineered EMT was achieved by culturing HepaRG as 3D spheroids (SP-3D) or 3D stretched cells (ST-3D) in non-adherent and adherent micro-scaffold respectively. In SP-3D, constrained EMT of HepaRG, a hepatic stem cell line, as represented by increased epithelial markers and decreased mesenchymal markers, was echoed by improved hepatic functions. To investigate the relationship between EMT status and hepatic functions, time-series RNA-Seq and gene network analysis were used for comparing different cell culture models, which identified histone deacetylases (HDACs) as key mediating factors. Protein analysis confirmed that high HDAC activity was correlated with high expression of Cadherin-1 (CDH1) and hepatic function genes, which were decreased upon HDAC inhibitor treatment in SP-3D, suggesting HDACs may play positive role in regulating EMT and hepatic functions. To illustrate the application of 3D micro-scaffold culture in drug safety evaluation, hepatotoxicity and metabolism assays of two hepatotoxins (i.e. N-acetyl-p-aminophenol and Doxorubicin) were performed and SP-3D showed more biomimetic toxicity response, indicating regulation of EMT as a vital consideration in designing 3D hepatocyte culture configuration.
authors.affiliation: Department of Biomedical Engineering, School of Medicine, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Jingyu
authors.initials: J
authors.lastname: Wang
authors.affiliation: MOE Key Laboratory of Bioinformatics, Bioinformatics Division & Center for Synthetic and Systems Biology, Department of Automation, TNLIST, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Fengling
authors.initials: F
authors.lastname: Chen
authors.affiliation: The School of Life Science, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Longwei
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of Biomedical Engineering, School of Medicine, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Chunxiao
authors.initials: C
authors.lastname: Qi
authors.affiliation: The School of Life Science, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Bingjie
authors.initials: B
authors.lastname: Wang
authors.affiliation: Department of Biomedical Engineering, School of Medicine, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Yan
authors.affiliation: Department of Plastic, Reconstructive and Aesthetic Surgery, Beijing Tsinghua Changgung Hospital, Medical Center, Tsinghua University, Beijing, 102218, PR China.
authors.forename: Chenyu
authors.initials: C
authors.lastname: Huang
authors.affiliation: Tianjin Second People's Hospital and Tianjin Institute of Hepatology, Tianjin, 300192, PR ​China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hou
authors.affiliation: MOE Key Laboratory of Bioinformatics, Bioinformatics Division & Center for Synthetic and Systems Biology, Department of Automation, TNLIST, Tsinghua University, Beijing, 100084, PR China.
authors.forename: Michael Q
authors.initials: MQ
authors.lastname: Zhang
authors.affiliation: MOE Key Laboratory of Bioinformatics, Bioinformatics Division & Center for Synthetic and Systems Biology, Department of Automation, TNLIST, Tsinghua University, Beijing, 100084, PR China. Electronic address: yc@tsinghua.edu.cn.
authors.forename: Yang
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Biomedical Engineering, School of Medicine, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, Beijing, 100084, PR China; China Orthopedic Regenerative Medicine Group (CORMed), Hangzhou, Zhejiang, 310058, PR ​China. Electronic address: duyanan@tsinghua.edu.cn.
authors.forename: Yanan
authors.initials: Y
authors.lastname: Du
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D058750:Epithelial-Mesenchymal Transition; D005786:Gene Expression Regulation; D053263:Gene Regulatory Networks; D022781:Hepatocytes / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D006655:Histone Deacetylases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D054457:Tissue Scaffolds / Q000737:chemistry*; D018675:Toxicity Tests / Q000379:methods*; D059467:Transcriptome
chemical_list: D006655:Histone Deacetylases
keywords: 3D micro-scaffold; Drug hepatotoxicity evaluation; EMT; HDACs; Hepatic functions