Abstract
Liver fibrosis is the late consequence of chronic liver inflammation which could eventually lead to cirrhosis, and liver failure. Among various etiological factors, activated hepatic stellate cells (aHSCs) are the major players in liver fibrosis. To date, various in vitro liver fibrosis models have been introduced to address biological and medical questions. Availability of traditional in vitro models could not fully recapitulate complicated pathology of liver fibrosis. The purpose of this study was to develop a simple and robust model to investigate the role of aHSCs on the progression of epithelial to mesenchymal transition (EMT) in hepatocytes during liver fibrogenesis. Therefore, we applied a micropatterning approach to generate 3D co-culture microtissues consisted of HepaRG and human umbilical cord endothelial cells (HUVEC) which co-cultured with inactivated LX-2 cells or activated LX-2 cells, respectively, as normal or fibrotic liver models in vitro. The result indicated that the activated LX-2 cells could induce EMT in HepaRG cells through activation of TGF-β/SMAD signaling pathway. Besides, in the fibrotic microtissue, physiologic function of HepaRG cells attenuated compared to the control group, e.g., metabolic activity and albumin secretion. Moreover, our results showed that after treatment with Galunisertib, the fibrogenic properties decreased, in the term of gene and protein expression. In conclusion, it is proposed that aHSCs could lead to EMT in hepatocytes during liver fibrogenesis. Furthermore, the scalable micropatterning approach could provide enough required liver microtissues to prosper our understanding of the mechanisms involved in the progression of liver fibrosis as well as high throughput (HT) drug screening.
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Funding
This project was financially supported by grants from “The National Institute for Medical Research Development (NIMAD, 940231 and 982403), Tehran, Iran”, to M.V. In addition, the project was partially funded by institutional funding and a grant by the Federal Ministry of Education and Research (FZK: 01DK20020) to AN.
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EZ performed all the experiments and drafted the manuscript. EZ, MV, AN, MR, AO, RA, BB were involved in the bioassays and analysis and discussion of data. AP revised and approved the manuscript critically. MV & AN developed the idea, supported the project, and revised and finally approved the manuscript.
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Zahmatkesh, E., Othman, A., Braun, B. et al. In vitro modeling of liver fibrosis in 3D microtissues using scalable micropatterning system. Arch Toxicol 96, 1799–1813 (2022). https://doi.org/10.1007/s00204-022-03265-7
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DOI: https://doi.org/10.1007/s00204-022-03265-7