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3D human nonalcoholic hepatic steatosis and fibrosis models

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Abstract

This study presents a simple and robust three-dimensional human hepatic tissue model to emulate steatotic and fibrotic conditions and provide an in vitro model for drug testing and mechanistic studies. Using a photolithographic biofabrication method with a photomask featuring hexagonal units, liver cells, including a human hepatic cell line (HepG2-C3A) and a human hepatic stellate cell line (LX-2) were embedded in gelatin methacryloyl hydrogel. Hepatic steatosis was induced by supraphysiological concentration of free fatty acids; hepatic fibrosis was induced by transforming growth factor-β1. Induction of steatosis was confirmed by Oil Red O and BODIPY staining and was inhibited with toyocamycin and obeticholic acid. Induction of fibrosis was confirmed by immunostaining for collagen type I and alpha smooth muscle actin and inhibited by rapamycin and curcumin treatment. This model was further preliminarily validated using primary human hepatocytes in a similar setup. These constructs provide a viable, biologically relevant, and higher throughput model of hepatic steatosis and fibrosis and may facilitate the study of the mechanisms of disease and testing of liver-directed drugs.

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Availability of data and materials

The datasets that support the findings of this study are available from the corresponding authors upon reasonable request. All requests for raw and analyzed data and materials will be promptly reviewed by the Brigham and Women’s Hospital and University of Florida to verify whether the request is subject to any intellectual property or confidentiality obligations. Any data and materials that can be shared will be released via a Material Transfer Agreement.

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Funding

YSZ received funding from National Institutes of Health (K99CA201603, R00CA201603, R21EB025270, R21EB026175, R01EB028143, R03EB027984), National Science Foundation (1935105), Brigham Research Institute New England Anti-Vivisection Foundation, and American Fund for Alternatives to Animal Research (AFAAR). AZ received funding from National Institutes of Health (K08DK113244, R01MD012579). SD received funding from National Institutes of Health (R01MD012579-UT20664DS).

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Authors and Affiliations

  1. Division of Engineering in Medicine, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, 65 Landsdowne St, Cambridge, MA, 02139, USA

    Sushila Maharjan, Diana Bonilla, Princy Sindurakar, Hongbin Li, Wanlu Li & Y. Shrike Zhang

  2. Department of Biology, College of the Holy Cross, Worcester, MA, 01610, USA

    Princy Sindurakar

  3. Department of Surgery, University of Florida, 1600 SW Archer Rd, Gainesville, FL, 32610, USA

    Sergio Duarte & Ali Zarrinpar

  4. Department of Biomedical Engineering, University of Florida, Gainesville, FL, 32610, USA

    Ali Zarrinpar

  5. Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, FL, 32610, USA

    Ali Zarrinpar

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  1. Sushila Maharjan
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  2. Diana Bonilla
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  3. Princy Sindurakar
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  4. Hongbin Li
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  5. Wanlu Li
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  6. Sergio Duarte
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  7. Ali Zarrinpar
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  8. Y. Shrike Zhang
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Contributions

SM, SD, AZ, and YSZ were involved in conceptualization. SM, DB, and PS were involved in data curation. SM, DB, and YSZ performed formal analysis. SM, DB, PS, HL, WL, AZ, and YSZ conducted investigation. SM, DB, SD, and YSZ contributed to methodology. AZ and YSZ contributed to project administration. AZ and YSZ contributed resources. AZ and YSZ conducted supervision. SM, AZ, and YSZ were involved in validation. SM and YSZ contributed to visualization. SM, AZ, and YSZ were involved in writing original draft. SM, DB, PS, HL, WL, SD, AZ, and YSZ were involved in writing review and editing.

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Correspondence to Ali Zarrinpar or Y. Shrike Zhang.

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Maharjan, S., Bonilla, D., Sindurakar, P. et al. 3D human nonalcoholic hepatic steatosis and fibrosis models. Bio-des. Manuf. 4, 157–170 (2021). https://doi.org/10.1007/s42242-020-00121-4

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