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Biofabrication of size-controlled liver microtissues incorporated with ECM-derived microparticles to prolong hepatocyte function

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Abstract

Multicellular microtissues of primary human hepatocytes (PHHs) co-cultured with other supporting cell types are a promising model for drug screening and toxicological studies. However, these liver microtissues (LMs) rapidly lose their functions during ex vivo culture. Here, in order to mimic the cellular and structural hepatic microenvironment, we co-cultured PHHs with human mesenchymal stromal cells (MSCs) and human umbilical vein endothelial cells (HUVECs) in the presence of cell-sized microparticles (MPs) derived from liver extracellular matrix (LEMPs). The microwell culture platform enabled biofabrication of size-controlled multicellular microtissues (PHH:HUVEC:MSC = 3:2:1) with efficient LEMP incorporation (about 70% at a 2:1 ratio of cells:MP). The biofabricated liver microtissues (BLMs) were cultured ex vivo for 14 days and compared to the cell-only LM in terms of gene and protein expression, functional activity, cytochrome P450 (CYP450) enzyme inducibility, and drug sensitivity. The results supported superior hepatic-related gene expression, functional activity, and polarity for PHH in BLM compared to LM. CYP450 enzyme inducibility and dose-responsive sensitivity to toxic drugs were significantly higher in the BLM group. In conclusion, microtissue engineering by incorporation of tissue-specific microparticles within a multicellular microtissue can offer some advantages for drug discovery studies and cell transplantation applications. In the near future, this approach could generate a scalable platform of several functional biofabricated microtissues representing different organs.

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Acknowledgements

This project was financially supported by Grants from Royan Institute (No. 96000165) to MV and HB; Bahar Tashkhis Teb Co. (Nos. BTT, 9702, and 9802), Iran National Science Foundation (No. 97014445) to MV; the Ministry of Health and Medical Education (No. 56700/147) to HB. We would like to express our sincere gratitude to colleagues in Royan Institute, Regenerative Medicine Department.

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Authors and Affiliations

  1. Department of Developmental Biology, University of Science and Culture, ACECR, 14155-4364, Tehran, Iran

    Zahra Heydari, Ensieh Zahmatkesh & Hossein Baharvand

  2. Department of Regenerative Medicine, Cell Science Research Centre, Royan Institute for Stem Cell Biology and Technology, ACECR, 14155-4364, Tehran, Iran

    Zahra Heydari, Ensieh Zahmatkesh, Zahra Farzaneh, Iman Akbarzadeh & Massoud Vosough

  3. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, 14155-4364, Tehran, Iran

    Zahra Heydari, Ensieh Zahmatkesh, Zahra Farzaneh, Hossein Baharvand & Massoud Vosough

  4. Department of Cell Engineering, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, 14155-4364, Tehran, Iran

    Ibrahim Zarkesh & Mohammad-Hossein Ghanian

  5. Transplant Research Center, Shiraz University of Medical Sciences, 71936-13311, Shiraz, Iran

    Mahdokht H. Aghdaei

  6. N.N. Semenov Federal Research Center for Chemical Physics, Russian Academy of Sciences, Moscow, 117977, Russia

    Svetlana Kotova

  7. Department of Biology and Anatomical Sciences, School of Medicine, Shahid Beheshti University of Medical Sciences, 14155-4364, Tehran, Iran

    Abbas Piryaei

  8. Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, 14155-4364, Tehran, Iran

    Abbas Piryaei

  9. Institute for Regenerative Medicine, Sechenov University, Moscow, 117977, Russia

    Anastasia Shpichka & Peter Timashev

  10. Division of Pathology, Department of Laboratory Medicine, Karolinska Institute, 17177, Stockholm, Sweden

    Roberto Gramignoli

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  1. Zahra Heydari
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  2. Ibrahim Zarkesh
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  3. Mohammad-Hossein Ghanian
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  4. Mahdokht H. Aghdaei
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Contributions

ZH performed the experiments, collected the data, conducted the data analysis and interpretation, and wrote the manuscript. IZ, MG, MA, SK, EZ, ZF, AP, IA, AS, RG, and PT helped in some experiments and analysis for the results, data management and statistical analysis and made final approval of the manuscript. HB and MV generated the concept, designed the study, supported the study and involved in writing and critical editing and proofreading the manuscript.

Corresponding author

Correspondence to Massoud Vosough.

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The authors declare that there is no conflict of interest.

Ethical approval

This project was approved by Ethics Committee, Royan Institute. The approval code number is IR.ACECR.ROYAN.REC.1397.044. All institutional and national guidelines for the care and use of laboratory animals were followed.

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Heydari, Z., Zarkesh, I., Ghanian, MH. et al. Biofabrication of size-controlled liver microtissues incorporated with ECM-derived microparticles to prolong hepatocyte function. Bio-des. Manuf. 4, 790–805 (2021). https://doi.org/10.1007/s42242-021-00137-4

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