Long-term maintenance of functional primary human hepatocytes in 3D gelatin matrices produced by solution blow spinning

Mariliis Klaas,Martin Järvekülg,Viljar Jaks,Kristina Mäemets-Allas,Kaidi Möll,Mart Loog

doi:10.1038/s41598-021-99659-1

Mariliis Klaas, Martin Järvekülg + Show 4 more

Open Access

https://doi.org/10.1038/s41598-021-99659-1

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Journal: Scientific Reports	Publication Date: Oct 11, 2021
Citations: 10	License type: open-access

Affiliation: University of Tartu, Tartu University Hospital

Abstract

Solution blow spinning (SBS) has recently emerged as a novel method that can produce nano- and microfiber structures suitable for tissue engineering. Gelatin is an excellent precursor for SBS as it is derived mainly from collagens that are abundant in natural extracellular matrices. Here we report, for the first time the successful generation of 3D thermally crosslinked preforms by using SBS from porcine gelatin. These SBS mats were shown to have three-dimensional fibrous porous structure similar to that of mammalian tissue extracellular matrix. In pharma industry, there is an urgent need for adequate 3D liver tissue models that could be used in high throughput setting for drug screening and to assess drug induced liver injury. We used SBS mats as culturing substrates for human hepatocytes to create an array of 3D human liver tissue equivalents in 96-well format. The SBS mats were highly cytocompatible, facilitated the induction of hepatocyte specific CYP gene expression in response to common medications, and supported the maintenance of hepatocyte differentiation and polarization status in long term cultures for more than 3 weeks. Together, our results show that SBS-generated gelatin scaffolds are a simple and efficient platform for use in vitro for drug testing applications.

Highlights

Solution blow spinning (SBS) has recently emerged as a novel method that can produce nano- and microfiber structures suitable for tissue engineering
When the fibers in SBS mats had a diameter of 1.52 ± 0.54 μm, the decellularized liver microstructure was denser and the matrix fibers were considerably thinner with a diameter of 19.6 ± 7 nm (Supplementary Fig. S1C)
We found that the enzymatic activity of CYP1A2 and 2B6 was even higher in the hepatocytes grown on SBS mats at both basal level and when induced by omeprazole or rifampicin, respectively, when compared to the hepatocytes grown in MG bilayer

Summary

Introduction

Solution blow spinning (SBS) has recently emerged as a novel method that can produce nano- and microfiber structures suitable for tissue engineering. For the first time the successful generation of 3D thermally crosslinked preforms by using SBS from porcine gelatin These SBS mats were shown to have three-dimensional fibrous porous structure similar to that of mammalian tissue extracellular matrix. More recent techniques in hepatocyte culture include 3D liver microtissue biochips coupled to a microfluidic d evice[15] and bioprinting[16,17] Despite of these advances there is a continuous need for a cost-efficient, simple and reliable long-term 3D culture for primary human hepatocytes to assess DILI. SBS has been successfully applied for creating 3D structures from fish gelatin; the single published attempt to utilize mammalian (porcine) gelatin as the input was unsuccessful[26]

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Conclusion