Simple and Quick Method to Obtain a Decellularized, Functional Liver Bioscaffold
The development of new approaches for organ transplantation has become crucial in the last years. In particular, organ engineering, involving the preparation of acellular matrices that provide a natural habitat for reseeding with an appropriate population of cells, is an attractive although technically demanding approach. We here describe a method that allows for the derivation of functional in vitro hepatic organoids and that does not require a previous selection of all the parenchymal hepatocytes and non-parenchymal cells, namely, Kupffer cells, liver endothelial cells, and hepatic stellate cells. The procedure also replaces the costly standard collagenase perfusion step with a trypsin-based enzymatic digestion that results in high-yield decellularization. A combination of physical and chemical treatments through deep immersion and intraluminal infusion of two different consecutive solutions is used: (1) deionized water (DI) and (2) DI + Triton X 1% + ammonium hydroxide (NH4OH) 0.1%. This ensures the isolation of the hepatic constructs that reliably maintain original architecture and ECM components while completely removing cellular DNA and RNA. The procedure is fast, simple, and cheap and warrants an optimal organoid functionality that may find applications in both toxicological and transplantation studies.
KeywordsHepatocyte Scaffold Decellularization Organoid Liver bioengineering
This work was supported by Carraresi Foundation and by Fondazione CMT Cura Mini-invasiva Tumori ONLUS. T.A.L.B., F.G., M.G., and A.Z. are members of the COST Actions CA16119.
- 2.Badylak SF, Taylor D, Uygun K (2011) Whole-organ tissue engineering: Decellularization and recellularization of three-dimensional matrix scaffolds. Annu Rev Biomed Eng 13:27–53. https://doi.org/10.1146/annurev-bioeng-071910-124743CrossRefPubMedGoogle Scholar
- 3.Gilbert TW, Sellaro TL, Badylak TF (2006) Decellularization of tissues and organs. Biomaterials 27(19):3675–3683. https://doi.org/10.1016/j.biomaterials.2006.02.014CrossRefPubMedGoogle Scholar
- 7.Uygun BE, Soto-Gutierrez A, Yagi H, Izamis ML, Guzzardi MA, Shulman C, Milwid J, Kobayashi N, Tilles A, Berthiaume F, Herti M, Nahmias Y, Yarmush ML, Uygun K (2010) Organ reengineering through development of a transplantable recellularized liver graft using decellularized liver matrix. Nat Med 16(7):814–820. https://doi.org/10.1038/nm.2170CrossRefPubMedPubMedCentralGoogle Scholar