TY - JOUR
T1 - Simple and Quick Method to Obtain a Decellularized, Functional Liver Bioscaffold
AU - Ghiringhelli, Matteo
AU - Zenobi, Alessandro
AU - Brizzola, Stefano
AU - Gandolfi, Fulvio
AU - Bontempo, Valentino
AU - Rossi, Sandro
AU - Brevini, Tiziana A.L.
AU - Acocella, Fabio
PY - 2018/1/1
Y1 - 2018/1/1
N2 - 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.
AB - 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.
KW - Decellularization
KW - Hepatocyte
KW - Liver bioengineering
KW - Organoid
KW - Scaffold
UR - http://www.scopus.com/inward/record.url?scp=85057557469&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85057557469&partnerID=8YFLogxK
U2 - 10.1007/7651_2017_97
DO - 10.1007/7651_2017_97
M3 - Article
C2 - 29101679
AN - SCOPUS:85057557469
VL - 1577
SP - 283
EP - 292
JO - Methods in Molecular Biology
JF - Methods in Molecular Biology
SN - 1064-3745
ER -