Table 1.

Current modeling strategies and applications of human multi-cellular liver organoids

Model/application	Multiple cells (ratio)	Culture system	Advances/significance	Limitations	Ref.
ALF	iPSC-HEs, HUVECs, BM-MSCs (10:8:2)	3D, Matrigel	Multi-cellular LOs with vascularization	Low reproducibility; Time-consuming	2013[65]
ALF	HEs, MCs, ECs (all from iPSCs) (10:8:2)	3D, ULA	All-iPSC-based strategy	Time-consuming; High cost	2017[16]
ALF	iPSC endoderm cells, HUVECs, UC-MSCs (10:7:1)	3D, ULA	LOs generated from single donor-derived cells	Low reproducibility; Time-consuming	2018[75]
ALF	iPSCs, HAMECs (3:1)	EB, Agarose	HAMECs improved hepatic functions	Unable to reflect the nature cellular composition of liver	2019[102]
Liver fibrosis	HepaRG, THP-1, hTERT-HSC	3D, Hanging drop	LOs derived from cell lines	Functional deficiency	2017[103]
Liver fibrosis and steatohepatitis	PHHs, KCs, HSCs, SECs (16:2:1:1)	3D, ULA	LOs derived from primary cell sources	Low reproducibility; High cost	2018[104]
Steatohepatitis	Hepatocytes, HSCs, BCs, KCs (all from iPSC)	3D, ULA	Co-differentiation of multiple cell lineages for iPSC- LOs	Functions undetermined; Potential inter/intra-batch variability	2019[51]
HBV infection ex vivo	iPSC endoderm cells, HUVECs, BM- MSCs (10:7:1)	3D, ULA	Validation of advantages of iPSC-LOs in HBV modeling	Low reproducibility; Time-consuming	2018[15]
Hepatic differentiation	iPSC-HEs, MSCs, HUVECs (10:2:7)	3D, Matrigel	Platform to identify developmental paracrine signals involved in hepatocyte differentiation	Low reproducibility; Time-consuming	2017[105]
Hepatic differentiation	iHEPs, ECs, HSCs, cholangiocytes (10:7:2:1)	3D, ULA	Cholangiocytes impaired the hepatic functions in LOs and were associated with the liver disease relevant phenotype	Cholangiocyte activation in LOs was unclear	2019[106]
Liver development and angiogenesis	Hepatocytes, BCs, ECs, HSCs (all from iPSC)	3D, Matrigel	Engineered iPSC-LOs by programming of the gene regulatory network	Not completed for liver functions	2021[66]

ALF: Acute liver failure; BCs: Biliary cells; BM: Bone marrow; EB: Embryoid body; ECs: Endothelial cells; HAMECs: Human adipose microvascular endothelial cells; HBV: Hepatitis B virus; HEs: Hepatic endoderm cells; HSCs: Hepatic stellate cells; HUVECs: Human umbilical vein endothelial cells; iHEPs: Induced hepatocytes; KCs: Kupffer cells; Los: Liver organoids; MCs: Mesenchymal cells; MSCs: Mesenchymal stem cells; NPCs: Non-parenchymal cells; PHHs: Primary human hepatocytes; SECs: Sinusoidal endothelial cells; UC: Umbilical cord; ULA: Ultralow adhesion microwell plate.