Organoids as preclinical platforms for metabolic dysfunction-associated steatotic liver disease: Model construction, mechanistic dissection, and therapeutic translation

Table 1 Metabolic dysfunction-associated steatotic liver disease model construction and phenotypic recapitulation in liver organoids derived from human pluripotent stem cells

Cellular origin	Cell types	Model induction method	Phenotype reproduced	Ref.
hESCs	Hepatocyte-like, cholangiocyte-like	200/500 μM OA and PA for 4-14 days	Time- and dose-dependent elevation of lipid accumulation, ROS, and lipid peroxidation, accompanied by bile canaliculi network decay and ductular reactions	Ramli et al[11], 2020
hESCs	Hepatocyte-like	450 μM FFA (OA:PA = 2:1) for 2 days	Disrupted glucose/Lipid metabolism, oxidative stress, apoptosis, loss of polarization, impaired liver function, and ductular reaction	Qin et al[12], 2025
hiPSCs	Hepatocyte-like, KC-like, HSC-like	200/400/800 μM OA for 3-7 days	Steatosis, inflammation, and fibrosis	Ouchi et al[14], 2019
hESCs	Hepatocyte-like, liver sinusoidal endothelial cell-like	450 μM OA and 150 μM PA for 24/48 hours	Pericentral organoids from liver lobules exhibit faster and greater lipid droplet accumulation than periportal organoids	Zhang et al[15], 2025
hPSCs	Hepatocyte-like, cholangiocyte-like, HSC-like	10 ng/mL and 25 ng/mL TGF-β1/500 μM OA/500 μM PA for 4 days	OA induces benign steatosis; PA induces inflammatory changes of steatohepatitis; TGF-β1 induces steatohepatitis and fibrosis most prominently	Hess et al[16], 2023
hiPSCs	Hepatocyte-like, HSC-like, biliary-like	500 μM OA for 4 days, followed by treatment with 500 μM OA combined with 25 ng/mL TGF-β for another 3 days	Severe steatosis, oxidative stress, mitochondrial dysfunction, inflammation, and fibrosis as progressive characteristics of human MASH	Wu et al[17], 2024
hiPSCs	Hepatocyte-like, cholangiocyte-like	500 μM OA for 3 days	Increased lipid droplet accumulation, upregulated expression of fibrosis and lipid metabolism markers in higher stiffness matrices	Liu et al[18], 2025
hiPSCs	Hepatocyte-like, cholangiocyte-like	600 μM FFA (OA:PA = 2:1) for 7 days	Lipid accumulation, ROS production and up-regulated expression of inflammatory and fibrotic markers	Wang et al[20], 2020
hiPSCs and liver stem cells	Hepatocytes, cholangiocyte-like, KC-like, HSC-like	450 μM OA, 150 μM PA, 100 mmol/L fructose for 3 days, followed by addition of 10 ng/mL lipopolysaccharide and 10 mmol/L phenylacetic acid another 3/5 days	Steatosis, hepatocellular ballooning, inflammatory and fibrogenic microenvironment	Balachander et al[21], 2025
hiPSCs	Hepatocyte-like, cholangiocyte-like	400 μM OA and 200 μM PA for 48 hours	Dysregulated lipid metabolism, increased inflammatory factor secretion, and fibrosis	You et al[22], 2026

hESCs: Human embryonic stem cells; OA: Oleic acid; PA: Palmitic acid; ROS: Reactive oxygen species; FFA: Free fatty acids; hiPSCs: Human induced pluripotent stem cells; KC: Kupffer cell; HSC: Hepatic stellate cell; TGF-β: Transforming growth factor-β; MASH: Metabolic dysfunction-associated steatohepatitis; hPSCs: Human pluripotent stem cells.

Table 2 Metabolic dysfunction-associated steatotic liver disease model construction and phenotypic recapitulation in liver organoids derived from primary tissues

Cellular origin	Cell types	Model induction method	Phenotype reproduced	Ref.
Mice liver tissue	Mixture of cells from liver tissue	Mice fed with methionine-and choline-deficient diet for 4, 8, and 12 weeks	Liver fibrosis without lipid accumulation	Elbadawy et al[33], 2020
Mice liver tissue	Mixture of cells from liver tissue	245 μM OA and 122.5 μM PA for 48 hours	Lipid accumulation, increased liver fibrosis and inflammation	Zhou et al[34], 2024
Mice liver tissue and human fetal liver tissue	Mixture of cells from liver tissue	200 μM OA/150 μM OA and 150 μM PA for 3 days	Greater lipid accumulation in human vs mouse organoids	Liu et al[35], 2024
Adult stem cells from human liver tissue	Hepatocyte-like	600 μM OA for 3 days	Lipid accumulation, ROS production and inflammation	Zhu et al[36], 2025
Harbor residential bipotent progenitor cells from human liver tissue	Cholangiocyte-like, hepatocyte-like	Lactate/pyruvate/octanoic acid (10:1:2 mM; 15:1.5:3 mM) for 96 hours	Lipid accumulation, suppressed organoid growth, metabolic dysregulation, and mitochondrial impairment	Wang et al[37], 2022
Liver tissue from healthy individuals and MASLD patients and HLO: Primary human liver stellate cells = 5:1	Hepatocytes, cholangiocytes, stem cells, KC, HSC	750 μM PA for 72 hours	Steatosis, inflammation, and fibrosis	Goswami et al[38], 2025
Liver tissue from MASH patients	Hepatocytes, cholangiocytes	1.2 mmol/L and 2 mmol/L PA for 6 hours/2 mmol/L OA for 24 hours	MASH liver-derived organoids exhibit significantly reduced regenerative capacity, upregulated activity of multiple pathogenic pathways (inflammation, fibrosis, tumorigenesis), and greater susceptibility to PA-induced apoptosis and OA-induced lipid accumulation	McCarron et al[39], 2021

OA: Oleic acid; PA: Palmitic acid; ROS: Reactive oxygen species; MASLD: Metabolic dysfunction-associated steatotic liver disease; HLO: Human liver organoid; KC: Kupffer cell; HSC: Hepatic stellate cell; MASH: Metabolic dysfunction-associated steatohepatitis.

Table 3 Identification of potential metabolic dysfunction-associated steatotic liver disease biomarkers using liver organoid models

Biomarker	Cellular origin	Model	Expression	Function/Description	Ref.
26-hydroxycholesterol	hiPSCs	250 μM OA, 250 μM PA, 10 mmol/L fructose for 48 hours	Upregulated	26-hydroxycholesterol exerts an anti-inflammatory signal on quiescent HSCs	Kømurcu et al[27], 2023
ADGRG1	hPSCs	500 μM OA/500 μM PA/10 ng/mL TGF-β1 for 4 days	Upregulated	The transcriptional profile of ADGRG1 is upregulated in cells of the fibrotic and inflammatory niche in response to injury	Pirola et al[29], 2026
lncRNA ENSMUST00000244884	Primary biliary duct-derived LOs from mice liver tissue	2/10 ng/mL FSH for 48 hours	Upregulated	Knocking down this long non-coding RNA promotes liver cholesterol excretion and accelerating lipid metabolism	Yu et al[42], 2026
Tryptophan-derived metabolites	Primary ductal organoids-derived LOs from mice liver tissue	200 μM OA and 100 μM PA for 3 days	Upregulated	Tryptophan-derived metabolites promote lipid droplet formation and enhance the inflammatory response	Zhan et al[46], 2025
sCD46	Human liver tissue	2 mmol/L FFA (OA:PA = 2:1) for 24 hours	Upregulated	Lipid accumulation in human hepatocytes reduces cell-surface CD46 via matrix metalloproteinases, thereby releasing sCD46 and promoting the differentiation of IL-4+ iNKT cells	Bitterer et al[54], 2024
CES1/IL18R1	Harbor residential bipotent progenitor cells from human liver tissue	Lactate/pyruvate/octanoic acid (10:1:2 mM; 15:1.5:3 mM) for 96 hours	Upregulated	CES1, IL-18R1: Population-based proteomics biomarkers validated in human LOs for MASLD/fibrosis	Abozaid et al[55], 2023
HAMP1	Human liver tissue	400 μM OA and 200 μM PA	Upregulated	Elevated pre-treatment HAMP1 predicts better response to saponin therapy in MASLD, and reduced HAMP1 post-treatment links to histological improvement	Kim et al[62], 2025

hiPSCs: Human induced pluripotent stem cells; OA: Oleic acid; PA: Palmitic acid; HSC: Hepatic stellate cell; ADGRG1: Adhesion G-protein-coupled receptor G1; hPSCs: Human pluripotent stem cells; TGF-β: Transforming growth factor-β; LO: Liver organoid; sCD46: Soluble CD46; FSH: Follicle-stimulating hormone; FFA: Free fatty acids; iNKT: Invariant natural killer T; CES1: Carboxylesterase 1; IL18R1: Interleukin-18 receptor 1; MASLD: Metabolic dysfunction-associated steatotic liver disease; HAMP1: Hepcidin antimicrobial peptide 1.

Table 4 Metabolic dysfunction-associated steatotic liver disease model construction and phenotypic recapitulation in liver organoids derived from immortalized cell lines

Cellular origin	Cell types	Model induction method	Phenotype reproduced	Ref.
HepG2 (PNPLA3 I148M homozygous):LX-2 = 24:1	Hepatocytes, HSC	500 μM FFA (OA:PA = 2:1) for 48 hours	Lipid accumulation and fibrosis	Pingitore et al[68], 2019
HepaRG	Hepatocytes, cholangiocytes	500 μM FFA (OA:PA = 1:1) for 48 hours	Lipid accumulation, impaired glucose regulation, and reduced Akt phosphorylation	Teng et al[69], 2021
HepaRG:primary human macrophages:LX-2 = 8:1:1	Hepatocytes, cholangiocytes, HSC, KC	150 μM stearic acid and 300 μM OA for 9 days	Steatosis, increased expression of fibrosis markers and pro-inflammatory cytokines	Bronsard et al[70], 2024; Duclos et al[71], 2026

HSC: Hepatic stellate cell; FFA: Free fatty acids; OA: Oleic acid; PA: Palmitic acid; KC: Kupffer cell.