Skip to main content
. 2022 Sep 28;23(19):11428. doi: 10.3390/ijms231911428

Table 2.

Cellular models for studying chronic hepatotoxicity.

Cellular Model Long-Term Stability Characterization Hepatotoxicity Assessments Ref.
2D cultures
PHH collagen
(serum free conditions)		 Up to 4 weeks
  • -

    Maintenance of functionality (urea and albumin production, CYP activities), morphology and expression of hepatic markers
  • -

    6 drugs (cyclophosphamide, acrolein, rifampicin, loratidine, atorvastatin, APAP) for acute (48 h) or chronic (23 days) exposure

 [86]
PHH sandwich Up to 2 weeks
  • -

    Maintenance of functionality (urea and albumin production, CYP activities), polarisation and expression of hepatic markers up to 7–10 days
  • -

    Cholestatic compounds (+/−BA) incubated for 24 h. Assessment of CIx and urea production

  • -

    Repeated dose of 7 model compounds (14 days). Measurement of ATP content

 [87]
[88]
HepaRG cells Up to 4 weeks (+2 weeks-treatment)
  • -

    Maintenance of phase I and phase II enzymes activity and expression levels.

  • -

    Stable levels of antioxidant enzymes

  • -

    Maintenance of morphology
  • -

    Assessment of drug-induced steatosis after acute or chronic exposure)

  • -

    Long-term repeated dose up to 14 days. Evaluation by HCS

 [89,90]
[91,92,93,94]
Upcyte Human Hepatocytes Up to 3 weeks
  • -

    Maintenance of expression and activity levels of drug-metabolizing enzymes

  • -

    Stable levels of antioxidant enzymes
  • -

    Long-term repeated dose up to 21 days. Evaluation by HCS (mechanistic studies including drug-induced phospholipidosis and steatosis)

 [92,95]
HLC Up to 2 weeks
  • -

    Commercially available HLCs

  • -

    Maintenance of key hepatic markers
  • -

    Repeated dose (2, 7, or 14 days) of 4 model compounds (amiodarone, aflatoxin B, troglitazone and ximelagatran).

  • -

    Drug-induced phospholipidosis and steatosis

 [96]
[96,97]
3D cultures
PHH spheroids Up to 5 weeks
  • -

    Maintenance of expression and activity levels of drug-metabolizing enzymes

  • -

    Proteomic analysis revealed resemblance with liver in vivo.
  • -

    123 drugs (14-day repeated-dose exposure). ATP content.

  • -

    Fialuridine (32-day repeated-dose exposure) Changes in ATP, viability, lipid content and ROS production

  • -

    Drug-induced cholestasis (cholestatic compounds +/BA) at repeated-dose exposure (up to 28 days)

 [98]
[99]
[100]
HepG2 spheroids Up to 1 week
  • -

    Histological characterization, CYP activities, albumin secretion, expression hepatic markers and CLF (transport)
  • -

    Repeated-dose exposure (6 days)

 [101]
HepaRG Spheroids >4 weeks
  • -

    Higher levels of liver-specific genes involved in drug metabolism, BA transport, and energetic pathways and secreted more albumin, glucose, and urea compared to those in 2D cultures
  • -

    7-day exposure to model compounds. Multiplex hepatotoxicity and CYP induction assay.

  • -

    14-day repeated-dose study

  • -

    Evaluation of drug-induced cholestasis and BA effects.

 [102]
[103]
[100,103]
HLC spheroids
  • -

    RNAseq and functional profiling. Differentiation of immature vs. mature zonal hepatocytes

  • -

    BA production and transport
  • -

    Screening of 238 compounds (24 h). Measurement of viability and CLF uptake, and mitochondrial-induced toxicity

  • -

    Modeling CYP2C9*2 iPSC-liver organoids and susceptibility to bosentan-induced cholestasis

 [104]
Co-cultures
Micropatterned co-cultures fibroblast + PHH Up to 4 weeks
  • -

    Secretion of urea and albumin, functional bile canaliculi; metabolisation of compounds using active Phase I and Phase II drug metabolism enzymes
  • -

    45 drugs (14 days exposure, 4 concentrations up to 100× Cmax).

  • -

    Evaluation of urea, albumin and GSH.

 [105]
Micropatterned co-cultures HLC+ fibroblasts Up to 4 weeks
  • -

    Improved functionality (polarity, albumin and urea secretion, Phase I and II activities, induction, down+-regulation of fetal markers)
  • -

    47 drugs (6-day treatment) Evaluation of albumin, urea, ATP

 [106]
3D scaffold (PHH, stellate, KC and endothelial cells) Up to 3 months
  • -

    Maintenance of the production of albumin, fibrinogen, transferrin and urea; CYP inducibility, bile canaliculi-like structures and response to inflammatory stimuli
  • -

    3 and 15 days (troglitazone, APAP, trovafloxacin). LDH release.

 [107]
3D InsightTM Human Liver Microtissues (PHHs, endothelial, KCs) Up to 5 weeks
  • -

    Morphological characterization, glycogen accumulation, polarized expression of transporters (BSEP, MDR1). Responsive to LPS treatment
  • -

    APAP, diclofenac and (14 days treatment (3 re-dosing)

  • -

    Trovafloxacin +/LPS

 [108]
Bioprinted 3D Primary liver tissues (human stellate cells, HUVECs, PHHs) Up to 4 weeks
  • -

    Maintenance of ATP levels, albumin as well as expression and drug-induced CYPs. In vivo relevant architecture
  • -

    Trovafloxacin and levofloxacin (repeated-dose 7 days). Evaluation of albumin and ATP content. Analysis of histological effects

 [109]
Organ-on-a-Chip Platforms
Liver on a chip: bioprinted HepG2 spheroids Up to 4 weeks
  • -

    GelMa hydrogels containing HepG2/C3A maintained the expression of key hepatic markers. Monitoring of biomarkers.
  • -

    Toxicity APAP (6 days). Monitoring levels of albumin, A1AT, transferrin and ceruloplasmin.

 [110]
Liver-Chip (PHH KC + endothelial) Up to 2 weeks
  • -

    Drug-metabolising capacity, albumin secretion, transporters functionality
  • -

    Toxicity of APAP (7 days) Human and cross-species toxicities

  • -

    Methotrexateand fialuridineinduced fibrosis-steatosis (7 and 10 days, respectively)

 [111]
Biomimetic array chip (collagen 3D PHHs) Up to 12 days
  • -

    Improved and stabilized liver functionality (viability, albumin and urea production). CYP functionality.
  • -

    Toxicity of 122 clinical drugs (treatment for 7 and 14 days). ATP content.

 [112]
Multi-organ-chip (PHHs + stellate + skin) Up to 4 weeks
  • -

    Expression of key hepatic markers. Maintenance of metabolic activities.
  • -

    Toxicity of troglitazone (6 days). Cell viability and transcriptomics analysis.

 [113]