Table 2.

Key differences between 2D and 3D culture, regarding morphology, response to materials, in vivo likeness, and other key parameters

Property	2D liver cell model	3D liver cell model
Morphology and architecture	Sheet-like, flat, stretched cells grown in monolayers; do not mimic natural architecture of liver	In vivo-like cell shape; high similarities to in vivo liver architecture
Cell proliferation	Cells proliferate at a higher rate than in vivo	Cells proliferate at faster or slower rate than 2D culture, depending on cell type/3D system
Protein/gene expression	Often display different expression levels to human liver tissues	Protein/gene expression levels similar to those found in human liver tissues
Access to oxygen, metabolites, nutrients, and signalling molecules	Unlimited access	Access is defined by the 3D morphology of the cultures as per in vivo conditions
Cell–cell interactions	Cannot recapitulate cell–cell and/or cell-ECM interactions due to flat morphology	Appropriate interactions between cell–cell and cell-ECM are established
Multicellular composition	Co-cultures can be formed; number of cell types co-cultured is limited	Tissue composition can be fully replicated (e.g., organoids)
Sensitivity to stimuli/hepatotoxins	Sensitivity is often not comparable to the in vivo liver tissue	Better predictors of in vivo responses
Exposure to NBMs	All cells are equally exposed to NBMs	Depending on culture morphology, NBMs may not penetrate the core and reach all cells, as per in vivo conditions
Reproducibility	Reproducible high-performance and simple but highly reductionist	Reproducibility depends on method, can be user-dependent, but it can be optimised
Cost of maintaining culture	Cheap, all reagents/materials commercially available	Often more expensive, time consuming, increased batch-to-batch variation