On the adhesion-cohesion balance and oxygen consumption characteristics of liver organoids

Abstract

Liver organoids (LOs) are of interest in tissue replacement, hepatotoxicity and pathophysiological studies. However, it is still unclear what triggers LO self-assembly and what the optimal environment is for their culture. Hypothesizing that LO formation occurs as a result of a fine balance between cell-substrate adhesion and cell-cell cohesion, we used 3 cell types (hepatocytes, liver sinusoidal endothelial cells and mesenchymal stem cells) to investigate LO self-assembly on different substrates keeping the culture parameters (e.g. culture media, cell types/number) and substrate stiffness constant. As cellular spheroids may suffer from oxygen depletion in the core, we also sought to identify the optimal culture conditions for LOs in order to guarantee an adequate supply of oxygen during proliferation and differentiation. The oxygen consumption characteristics of LOs were measured using an O2 sensor and used to model the O2 concentration gradient in the organoids. We show that no LO formation occurs on highly adhesive hepatic extra-cellular matrix-based substrates, suggesting that cellular aggregation requires an optimal trade-off between the adhesiveness of a substrate and the cohesive forces between cells and that this balance is modulated by substrate mechanics. Thus, in addition to substrate stiffness, physicochemical properties, which are also critical for cell adhesion, play a role in LO self-assembly.

Injectable Cardiac Cell Microdroplets for Tissue Regeneration

“…As we hypothesized that cell encapsulation within microscale droplets could overcome this challenge, we initially modeled oxygen concentration by using COMSOL multiphysics modeling software. We have taken into consideration oxygen diffusion according to Fick's second law and consumption by the cells according to Michaelis–Menten kinetics . The external concentration of oxygen was considered as the concentration within arterial blood and oxygen diffusion coefficient was considered as the one in a muscle.…”

“…We have taken into consideration oxygen diffusion according to Fick's second law and consumption by the cells according to Michaelis-Menten kinetics. [18] The external concentration of oxygen was considered as the concentration within arterial blood and oxygen diffusion coefficient was considered as the one in a muscle. Two assumptions were made during our mathematical modelling: 1) Twenty minutes is considered the maximal period of time, in which cardiac cells can survive without oxygen.…”

Allometric Scaling of physiologically-relevant organoids

“…Developing physiologically relevant in vitro models to mimic cell and tissue organ response is one of the main challenges in experimental cell biology and tissue engineering. Organoid technology has emerged as a novel method to generate structural and functional tissue models 15,[32][33][34][35] . Luminal organoids were first generated from intestinal stem cells by Clevers and coworkers 36 .…”

“…The results are confirmed by experimental studies on liver buds by Ramachandran et al 22 ; the authors found no evidence of cell death in liver buds cultured in well-mixed fluidic bioreactors while those in static conditions developed a necrotic core. Mattei et al 43 demonstrated that the minimum oxygen level in liver buds with an average mass of a few mg cultured in bioreactors is 0.04 mol/m 3 , compared with 0.01 mol/m 3 in static culture plates. As shown in Table 4, the gel-free structures resembling liver buds maintain the criteria of quarter power scaling (b ≈ -1/4) and viable oxygen concentrations only if their radii are maintained in a narrow range (between 654 and 712 µm in the models).…”

“…allometric scaling and minimum oxygen concentration above Ccrit) are satisfied are smaller than those for the gel-free constructs. Interestingly, in most studies, the sizes of organoids surrounded by a shell of ECM are generally smaller than those without a shell 13,14,22,35 .…”