Speaker
Description
Introduction. The liver is a key organ that plays a crucial role in metabolism and is responsible for various functions in the body, including homeostasis, synthesis of essential components, nutrient storage, and detoxification [1]. With the growing need for reliable and effective in vitro liver models for toxicological studies, recent advances have been made in tissue engineering, biomaterials, microfabrication, and cell biology. Among the models developed using bioengineering techniques, induced pluripotent stem cells (iPSCs) derived three-dimensional (3D) hepatic organoids and liver-on-a-chip platforms have shown promising results by mimicking in vivo liver tissue behavior, providing a controlled physiological microenvironment, and eliciting cellular metabolic responses [2,3]. However, a significant limitation of these models is their retention of immature, fetal hepatoblast characteristics. Additionally, they fail to fully recapitulate the complete spectrum of mature hepatic cell types and exhibit substantial inter-culture variability, hindering their capacity to accurately model mature liver physiology [4]. So, in this study, we aimed to evaluate the molecular and functional maturation of hepatic organoids on a newly designed microfluidic chip platform that simulates low shear stress conditions with in vivo-like physiological dynamic laminar flow. The goal was to develop a model that closely approximates human liver functionality, as assessed through various functional analyses, including albumin secretion.
Methods. To this end, the Box-Behnken module in the Design-Expert v7 software was employed to identify the independent variables; organoid passage number (P/3-15), perfusion flow rate (5-15 µl/min), and maturation duration (7-13 days) that would optimize albumin level, a key functional marker of hepatic organoid maturation. These variables were selected as inputs, and a predicted experimental design was generated.
Results. Based on this design, iPSCs-derived hepatic organoids were cultured on-chip platforms during both expansion and maturation period after embedding in hydrogel (Figure 1), with albumin concentration data serving as output for statistical analysis. Using response surface methodology, the program determined the optimal parameters as independent variables, and further characterization of the optimal on-chip maturation experiments were carried out based on fat accumulation, ammonia and urea levels, CYP3A4 cytochrome activity and confocal imaging for specific markers after FunGI tissue clearing.
Figure 1. a. Hepatic organoids in culture and on b,c. layer-by-layer designed perfusable microfluidic platform
Discussion. Our newly designed perfusable liver organoid-on-a-chip model holds significant potential for preclinical pharmacokinetic high-throughput drug development and screening studies, due to its ability to serve as a functional perfused interface compatible with other organ-specific organoid-on-a-chip systems and to be integrated into humanoid-on-chip platforms.
Acknowledgments
This study was financially supported by the Scientific and Technological Research Council of Turkey (TUBITAK) – Science Fellowships and Grant Programmes (BIDEB) 2218 through number of 123C325.
References
[1] Karabicici, M., Akbari, S., Ertem, O., Gumustekin, M., & Erdal, E. (2023). Endocrine, Metabolic & Immune Disorders-Drug Targets, 23(14), 1713-1724.
[2] Ingber, D. E. (2022). Nature Reviews Genetics, 23(8), 467-491.
[3] Saglam-Metiner, P., Gulce-Iz, S., & Biray-Avci, C. (2019). Gene, 686, 203-212.
[4] Akbari, S., Sevinç, G. G., Ersoy, N., Basak, O., Kaplan, K., Sevinç, K., ... & Erdal, E. (2019). Stem Cell Reports, 13(4), 627-641.
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