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The analyses of various techniques to characterise HepG2 monolayer and spheroid cultured for 17 days and 21 days, respectively
The aim of this study was to characterise HepG2 cells differentially cultured as monolayer or three-dimensional spheroids, in the presence and absence of chronic enzyme-inducing drug cocktail exposure. Also, to evaluate the models’ feasibility over an extended culture time and compare their metabolic capabilities as candidates for hepatotoxicity screening platforms. This was done by generating HepG2 spheroids using the liquid overlay method for up to 21 days and culturing same origin HepG2 monolayers for 17 days. Cells were evaluated for morphology, viability, protein content, monolayer cell cycle profile, proteins mass profile differences, and the presence of hepatic markers in spheroids. The metabolic activity was assessed using liquid chromatography tandem mass spectrometry.
Furthermore, the growth plateau observed in spheroid cultures’ protein levels after Day 4 and the hepatic markers (AFP, HNF-4α, CK18 and albumin) expression observed from Day 7, would be desirable for repeated hepatotoxicity testing. Other studies have shown HepG2 spheroid cell viability and increased hepatic marker expression for more than 21 days, with a relatively consistent metabolic profile after 21 days, suggesting a stable differentiated phenotype. This is beneficial for repeated, long-term drug exposure for acute and chronic hepatotoxicity screening.
National Research Foundation Innovation Master’s Scholarship, Grant Number:120972
Roche Products Bursary (2021)
Sustainable Development Goals
- 3 Good Health and Well-Being