Expression of Drug-metabolizing genes and Acetaminophen drug toxicity studies on 3D scaffold culture of Huh-7 cell line

• Published in: Research journal of pharmacy and technology Vol. 13; no. 5; pp. 2399 - 2406
• Main Authors: Suma, M S, Jamuna, K S, Ramesh, C.K., Mahmood, Riaz
• Format: Journal Article
• Language: English
• Published: Raipur A&V Publications 2020
• Subjects: Analgesics; Apoptosis; Cell culture; Collagen; Cytochrome; Cytotoxicity; Enzymes; Gene expression; Liver; Metabolism; Metabolites; Morphology
• ISSN: 0974-3618; 0974-360X; 0974-306X
• DOI: 10.5958/0974-360X.2020.00431.X

[...]for a wide range of biology applications, it is important to culture the cell for long periods of time[1]. Many reports have demonstrated that the micro-space cell culture plate method, a three-dimensional (3D) culture system, can induce hepatocyte-specific functions, including CYP2E1 expression, in HepG2 cells, which results in an increase in cytotoxicity by acetaminophen compared with a conventional two-dimensional (2D) culture system [16]. [...]it remains to be demonstrated whether cell injury induced by APAP in 3D-cultured HCC in vitro was produced by corresponding human and rodent mechanisms of in vivo APAP-induced hepatotoxicity and if antidotes such as N-acetylcysteine (NAC) or other agents also show protective potential against APAP-induced hepatotoxicity in the 3D-cultivation system [18-19]. Scaffolds were fixed in 4% paraformaldehyde for ten minutes at room temperature after culturing, at day 3 and day 7 followed by permeabilization with 0.5% TritonX-100 for 10min at 37°C. Actin filaments were stained with rhodamine-phalloidin (Molecular Probes) for 20min at 37 °C, and Nucleus was stained with DAPI (4,6-diamidino-2-phenylindole, dihydrochloride) (Molecular Probes) for 10min at 37°C. Images were acquired with a TCS SP5 II confocal microscope.