Figure 1.
Effect of Livogrit on development of NASH in hepatocyte-derived spheroids. (a) Representative brightfield image of 7-day old HepG2 spheroid with initial cell density of 20,000 cells/ 20 µL. Spheroids with a diameter of <600 µm on 7th day were used for all experiments. (b) Cytosafety analysis (72 h) of Livogrit (0–100 µg/mL) and Pioglitazone (10 µM) treatment on spheroid, as determined by colorimetric LDH quantification (Abs. 450 nm). (c) Intraspheroidal lipid accumulation (72 h) post incubation in methionine and cystine deficient media (MCDM) with treatment of Livogrit (0–30 µg/mL) and Pioglitazone (10 µM), as determined by Nile red stain (Ex 530/Em 600 nm) fluorescence measurement. (d) Reactive oxygen species (ROS) generation (72 h) in spheroid post incubation in MCDM with treatment of Livogrit (0–30 µg/mL) and Pioglitazone (10 µM) as determined by CellROX green (Ex 475/Em 520 nm) fluorescence measurement. (e) Fluorescent microscopy images of Hoechst 33342 (DAPI), CellROX green (FITC) and LipidSpot 610 (Cy5) stained HepG2 spheroids post 72 h incubation in MCDM with treatment of Livogrit (30 µg/mL) and Pioglitazone (10 µM). Scale bar = 100 µm.