Journal: Cell Death & Disease

Article Title: High-content imaging of human hepatic spheroids for researching the mechanism of duloxetine-induced hepatotoxicity

doi: 10.1038/s41419-022-05042-x

Figure Lengend Snippet: A Representative confocal images of the hepatic cellular alterations induced by DLX. Scale bar = 100 μm. B The dose dependent multiparametric curves of the hepatic cellular alterations induced by DLX. C Representative confocal fluorescent images of EHSs with DLX, DLX plus mito-TEMPO, and DLX plus NAC treatment stained by MitoSOX to detect the ROS generation in mitochondria or Live/Dead to detect the cell viability. Scale bar = 100 μm. D Cytochrome b (cytb), Cyclooxygenase 1 (COX1) and NADH dehydrogenase subunit 5 (ND5) gene expression in drug treated EHS cells. E Basal Oxygen consumption (OCR) metabolic profiling upon exposure to 0.2 mM DLX or DLX plus NAC for 200 min. F OCR metabolic profiling upon exposure to 0.06 mM DLX or DLX plus NAC in response to indicated inhibitors. G Flow cytometry quantification of the alteration of mitochondrial dysfunction in cells after drug treatments (left). The ratio of red and green AFI of JC-1 staining (right). H Representative transmission electron microscope (TEM) images of mitochondria in DLX or DLX plus NAC treated HepaRG spheroids. Scale bar = 200 nm. I Flow cytometry assays to detect apoptosis induction. ( J ) Cell cycle-based apoptosis assay for HepaRG cells in EHSs. The percentage of sub-G1 phase are shown in the images. DLX vs Ctrl: **, p < 0.01; ***, p < 0.001; two-tailed Student’s t tests.

Article Snippet: The spheroid culture system was developed by using human hepatoma HepaRG cells supported by a pulverized liver biomatrix scaffold.

Techniques: Staining, Gene Expression, Flow Cytometry, Transmission Assay, Microscopy, Apoptosis Assay, Two Tailed Test

Journal: Cell Death & Disease

Article Title: High-content imaging of human hepatic spheroids for researching the mechanism of duloxetine-induced hepatotoxicity

doi: 10.1038/s41419-022-05042-x

Figure Lengend Snippet: A Representative confocal fluorescent images with identified lipid spots of EHSs with DLX or S-071031B treatment stained by Nile Red (orange). The AFI of lipid spots were shown in the images (up) and their area was measured and normalized to control group (down). Scale bar = 20 μm. B Fluorescence images and fatty acid accumulation analysis of EHSs with DLX or S-071031B treatment stained by QBT kit to monitor the fatty acid uptake for 180 min. Scale bar = 200 μm. C Related gene expressions of pyruvate dehydrogenase kinase 4 (PDK4), acyl-CoA synthetase long-chain family member 4 (ACSL4), 1-acylglycerol-3-phosphate O-acyltransferase 1 (AGPAT1), solute carrier family 2 member 4 (SLC2A4), ATP citrate lyase (ACLY), fatty acid desaturase 2 (FADS2) and elongation of very long chain fatty acids 1 (ELOVL1) in DLX or S-071031B treated HepaRG cells estimated by RT-qPCR. D and E The enzyme activity of MRC complex I ( D ) and the ratio of NADH/NAD + ( E ) in EHS cells treated with DLX or S-071031B. F Related gene expressions of apolipoprotein B (ApoB) in DLX or S-071031B treated EHSs. G The schematic summary of mechanism about DLX induced steatosis. H Related gene expressions of CYP2E1 in DLX or S-071031B treated EHSs. I MDA content in DLX or S-071031B treated EHSs. J Representative confocal fluorescent images of EHSs with DLX or S-071031B treatment to detect the drug caused lipid peroxidation. Scale bar = 100 μm. *, p < 0.05; ***, p < 0.001; two-tailed Student’s t tests.

Article Snippet: The spheroid culture system was developed by using human hepatoma HepaRG cells supported by a pulverized liver biomatrix scaffold.

Techniques: Staining, Control, Fluorescence, Quantitative RT-PCR, Activity Assay, Two Tailed Test