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Cells were cultured in 96-well plates and then incubated with 200 μM OA alone or combined with various combinations of TC overnight. Cells were then stained, imaged (20X objective, 4 images/well), and analyzed in an automated fashion. A, B, and C show control cells visualized for nuclei (blue), lipid (green), and the lipid mask using the lipid droplet analysis algorithm (red). D, E, and F show cells treated with OA. G, H, and I show cells treated with OA plus 5.4 μM TC. Images in B and E were optimized equally for display. J shows the ratios of lipid mask to cell area. Z′ values were calculated for the effect of OC (Z′ = 0.86 between columns 1 and 2) and for the antagonistic effect of TC (Z′ = 0.56 between columns 2 and 12). Each bar is the mean ± SD for n = 8 wells.

(A) Lipid droplet accumulation in mouse 3T3-L1 cells (AID 652180) showed dose response inhibition of ML360 (green circle) but not with the negative control compound 1 (blue circle). (B) 3T3-L1 differentiation assay (AID 652183) showed dose response reduction of the lipid droplets (blue square) without any cytotoxic effect as the cell number per treatment remained the same (orange circle).

(A) Image obtained from a human liver biopsy stained with hematoxylin and eosin. The circular “holes” in the sample image correspond to lipid droplets (steatosis). (B) General progression of fatty liver diseases. NAFLD, alcoholic liver disease, and hepatitis C mediated by HCV/HBV are all thought to progress through similar etiologies, initiating with excessive lipid droplet formation within the hepatocytes (steatosis), which progresses to damage to the hepatocytes (steatohepatitis) and fibrosis, cirrhosis and development of hepatocellular carcinoma.