Fig. S6

Ethanol-induced HSC activation is partially rescued by CMZ assessed by laminin deposition. (A-D) show confocal single plane images of Tg(hand2:EGFP) expression in the HSCs of larvae treated under different conditions from 96 hpf to 120 hpf. The larvae were fixed immediately after the treatment at 120 hpf. Arrows point to individual HSCs. In control and CMZ-treated larvae, HSCs send out long and multiple cellular processes. Moreover, these HSCs are well segregated from one another. In ethanol (EtOH)-treated larvae, HSCs lose the complex processes, become elongated, and tend to cluster together. In larvae treated with both CMZ and ethanol, the HSC phenotypes seem to be partially suppressed as these cells form short and multiple cellular processes (arrows in D). (A′-D′) show the same images of (A-D), but with laminin staining. Ethanol treatment triggers laminin deposition (arrows in C′), whereas co-treatment with ethanol and CMZ suppresses laminin deposition. The blue staining in A′ and B′ is due to autofluorescence of the red blood cells. (A"-D") show three-dimensional projection of the confocal Z-stack that scanning through the whole livers in (A-D). In ethanoltreated liver, HSCs are clustered together, whereas in control, CMZ, and CMZ and ethanol cotreated liver, HSCs were well separated from one another. In (A-D") white dashed lines outline the periphery of the liver. Scale bar: 20 μm shown in (A). (E) Quantification of whole mount oil red O staining in 120 hpf larvae that were either untreated (N/T), or pre-treated with 3 mM CYA, 40 mM OAc at 94 hpf and then exposed to 0 or 350 mM ethanol at 96 hpf for 24 hours. Note that neither of these treatments affected the ethanol-induced steatosis.