Fig. 4. IFNγ does not affect early phases of hepatocyte infection but suppresses LS development.
A Experimental layout. HepG2-CD81 hepatoma cells were treated with 10 U/ml human recombinant IFNγ at 16 h, or 3 h pre-infection or during Py or PyUIS4-GFP sporozoite addition to the cells for up to 3 h post infection (hpi). The suppressive effect of IFNγ was determined at 6 hpi (early LS), which encompasses sporozoite invasion of hepatocytes and transformation of intracellular sporozoites into LS trophozoites and 48 hpi, which measures near full LS development. B, C The number of infected cells between control and IFNγ treated groups at 6 hpi (early LS) determined by UIS4 staining. Scale bar, 4 μm. n = 3 independent experiments per group. D, E Quantification of UIS4-GFP positive infected cells between control and IFNγ treated groups at 48 hpi (developed LS) determined by live imaging after staining with Hoechst 33342. Scale bar, 100 μm. n = 3 independent experiments per group. ****P < 0.0001. F, G Quantification of LS size at 48 hpi by measuring area of UIS4-GFP after staining with Phalloidin rhodamine and Hoechst 33342. Scale bar, 10 μm. n = 100 Py LS count per group. ****P < 0.0001. B, E, G One-way ANOVA with Tukey’s multiple comparison test. Results are combined and represented as means ± SD from three independent experiments (B–E) or represented from one of the three independent experiments with similar results obtained (F, G). Source data are provided as a Source data file.