43 2 Organoids as a model to study intestinal and liver dysfunction in severe malnutrition Figure 4. Impact of amino acid starvation and re-supplementation on mitochondria and peroxisomes in mature hepatic organoids (A) Representative immunoblot images. (B) Peroxisomal protein levels relative to β-actin. Quantification of data shown in (A) Organoids were grown in complete culture medium throughout (control, grey), amino-acid-free medium for 48 hours (starved, blue), ), starved from 0h to 48h and re-supplemented with complete medium from 48h to 96h (re-supplementation, orange) or in amino-acid-free medium for 48 h supplemented with 100uM fenofibrate (fenofibrate, green). Data represent mean ± SEM from 7-10 biological replicates and 3 biological replicates for Fenofibrate treatment (biological replicates are obtained from independent experiments) (*P<0.05, ** P< 0.01, two-way ANOVA with Tukey’s posthoc test). (C) Representative scheme of the peroxisomal pathway of phytol into phytanoyl-CoA and Pristanoyl-CoA. (D) Percentage of phytanic acid and pristanic acid in the supernatant of organoids grown in complete culture medium (control, grey), amino-acid-free medium (48h starvation, blue), amino-acid-free medium for 48 h followed by complete culture medium for 48 h (re-supplementation, orange) or in amino-acid-free medium for 48 h supplemented with 10 μM fenofibrate (fenofibrate, green). Data represents mean of 4 biological replicates (from independent experiments) ± SEM (*P<0.05, ** P< 0.01. One-way ANOVA with Tukey’s post-hoc test) (E) Ratios of very-long chain fatty acids C24/C22 and C26/C22 measured in the supernatant of organoids grown in complete culture medium (control), amino-acid-free medium (starved), amino-acid-free medium for 48 h followed by complete culture medium for 48 h (re-supplementation) or in amino-acid-free medium for 48 h supplemented with 10 μM fenofibrate Data represents mean of 4 biological replicates (from independent experiments)
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