Franny Jongbloed

85 4 A PROTEIN-FREE DIET PROTECTS AGAINST RENAL IRI Compared to 3-days of fasting, a 3-day CHO-free diet is the least clustered with the other dietary groups. The 3-day protein-free diet clusters closely with the non-protective 3-day fat-free diet, based on the number of probe sets as well as the expression levels. Three days 30% DR had a similar cluster pattern as two weeks of 30% DR. Table 1. Comparison of the overlapping DEPS between the five dietary interventions and their corresponding P- value and enrichment factor Comparison Overlapping DEPS Overlap (%)* Enrichment factor P- value Protein-free vs. Fasting 222 53.3 9.8 5.27 E -169 2wks 30% DR vs. Fasting 247 45.9 8.7 8.88 E -171 3d 30% DR vs. Fasting 208 42.0 8.7 3.12 E -156 2wks 30% DR vs. 3d 30% DR 195 28.1 39.4 3.44 E -270 Protein-free vs. CHO-free 113 24.9 6.0 5.84 E -55 Fasting vs. CHO-free 584 23.7 5.9 3.73 E -300 2wks 30% DR vs. CHO-free 122 20.4 8.2 5.71 E -76 3d 30% DR vs. CHO-free 107 19.6 5.8 8.66 E -48 Protein-free vs. 3d 30% DR 116 18.5 29.5 2.22 E -138 Protein-free vs. 2wks 30% DR 95 15.2 22.3 1.15 E -99 The highest percentage of relative overlap was found between three days of protein-free diet and three days of fasting. The enrichment factor indicates the number of times the overlapping DEPS is higher than expected by chance. All diets showed a significantly overlapping number of DEPS, as shown by the corresponding p- values in the last column. *Relative overlap is calculated by the number of DEPS in common between the two groups, divided by the total number of unique DEPS across both groups, relative to the theoretical maximum overlap according to this formula. Subsequently, the same set of 2,604DEPS significantly regulated after fasting compared toAL fed controls, was used for a principal component analysis (PCA) among the macronutrient- free diets, two weeks- and three days 30% DR, and their control diets (Figure 2B). Samples from the control diets and the 3-day fat-free diet clustered together, with high overlap and corresponding directionality between the groups. With a large distance on the principal component (PC) 2 axis, a similar clustering was seen among the 3-day 30% DR and two weeks 30%DR samples. The protein-free diet had its own cluster, separated on both PC axes from the other groups. The 3-day CHO-free diet resembled the DR diets, but showed no overlap with these diets and showed a large dissimilarity to the other groups. To compare the transcriptomic responses between all dietary interventions, all DEPS were visualized in a Venn diagram (Figure 3A).This revealed a total of 40 overlapping DEPS in the three protective diets, while 30 overlapping DEPS were also present in the non-protective