Research Paper Volume 13, Issue 21 pp 24017—24036

Low-protein diet applied as part of combination therapy or stand-alone normalizes lifespan and tumor proliferation in a model of intestinal cancer


Figure 5. Effects of DR and afatinib on established phenotypes. (A) Control animals (esg > w1118) and animals with Egfr-induced over-proliferation (esg > EgfrCA) in intestinal stem cells and enteroblasts (esg+ cells) were given interventions (100 μM afatinib, DR, or a combination of afatinib and DR) either immediately for 15 days (early application, upper row) or 5 days after the onset of over-proliferation for 10 consecutive days (late application, lower row). (BD) Anterior midguts of animals exposed immediately (early application) to DR, afatinib, or the combination of DR and afatinib. (B’D’) Anterior midguts of animals exposed to DR, afatinib, or a combination of DR and afatinib for 10 days after the onset of over-proliferation to simulate a real-life situation. (E) Quantification of the area covered by GFP-positive cells indicating the number and phenotype of esg+ cells in the midgut. n = 10–12. (F) Luciferase and GFP were expressed simultaneously and quantified in whole animals to measure the over-proliferation phenotype. n = 5–6. (GI) The lifespan of animals exposed to an early application or late application. n = 32–69. Statistical significance was tested by one-way ANOVA and the Tukey test. The lifespan significance was tested by the log-rank (Mantel-Cox) test. Significances are marked with lines or corresponding color. ns = not significant, *** = p < 0.001, **** = p < 0.0001. Scale bar: 100 μm.