Journal: Current Biology
Article Title: Short-Term, Intermittent Fasting Induces Long-Lasting Gut Health and TOR-Independent Lifespan Extension
doi: 10.1016/j.cub.2018.04.015
Figure Lengend Snippet: Early-Life Fasting Leads to Long-Lasting Improvements in Gut Health and Reduced Bacterial Load (A) ISC activity. Representative confocal fluorescence z projections of midgut regions 4/5 from 70-day-old ad libitum , 5-day IF, and IF switch flies stained with DAPI (magenta) and anti-phospho-H3 antibody (pH3, green) are shown. The scale bar represents 100 μm. (B) Quantification of pH3-positive (pH3+) cells per gut. After 5 days of fasting, 70-day-old 5-day IF guts had significantly fewer pH3+ cells compared to ad libitum controls (p < 0.0001; one-way ANOVA). IF switch guts, after 40 days on food, also had significantly fewer pH3+ cells compared to ad libitum controls (p < 0.0001; one-way ANOVA). 19 or 20 guts per condition are shown. Data are shown as mean ± SEM. (C) Quantification of gut length. After 5 days of fasting, 70-day-old 5-day IF guts were significantly shorter than ad libitum controls (p = 0.038; one-way ANOVA). IF switch guts were not significantly different from ad libitum controls. 11–13 guts per condition are shown. Data are shown as mean ± SEM. (D) Quantification of gut width at R4 section. After 5 days of fasting, 70-day-old 5-day IF guts were significantly narrower than ad libitum controls (p = 0.025; one-way ANOVA). IF switch guts were not significantly different compared to ad libitum controls. 16–18 guts per condition are shown. Data are shown as mean ± SEM. (E) Midgut pathology. Representative confocal fluorescence sections of R2 (top row) and R4 (bottom row) midgut regions from 70-day-old ad libitum , 5-day IF, and IF switch flies stained with DAPI (white) to visualize nuclei and anti-βPS-integrin antibody (βPS, green) to visualize the basement membrane/muscle are shown. (F and G) Pathologies were binned into scaled categories and quantified. R2 (F, n = 10–12 per condition) and R4 (G, n = 7–10 per condition) categories were defined as follows: I, wild-type (WT), nuclei form single layer epithelium; II, sporadic pathology of small nuclei “nests”; III, intermediate pathology, sections of epithelium have several layers of nuclei; and IV, widespread pathology, majority of epithelium has several layers of nuclei. (H) Barrier function at day 60, as measured by “smurf” flies with leaky intestines, was significantly improved in IF switch females compared to ad libitum controls (p = 0.0013; Fisher’s exact; n numbers on panel). (I) Quantification of relative bacterial abundance in whole females by qPCR of the bacterial 16S rRNA gene with pan-bacterial primers and Lactobacillus - plantarum -specific primers. IF switch flies at day 40 (i.e., day 10 post-IF) had significantly reduced bacterial abundance compared to ad libitum controls (pan bacterial, p = 0.00012; L. plantarum , p = 0.0004; Student’s t test). Values were normalized to the Drosophila GAPDH gene. Data are shown as mean ± SEM (n = 8 biological replicates per condition).
Article Snippet: Mouse monoclonal anti-integrin βPS (myospheroid) antibody , Developmental Studies Hybridoma Bank , Cat# # CF. 6G11 RRID: AB_528310.
Techniques: Activity Assay, Fluorescence, Staining, Membrane
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