Body weight and blood glucose levels of male zebrafish. (a) Zebrafish were fed 7 times per day (H) to induce obesity or twice daily (L) to induce weight loss, and the feeding frequency was switched every two weeks to induce weight cycling. ^ (H vs. L), * (H vs. LH), ^& (L vs. LH), ^$ (L vs. HL), and p ≤ 0.05. Blood glucose levels at weeks 8 (b) and 20 (c) were analyzed. ^ab In the graphs, lines, and bars sharing different letters are significantly different (p ≤ 0.05). Data are presented as the mean ± SEM (n = 5–10).

Liver histology. The liver sections were individually stained with H&E, Sirius red, and Oil Red O (a). The positive area of oil red-O staining at week 8 and week 20 was quantified (b). The scale bar for H&E and Sirius red is 100 µm, and that for Oil Red O is 500 µm. Black arrows indicate the ballooning hepatocytes; blue arrows indicate the cholestasis site; yellow arrows indicate hyperemia; and white arrows indicate fibrosis. ^ab In the graphs, lines, and bars sharing different letters are significantly different (p ≤ 0.05). Data are presented as the mean ± SEM (n = 3–6).

Gene expression in the liver. Data were analyzed by one-way ANOVA followed by Tukey’s test for multiple comparisons and are presented as the mean ± SEM (n = 3–7). ^abc In the graphs, lines and bars sharing different letters are significantly different (p ≤ 0.05). cpt1a1a: carnitine palmitoyltransferase 1Aa; hadhaa: hydroxy acyl-CoA dehydrogenase trifunctional multienzyme complex subunit alpha a; fasn: fatty acid synthase; gnpat: glycerophosphate O-acyltransferase; gck: glucokinase; g6pc1a: glucose-6-phosphatase catalytic subunit 1a.

Swimming performance and muscle histology. The speeds of burst swimming and ramp-U_crit swimming at week 8 (a) and week 20 (b) were determined. The speed of the burst swimming test was ramped up every minute, and the speed of ramp-Ucrit swimming was ramped up every ten minutes until the fish became fatigued. The cross-section of the tail muscle was stained with H&E (c), and the fiber size (d) at week 8 and week 20 was quantified. Scale bar = 500 µm. ^ab In the graphs, lines, and bars sharing different letters are significantly different (p ≤ 0.05). Data are presented as the mean ± SEM (n = 3–7).

Gene expression in the muscle. Data were analyzed via one-way ANOVA followed by Tukey’s test for multiple comparisons and are presented as the mean ± SEM (n = 3–7). ^ab In the graphs, lines, and bars sharing different letters are significantly different (p ≤ 0.05). PCNA: Proliferating Cell Nuclear Antigen; myoD: Myoblast determination protein 1; myf5: Myogenic factor 5; HK1: hexokinase 1; pkmb: pyruvate kinase M1/2b; trim63b: E3 ubiquitin-protein ligase b; fbxo32: Atrogin1; map1c3b: tubule-associated protein 1 light chain 3 beta; atg4b: cysteine protease.

The histology and characteristics of adipose tissues. The sections of subcutaneous and abdominal adipose tissues were stained with H&E (a). Scale bar = 500 µm. The characteristics of adipose tissue, including fat percentages, average size, and distribution of abdominal adipocytes, were analyzed at week 8 (b) and week 20 (c). Data are presented as the mean ± SEM (n = 3–6). ^ab In the graphs, lines, and bars sharing different letters are significantly different (p ≤ 0.05).