Figure 8.

Melatonin facilitates SIRT1 expression by activating the AMPK pathway. (A) Immunoblot analysis of p-AMPK, AMPK, and SIRT1 in ileum sections of breastfed (BF) and NEC pups upon melatonin treatment (MEL) or treatment with vehicle (VEH); β-actin was used as an internal control. (B) Quantification of p-AMPK, AMPK, and SIRT1 in (A), n = 6 per group. (C) Kaplan-Meier estimates and log-rank tests were used to analyze the survival rate of pups following NEC induction upon treatment with vehicle (VEH) alone or with melatonin (MEL), or melatonin combined with Compound C (MEL + CC). ***P < 0.001. (D) Weight change of mouse pups in VEH, MEL, and MEL + CC groups. (E) Necrotizing enterocolitis (NEC) severity scores of the histopathological evaluation of mouse ilea (n = 15 for VEH, 20 for MEL, and 16 for MEL + CC groups). (F) Incidence of NEC (damage scores > 2) in VEH, MEL, and MEL + CC groups. (G) Fluorescence readings in plasma 4 h after gavage with FITC-dextran from VEH (n = 8), MEL (n = 17), and MEL +CC (n = 12) groups. (H) Immunoblot analysis of the expression of ZO-1, p-AMPK, AMPK, and SIRT1 in ileum sections of VEH, MEL, and MEL + CC pups. (I-L) Quantification of (H) (n = 6). Each symbol (B, D, E, G, I-L) represents an individual mouse and (f) shows the incidence of each independent experiment (n = 24); column graphs represent the mean with error bars indicating standard deviation (SD), **P < 0.05; ***P < 0.001; ns: not significant. P values were derived through one-way ANOVA followed by the Bonferroni multiple comparison test (B-G, I-L). Data are representative of three independent experiments (D-G, I-L).