Propionate promotes intestinal lipolysis and metabolic benefits via AMPK/LSD1 pathway in mice

in Journal of Endocrinology
Correspondence should be addressed to L Zhang or Y-L Yin: linzhang@scau.edu.cn or yinyulong@isa.ac.cn
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Dietary fibers and their microbial fermentation products short-chain fatty acids promote metabolic benefits, but the underlying mechanisms are still unclear. Recent studies indicate that intestinal lipid handling is under regulatory control and has broad influence on whole body energy homeostasis. Here we reported that dietary inulin and propionate significantly decreased whole body fat mass without affecting food intake in mice fed with chow diet. Meanwhile, triglyceride (TG) content was decreased and lipolysis gene expression, such as adipose triglyceride lipase (A tgl), hormone-sensitive lipase (H sl) and lysosomal acid lipase (L al) was elevated in the jejunum and ileum of inulin- and propionate-treated mice. In vitro studies on Caco-2 cells showed propionate directly induced enterocyte Atgl, Hsl and Lal gene expression and decreased TG content, via activation of phosphorylation of AMP-activated protein kinase (p-AMPK) and lysine-specific demethylase 1 (LSD1). Moreover, inulin and propionate could increase intestinal lipolysis under high-fat diet (HFD)-fed condition which contributed to the prevention of HFD-induced obesity. Our study suggests that dietary fiber inulin and its microbial fermentation product propionate can regulate metabolic homeostasis through regulating intestinal lipid handling, which may provide a novel therapeutic target for both prevention and treatment of obesity.

Downloadable materials

  • Supplementary Table: Primers used for quantitative real time-PCR
  • Supplementary Figure 1. Effect of SCFAs on metabolic phenotype in chow diet fed mice. (A-B) Body weight gain (A) and fat mass (B) of mice treated with acetate or butyrate in chow diet for 2 weeks. (C-D) iWAT (C) and eWAT (D) fat-pad weights. (E) Hematoxylin-eosin staining of jejunum. (F) Villus length of jejunum. (G) Oil-red O staining of liver. (H) Triglyceride content in the liver. (I) Hematoxylin-eosin staining of eWAT. (J) Triglyceride content in the eWAT. Ctrl, control; NaA, acetate; NaB, butyrate; iWAT, inguinal white adipose tissue; eWAT, epididymal white adipose tissue. n=8 per group. Data are mean ±SEM and statistical analyses were performed using two-tailed Student’s t test; *P <0.05 versus control.
  • Supplementary Figure 2. Effects of propionate and inulin on lipid metabolism gene in the liver, WAT and small intestine. (A-B) The mRNA expression of ATGL, HSL, Cpt1α, CD36 and FATP4 in the liver (A) and eWAT (B). (C-D) The mRNA expression of CD36 and FATP4 in the jejunum (C) and ileum (D). n=6 per group. Data are mean ±SEM and statistical analyses were performed using two-tailed Student’s t test; *P <0.05 versus control.
  • Supplementary Figure 3. Effects of SCFAs on lipid metabolism and cell viability in Caco-2 cells. (A) Oil Red O staining of Caco-2 cells after incubated with 0.5 mmol/L SCFAs for 24 h, n=6 per group. (B) Cell viability was assessed in Caco-2 cells after incubated with (0, 10, 50, 100, 500 and 1000 μmol/L) propionate at 12, 24 and 48 h by MTT assay, n=6 per group. (C-D) The mRNA expression of CD36 (C) and DGAT1 (D) in Caco-2 cells incubation with 0.5 mmol/L SCFAs, n=6 per group. Data are mean ±SEM and statistical analyses were performed using one-way ANOVA. *P <0.05 indicates a significant difference. Scale bar: 50 μm.
  • Supplementary Figure 4. Effect of propionate and inulin on FFARs in the jejunum. (A-B) The mRNA expression of FFAR3 (A) and FFAR2 (B) in the jejunum of chow fed-mice treated with propionate or inulin for 2 weeks. n=6 per group, Data are mean ±SEM and statistical analyses were performed using two-tailed Student’s t test.
  • Supplementary Figure 5. Effects of compound C or GSK 2879552 on p-AMPK and LSD1 expression in Caco-2 cells. (A) Intracellular triglyceride content in Caco-2 cells when stimulated with 0, 1, 5,10 μmol/L compound C (Cc) for 24 h, n=6 per group. (B) Protein expression of p-AMPK and AMPK in Caco-2 cell stimulated with or without Cc in the presence of propionate for 24 h, n=4 per group. (C) Protein expression of LSD1 in Caco-2 cell treated with or without GSK2879552 (GSK552) in the presence of propionate for 24 h, n=4 per group. Data are mean ±SEM and statistical analyses were performed using one-way ANOVA. *P <0.05 indicates a significant difference.
  • Supplementary Figure 6. Effect of propionate or inulin on metabolic phenotype in HFD fed mice. (A-B) iWAT (A) and eWAT (B) fat-pad weights of HFD-fed mice treated with propionate or inulin for 2 weeks. (C-D) The unit length weight of jejunum (C) and ileum (D). (E) Triglyceride content in the liver. (F-G) The mRNA expression of MGAT2 and DGAT1 in the jejunum (F) and ileum (G). Data are mean ±SEM and statistical analyses were performed using two-tailed Student’s t test, n=6 per group; *P <0.05 versus control.
  • Supplementary Figure 7. Effects of propionate and inulin on amino acid absorption and inflammatory cytokines in the jejunum. (A) The mRNA expression of amino acid transporters (y+LAT1, B0,+AT and B0AT1) in the jejunum of mice treated with propionate or inulin in chow diet, n=6. (B) Plasma arginine of mice treated with propionate or inulin after 10% whey protein administration for 30 min, n=4. (C) The mRNA expression of inflammatory cytokines (TNFα, IL-6 and NFκB) in the jejunum of mice fed with propionate or inulin, n=6. Data are mean ±SEM and statistical analyses were performed using two-tailed Student’s t test; *P <0.05 versus control.

 

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    Effect of propionate and inulin on metabolic phenotype in mice fed a chow diet. (A, B and C) Body weight gain (A), fat mass (B) and food intake (C) of mice treated with propionate or inulin for 2 weeks. (D) Fecal triglyceride content. (E and F) The unit length weight of jejunum (E) and ileum (F). (G and H) Triglyceride content in the jejunum (G) and ileum (H). (I) Triglyceride content in serum. Data are mean ± s.e.m. and statistical analyses were performed using two-tailed Student’s t test, n = 8 per group. Ctrl, control; NaP, sodium propionate. *P < 0.05 versus control.

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    Effect of propionate and inulin on small intestinal lipid metabolism gene in mice. (A and B) Relative mRNA expression of A tgl, Hsl and Lal in the jejunum (A) and ileum (B). (C and D) Relative mRNA expression of Mgat 2 and Dgat1 in the jejunum (C) and ileum (D). (E and F) Relative mRNA expression of Cpt1α in the jejunum (E) and ileum (F). Data are mean ± s.e.m. and statistical analyses were performed using two-tailed Student’s t test, n = 8 per group. *P < 0.05 versus control.

  • View in gallery

    Effect of SCFAs on lipid metabolism in Caco-2 cells. (A) Triglyceride content was assessed in Caco-2 cells treated with 0.1 and 0.5 mmol/L SCFAs for 24 h by Oil Red O staining. (B) Triglyceride content of Caco-2 cells treated with 0.1 and 0.5 mmol/L SCFAs for 24 h. (C, D and E) Relative mRNA expression of A tgl (C), Hsl (D) and Lal (E) in Caco-2 cells by 24 h incubation with 0.1 and 0.5 mmol/L SCFAs. Data are means ± s.e.m. and statistical analyses were performed using one-way ANOVA, n = 6 per group. Ctrl, control; NaA, sodium acetate; NaB, sodium butyrate; NaP, sodium propionate. *P < 0.05 indicates a significant difference.

  • View in gallery

    Propionate regulates lipolysis dependent on p-AMPK in Caco-2 cells. (A and B) Protein expression of p-AMPK and AMPK in the jejunum (A) and ileum (B) of mice treated with propionate or inulin, n = 6 mice per group. (C) Protein expression of p-AMPK and AMPK in Caco-2 cells treated with 0.1 and 0.5 mmol/L propionate for 24 h, n = 4 per group. (D) Triglyceride content of Caco-2 cells when stimulated with compound C (Cc, 10μmol/L) in the presence of propionate for 24 h, n = 6 per group. (E, F and G) Relative mRNA expression of Atgl (E), Hsl (F) and Lal (G) in Caco-2 cells when stimulated with compound C (Cc, 10μmol/L) in the presence of propionate for 24 h, n = 6 per group. Data are means ± s.e.m. and and statistical analyses were performed using one-way ANOVA. *P < 0.05 indicates a significant difference.

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    LSD1 regulates propionate-induced lipolysis in Caco-2 cells. (A and B) Protein expression of LSD1 in the jejunum (A) and ileum (B) of mice treated with propionate or inulin, n = 6 mice per group. (C) Protein expression of LSD1 in Caco-2 cells treated with 0.1 and 0.5 mmol/L propionate for 24 h, n = 4 per group. (D) Triglyceride content of Caco-2 cells stimulated with or without GSK2879552 (GSK552, 5 μmol/L) in the presence of propionate for 24 h, n = 6 per group. (E, F and G) Relative mRNA expression of Atgl (E), Hsl (F) and Lal (G) of Caco-2 cells treated with or without GSK552 in the presence of propionate for 24 h, n = 9 per group. (H) Protein expression of LSD1 in Caco-2 cells treated with or without Cc in the presence of propionate for 24 h, n = 4 per group. (I) Protein expression of p-AMPK in Caco-2 cells stimulated with or without GSK552 in the presence of propionate for 24 h, n = 4 per group. Data are means ± s.e.m. and statistical analyses were performed using one-way ANOVA. *P < 0.05 indicates a significant difference.

  • View in gallery

    Effect of propionate and inulin on metabolic phenotype in HFD-fed mice. (A, B and C) Body weight gain (A), fat mass (B) and food intake (C) of HFD-fed mice treated with propionate or inulin for 2 weeks. (D) Fecal triglyceride content. (E and F) Triglyceride content in the jejunum (E) and ileum (F). (G) Triglyceride content in serum. (H and I) Relative mRNA expression of ATGL, HSL and LAL in the jejunum (H) and ileum (I). (J and K) Relative mRNA expression of Cpt1α in the jejunum (J) and ileum (K). (L and M) Protein expression of p-AMPK and AMPK in the jejunum (L) and ileum (M). (N and O) Protein expression of LSD1 in the jejunum (N) and ileum (O). n = 6 per group. Data are means ± s.e.m. and statistical analyses were performed using two-tailed Student’s t test. *P < 0.05 versus control.

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