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, 8 (1), 74

SCFAs Strongly Stimulate PYY Production in Human Enteroendocrine Cells

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SCFAs Strongly Stimulate PYY Production in Human Enteroendocrine Cells

P Larraufie et al. Sci Rep.

Abstract

Peptide-YY (PYY) and Glucagon-Like Peptide-1 (GLP-1) play important roles in the regulation of food intake and insulin secretion, and are of translational interest in the field of obesity and diabetes. PYY production is highest in enteroendocrine cells located in the distal intestine, mirroring the sites where high concentrations of short chain fatty acids (SCFAs) are produced by gut microbiota. We show here that propionate and butyrate strongly increased expression of PYY but not GCG in human cell line and intestinal primary culture models. The effect was predominantly attributable to the histone deacetylase inhibitory activity of SCFA and minor, but significant contributions of FFA2 (GPR43). Consistent with the SCFA-dependent elevation of PYY gene expression, we also observed increased basal and stimulated PYY hormone secretion. Interestingly, the transcriptional stimulation of PYY was specific to human-derived cell models and not reproduced in murine primary cultures. This is likely due to substantial differences in PYY gene structure between mouse and human. In summary, this study revealed a strong regulation of PYY production by SCFA that was evident in humans but not mice, and suggests that high fibre diets elevate plasma concentrations of the anorexigenic hormone PYY, both by targeting gene expression and hormone secretion.

Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Figure 1
Figure 1
SCFAs increase PYY expression in human EEC model cell lines. (a) Expression of EEC markers, SCFA transporters and receptors and specific enteroendocrine differentiation factors in NCI-h716 (dark grey) and HuTu-80 (light grey) were detected by RT-qPCR. Data are expressed relative to β-actin determined by the 2−ΔCt method, on at least 4 distinct experiments performed in duplicate, represented as means ± s.e.m. N.D.: Not Detected. (b,c) Relative expression to control of PYY (dark grey) and GCG (light grey) after treatment with SCFAs (2 mM, 24 h) on NCI-h716 cells (b) and HuTu-80 cells (c). (d,e) Effect of concentration (d) or time of incubation (e) of butyrate on PYY expression in NCI-h716 cells. PYY expression relative to control expression is determined by the 2−ΔΔCt method using β-actin as control gene. Data are means ± s.e.m. of at least three distinct experiments. (***P < 0.001; **P < 0.01; *P < 0.05, Dunn to control test).
Figure 2
Figure 2
Mechanisms involved in response to SCFAs. (a) Expression relative to control of PYY-producing EEC characteristic transcription factors after incubation with 2 mM butyrate for 24 h in NCI-h716 cells. (b) Effect of 24 h incubation with SCFAs, receptor agonists, tiglic acid (FFA2, 100 μM), 1-MCPC (FFA3, 100 μM) and niacin (GPR109a, 1 mM), PMA (200 nM), TSA (500 nM), Valproic acid (5 mM) or SAHA (5 μM) on PYY expression in NCI-h716 cells. (c) Effect of 24 h incubation with SCFAs and receptor agonists on PYY expression in FFA2 depleted NCI-h716 cells. (d) Expression of PYY relative to untreated cells in response to acetate (2 mM), tiglic acid (100 μM) or 1MCPC (100 μM) in Hutu-80 cells transfected with pCMV-FFAR2 or pCMV-eGFP. (e) Secretion relative to control of PYY (dark grey) or GLP-1 (light grey) by NCI-h716 pre-incubated with butyrate (2 mM) or TSA (500 nM) for 24 h, and subsequently incubated in non-stimulated conditions for 2 h; measured by ELISA. Mean basal secretion was 15.6 ± 1.6 pg/mL for PYY and 90.6 ± 4.3 pg/mL or GLP-1. Data are means ± s.e.m. of four distinct experiments. (***P < 0.001; **P < 0.01; *P < 0.05, Dunn to control test).
Figure 3
Figure 3
Effect of SCFAs is species dependent. (a,b) Expression relative to untreated control cultures of PYY (dark grey) and GCG (light grey) in human colonic (a) and duodenal (b) cultures after incubation with SCFAs (2 mM) or TSA (500 nM). (c,d) Secretion, relative to control, of PYY (c) or GLP-1 (d) by colonic primary cultures pre-incubated with butyrate (2 mM) or TSA (500 nM) for 24 h, and subsequently incubated for 2 h in the absence of stimuli (dark grey) or in the presence of forskolin (10 μM), IBMX (10 μM) and glucose (10 mM) (light grey); measured by ELISA. Secretion is normalized to the protein content in the lysed cells, measured by BCA assay. Mean basal secretion was 0.42 pg/mL/μg of tissue and 0.019 pg/mL/μg of tissue for PYY and GLP-1, respectively. Analysis was performed comparing cells incubated with either butyrate or TSA to control-incubated cells, in both secretion conditions separately (***P < 0.001; **P < 0.01; *P < 0.05, Dunn to control test). (e) Expression of Pyy and Gcg in primary mouse colonic cultures after incubation with SCFAs (2 mM), relative to control-treated cultures. Data are means ± s.e.m. of four distinct experiments. (f) Effect of TSA (500 nM, 24 h) on the expression of genes known to be regulated by HDAC inhibition and on Pyy. (***P < 0.001; **P < 0.01; *P < 0.05, Dunn to control test). (g) Schematic representation of human and mouse PYY gene structure, indicating the exons and introns and highlighting the presence of GC boxes in the first 100 bases of the promoter.

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