Dairy protein and leucine alter GLP-1 release and mRNA of genes involved in intestinal lipid metabolism in vitro

Abstract

Objective: A growing body of evidence supports an antiobesity effect of dairy products; however, the mechanisms remain unclear. The objective of this study was to explore possible intestinal mechanisms by which dairy delivers an antiobesity effect. The human intestinal cell line, NCI-H716, was used to test the hypothesis that branched-chain amino acids and dairy proteins regulate satiety hormone secretion and modulate genes involved in fatty acid and cholesterol metabolism.

Methods: In dose-response (0.5%, 1.0%, 2.0%, and 3.0%) studies, the effect of leucine, isoleucine, valine, skim milk, casein, and whey on glucagon-like peptide-1 release and the expression of selected genes were tested.

Results: Leucine, isoleucine, skim milk, and casein stimulated glucagon-like peptide-1 release (P < 0.05). Isoleucine and whey downregulated the expression of intestinal-type fatty acid binding protein (i-FABP), fatty acid transport protein 4 (FATP4), Niemann-Pick C-1-like-1 protein (NPC1L1), acetyl-coenzyme A carboxylase (ACC), fatty acid synthase (FAS), sterol regulatory element-binding protein-2 (SREBP-2), and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR; P < 0.05). Leucine and valine downregulated the expression of NPC1L1, ACC, FAS, SREBP-2, and HMGCR (P < 0.05). Casein downregulated the expression of i-FABP, FATP4, ACC, FAS, SREBP-2, and HMGCR (P < 0.05). Skim milk downregulated the expression of ACC, FAS, and SREBP-2, but not i-FABP, FATP4, and NPC1L1.

Conclusion: This work suggests that the antiobesity effect of dairy may be mediated, at least in part, by integration of events that promote glucagon-like peptide-1 secretion and inhibit expression of genes involved in intestinal fatty acid and cholesterol absorption and synthesis.

Fig. 1

Effects of branched-chain amino acids, skim milk, casein, and whey on GLP-1 release in NCI-H716 cells. Cells (1 × 10⁶) were incubated for 2 h with increasing concentrations of (A) leucine, isoleucine, valine and (B) skim milk, casein, and whey. Secretion into the medium was normalized to the total cellular content and expressed as a percentage of the control value. Due to lower solubility, leucine concentrations were 1.5% and 2.0% instead of 2.0% and 3.0%. Values are means ± SEMs of four independent experiments. Treatments with different letters are significantly different (P < 0.05). GLP-1, glucagon-like peptide-1.

Fig. 2

Effects of leucine, isoleucine, and valine on in vitro mRNA expression of (A) i-FABP, (B) FATP4, and (C) NPC1L1. NCI-H716 cells (2 × 10⁶) were incubated for 42 h with Dulbecco’s Minimal Eagle’s Medium plus 0.2% bovine serum albumin with or without one of the treatments (w/v). Polymerase chain reaction products for genes of interest were normalized to β-actin mRNA as a control. Due to lower solubility, leucine concentrations are 1.5% and 2.0% instead of 2.0% and 3.0%. Values are means ± SEMs of four experiments. Treatments with different letters are significantly different (P < 0.05). FATP4, fatty acid transport protein-4; i-FAB, intestinal-type fatty acid binding protein; NPC1L1, Niemann-Pick C-1–like-1 protein.

Fig. 3

Effect of leucine, isoleucine, and valine on in vitro mRNA expression of (A) ACC, (B) FAS, (C) SREBP-2, and (D) HMGCR. NCI-H716 cells (2 × 10⁶) were incubated for 42 h with Dulbecco’s Minimal Eagle’s Medium plus 0.2% bovine serum albumin with or without one of the treatments (w/v). Due to lower solubility, leucine concentrations are 1.5% and 2.0% instead of 2.0% and 3.0%. Values are means ± SEMs of four experiments. Treatments with different letters are significantly different (P < 0.05). ACC, acetyl-coenzyme A carboxylase; FAS, fatty acid synthase; HMGCR, 3-hydroxy-3-methylglutaryl-coenzyme A reductase; SREBP-2, sterol regulatory element binding protein-2.

Fig. 4

Effects of skim milk, casein, and whey on in vitro mRNA expression of (A) i-FABP, (B) FATP4, and (C) NPC1L1. NCI-H716 cells (2 × 10⁶) were incubated for 42 h with Dulbecco’s Minimal Eagle’s Medium plus 0.2% bovine serum albumin with or without one of the treatments (w/v). Values are means ± SEMs of four experiments. Treatments with different letters are significantly different (P < 0.05). FATP4, fatty acid transport protein-4; i-FABP, intestinal-type fatty acid binding protein; NPC1L1, Niemann-Pick C-1–like-1 protein.

Fig. 5

Effects of skim milk, casein, and whey on in vitro mRNA expression of (A) ACC, (B) FAS, (C) SREBP-2, and (D) HMGCR. NCI-H716 cells (2 × 10⁶) were incubated for 42 h with Dulbecco’s Minimal Eagle’s Medium plus 0.2% bovine serum albumin with or without one of the treatments (w/v). Values are means ± SEMs of four experiments. Treatments with different letters are significantly different (P < 0.05). ACC, acetyl-coenzyme A carboxylase; FAS, fatty acid synthase; HMGCR, 3-hydroxy-3-methylglutaryl-cenzyme A reductase; SREBP-2, sterol regulatory element binding protein-2.