Effects of oral glutamine supplementation on exercise-induced gastrointestinal permeability and tight junction protein expression

Abstract

The objectives of this study are threefold: 1) to assess whether 7 days of oral glutamine (GLN) supplementation reduces exercise-induced intestinal permeability; 2) whether supplementation prevents the proinflammatory response; and 3) whether these changes are associated with upregulation of the heat shock response. On separate occasions, eight human subjects participated in baseline testing and in GLN and placebo (PLA) supplementation trials, followed by a 60-min treadmill run. Intestinal permeability was higher in the PLA trial compared with baseline and GLN trials (0.0604 ± 0.047 vs. 0.0218 ± 0.008 and 0.0272 ± 0.007, respectively; P < 0.05). IκBα expression in peripheral blood mononuclear cells was higher 240 min after exercise in the GLN trial compared with the PLA trial (1.411 ± 0.523 vs. 0.9839 ± 0.343, respectively; P < 0.05). In vitro using the intestinal epithelial cell line Caco-2, we measured effects of GLN supplementation (0, 4, and 6 mM) on heat-induced (37° or 41.8°C) heat shock protein 70 (HSP70), heat shock factor-1 (HSF-1), and occludin expression. HSF-1 and HSP70 levels increased in 6 mM supplementation at 41°C compared with 0 mM at 41°C (1.785 ± 0.495 vs. 0.6681 ± 0.290, and 1.973 ± 0.325 vs. 1.133 ± 0.129, respectively; P < 0.05). Occludin levels increased after 4 mM supplementation at 41°C and 6 mM at 41°C compared with 0 mM at 41°C (1.236 ± 0.219 and 1.849 ± 0.564 vs. 0.7434 ± 0.027, respectively; P < 0.001). GLN supplementation prevented exercise-induced permeability, possibly through HSF-1 activation.

Keywords: exercise; permeability; tight junction.

Fig. 1.

Experimental protocol.

Fig. 2.

End exercise core temperature for each subject during the placebo (PLA) and glutamine (GLN) trials. Data are expressed as individual subject's core temperature, n = 8.

Fig. 3.

Time course effect of 60 min of treadmill exercise on heart rate (A) and rectal temperature (B). Both heart rate and rectal temperature were not different between trials at any 5-min interval. Data are means ± SD, n = 8.

Fig. 4.

Oral GLN supplementation increased plasma GLN levels. Plasma GLN was significantly higher at the preexercise time point in the GLN trial compared with preexercise in PLA trial. Plasma GLN was significantly lower at 20 min postexercise in the GLN trial compared with preexercise in the GLN trial. *P < 0.05 statistically significant from the same time point in the PLA trial; **P < 0.05 statistically significant from the GLN preexercise time point. Data are means ± SD, n = 8.

Fig. 5.

Glutamine prevented a rise in intestinal permeability as measured by the urinary excretion ratio of lactulose and rhamnose (L/R ratio). The L/R ratio was higher in the PLA trial compared with rest and GLN trials. *P < 0.05, statistically significant from the rest trial; **P < 0.05, statistically significant from the PLA. Data are means ± SD, n = 8.

Fig. 6.

Effect of GLN supplementation on IκBα levels in PBMCs. The 240-min postexercise time point was higher in the GLN trial compared with the 240-min postexercise time point in the PLA trial. A: protein expression of IκBα and β-actin (loading control) were measured in PBMCs of subjects after 7 days of PLA and GLN supplementation. B: densitometric values of protein content were obtained using Photoshop software and normalized to β-actin and set to 1. *P < 0.05, statistically significant from the same time point in the PLA trial. Data are means ± SD, n = 8 for each time point.

Fig. 7.

Effect of GLN supplementation and heat stress on HSF-1, HSP70, and occludin protein expression in Caco-2 epithelial cells. HSF-1 (A and B) was significantly higher in the 4 and 6 mM 41°C trial compared with the 0 mM 37°C trial. A: HSF-1 and β-actin (loading control) protein expression. B: densitometry of protein content of the corresponding blots (A) was corrected for loading with β-actin and expressed as a ratio. *P < 0.05, statistically significant from the 0 mM 37°C trial; +P < 0.05, statistically significant from the 0 mM 41°C trial. Data are means ± SD, n = 4. HSP70 (C and D) protein expression was higher in the 4 and 6 mM 41°C trial compared with 0 mM 37°C trial. C: HSP70 and β-actin (loading control) protein expression. D: densitometry of protein content of the corresponding blots (C) was corrected for loading with β-actin and expressed as a ratio. *P < 0.05, statistically significant from the 0 mM 37°C trial; +P < 0.05, statistically significant from the 0 mM 41°C trial. Data are means ± SD, n = 4. Occludin expression (E and F) was significantly decreased in the 0 mM 41°C trial compared with the 0 mM 37°C trial. Occludin levels were higher in the 4 and 6 mM 41°C trials compared with the 0 mM 41°C trial. E: occludin and β-actin (loading control) protein expression. F: densitometry of protein content of the corresponding blots (E) was corrected for loading with β-actin and expressed as a ratio. *P < 0.05, statistically significant from the 0°C mM 37°C trial; +P < 0.05, statistically significant from the 0 mM 41°C trial. Data are means ± SD, n = 4.