Glutamine stimulates amino acid transport during ischemia-reperfusion in human intestinal epithelial cells.
BACKGROUND
The potential mechanism of intestinal ischemia-reperfusion (I/R) injury includes oxygen-derived toxic free radicals. We tested the hypothesis that glutamine increases intracellular glutathione, a protective substrate against oxidative stress, by stimulating membrane amino acid transport during I/R using human intestinal epithelial cell line Caco-2.


METHODS
Ischemic conditions were obtained by combining both hypoxic (1%O2-5%CO2-94% N2) and nutrient-deprived (Phosphate-Buffered Saline; PBS) conditions. After 2 h of ischemia, re-oxygenation (5%CO2-95% air) was initiated and the culture medium was changed to PBS, PBS supplemented with amino acids (A.A.), and PBS supplemented with 2 mm glutamine plus amino acids (Gln) (reperfusion). After 4 h of reperfusion, the transport of 3H-glutamine, 3H-glutamate, and 3H-leucine was assayed and intracellular glutathione was measured. 3H-thymidine incorporation was measured for the determination of DNA synthesis. Data (mean +/- SD) were analyzed by ANOVA.


RESULTS
Ischemia decreased Na+-dependent glutamine, Na+-dependent glutamate, and Na+-independent leucine transport compared with control (P < 0.01). After reperfusion, glutamine and glutamate transport in the PBS and A.A. groups decreased significantly compared with control (P < 0.01), whereas glutamine supplementation increased glutamine transport to the levels in control (P < 0.01) and partially increased glutamate transport (P < 0.01). Leucine transport significantly increased in the A.A. and Gln groups compared with the PBS group. Glutamine significantly increased intracellular glutathione and DNA synthesis compared with the PBS and A.A. groups (P < 0.01).


CONCLUSIONS
This study demonstrated that glutamine up-regulates amino acid transport during I/R in human intestinal epithelial cells, possibly resulting in increased intracellular glutathione and DNA synthesis.
Wasa
Masafumi
M
Department of Pediatric Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka, Japan. wasa@pedsurg.med.osaka-u.ac.jp
Soh
Hideki
H
Shimizu
Yoshiyuki
Y
Fukuzawa
Masahiro
M
D000596:Amino Acids / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D018938:Caco-2 Cells; D004247:DNA / Q000096:biosynthesis; D005973:Glutamine / Q000494:pharmacology*; D005978:Glutathione / Q000096:biosynthesis; D006801:Humans; D007413:Intestinal Mucosa / Q000098:blood supply*; D007511:Ischemia / Q000378:metabolism*; D015424:Reperfusion
D000596:Amino Acids; D005973:Glutamine; D004247:DNA; D005978:Glutathione