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- Title
L-Glutamate deficiency can trigger proliferation inhibition via down regulation of the mTOR/S6K1 pathway in pig intestinal epithelial cells.
- Authors
Li, X.-G.; Sui, W.-G.; Gao, C.-Q.; Yan, H.-C.; Yin, Y.-L.; Li, H.-C.; Wang, X.-Q.
- Abstract
The objective of this study was to investigate the effects of L-glutamate (Glu) deficiency or L-trans pyrrolidine-2,4-dicarboxylic acid (PDC) supplementation on the proliferation of pig intestinal epithelial cells (IPEC-1). First, IPEC-1 cells were cultured in normal growing medium supplemented with 0 (Control), 50, 100, or 200 μmol/L PDC to determine an appropriate concentration of PDC supplementation. Second, IPEC-1 cells were cultured in Glu-deficient medium supplemented with 0 μmol/L Glu (Glu deficiency), 50 mmol/L Glu (Control), or 50 μmol/L Glu plus 100 μmol/L PDC (PDC supplementation). Cell proliferation (n = 24), cell cycle distribution (n = 6), cell apoptosis (n = 6), and expression levels of proteins of interest (n = 4) were determined by MTT assay, flow cytometry, or western blot. The results showed that cell proliferation was inhibited (P < 0.05) by 50, 100, and 200 mmol/L PDC supplementation at 24 and 48 h after treatment. Variance analysis was performed using the GLM procedure, and the results demonstrated that Glu deficiency or PDC supplementation led to the inhibition (P < 0.05) of cell proliferation, a greater (P < 0.05) percentage of cells in the G1 phase, and a lower (P < 0.05) percentage of cells in the S phase. Moreover, Glu deficiency or PDC supplementation reduced (P < 0.05) the expression levels of excitatory AA transporter 3 (EAAT3), phosphormammalian target of rapamycin (p-mTOR; Ser2448), p-ribosomal protein S6 kinase 1 (S6K1; Thr389), and p-S6 (Ser235/236). This study demonstrates that Glu deficiency or PDC supplementation inhibits proliferation of IPEC-1 cells via downregulation of the mTOR/ S6K1 pathway and EAAT3 expression indicating that Glu deficiency may lead to the disturbances of intestinal epithelial renewal in pigs, particularly in neonates.
- Subjects
GLUTAMIC acid; EPITHELIAL cells; CELL proliferation; GENE expression; SWINE genetics; LIVESTOCK genetics
- Publication
Journal of Animal Science, 2016, Vol 94, Issue 4, p1541
- ISSN
0021-8812
- Publication type
Article
- DOI
10.2527/jas.2015-9432