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l-Glutamine enhances enterocyte growth via activation of the mTOR signaling pathway independently of AMPK

Abstract

Neonates (including human infants) require l-glutamine (Gln) for optimal intestinal health. This study tested the hypothesis that Gln enhances enterocyte growth via both mammalian target of rapamycin (mTOR) and AMP-activated kinase (AMPK) signaling pathways. Intestinal porcine epithelial cells (IPEC-1) were cultured for 3 days in Gln-free Dulbecco’s modified Eagle medium containing 0 or 2 mM Gln. To determine the role of mTOR and AMPK on cell growth, additional experiments were conducted where medium contained 2 mM Gln and 10 nM rapamycin (Rap, an inhibitor of mTOR) or 1 μM compound C (an inhibitor of AMPK). IPEC-1 cell growth increased with increasing concentrations of Gln from 0 to 2 mM. Compared with 0 mM Gln, 2 mM Gln increased (P < 0.05) the amounts of phosphorylated 4E-binding protein 1 (4E-BP1) and ribosomal protein S6 kinase (p70S6 kinase) proteins but did not affect abundances of total or phosphorylated AMPK protein. Gln also increased mRNA levels for Bcl-2, mTOR, p70S6 kinase, 4E-BP1, COX7C, ASCT2, ODC, SGLT-1, CFTR, Na+/K+-ATPase, HSP70, and ZO-1. Similarly, cells cultured with Rap and Gln exhibited higher (P < 0.05) abundances of phosphorylated 4E-BP1 and p70S6 kinase proteins than the Rap-only group, whereas abundances of phosphorylated mTOR and 4E-BP1 proteins were increased when AMPK was inhibited by compound C. Conversely, the amount of phosphorylated AMPK increased when mTOR was inhibited by Rap, suggesting a negative cross-talk between mTOR and AMPK. Collectively, these results indicate that Gln stimulates enterocyte growth by activating the mTOR signaling pathway independently of AMPK.

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Abbreviations

AMPK:

Adenosine 5′-monophosphate (AMP)-activated protein kinase

ASCT2:

Na+-neutral AA exchanger 2

Bcl-2:

B-cell lymphoma 2

Cc:

Compound C

CFTR:

Cystic fibrosis transmembrane conductance regulator

COX7C:

Cytochrome c oxidase subunit I

HSP70:

Heat shock protein 70

MAPK6:

Mitogen-activated protein kinase 6

mTOR:

Mammalian target of rapamycin

ODC:

Ornithine decarboxylase

pBD-1:

Porcine β-defense 1

Rap:

Rapamycin

SGLT-1:

Sodium/glucose co-transporter-1

Sirt 1:

Sirtuin 1

4E-BP1:

4E-binding protein-1

ZO-1:

Zonula occludens-1

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Acknowledgments

This research was jointly supported by National Basic Research Program of China (No. 2012CB126305), National Natural Science Foundation of China (No. 31372319 and 31402084), Hubei Provincial Research and Development Program (No. 2010BB023), Natural Science Foundation of Hubei Province (No. 2013CFA097, 2013CFB325, 2012FFB04805, 2011CDA131), the Hubei Hundred Talent program, Agriculture and Food Research Initiative Competitive Grant (2014-67015-21770) of the USDA National Institute of Food and Agriculture, and Texas AgriLife Research (H-82000). All these funding agencies had no role in the design, analysis or writing of this study.

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The authors declare that they have no conflict of interest.

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Correspondence to Yongqing Hou.

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Yi, D., Hou, Y., Wang, L. et al. l-Glutamine enhances enterocyte growth via activation of the mTOR signaling pathway independently of AMPK. Amino Acids 47, 65–78 (2015). https://doi.org/10.1007/s00726-014-1842-8

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  • DOI: https://doi.org/10.1007/s00726-014-1842-8

Keywords

  • l-Glutamine
  • IPEC-1
  • mTOR
  • AMPK