Abstract
Dietary supplementation with N-acetylcysteine (NAC) has been reported to improve intestinal health and treat gastrointestinal diseases. However, the underlying mechanisms are not fully understood. According to previous reports, NAC was thought to exert its effect through glutathione synthesis. This study tested the hypothesis that NAC enhances enterocyte growth and protein synthesis independently of cellular glutathione synthesis. Intestinal porcine epithelial cells were cultured for 3 days in Dulbecco’s modified Eagle medium containing 0 or 100 μM NAC. To determine a possible role for GSH (the reduced form of glutathione) in mediating the effect of NAC on cell growth and protein synthesis, additional experiments were conducted using culture medium containing 100 μM GSH, 100 μM GSH ethyl ester (GSHee), diethylmaleate (a GSH-depletion agent; 10 μM), or a GSH-synthesis inhibitor (buthionine sulfoximine, BSO; 20 μM). NAC increased cell proliferation, GSH concentration, and protein synthesis, while inhibiting proteolysis. GSHee enhanced cell proliferation and GSH concentration without affecting protein synthesis but inhibited proteolysis. Conversely, BSO or diethylmaleate reduced cell proliferation and GSH concentration without affecting protein synthesis, while promoting protein degradation. At the signaling level, NAC augmented the protein abundance of total mTOR, phosphorylated mTOR, and phosphorylated 70S6 kinase as well as mRNA levels for mTOR and p70S6 kinase in IPEC-1 cells. Collectively, these results indicate that NAC upregulates expression of mTOR signaling proteins to stimulate protein synthesis in enterocytes independently of GSH generation. Our findings provide a hitherto unrecognized biochemical mechanism for beneficial effects of NAC in intestinal cells.
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Abbreviations
- BSO:
-
Buthionine sulfoximine
- DEM:
-
Diethylmaleate
- GSH:
-
Glutathione
- GSHee:
-
Glutathione ethyl esters
- HMBS:
-
Hydroxymethylbilane synthase
- HPRT1:
-
Hypoxanthine phosphoribosyltransferase 1
- mTOR:
-
Mammalian target of rapamycin
- RPL19:
-
Ribosomal protein L19
- 4E-BP1:
-
4E-binding protein-1
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Acknowledgments
This research was jointly supported by the National Natural Science Foundation of China (Nos. 31372319, 31572416, 31402084), the Hubei Provincial Key Project for Scientific and Technical Innovation (2014ABA022), Hubei Provincial Research and Development Program (No. 2010BB023), Natural Science Foundation of Hubei Province (Nos. 2013CFA097, 2012FFB04805, 2011CDA131), the Hubei Hundred Talent Program, Agriculture and Food Research Initiative Competitive Grants of (2014-67015-21770) of the USDA National Institute of Food and Agriculture, and Texas A&M AgriLife Research (H-82000).
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Yi, D., Hou, Y., Wang, L. et al. N-acetylcysteine stimulates protein synthesis in enterocytes independently of glutathione synthesis. Amino Acids 48, 523–533 (2016). https://doi.org/10.1007/s00726-015-2105-z
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DOI: https://doi.org/10.1007/s00726-015-2105-z