Abstract
Calorie restriction (CR) extends lifespan in a remarkable range of organisms. However, the mechanisms of CR related to the longevity effects are not fully elucidated to date. Using human fibroblast Hs68 (Hs68) cells cultured at a lower level of medium glucose (i.e., glucose restriction; GR) to mimic CR, we investigated the crucial role of nicotinamide phosphoribosyltransferase (Nampt), nicotinamide adenine dinucleotide (NAD+), and nicotinamide (NAM) in GR-extended replicative lifespan of Hs68 cells. We found that GR extended the lifespan of Hs68 cells, in parallel to significantly increased expression of Nampt, intracellular NAD+ levels, and SIRT1 activities, and to significantly decreased NAM levels. The lifespan-extending effects of GR were profoundly diminished by FK866 (a noncompetitive inhibitor of Nampt) and blocked by sirtinol (a noncompetitive inhibitor of sirtuins). However, the steady-state intracellular NAM level (averaged 2.5 μM) was much lower than the IC50 of NAM on human SIRT1 (about 50 μM). All these results suggest that up-regulation of Nampt play an important role in GR-extended lifespan of Hs68 cells by increasing the intracellular NAD+ levels followed by activating SIRT1 activity in Hs68 cells. In contrast, the role of NAM depletion is limited.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Anderson RM, Bitterman KJ, Wood JG, Medvedik O, Cohen H, Lin SS, Manchester JK, Gordon J, Sinclair DA (2002) Manipulation of a nuclear NAD + salvage pathway delays aging without altering steady-state NAD + levels. J Biol Chem 277:18881–18890. doi:10.1074/jbc.M111773200
Anderson RM, Bitterman KJ, Wood JG, Medvedik O, Sinclair DA (2003a) Nicotinamide and PNC1 govern lifespan extension by calorie restriction in Saccharomyces cerevisiae. Nature 423:181–185
Anderson RM, Latorre-Esteves M, Neves AR, Lavu S, Medvedik O, Taylor C, Howitz KT, Santos H, Sinclair DA (2003b) Yeast life-span extension by calorie restriction is independent of NAD fluctuation. Science 302:2124–2126. doi:10.1126/science.1088697
Billington RA, Travelli C, Ercolano E, Galli U, Roman CB, Grolla AA, Canonico PL, Condorelli F, Genazzani AA (2008) Characterization of NAD uptake in mammalian cells. J Biol Chem 283:6367–6374. doi:10.1074/jbc.M706204200
Bogan KL, Brenner C (2008) Nicotinic acid, nicotinamide, and nicotinamide riboside: a molecular evaluation of NAD+ precursor vitamins in human nutrition. Annu Rev Nutr 28:115–130. doi:10.1146/annurev.nutr.28.061807.155443
Borradaile NM, Pickering JG (2009) Nicotinamide phosphoribosyltransferase imparts human endothelial cells with extended replicative lifespan and enhanced angiogenic capacity in a high glucose environment. Aging Cell 8:100–112. doi:10.1111/j.1474-9726.2009.00453.x
Colman RJ, Anderson RM (2011) Nonhuman primate calorie restriction. Antioxid Redox Signal 14:229–239. doi:10.1089/ars.2010.3224
Dahl TB, Holm S, Aukrust P, Halvorsen B (2012) Visfatin/NAMPT: a multifaceted molecule with diverse roles in physiology and pathophysiology. Annu Rev Nutr 32:229–243. doi:10.1146/annurev-nutr-071811-150746
David Adams J Jr, Klaidman LK (2007) Sirtuins, nicotinamide and aging: a critical review. Lett Drug Des Discov 4:44–48. doi:10.2174/157018007778992892
Donmez G, Guarente L (2010) Aging and disease: connections to sirtuins. Aging Cell 9:285–290. doi:10.1111/j.1474-9726.2010.00548.x
Dragovic J, Kim SH, Brown SL, Kim JH (1995) Nicotinamide pharmacokinetics in patients. Radiother Oncol 36:225–228. doi:10.1016/0167-8140(95)01581-Z
Everitt AV, Le Couteur DG (2007) Life extension by calorie restriction in humans. Ann N Y Acad Sci 1114:428–433. doi:10.1196/annals.1396.005
Fulco M, Cen Y, Zhao P, Hoffman EP, McBurney MW, Sauve AA, Sartorelli V (2008) Glucose restriction inhibits skeletal myoblast differentiation by activating SIRT1 through AMPK-mediated regulation of Nampt. Dev Cell 14:661–673. doi:10.1016/j.devcel.2008.02.004
Grozinger CM, Chao ED, Blackwell HE, Moazed D, Schreiber SL (2001) Identification of a class of small molecule inhibitors of the sirtuin family of NAD-dependent deacetylases by phenotypic screening. J Biol Chem 276:38837–38843. doi:10.1074/jbc.M106779200
Haigis MC, Sinclair DA (2010) Mammalian sirtuins: biological insights and disease relevance. Annu Rev Pathol 5:253–295. doi:10.1146/annurev.pathol.4.110807.092250
Hara N, Yamada K, Shibata T, Osago H, Tsuchiya M (2011) Nicotinamide phosphoribosyltransferase/visfatin does not catalyze nicotinamide mononucleotide formation in blood plasma. PLoS One 6:e22781. doi:10.1371/journal.pone.0022781
Hasmann M, Schemainda I (2003) FK866, a highly specific noncompetitive inhibitor of nicotinamide phosphoribosyltransferase, represents a novel mechanism for induction of tumor cell apoptosis. Cancer Res 63:7436–7442
Hayashida S, Arimoto A, Kuramoto Y, Kozako T, Honda S, Shimeno H, Soeda S (2010) Fasting promotes the expression of SIRT1, an NAD+ -dependent protein deacetylase, via activation of PPARalpha in mice. Mol Cell Biochem 339:285–292. doi:10.1007/s11010-010-0391-z
Hayflick L, Moorehead PS (1961) The serial cultivation of human diploid cell strains. Exp Cell Res 25:585–621. doi:10.1016/0014-4827(61)90192-6
Horsman MR, Høyer M, Honess DJ, Dennis IF, Overgaard J (1993) Nicotinamide pharmacokinetics in humans and mice: a comparative assessment and the implications for radiotherapy. Radiother Oncol 27:131–139. doi:10.1016/0167-8140(93)90133-S
Hoskin PJ, Stratford MR, Saunders MI, Hall DW, Dennis MF, Rojas A (1995) Administration of nicotinamide during chart: pharmacokinetics, dose escalation, and clinical toxicity. Int J Radiat Oncol Biol Phys 32:1111–1119. doi:10.1016/0360-3016(95)00022-Q
Imai S (2010) A possibility of nutriceuticals as an anti-aging intervention: activation of sirtuins by promoting mammalian NAD biosynthesis. Pharmacol Res 62:42–47. doi:10.1016/j.phrs.2010.01.006
Lee PD, Chang YJ, Lin KL, Chang YZ (2012) Simultaneous determination of Δ9-tetrahydrocannabinol and 11-nor-9-carboxy-Δ9-tetrahydrocannabinol in oral fluid using isotope dilution liquid chromatography tandem mass spectrometry. Anal Bioanal Chem 402:851–859. doi:10.1007/s00216-011-5439-8
Li Y, Tollefsbol T (2011) p16INK4a suppression by glucose restriction contributes to human cellular lifespan extension through SIRT1-Mediated epigenetic and genetic mechanisms. PLoS One 6:e17421. doi:10.1371/journal.pone.0017421
Li Y, Liu L, Tollefsbol T (2010) Glucose restriction can extend normal cell lifespan and impair precancerous cell growth through epigenetic control of hTERT and p16 expression. FASEB J 24:1442–1453. doi:10.1096/fj.09-149328
Lin SJ, Guarente L (2003) Nicotinamide adenine dinucleotide, a metabolic regulator of transcription, longevity and disease. Curr Opin Cell Biol 15:241–246. doi:10.1016/S0955-0674(03)00006-1
Lin SS, Manchester JK, Gordon JI (2001) Enhanced gluconeogenesis and increased energy storage as hallmarks of aging in Saccharomyces cerevisiae. J Biol Chem 276:36000–36007. doi:10.1074/jbc.M103509200
Lin SJ, Ford E, Haigis M, Liszt G, Guarente L (2004) Calorie restriction extends yeast life span by lowering the level of NADH. Genes Dev 18:12–16. doi:10.1101/gad.1164804
Lu SP, Lin SJ (2010) Regulation of yeast sirtuins by NAD(+) metabolism and calorie restriction. Biochim Biophys Acta 1804:1567–1575. doi:10.1016/j.bbapap.2009.09.030
Ota H, Tokunaga E, Chang K, Hikasa M, Iijima K, Eto M, Kozaki K, Akishita M, Ouchi Y, Kaneki M (2006) Sirt1 inhibitor, Sirtinol, induces senescence-like growth arrest with attenuated Ras-MAPK signaling in human cancer cells. Oncogene 25:176–185. doi:10.1038/sj.onc.1209049
Revollo JR, Grimm AA, Imai S (2004) The NAD biosynthesis pathway mediated by nicotinamide phosphoribosyltransferase regulates Sir2 activity in mammalian cells. J Biol Chem 279:50754–50763. doi:10.1074/jbc.M408388200
Sauve AA (2008) NAD+ and vitamin B3: from metabolism to therapies. J Pharmacol Exp Ther 324:883–893. doi:10.1124/jpet.107.120758
Sauve AA, Wolberger C, Schramm VL, Boeke JD (2006) The biochemistry of sirtuins. Annu Rev Biochem 75:435–465. doi:10.1146/annurev.biochem.74.082803.133500
Schmidt MT, Smith BC, Jackson MD, Denu JM (2004) Coenzyme specificity of Sir2 protein deacetylases: implications for physiological regulation. J Biol Chem 279:40122–40129. doi:10.1074/jbc.M407484200
Srivastava DK, Bernhard SA (1987) Biophysical chemistry of metabolic reaction sequences in concentrated enzyme solution and in the cell. Annu Rev Biophys Biophys Chem 16:175–204. doi:10.1146/annurev.bb.16.060187.001135
Stratford MR, Dennis MF (1992) High-performance liquid chromatographic determination of nicotinamide and its metabolites in human and murine plasma and urine. J Chromatogr 582:145–151. doi:10.1016/0378-4347(92)80313-F
Stratford MR, Rojas A, Hall DW, Dennis MF, Dische S, Joiner MC, Hodgkiss RJ (1992) Pharmacokinetics of nicotinamide and its effect on blood pressure, pulse and body temperature in normal human volunteers. Radiother Oncol 25:37–42. doi:10.1016/0167-8140(92)90193-X
Takeuchi M, Niimi T, Masumoto M, Orita M, Yokota H, Yamamoto T (2014) Discovery of a novel nicotinamide phosphoribosyl transferase (NAMPT) inhibitor via in silico screening. Biol Pharm Bull 37:31–36. doi:10.1248/bpb.b13-00495
van der Veer E, Ho C, O’Neil C, Barbosa N, Scott R, Cregan SP, Pickering JG (2007) Extension of human cell lifespan by nicotinamide phosphoribosyltransferase. J Biol Chem 282:10841–10845. doi:10.1074/jbc.C700018200
Yang H, Lavu S, Sinclair DA (2006) Nampt/PBEF/Visfatin: a regulator of mammalian health and longevity? Exp Gerontol 41:718–726. doi:10.1016/j.exger.2006.06.003
Yang NC, Song TY, Chen MY, Hu ML (2011) Effects of 2-deoxyglucose and dehydroepiandrosterone on intracellular NAD(+) level, SIRT1 activity and replicative lifespan of human Hs68 cells. Biogerontology 12:527–536. doi:10.1007/s10522-011-9342-7
Acknowledgments
This work was supported by grants from Chung-San Medical University (G100N0004) and the National Science Foundation (NSC 101-2320-B-040-010). Taiwan ROC. The microplate fluorometer and Chemiluminescence Imaging System was performed in the Instrument Center of Chung Shan Medical University, which is supported by National Science Council, Ministry of Education and Chung Shan Medical University.
Author information
Authors and Affiliations
Corresponding authors
Rights and permissions
About this article
Cite this article
Yang, NC., Song, TY., Chang, YZ. et al. Up-regulation of nicotinamide phosphoribosyltransferase and increase of NAD+ levels by glucose restriction extend replicative lifespan of human fibroblast Hs68 cells. Biogerontology 16, 31–42 (2015). https://doi.org/10.1007/s10522-014-9528-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10522-014-9528-x