Format

Send to

Choose Destination
Lipids Health Dis. 2017 Oct 3;16(1):191. doi: 10.1186/s12944-017-0583-6.

Oleate ameliorates palmitate-induced reduction of NAMPT activity and NAD levels in primary human hepatocytes and hepatocarcinoma cells.

Author information

1
Center for Pediatric Research Leipzig (CPL), University Hospital for Children & Adolescents, University of Leipzig, Liebigstraße 21, 04103, Leipzig, Germany. melanie.penke@medizin.uni-leipzig.de.
2
Center for Pediatric Research Leipzig (CPL), University Hospital for Children & Adolescents, University of Leipzig, Liebigstraße 21, 04103, Leipzig, Germany.
3
Institute of Biochemistry, Faculty of Medicine, University of Leipzig, Johannisallee 30, 04103, Leipzig, Germany.

Abstract

BACKGROUND:

Nicotinamide phosphoribosyltransferase (NAMPT) and nicotinamide adenine dinucleotide (NAD) levels are crucial for liver function. The saturated fatty acid palmitate and the unsaturated fatty acid oleate are the main free fatty acids in adipose tissue and human diet. We asked how these fatty acids affect cell survival, NAMPT and NAD levels in HepG2 cells and primary human hepatocytes.

METHODS:

HepG2 cells were stimulated with palmitate (0.5mM), oleate (1mM) or a combination of both (0.5mM/1mM) as well as nicotinamide mononucleotide (NMN) (0.5 mM) or the specific NAMPT inhibitor FK866 (10nM). Cell survival was measured by WST-1 assay and Annexin V/propidium iodide staining. NAD levels were determined by NAD/NADH Assay or HPLC. Protein and mRNA levels were analysed by Western blot analyses and qPCR, respectively. NAMPT enzyme activity was measured using radiolabelled 14C-nicotinamide. Lipids were stained by Oil red O staining.

RESULTS:

Palmitate significantly reduced cell survival and induced apoptosis at physiological doses. NAMPT activity and NAD levels significantly declined after 48h of palmitate. In addition, NAMPT mRNA expression was enhanced which was associated with increased NAMPT release into the supernatant, while intracellular NAMPT protein levels remained stable. Oleate alone did not influence cell viability and NAMPT activity but ameliorated the negative impact of palmitate on cell survival, NAMPT activity and NAD levels, as well as the increased NAMPT mRNA expression and secretion. NMN was able to normalize intracellular NAD levels but did not ameliorate cell viability after co-stimulation with palmitate. FK866, a specific NAMPT inhibitor did not influence lipid accumulation after oleate-treatment.

CONCLUSIONS:

Palmitate targets NAMPT activity with a consequent cellular depletion of NAD. Oleate protects from palmitate-induced apoptosis and variation of NAMPT and NAD levels. Palmitate-induced cell stress leads to an increase of NAMPT mRNA and accumulation in the supernatant. However, the proapoptotic action of palmitate seems not to be mediated by decreased NAD levels.

KEYWORDS:

FK866; Liver; NAD; NAMPT; NMN; Oleate; Palmitate; Steatosis

PMID:
28974242
PMCID:
PMC5627432
DOI:
10.1186/s12944-017-0583-6
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for BioMed Central Icon for PubMed Central
Loading ...
Support Center