Nicotinamide Phosphoribosyltransferase Knockdown Leads to Lipid Accumulation in HepG2 Cells through The SIRT1-AMPK Pathway

(Pages: 125-132)
Davod Ilbeigi, Ph.D, 1Mitra Nourbakhsh, Ph.D, 2,3,*Parvin Pasalar, Ph.D, 1,3,*Reza Meshkani, Ph.D, 1Hajar Shokri Afra, Ph.D., 1Ghodratollah Panahi, Ph.D., 1Mohammad Borji, Ph.D., 4Roya Sharifi, Ph.D., 5
Department of Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran
Department of Biochemistry, School of Medicine, Iran University of Medical Sciences, Tehran, Iran
Metabolic Disorders Research Center, Endocrinology and Metabolism Molecular, Cellular Sciences Institute, Tehran University of Medical Sciences, Tehran, Iran
Department of Biochemistry, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Department of Medical Laboratory Sciences, School of Allied Medical sciences, Iran University of Medical Sciences, Tehran, Iran
Department of Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran
Department of Biochemistry, School of Medicine, Iran University of Medical Sciences, Tehran, Iran
Metabolic Disorders Research Center, Endocrinology and Metabolism Molecular, Cellular Sciences Institute, Tehran University of Medical Sciences, Tehran, Iran
Department of Biochemistry, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
Department of Medical Laboratory Sciences, School of Allied Medical sciences, Iran University of Medical Sciences, Tehran, Iran
*Corresponding Addresses: P.O.Box: 1449614535 Department of Biochemistry School of Medicine Iran University of Medical Sciences Tehran Iran P.O.Box: 1417614418 Department of Biochemistry School of Medicine Tehran University of Medical Sciences Tehran Iran Emails:nourbakhsh.m@iums.ac.ir,pasalar@sina.tums.ac.ir
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Ilbeigi Davod, Nourbakhsh Mitra, Pasalar Parvin, Meshkani Reza, Shokri Afra Hajar, Panahi Ghodratollah, Borji Mohammad, Sharifi Roya. Nicotinamide Phosphoribosyltransferase Knockdown Leads to Lipid Accumulation in HepG2 Cells through The SIRT1-AMPK Pathway . Cell J. 2020; 22(): 125-132.

Abstract

Objective

Nicotinamide phosphoribosyltransferase (NAMPT), which is responsible for biosynthesis of nicotinamide adenine dinucleotide (NAD), has a regulatory role in cellular metabolism and thus, might be implicated in non-alcoholic fatty liver disease (NAFLD). This study aimed to show how NAMPT down-regulation in liver cells influences lipid metabolism and sirtiun 1 (SIRT1), as the main NAD-dependent deacetylase enzyme.

Materials and Methods

In this experimental study, HepG2 cells were transfected with NAMPT siRNA and hepatic triglyceride (TG) content and SIRT1 deacetylase activity were measured by colorimetric and fluorometric methods, respectively. Gene expression of fatty acid synthase (FAS) and sterol regulatory element-binding protein-1c (SREBP- 1c) was evaluated by real-time polymerase chain reaction (PCR). Total protein level and the phosphorylated form of acetyl-CoA carboxylase (ACC) and AMP-activated protein kinase (AMPK) were also investigated by western blotting.

Results

Knockdown of NAMPT significantly promoted the accumulation of TG in HepG2 cells, accompanied by a remarkable decline in SIRT1 deacetylase activity. A significant rise in the gene expression of two key lipogenic factors, FAS and SREBP-1c was also observed. These effects were also accompanied by decreased phosphorylation of ACC and AMPK. On the other hand, treatment of transfected cells with either NAD, as the SIRT1 substrate or resveratrol, as the SIRT1 activator reversed the outcomes.

Conclusion

These results demonstrated a protective role for NAMPT against NAFLD and its involvement in the regulation of de novo lipogenesis through the SIRT1/AMPK pathway.