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Original Article

Korean J Physiol Pharmacol 2022; 26(4): 263-275

Published online July 1, 2022 https://doi.org/10.4196/kjpp.2022.26.4.263

Copyright © Korean J Physiol Pharmacol.

Comprehensive investigations of key mitochondrial metabolic changes in senescent human fibroblasts

Hazem K. Ghneim, Mohammad A. Alfhili*, Sami O. Alharbi, Shady M. Alhusayni, Manal Abudawood, Feda S. Aljaser, and Yazeed A. Al-Sheikh

Chair of Medical and Molecular Genetics Research, Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Saud University, Riyadh 12372, Saudi Arabia

Correspondence to:Mohammad A. Alfhili
E-mail: malfeehily@ksu.edu.sa

Author contributions: H.K.G. and M.A.A. performed the Conception and design of the study. S.O.A. and S.M.A. contributed to data acquisition. H.K.G., S.O.A., S.M.A., M.A., F.S.A., Y.A.A., and M.A.A. performed the analysis and/or interpretation of data. H.K.G., M.A.A., and M.A. wrote the draft the manuscript. M.A.A., M.A., and Y.A.A. revised the manuscript critically for important intellectual content. All authors have read and approved the final manuscript.

Received: December 17, 2021; Revised: March 23, 2022; Accepted: April 7, 2022

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

There is a paucity of detailed data related to the effect of senescence on the mitochondrial antioxidant capacity and redox state of senescent human cells. Activities of TCA cycle enzymes, respiratory chain complexes, hydrogen peroxide (H2O2), superoxide anions (SA), lipid peroxides (LPO), protein carbonyl content (PCC), thioredoxin reductase 2 (TrxR2), superoxide dismutase 2 (SOD2), glutathione peroxidase 1 (GPx1), glutathione reductase (GR), reduced glutathione (GSH), and oxidized glutathione (GSSG), along with levels of nicotinamide cofactors and ATP content were measured in young and senescent human foreskin fibroblasts. Primary and senescent cultures were biochemically identified by monitoring the augmented cellular activities of key glycolytic enzymes including phosphofructokinase, lactate dehydrogenase, and glycogen phosphorylase, and accumulation of H2O2, SA, LPO, PCC, and GSSG. Citrate synthase, aconitase, α-ketoglutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, isocitrate dehydrogenase, and complex I-III, IIIII, and IV activities were significantly diminished in P25 and P35 cells compared to P5 cells. This was accompanied by significant accumulation of mitochondrial H2O2, SA, LPO, and PCC, along with increased transcriptional and enzymatic activities of TrxR2, SOD2, GPx1, and GR. Notably, the GSH/GSSG ratio was significantly reduced whereas NAD+/NADH and NADP+/NADPH ratios were significantly elevated. Metabolic exhaustion was also evident in senescent cells underscored by the severely diminished ATP/ADP ratio. Profound oxidative stress may contribute, at least in part, to senescence pointing at a potential protective role of antioxidants in aging-associated disease.

Keywords: Aging, Antioxidants, Mitochondria, Oxidative stress, Senescence