Potential anti-aging agents suppress the level of constitutive mTOR- and DNA damage- signaling.
Two different mechanisms are considered to be the primary cause of aging. Cumulative DNA damage caused by reactive oxygen species (ROS), the by-products of oxidative phosphorylation, is one of these mechanisms (ROS concept). Constitutive stimulation of mitogen- and nutrient-sensing mTOR/S6 signaling is the second mechanism (TOR concept). The flow- and laser scanning- cytometric methods were developed to measure the level of the constitutive DNA damage/ROS- as well as of mTOR/S6- signaling in individual cells. Specifically, persistent activation of ATM and expression of γH2AX in untreated cells appears to report constitutive DNA damage induced by endogenous ROS. The level of phosphorylation of Ser235/236-ribosomal protein (RP), of Ser2448-mTOR and of Ser65-4EBP1, informs on constitutive signaling along the mTOR/S6 pathway. Potential gero-suppressive agents rapamycin, metformin, 2-deoxyglucose, berberine, resveratrol, vitamin D3 and aspirin, all decreased the level of constitutive DNA damage signaling as seen by the reduced expression of γH2AX in proliferating A549, TK6, WI-38 cells and in mitogenically stimulated human lymphocytes. They all also decreased the level of intracellular ROS and mitochondrial trans-membrane potential ΔΨm, the marker of mitochondrial energizing as well as reduced phosphorylation of mTOR, RP-S6 and 4EBP1. The most effective was rapamycin. Although the primary target of each on these agents may be different the data are consistent with the downstream mechanism in which the decline in mTOR/S6K signaling and translation rate is coupled with a decrease in oxidative phosphorylation, (revealed by ΔΨm) that leads to reduction of ROS and oxidative DNA damage. The decreased rate of translation induced by these agents may slow down cells hypertrophy and alleviate other features of cell aging/senescence. Reduction of oxidative DNA damage may lower predisposition to neoplastic transformation which otherwise may result from errors in repair of DNA sites coding for oncogenes or tumor suppressor genes. The data suggest that combined assessment of constitutive γH2AX expression, mitochondrial activity (ROS, ΔΨm) and mTOR signaling provides an adequate gamut of cell responses to evaluate effectiveness of gero-suppressive agents.
Brander Cancer Research Institute, Department of Pathology, New York Medical College, Valhalla, NY 10595, USA.
H Dorota
HD
Halicka
Hong
H
Zhao
Jiangwei
J
Li
Yong-Syu
YS
Lee
Tze-Chen
TC
Hsieh
Joseph M
JM
Wu
Zbigniew
Z
Darzynkiewicz
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D001241:Aspirin / Q000494:pharmacology; D064007:Ataxia Telangiectasia Mutated Proteins; D001599:Berberine / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D016922:Cellular Senescence / Q000187:drug effects*; D002762:Cholecalciferol / Q000378:metabolism; D004249:DNA Damage / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000378:metabolism; D003847:Deoxyglucose / Q000494:pharmacology; D015536:Down-Regulation; D004789:Enzyme Activation; D005434:Flow Cytometry; D006657:Histones / Q000378:metabolism; D006801:Humans; D047569:Laser Scanning Cytometry; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008687:Metformin / Q000494:pharmacology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D000077185:Resveratrol; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D020123:Sirolimus / Q000494:pharmacology; D013267:Stilbenes / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C087000:EIF4EBP1 protein, human; C468783:H2AX protein, human; D006657:Histones; D010750:Phosphoproteins; D017382:Reactive Oxygen Species; D013267:Stilbenes; D025521:Tumor Suppressor Proteins; D001599:Berberine; D002762:Cholecalciferol; D008687:Metformin; D003847:Deoxyglucose; C546842:MTOR protein, human; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D000077185:Resveratrol; D001241:Aspirin; D020123:Sirolimus