Fucoxanthin Pretreatment Ameliorates Visible Light-Induced Phagocytosis Disruption of RPE Cells under a Lipid-Rich Environment via the Nrf2 Pathway

Fucoxanthin, a special xanthophyll derived from marine algae, has increasingly attracted attention due to its diverse biological functions. However, reports on its ocular benefits are still limited. In this work, the ameliorative effect of fucoxanthin on visible light and lipid peroxidation-induced phagocytosis disruption in retinal pigment epithelium (RPE) cells was investigated in vitro. Marked oxidative stress, inflammation, and phagocytosis disruption were evident in differentiated RPE cells following their exposure to visible light under a docosahexaenoic acid (DHA)-rich environment. Following pretreatment with fucoxanthin, however, the activated nuclear factor erythroid-derived-2-like 2 (Nrf2) signaling pathway was observed and, furthermore, when the fucoxanthin -pretreated RPE cells were irradiated with visible light, intracellular reactive oxygen species (ROS), malondialdehyde (MDA) levels and inflammation were obviously suppressed, while phagocytosis was significantly improved. However, following the addition of Nrf2 inhibitor ML385, the fucoxanthin exhibited no ameliorative effects on the oxidative stress, inflammation, and phagocytosis disruption in the RPE cells, thus indicating that the ameliorative effect of fucoxanthin on the phagocytosis of RPE cells is closely related to the Nrf2 signaling pathway. In conclusion, these results suggest that fucoxanthin supplementation might be beneficial to the prevention of visible light-induced retinal injury.

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JAM-C maintains VEGR2 expression to promote retinal pigment epithelium cell survival under oxidative stress

Thrombosis and Haemostasis ◽

10.1160/th16-11-0885 ◽

2017 ◽

Vol 117 (04) ◽

pp. 750-757

Author(s):

Xin Jia ◽

Chen Zhao ◽

Qishan Chen ◽

Yuxiang Du ◽

Lijuan Huang ◽

...

Keyword(s):

Oxidative Stress ◽

Retinal Pigment Epithelium ◽

Cell Survival ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Retinal Pigment Epithelium Cell ◽

Photoreceptor Cells ◽

Rpe Cells

SummaryJunctional adhesion molecule-C (JAM-C) has been shown to play critical roles during development and in immune responses. However, its role in adult eyes under oxidative stress remains poorly understood. Here, we report that JAM-C is abundantly expressed in adult mouse retinae and choroids in vivo and in cultured retinal pigment epithelium (RPE) and photoreceptor cells in vitro. Importantly, both JAM-C expression and its membrane localisation are downregulated by H2O2-induced oxidative stress. Under H2O2-induced oxidative stress, JAM-C is critically required for the survival of human RPE cells. Indeed, loss of JAM-C by siRNA knockdown decreased RPE cell survival. Mechanistically, we show that JAM-C is required to maintain VEGFR2 expression in RPE cells, and VEGFR2 plays an important role in keeping the RPE cells viable since overexpression of VEGFR2 partially restored impaired RPE survival caused by JAM-C knockdown and increased RPE survival. We further show that JAM-C regulates VEGFR2 expression and, in turn, modulates p38 phosphorylation. Together, our data demonstrate that JAM-C plays an important role in maintaining VEGR2 expression to promote RPE cell survival under oxidative stress. Given the vital importance of RPE in the eye, approaches that can modulate JAM-C expression may have therapeutic values in treating diseases with impaired RPE survival.

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Overexpression of CERKL Protects Retinal Pigment Epithelium Mitochondria from Oxidative Stress Effects

Antioxidants ◽

10.3390/antiox10122018 ◽

2021 ◽

Vol 10 (12) ◽

pp. 2018

Author(s):

Rocío García-Arroyo ◽

Aleix Gavaldà-Navarro ◽

Francesc Villarroya ◽

Gemma Marfany ◽

Serena Mirra

Keyword(s):

Oxidative Stress ◽

Retinal Pigment Epithelium ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Stress Conditions ◽

Phenotypic Traits ◽

Mitochondrial Network ◽

Causative Gene ◽

Rpe Cells

The precise function of CERKL, a Retinitis Pigmentosa (RP) causative gene, is not yet fully understood. There is evidence that CERKL is involved in the regulation of autophagy, stress granules, and mitochondrial metabolism, and it is considered a gene that is resilient against oxidative stress in the retina. Mutations in most RP genes affect photoreceptors, but retinal pigment epithelium (RPE) cells may be also altered. Here, we aimed to analyze the effect of CERKL overexpression and depletion in vivo and in vitro, focusing on the state of the mitochondrial network under oxidative stress conditions. Our work indicates that the depletion of CERKL increases the vulnerability of RPE mitochondria, which show a shorter size and altered shape, particularly upon sodium arsenite treatment. CERKL-depleted cells have dysfunctional mitochondrial respiration particularly upon oxidative stress conditions. The overexpression of two human CERKL isoforms (558 aa and 419 aa), which display different protein domains, shows that a pool of CERKL localizes at mitochondria in RPE cells and that CERKL protects the mitochondrial network—both in size and shape—against oxidative stress. Our results support CERKL being a resilient gene that regulates the mitochondrial network in RPE as in retinal neurons and suggest that RPE cell alteration contributes to particular phenotypic traits in patients carrying CERKL mutations.

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Celastrol Protects RPE Cells from Oxidative Stress-Induced Cell Death via Activation of Nrf2 Signaling Pathway

Current Molecular Medicine ◽

10.2174/1566524019666190424131704 ◽

2019 ◽

Vol 19 (3) ◽

pp. 172-182 ◽

Cited By ~ 2

Author(s):

Yeqi Zhou ◽

Linbin Zhou ◽

Kewen Zhou ◽

Jingyue Zhang ◽

Fu Shang ◽

...

Keyword(s):

Oxidative Stress ◽

Signaling Pathway ◽

Western Blotting ◽

Retinal Pigment ◽

Age Related Macular Degeneration ◽

Protective Effects ◽

Rpe Cells ◽

Age Related ◽

Nrf2 Signaling Pathway ◽

Chemical Inhibition

Purpose: Oxidative stress to retinal pigment epithelial (RPE) cells and inflammation are closely related to the pathogenesis of age-related macular degeneration (AMD). Celastrol is a natural compound isolated from the root of Tripterygium wilfordii. Celastrol has been shown to have potent anti-inflammatory and anti-tumor effects in multiple disease models. The objective of this study was to test the anti-oxidative effects of celastrol in RPE cells and to investigate the underlying mechanisms. Methods: ARPE-19 cells were treated with hydrogen peroxide (H2O2) and menadione alone or in combination with celastrol. Cell viability and apoptosis were examined by CCK-8 and TUNEL assay, respectively. The expression of Nrf2 and its target genes, such as GCLM and HO-1 was determined by Western blotting. The knockdown of Nrf2 was done by transfecting ARPE-19 cells with lentivirus encoding shRNA against Nrf2. The knockdown efficiency was determined by real-time quantitative PCR and Western blotting. Results: Treatment of ARPE-19 cells with celastrol significantly attenuated the toxic effects of both H2O2 and menadione. Treatment with celastrol enhanced the expression of transcription factor Nrf2 and its targets, GCLM and HO-1. Knockdown of Nrf2 expression by shRNA partially abolished the protective effects of celastrol. Chemical inhibition of glutathione synthesis by L-buthionine-S,R-sulfoximine (BSO) completely abolished the protective effects of celastrol against H2O2 and menadione-induced damage. However, chemical inhibition of HO-1 activity by ZnPPIX did not reduce the protective effects of celastrol. Conclusion: This study provides evidence that treatment of RPE cells with celastrol shows potent protective effects against oxidative insults via activation of Nrf2 signaling pathway and upregulation of GCLM expression. This finding suggests that celastrol might be used as a potential therapeutic agent for oxidative stress-related eyes diseases, such as AMD.

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Phillyrin Mitigates Apoptosis and Oxidative Stress in Hydrogen Peroxide-Treated RPE Cells through Activation of the Nrf2 Signaling Pathway

Oxidative Medicine and Cellular Longevity ◽

10.1155/2020/2684672 ◽

2020 ◽

Vol 2020 ◽

pp. 1-16

Author(s):

Yuanyuan Du ◽

Longtai You ◽

Boran Ni ◽

Na Sai ◽

Wenping Wang ◽

...

Keyword(s):

Oxidative Stress ◽

Hydrogen Peroxide ◽

Signaling Pathway ◽

Retinal Pigment ◽

Age Related Macular Degeneration ◽

Heme Oxygenase 1 ◽

Related Factor ◽

Apoptosis Pathway ◽

Rpe Cells ◽

Nrf2 Signaling Pathway

Oxidative stress-induced dysfunction or apoptosis in retinal pigment epithelial (RPE) cells is an important cause of dry age-related macular degeneration (AMD). Although phillyrin has been shown to exert significant antioxidant effects, the underlying mechanism of action remains unclear. The purpose of this study was to investigate the protective effect of phillyrin on hydrogen peroxide- (H2O2-) induced oxidative stress damage in RPE cells and the potential mechanism involved. It was found that phillyrin significantly protected RPE cells from H2O2 cytotoxicity. Furthermore, phillyrin alleviated oxidative stress-induced apoptosis via inhibition of endogenous and exogenous apoptotic pathways. Compared with the H2O2-treated group, the expressions of cleaved caspase-3, cleaved caspase-9, cleaved polymerase (PARP), death receptor Fas, and cleaved caspase-8, as well as Bax/Bcl-2 ratio were decreased in RPE cells after the phillyrin intervention. In addition, phillyrin reversed the oxidative stress-induced reductions in superoxide dismutase (SOD) and glutathione (GSH) levels and annulled the elevations in reactive oxygen species (ROS) and malondialdehyde (MDA), thereby restoring oxidant-antioxidant homeostasis. Phillyrin treatment upregulated the expressions of cyclin E, cyclin-dependent kinase 2 (CDK2), and cyclin A and downregulated the expressions of p21 and p-p53, thereby reversing the G0/G1 cell cycle arrest in H2O2-treated RPE cells. Pretreatment with phillyrin also increased the expressions of nuclear factor-erythroid 2-related factor 2 (Nrf2), total Nrf2, heme oxygenase-1 (HO-1), and NAD(P)H: quinone oxidoreductases-1 (NQO-1) in RPE cells and inhibited the formation of Kelch-like ECH-associated protein 1 (Keap1)/Nrf2 protein complex. Thus, phillyrin effectively protected RPE cells from oxidative stress through activation of the Nrf2 signaling pathway and inhibition of the mitochondria-dependent apoptosis pathway.

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Melatonin Promotes In Vitro Maturation of Vitrified-Warmed Mouse Germinal Vesicle Oocytes, Potentially by Reducing Oxidative Stress through the Nrf2 Pathway

Animals ◽

10.3390/ani11082324 ◽

2021 ◽

Vol 11 (8) ◽

pp. 2324

Author(s):

Shichao Guo ◽

Jinyu Yang ◽

Jianpeng Qin ◽

Izhar Hyder Qazi ◽

Bo Pan ◽

...

Keyword(s):

Oxidative Stress ◽

Signaling Pathway ◽

In Vitro Maturation ◽

Germinal Vesicle ◽

Melatonin Receptors ◽

Development Rates ◽

Nrf2 Signaling Pathway ◽

Mouse Gv Oocytes ◽

Intracellular Oxidative Stress

Previously it was reported that melatonin could mitigate oxidative stress caused by oocyte cryopreservation; however, the underlying molecular mechanisms which cause this remain unclear. The objective was to explore whether melatonin could reduce oxidative stress during in vitro maturation of vitrified-warmed mouse germinal vesicle (GV) oocytes through the Nrf2 signaling pathway or its receptors. During in vitro maturation of vitrified-warmed mouse GV oocytes, there were decreases (p < 0.05) in the development rates of metaphase I (MI) oocytes and metaphase II (MII) and spindle morphology grades; increases (p < 0.05) in the reactive oxygen species (ROS) levels; and decreases (p < 0.05) in expressions of Nrf2 signaling pathway-related genes (Nrf2, SOD1) and proteins (Nrf2, HO-1). However, adding 10−7 mol/L melatonin to both the warming solution and maturation solutions improved (p < 0.05) these indicators. When the Nrf2 protein was specifically inhibited by Brusatol, melatonin did not increase development rates, spindle morphology grades, genes, or protein expressions, nor did it reduce vitrification-induced intracellular oxidative stress in GV oocytes during in vitro maturation. In addition, when melatonin receptors were inhibited by luzindole, the ability of melatonin to scavenge intracellular ROS was decreased, and the expressions of genes (Nrf2, SOD1) and proteins (Nrf2, HO-1) were not restored to control levels. Therefore, we concluded that 10−7 mol/L melatonin acted on the Nrf2 signaling pathway through its receptors to regulate the expression of genes (Nrf2, SOD1) and proteins (Nrf2, HO-1), and mitigate intracellular oxidative stress, thereby enhancing in vitro development of vitrified-warmed mouse GV oocytes.

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The cytoskeletal elements of human retinal pigment epithelium: in vitro and in vivo

Journal of Cell Science ◽

10.1242/jcs.91.2.303 ◽

1988 ◽

Vol 91 (2) ◽

pp. 303-312

Author(s):

N.M. McKechnie ◽

M. Boulton ◽

H.L. Robey ◽

F.J. Savage ◽

I. Grierson

Keyword(s):

Retinal Pigment Epithelium ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Cytokeratin 18 ◽

Rpe Cells ◽

Human Retinal Pigment Epithelium ◽

Cytoskeletal Elements

The cytoskeletal elements of normal (in situ) and cultured human retinal pigment epithelium (RPE) were studied by a variety of immunocytochemical techniques. Primary antibodies to vimentin and cytokeratins were used. Positive immunoreactivity for vimentin was obtained with in situ and cultured material. The pattern of reactivity obtained with antisera and monoclonals to cytokeratins was more complex. Cytokeratin immunoreactivity could be demonstrated in situ and in cultured cells. The pattern of cytokeratin expression was similar to that of simple or glandular epithelia. A monoclonal antibody that specifically recognizes cytokeratin 18 identified a population of cultured RPE cells that had particularly well-defined filamentous networks within their cytoplasm. Freshly isolated RPE was cytokeratin 18 negative by immunofluorescence, but upon culture cytokeratin 18 positive cells were identifiable. Cytokeratin 18 positive cells were identified in all RPE cultures (other than early primaries), regardless of passage number, age or sex of the donor. In post-confluent cultures cytokeratin 18 cells were identified growing over cytokeratin 18 negative cells, suggesting an association of cytokeratin 18 immunoreactivity with cell proliferation. Immunofluorescence studies of retinal scar tissue from two individuals revealed the presence of numerous cytokeratin 18 positive cells. These findings indicate that RPE cells can be identified by their cytokeratin immunoreactivity and that the overt expression of cytokeratin 18 may be associated with proliferation of human RPE both in vitro and in vivo.

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PGC-1α Protects RPE Cells of the Aging Retina against Oxidative Stress-Induced Degeneration through the Regulation of Senescence and Mitochondrial Quality Control. The Significance for AMD Pathogenesis

International Journal of Molecular Sciences ◽

10.3390/ijms19082317 ◽

2018 ◽

Vol 19 (8) ◽

pp. 2317 ◽

Cited By ~ 32

Author(s):

Kai Kaarniranta ◽

Jakub Kajdanek ◽

Jan Morawiec ◽

Elzbieta Pawlowska ◽

Janusz Blasiak

Keyword(s):

Oxidative Stress ◽

Cellular Senescence ◽

Mitochondrial Biogenesis ◽

Vision Loss ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Sirtuin 1 ◽

Age Related Macular Degeneration ◽

Rpe Cells ◽

Age Related

PGC-1α (peroxisome proliferator-activated receptor gamma coactivator 1-alpha) is a transcriptional coactivator of many genes involved in energy management and mitochondrial biogenesis. PGC-1α expression is associated with cellular senescence, organismal aging, and many age-related diseases, including AMD (age-related macular degeneration), an important global issue concerning vision loss. We and others have developed a model of AMD pathogenesis, in which stress-induced senescence of retinal pigment epithelium (RPE) cells leads to AMD-related pathological changes. PGC-1α can decrease oxidative stress, a key factor of AMD pathogenesis related to senescence, through upregulation of antioxidant enzymes and DNA damage response. PGC-1α is an important regulator of VEGF (vascular endothelial growth factor), which is targeted in the therapy of wet AMD, the most devastating form of AMD. Dysfunction of mitochondria induces cellular senescence associated with AMD pathogenesis. PGC-1α can improve mitochondrial biogenesis and negatively regulate senescence, although this function of PGC-1α in AMD needs further studies. Post-translational modifications of PGC-1α by AMPK (AMP kinase) and SIRT1 (sirtuin 1) are crucial for its activation and important in AMD pathogenesis.

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Mesenchymal Stem Cell Secretome Enhancement by Nicotinamide and Vasoactive Intestinal Peptide: A New Therapeutic Approach for Retinal Degenerative Diseases

Stem Cells International ◽

10.1155/2020/9463548 ◽

2020 ◽

Vol 2020 ◽

pp. 1-14

Author(s):

Maria L. Alonso-Alonso ◽

Girish K. Srivastava ◽

Ricardo Usategui-Martín ◽

Maria T. García-Gutierrez ◽

José Carlos Pastor ◽

...

Keyword(s):

Oxidative Stress ◽

Vasoactive Intestinal Peptide ◽

Cell Activation ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Glial Cell Activation ◽

Oxidative Stress Status ◽

Experimental Approaches ◽

Exogenous Factors

Mesenchymal stem cells (MSC) secrete neuroprotective molecules that may be useful as an alternative to cell transplantation itself. Our purpose was to develop different pharmaceutical compositions based on conditioned medium (CM) of adipose MSC (aMSC) stimulated by and/or combined with nicotinamide (NIC), vasoactive intestinal peptide (VIP), or both factors; and to evaluate in vitro their proliferative and neuroprotective potential. Nine pharmaceutical compositions were developed from 3 experimental approaches: (1) unstimulated aMSC-CM collected and combined with NIC, VIP, or both factors (NIC+VIP), referred to as the aMSC-CM combined composition; (2) aMSC-CM collected just after stimulation with the mentioned factors and containing them, referred to as the aMSC-CM stimulated-combined composition; and (3) aMSC-CM previously stimulated with the factors, referred to as the aMSC stimulated composition. The potential of the pharmaceutical compositions to increase cell proliferation under oxidative stress and neuroprotection were evaluated in vitro by using a subacute oxidative stress model of retinal pigment epithelium cells (line ARPE-19) and spontaneous degenerative neuroretina model. Results showed that oxidatively stressed ARPE-19 cells exposed to aMSC-CM stimulated and stimulated-combined with NIC or NIC+VIP tended to have better recovery from the oxidative stress status. Neuroretinal explants cultured with aMSC-CM stimulated-combined with NIC+VIP had better preservation of the neuroretinal morphology, mainly photoreceptors, and a lower degree of glial cell activation. In conclusion, aMSC-CM stimulated-combined with NIC+VIP contributed to improving the proliferative and neuroprotective properties of the aMSC secretome. Further studies are necessary to evaluate higher concentrations of the drugs and to characterize specifically the aMSC-secreted factors related to neuroprotection. However, this study supports the possibility of improving the potential of new effective pharmaceutical compositions based on the secretome of MSC plus exogenous factors or drugs without the need to inject cells into the eye, which can be very useful in retinal pathologies.

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A Higher Proportion of Eicosapentaenoic Acid (EPA) When Combined with Docosahexaenoic Acid (DHA) in Omega-3 Dietary Supplements Provides Higher Antioxidant Effects in Human Retinal Cells

Antioxidants ◽

10.3390/antiox9090828 ◽

2020 ◽

Vol 9 (9) ◽

pp. 828 ◽

Cited By ~ 1

Author(s):

Manuel Saenz de Viteri ◽

María Hernandez ◽

Valentina Bilbao-Malavé ◽

Patricia Fernandez-Robredo ◽

Jorge González-Zamora ◽

...

Keyword(s):

Docosahexaenoic Acid ◽

Eicosapentaenoic Acid ◽

Cell Viability ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Ethyl Esters ◽

Omega 3 ◽

Rpe Cells ◽

Antioxidant Effects

Retinal pigment epithelium (RPE) is a key regulator of retinal function and is directly related to the transport, delivery, and metabolism of long-chain n-3 polyunsaturated fatty acids (n3-PUFA), in the retina. Due to their functions and location, RPE cells are constantly exposed to oxidative stress. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have shown to have antioxidant effects by different mechanisms. For this reason, we designed an in vitro study to compare 10 formulations of DHA and EPA supplements from different origins and combined in different proportions, evaluating their effect on cell viability, cell proliferation, reactive oxygen species production, and cell migration using ARPE-19 cells. Furthermore, we assessed their ability to rescue RPE cells from the oxidative conditions seen in diabetic retinopathy. Our results showed that the different formulations of n3-PUFAs have a beneficial effect on cell viability and proliferation and are able to restore oxidative induced RPE damage. We observed that the n3-PUFA provided different results alone or combined in the same supplement. When combined, the best results were obtained in formulations that included a higher proportion of EPA than DHA. Moreover, n3-PUFA in the form of ethyl-esters had a worse performance when compared with triglycerides or phospholipid based formulations.

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Ganoderic Acid D Protects Human Amniotic Mesenchymal Stem Cells against Oxidative Stress-Induced Senescence through the PERK/NRF2 Signaling Pathway

Oxidative Medicine and Cellular Longevity ◽

10.1155/2020/8291413 ◽

2020 ◽

Vol 2020 ◽

pp. 1-18

Author(s):

Yan Xu ◽

Huan Yuan ◽

Yi Luo ◽

Yu-Jie Zhao ◽

Jian-Hui Xiao

Keyword(s):

Oxidative Stress ◽

Stem Cells ◽

Signaling Pathway ◽

Adult Stem Cells ◽

Telomerase Activity ◽

Related Factor ◽

Dependent Manner ◽

Ganoderic Acid ◽

Nrf2 Signaling Pathway

Aging is an important risk factor in the occurrence of many chronic diseases. Senescence and exhaustion of adult stem cells are considered as a hallmark of aging in organisms. In this study, a senescent human amniotic mesenchymal stem cell (hAMSC) model subjected to oxidative stress was established in vitro using hydrogen peroxide. We investigated the effects of ganoderic acid D (GA-D), a natural triterpenoid compound produced from Ganoderma lucidum, on hAMSC senescence. GA-D significantly inhibited β-galactosidase (a senescence-associated marker) formation, in a dose-dependent manner, with doses ranging from 0.1 μM to 10 μM, without inducing cytotoxic side-effects. Furthermore, GA-D markedly inhibited the generation of reactive oxygen species (ROS) and the expression of p21 and p16 proteins, relieved the cell cycle arrest, and enhanced telomerase activity in senescent hAMSCs. Furthermore, GA-D upregulated the expression of phosphorylated protein kinase R- (PKR-) like endoplasmic reticulum kinase (PERK), peroxidase III (PRDX3), and nuclear factor-erythroid 2-related factor (NRF2) and promoted intranuclear transfer of NRF2 in senescent cells. The PERK inhibitor GSK2656157 and/or the NRF2 inhibitor ML385 suppressed the PERK/NRF2 signaling, which was activated by GA-D. They induced a rebound for the generation of ROS and β-galactosidase-positive cells and attenuated the differentiation capacity. These findings suggest that GA-D retards hAMSC senescence through activation of the PERK/NRF2 signaling pathway and may be a promising candidate for the discovery of antiaging agents.

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