Elsevier

Chemico-Biological Interactions

Volume 300, 25 February 2019, Pages 151-158
Chemico-Biological Interactions

SIRT6 protects retinal ganglion cells against hydrogen peroxide-induced apoptosis and oxidative stress by promoting Nrf2/ARE signaling via inhibition of Bach1

https://doi.org/10.1016/j.cbi.2019.01.018Get rights and content

Highlights

  • SIRT6 expression was downregulated in RGCs with H2O2 treatment.

  • SIRT6 alleviates H2O2-induced apoptosis and oxidative stress.

  • SIRT6 protects RGCs from H2O2-induced injury by Nrf2/ARE signaling.

  • SIRT6 increases Nrf2/ARE signaling by targeting Bach1.

Abstract

Oxidative stress-induced damage of retinal ganglion cells (RGCs) is a major contributor to retinal degenerative diseases, such as glaucoma. Sirtuin 6 (SIRT6) has emerged as a cytoprotective protein against various insults. However, whether SIRT6 exerts a protective effect against oxidative stress-damaged RGCs remains unknown. In this study, we aimed to investigate the potential role and regulatory mechanism of SIRT6 in hydrogen peroxide (H2O2)-induced oxidative damage of RGCs in vitro. We found that SIRT6 expression was significantly downregulated in RGCs with H2O2 treatment. Functional experiments showed that overexpression of SIRT6 improved survival and reduced apoptosis and the production of reactive oxygen species (ROS) in H2O2-treated RGCs. In contrast, SIRT6 knockdown had the opposite effect. Moreover, we found that SIRT6 overexpression promoted the nuclear accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and increased the activity of antioxidant response element (ARE). In addition, we found that the promotional effect of SIRT6 on Nrf2/ARE signaling was associated with inhibition of BTB and CNC homology 1 (Bach1), an inhibitor of Nrf2. However, overexpression of Bach1 or inhibition of Nrf2/ARE signaling partially reversed the SIRT6-mediated protective effect. Taken together, these results demonstrate that SIRT6 protects RGCs from oxidative stress-induced damage by promoting the activation of Nrf2/ARE signaling via inhibition of Bach1, suggesting a potential role of SIRT6 in retinal degenerative diseases.

Introduction

Oxidative stress-induced death of retinal ganglion cells (RGCs) is a major pathomechanism of multiple retinal degenerative diseases, such as glaucoma [1]. Glaucoma is characterized by excessive RGC death, which leads to irreversible vision loss [2]. Under pathologic conditions, reactive oxygen species (ROS) are generated at high levels causing DNA damage, lipid peroxidation, and calcium overload, which eventually result in RGC death and apoptosis [3]. However, the precise molecular mechanism underlying oxidative stress-induced damage of RGCs remains largely unknown. Therefore, it is essential to identify new regulators involved in oxidative stress-induced damage of RGCs, which may aid the development of promising targeted therapies for retinal degenerative diseases.

Mammalian sirtuins, the homologues of the yeast silent information regulator 2, are a group of NAD+-dependent histone deacetylases consisting of seven members (SIRT1-SIRT7) [4]. Among these sirtuins, SIRT6 has attracted particular interest due to its importance in various physiological and pathological processes [5]. SIRT6 is mainly a nuclear protein that preferentially deacetylates histone H3 lysine 9 and lysine 56 and regulates the transcriptional activities of multiple transcription factors, including p53, nuclear factor-κB and hypoxia inducible factor-1α [[5], [6], [7]]. Therefore, SIRT6 is a multifunctional protein involved in diverse cellular processes, including cell proliferation, apoptosis, the cell cycle, and differentiation [8]. Notably, the dysregulation of SIRT6 has been implicated in various diseases, including cancer, neurological disorders, and cardiovascular diseases [[9], [10], [11]]. Therefore, SIRT6 has emerged as a promising therapeutic target for treating related diseases.

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that regulates cellular defenses against oxidative stress [12]. Under the stimuli of oxidative stress, Nrf2 translocates to the nucleus from the cytoplasm and binds to antioxidant response element (ARE) to activate the production of antioxidants [13]. However, targeting between Nrf2 and ARE sequences is negatively regulated by BTB and CNC homology 1 (Bach1), because Bach1 competitively binds to ARE-like sequences [14]. Therefore, Bach1 functions as an inhibitor of Nrf2/ARE signaling. Nrf2/ARE has been suggested as a therapeutic target for overcoming oxidative stress-induced damage in various diseases [[15], [16], [17]]. Interestingly, Nrf2/ARE signaling is involved in ocular neurodegenerative diseases through regulation of oxidative stress-induced damage of RGCs [18]. Therefore, a better understanding of the regulatory mechanism of Nrf2/ARE signaling in oxidative stress-induced damage of RGCs will aid the development of novel strategies for RGC protection.

SIRT6 has been reported to confer a cytoprotective effect against oxidative stress-induced cellular damage of various cell types [11,[19], [20], [21]]. However, whether SIRT6 is involved in oxidative stress-induced damage of RGCs remains unclear. In the present study, we aimed to investigate the potential role and regulatory mechanism of SIRT6 in hydrogen peroxide (H2O2)-induced oxidative damage of RGCs in vitro.

Section snippets

Cell culture

Primary RGCs were isolated from rat retinal cells according to a previously described method [22]. In brief, retinas isolated from neonatal SD rats were digested by papain and collagenase in Eagle's balanced salt solution for 30 min at 37 °C. Then, the retinas were triturated sequentially by a narrow-bore Pasteur pipette. Therefore, the yielded single cells were collected after centrifugation and resuspended in phosphate-buffered saline (PBS) containing 0.1% bovine serum albumin (BSA). Cells

SIRT6 expression is decreased in H2O2-treated RGCs

To explore the potential role of SIRT6 in regulating oxidative damage in RGCs, we treated RGCs with H2O2 to establish oxidative damage and then detected the change in expression of SIRT6 in response to oxidative stress. Our results showed reduced SIRT6 expression in RGCs after H2O2 treatment (Fig. 1A and B), indicating that SIRT6 may play a critical role in regulating oxidative damage in RGCs.

SIRT6 overexpression reduces H2O2-induced apoptosis and ROS production in RGCs

To investigate whether SIRT6 exerts a protective effect against H2O2-induced oxidative damage, we

Discussion

In this study, we reported an important role of SIRT6 in regulating oxidative stress-induced damage of RGCs. Our results showed that SIRT6 overexpression repressed apoptosis and ROS production in RGCs induced by H2O2 treatment. We elucidated that the molecular mechanism underlying the SIRT6-mediated protective effect is associated with its promotional effect on activation of Nrf2/ARE signaling via inhibition of Bach1 (Fig. 6F). Our study suggests that SIRT6 may serve as a potential target for

Conflicts of interest

The authors declare that they have no conflict of interest.

References (46)

  • T. Masuda et al.

    Retinal diseases associated with oxidative stress and the effects of a free radical scavenger (Edaravone)

    Oxid. Med. Cell Longev.

    (2017)
  • R.H. Houtkooper et al.

    Sirtuins as regulators of metabolism and healthspan

    Nat. Rev. Mol. Cell Biol.

    (2012)
  • A.A. Gertler et al.

    SIRT6, a protein with many faces

    Biogerontology

    (2013)
  • E. Michishita et al.

    Evolutionarily conserved and nonconserved cellular localizations and functions of human SIRT proteins

    Mol. Biol. Cell

    (2005)
  • J.M. Beauharnois et al.

    Sirtuin 6: a review of biological effects and potential therapeutic properties

    Mol. Biosyst.

    (2013)
  • B. Lerrer et al.

    The complex role of SIRT6 in carcinogenesis

    Carcinogenesis

    (2016)
  • X.X. Wang et al.

    SIRT6 protects cardiomyocytes against ischemia/reperfusion injury by augmenting FoxO3alpha-dependent antioxidant defense mechanisms

    Basic Res. Cardiol.

    (2016)
  • K.W. Kang et al.

    Molecular mechanism of nrf2 activation by oxidative stress

    Antioxid. Redox Signal

    (2005)
  • J. Sun et al.

    Heme regulates the dynamic exchange of Bach1 and NF-E2-related factors in the Maf transcription factor network

    Proc. Natl. Acad. Sci. U. S. A.

    (2004)
  • H.Y. Cho et al.

    Nrf2 defends the lung from oxidative stress

    Antioxid. Redox Signal

    (2006)
  • W.S. Jeong et al.

    Nrf2: a potential molecular target for cancer chemoprevention by natural compounds

    Antioxid. Redox Signal

    (2006)
  • S. Magesh et al.

    Small molecule modulators of Keap1-Nrf2-ARE pathway as potential preventive and therapeutic agents

    Med. Res. Rev.

    (2012)
  • X.F. Liu et al.

    The Nrf2 signaling in retinal ganglion cells under oxidative stress in ocular neurodegenerative diseases

    Int. J. Biol. Sci.

    (2018)
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