An Integrated Imaging Approach to the Study of Oxidative Stress Generation by Mitochondrial Dysfunction in Living Cells

Cheng, Wan Yun; Tong, Haiyan; Miller, Evan W.; Chang, Christopher J.; Remington, J.; Zucker, Robert M.; Bromberg, Philip A.; Samet, James M.; Höfer, Thomas

doi:10.1289/ehp.0901811

Cited by 50 publications

(32 citation statements)

References 45 publications

(66 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Excessive ROS production and accumulation damage the structure of cell membranes and the biological functions of lipids, proteins and DNA, and ultimately cause the initiation of cell apoptosis (Ma, 2012). Additionally, hydrogen peroxide (H 2 O 2 )-induced ROS production and release can cause a series of oxidative stress responses, which in turn lead to mitochondrial dysfunction, cell damage, and death (Cheng et al, 2010). Apoptosis induced by mitochondrial dysfunction is regulated by the anti-apoptotic protein, Bcl-2, and the pro-apoptotic protein Bax (D'orsi et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

α-Cyperone Attenuates H2O2-Induced Oxidative Stress and Apoptosis in SH-SY5Y Cells via Activation of Nrf2

Huang

Liu

et al. 2020

Front. Pharmacol.

View full text Add to dashboard Cite

a-Cyperone, extracted from Cyperus rotundus, has been reported to inhibit microgliamediated neuroinflammation. Oxidative stress and apoptosis play crucial roles in the course of Parkinson's disease (PD). PD is a common neurodegenerative disease characterized by selective death of dopaminergic neurons. This study was designed to investigate the neuroprotective effects of a-cyperone against hydrogen peroxide (H 2 O 2)induced oxidative stress and apoptosis in dopaminergic neuronal SH-SY5Y cells. Neurotoxicity was assessed by MTT assay and the measurement of lactic dehydrogenase (LDH) release. The level of reactive oxygen species (ROS) was measured by dichlorodihydrofluorescin diacetate (DCFH-DA) staining. The apoptosis of SH-SY5Y cells was evaluated by annexin-V-FITC staining. The translocation of NF-E2related factor 2 (Nrf2) was determined by western blot and immunofluorescence staining. Western blot analysis was conducted to determine the expression level of cleavedcaspase-3, the pro-apoptotic factor Bax, and the anti-apoptotic factor, Bcl-2. The results showed that a-cyperone substantially decreased H 2 O 2-induced death, release of LDH, and the production of ROS in SH-SY5Y cells. In addition, we found that a-cyperone attenuated H 2 O 2-induced cellular apoptosis. Moreover, a-cyperone remarkably reduced the expression of cleaved-caspase-3 and Bax, and upregulated Bcl-2. Furthermore, acyperone enhanced the nuclear translocation of Nrf2. Pretreatment with brusatol (BT, an Nrf2 inhibitor) attenuated a-cyperone-mediated suppression of ROS, cleaved-caspase-3, and Bax, as well as a-cyperone-induced Bcl-2 upregulation in H 2 O 2-treated SH-SY5Y cells. a-cyperone neuroprotection required Nrf2 activation. In conclusion, a-cyperone attenuated H 2 O 2-induced oxidative stress and apoptosis in SH-SY5Y cells via the activation of Nrf2, suggesting the potential of this compound in the prevention and treatment of PD.

show abstract

Section: Introductionmentioning

confidence: 99%

α-Cyperone Attenuates H2O2-Induced Oxidative Stress and Apoptosis in SH-SY5Y Cells via Activation of Nrf2

Huang

Liu

et al. 2020

Front. Pharmacol.

View full text Add to dashboard Cite

show abstract

“…Mitochondrial oxidant production, frequently associated with increased oxidation of mitochondrial glutathione, has been implicated as a contributing factor in the cellular response to xenobiotics (Cheng et al 2010, 2012; Hanson et al 2004). Therefore, we next used mitochondrially targeted roGFP2 (roGFP2-mito) to assess the impact of O 3 exposure on the mitochondrial E GSH of BEAS-2B cells.…”

Section: Resultsmentioning

confidence: 99%

Monitoring Intracellular Redox Changes in Ozone-Exposed Airway Epithelial Cells

Gibbs-Flournoy¹,

Simmons²,

Bromberg³

et al. 2013

Environ Health Perspect

Self Cite

View full text Add to dashboard Cite

Background: The toxicity of many xenobiotic compounds is believed to involve oxidative injury to cells. Direct assessment of mechanistic events involved in xenobiotic-induced oxidative stress is not easily achievable. Development of genetically encoded probes designed for monitoring intracellular redox changes represents a methodological advance with potential applications in toxicological studies.Objective: We tested the utility of redox-sensitive green fluorescent protein (roGFP)–based redox sensors for monitoring real-time intracellular redox changes induced by xenobiotics in toxicological studies.Methods: roGFP2, a reporter of the glutathione redox potential (EGSH), was used to monitor EGSH in cultured human airway epithelial cells (BEAS-2B cells) undergoing exposure to 0.15–1.0 ppm ozone (O3). Cells were imaged in real time using a custom-built O3 exposure system coupled to a confocal microscope.Results: O3 exposure induced a dose- and time-dependent increase of the cytosolic EGSH. Additional experiments confirmed that roGFP2 is not directly oxidized, but properly equilibrates with the glutathione redox couple: Inhibition of endogenous glutaredoxin 1 (Grx1) disrupted roGFP2 responses to O3, and a Grx1-roGFP2 fusion protein responded more rapidly to O3 exposure. Selenite-induced up-regulation of GPx (glutathione peroxidase) expression–enhanced roGFP2 responsiveness to O3, suggesting that (hydro)peroxides are intermediates linking O3 exposure to glutathione oxidation.Conclusion: Exposure to O3 induces a profound increase in the cytosolic EGSH of airway epithelial cells that is indicative of an oxidant-dependent impairment of glutathione redox homeostasis. These studies demonstrate the utility of using genetically encoded redox reporters in making reliable assessments of cells undergoing exposure to xenobiotics with strong oxidizing properties.

show abstract

“…was measured with a JC-1 Mitochondrial Membrane Potential Assay Kit (Yeasen, Shanghai, China) according to the manufacturer's instructions, as described previously 57 . In brief, HL-60 cells were treated with 0.1% DMSO, acetylcysteine (5 mM), celastrol (0.5 μM) and celastrol (0.5 μM) + acetylcysteine (5 mM) for 24 h. Then, cells were incubated with JC-1 for 20 min in the dark at 37 °C.…”

Section: Measurement Of the Mitochondrial Membrane Potential The Mitmentioning

confidence: 99%

Metabolomics Reveals that Cysteine Metabolism Plays a Role in Celastrol-Induced Mitochondrial Apoptosis in HL-60 and NB-4 Cells

Chen

Yang

et al. 2020

Sci Rep

View full text Add to dashboard Cite

Recently, celastrol has shown great potential for inducing apoptosis in acute myeloid leukemia cells, especially acute promyelocytic leukaemia cells. However, the mechanism is poorly understood. Metabolomics provides an overall understanding of metabolic mechanisms to illustrate celastrol's mechanism of action. We treated both nude mice bearing HL-60 cell xenografts in vivo and HL-60 cells as well as NB-4 cells in vitro with celastrol. Ultra-performance liquid chromatography coupled with mass spectrometry was used for metabolomics analysis of HL-60 cells in vivo and for targeted L-cysteine analysis in HL-60 and NB-4 cells in vitro. Flow cytometric analysis was performed to assess mitochondrial membrane potential, reactive oxygen species and apoptosis. Western blotting was conducted to detect the p53, Bax, cleaved caspase 9 and cleaved caspase 3 proteins. Celastrol inhibited tumour growth, induced apoptosis, and upregulated pro-apoptotic proteins in the xenograft tumour mouse model. Metabolomics showed that cysteine metabolism was the key metabolic alteration after celastrol treatment in HL-60 cells in vivo. Celastrol decreased L-cysteine in HL-60 cells. Acetylcysteine supplementation reversed reactive oxygen species accumulation and apoptosis induced by celastrol and reversed the dramatic decrease in the mitochondrial membrane potential and upregulation of pro-apoptotic proteins in HL-60 cells. In NB-4 cells, celastrol decreased L-cysteine, and acetylcysteine reversed celastrol-induced reactive oxygen species accumulation and apoptosis. We are the first to identify the involvement of a cysteine metabolism/reactive oxygen species/p53/Bax/caspase 9/ caspase 3 pathway in celastrol-triggered mitochondrial apoptosis in HL-60 and NB-4 cells, providing a novel underlying mechanism through which celastrol could be used to treat acute myeloid leukaemia, especially acute promyelocytic leukaemia.Acute leukemia is the most common malignant hematological malignancy, of which acute myeloid leukemia (AML) accounts for over 80% of all acute leukemias in individuals aged >18 years 1 . Chemotherapy is the main treatment of AML. There are still problems such as drug resistance and relapse of leukemia after drug withdrawal, which seriously affect the quality of life and long-term survival of patients. Therefore, it is of great significance to find effective new antineoplastic drugs and explore new therapeutic schemes.Acute promyelocytic leukaemia (APL), a unique subtype of acute myeloid leukaemia (AML), is a devastating disease characterized by neoplastic proliferation of cells in the bone marrow that have a promyelocytic phenotype and fail to terminally differentiate 2 . Unlike other forms of AML, APL can cause diffuse intravascular coagulopathy

show abstract

An Integrated Imaging Approach to the Study of Oxidative Stress Generation by Mitochondrial Dysfunction in Living Cells

Cited by 50 publications

References 45 publications

α-Cyperone Attenuates H2O2-Induced Oxidative Stress and Apoptosis in SH-SY5Y Cells via Activation of Nrf2

α-Cyperone Attenuates H2O2-Induced Oxidative Stress and Apoptosis in SH-SY5Y Cells via Activation of Nrf2

Monitoring Intracellular Redox Changes in Ozone-Exposed Airway Epithelial Cells

Metabolomics Reveals that Cysteine Metabolism Plays a Role in Celastrol-Induced Mitochondrial Apoptosis in HL-60 and NB-4 Cells

Contact Info

Product

Resources

About