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Nickel oxide (NiO) nanoparticles trigger caspase- and mitochondriadependent apoptosis in yeast Saccharomyces cerevisiae Cátia A. Sousa, Helena MVM Soares, and Eduardo V. Soares Chem. Res. Toxicol., Just Accepted Manuscript • DOI: 10.1021/acs.chemrestox.8b00265 • Publication Date (Web): 18 Jan 2019 Downloaded from http://pubs.acs.org on January 20, 2019
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Chemical Research in Toxicology
Nickel oxide (NiO) nanoparticles trigger caspase- and mitochondria-dependent apoptosis in yeast Saccharomyces cerevisiae
Cátia A. Sousa†,‡,§, Helena M.V.M. Soares§ and Eduardo V. Soares†,‡*
†Bioengineering Laboratory-CIETI,
Chemical Engineering Department, ISEP-School of Engineering
of Polytechnic Institute of Porto, Rua Dr António Bernardino de Almeida, 431, 4249-015 Porto, Portugal ‡CEB-Centre
of Biological Engineering, University of Minho, 4710-057 Braga, Portugal
§REQUIMTE/LAQV,
Departamento de Engenharia Química, Faculdade de Engenharia,
Universidade do Porto, rua Dr. Roberto Frias, 4200-465, Porto, Portugal
Keywords: apoptosis, caspase, mitochondrial dysfunction; regulated cell death; Saccharomyces cerevisiae; toxicity.
*Author
for correspondence:
Eduardo V. Soares e-mail:
[email protected]; Tel: 351-22-8340500
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Abstract The expansion of industrial use of nickel oxide (NiO) nanoparticles (NPs) raises concerns about their potential adverse effects. The present work aimed to investigate the mechanisms of toxicity induced by NiO NPs, using the yeast Saccharomyces cerevisiae, as cell model. Yeast cells exposed to NiO NPs exhibited typical hallmarks of regulated cell death (RCD) by apoptosis [loss of cell proliferation capacity (cell viability), exposure of phosphatidylserine at the outer cytoplasmic membrane leaflet, nuclear chromatin condensation and DNA damage] in a process which required de novo protein synthesis. The execution of yeast cell death induced by NiO NPs is Yca1p metacaspase-dependent. NiO NPs also induced the reduction of the mitochondrial membrane potential and the increase of the frequency of respiratory deficient mutants, which supports the involvement of mitochondria in the cell death process. Cells deficient in the apoptosis-inducing factor (aif1Δ) displayed higher tolerance to NiO NPs, which reinforces the involvement of mitochondria in the RCD by apoptosis. In resume, this study shows that NiO NPs induce caspaseand mitochondria-dependent apoptosis in yeast. The presented results warn to the possible harmful effects associated with the use of NiO NPs.
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Chemical Research in Toxicology
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INTRODUCTION Due to their unique physiochemical properties and reactivity, the metal oxide nanoparticles (NPs) are increasingly manufactured in the last decade. Nickel oxide (NiO) NPs are used for multiple applications, such as catalysts, memory cells, electrode materials in multilayer ceramic capacitors, diesel-fuel additives and pigments for ceramics and glasses.1 As the nanotechnology industry expands, the exposure to NiO NPs emerges as a significant occupational hazard. In addition, a deeper understanding of the potential impact of NiO NPs on the environment and human health is required. Adverse effects as consequence of the exposure to NiO NPs, such as, cytotoxicity, inflammation and genotoxicity in human pulmonary cells, has been reported.2-5 The International Agency for Research on Cancer classed nickel compounds as group 1 (carcinogenic to humans) and metallic nickel was classed as group 2B (possible carcinogenic to humans).6 Cell death can be classified into a not programed “accidental cell death” (ACD) and “regulated cell death” (RCD). The first subtype of cell death (ACD) occurs as a consequence of the exposure of cells to severe physical, chemical or mechanical stimuli; cells demise in an uncontrollable manner as a result of their immediate loss of structural integrity. RCD involves a genetically encoded machinery and can result from multiple signalling pathways, including apoptosis. The designation “programed cell death” (PCD) is used to refer RCD that is executed in specific physiological scenarios.7,
8
Many human diseases, such as several neurological disorders (for example,
Huntington’s, Parkinson’s and Alzheimer’s disease) and cancer are associated with the dysregulation of cell death pathways.9 In this context, it has been shown that the exposure to NiO NPs induce apoptosis in different cell lines, such as in human bronchial epithelial (BEAS-2B)
10,
lung epithelial (NCIH460) 11, liver (HepG2) 12, 13 and neuronal cells (SH-SY5Y) 14. The yeast S. cerevisiae is a relevant eukaryotic cell model in different fields, such as biochemistry, molecular and cellular biology. In the last decade, this unicellular microorganism has been used in the study of RCD.15,
16
This yeast displays some similarities (orthologues) with mammalian
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Chemical Research in Toxicology
(including human) cells, which makes it an effective cell model to human diseases.17 In a previous work, it was found that the exposure of yeast cells of S. cerevisiae to NiO NPs induced the loss of metabolic activity and cell viability in a dose-dependent manner.18 In a subsequent work, it was observed that the loss of cell viability induced by NiO could be attributed to the intracellular generation of reactive oxygen species (ROS) and glutathione depletion.19 It is described that oxidative stress is a common denominator of RCD initiation.20 The present work aimed to characterize the cell death observed in S. cerevisiae upon exposure to NiO NPs. For this purpose, yeast cell death induced by NiO NPs was assessed by monitoring the loss of plasma membrane integrity and cell viability. Subsequently, the presence of typical morphological and biochemical features of apoptotic cell death was examined in yeast cells exposed to a cytotoxic concentration of NiO NPs. In addition, the dependence of known apoptotic regulators/executors (Yca1p and Aif1p) in the RCD NiO NPs-induced was evaluated.
EXPERIMENTAL PROCEDURES Preparation of nickel oxide nanoparticles suspensions Commercial nickel oxide (NiO) nanoparticles (NPs) (Catalogue number 637130, with a particle size
