ZnO nanoparticles-associated mitochondrial stress-induced apoptosis and G2/M arrest in HaCaT cells: a mechanistic approach

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Abstract
Zinc oxide nanoparticles (ZnO NPs) with their wide range of consumer applications in day-to-day life received great attention to evaluate their effects in humans. This study has been attempted to elucidate the DNA damage response mechanism in a dermal model exposed to ZnO NPs through Ataxia Telangiectasia Mutated (ATM)-mediated ChK1-dependent G2/M arrest. Further, viability parameters and mechanism involved in the cell death with special reference to the consequences arising due to DNA damage were explored. Our study showed that ZnO NPs at concentrations 5 and 10 µg/ml induced significant cytotoxic effect in skin cell line. Moreover, the results confirmed generation of reactive oxygen species (ROS) induces the cell death by genotoxic insult, leading to mitochondrial membrane depolarisation and cell cycle arrest. Subsequently, ZnO NPs treatment created DNA damage as confirmed via Comet assay (increase in olive tail moment), micronucleus assay (increase in micronucleus formation), double-strand breaks (increase in ATM and Ataxia Telangiectasia and Rad3 related (ATR) expression), DNA fragmentation and cell cycle (G2/M arrest) studies. Finally, marker proteins analysis concluded the mechanistic approach by demonstrating the key marker expressions HMOX1 and HSP60 (for oxidative stress), cytochrome c, APAF1, BAX, Caspase 9, Caspase 3 and decrease in BCL2 (for activating apoptotic pathway), pATM, ATR and γH2AX (for double-strand breaks), DNA-PK (involved in DNA repair) and decrease in cell cycle regulators. In together, our data revealed the mechanism of ROS generation that triggers apoptosis and DNA damage in HaCaT cell lines exposed to ZnO NPs.
Funding Information
  • Department of Biotechnology, Government of India (BT/PR10414/PFN/20/961/2014)
  • Nanosensors for the Detection of Food Adulterants and Contaminants (EMR/2016/005286)
  • Gujarat Institute for Chemical Technology

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