Cobalt oxide nanoparticles disrupt cellular homeostasis through oxidative genomic instability and mitochondrial stress, driving necrosis in non-small lung cancer cells

Abstract

Non–small cell lung cancer (NSCLC) remains one of the most aggressive and treatment-resistant malignancies worldwide, with current therapies often limited by poor tumor specificity, resistance, and systemic toxicity. Nanoparticle-based therapeutics offer a promising strategy to overcome these challenges. Cobalt oxide nanoparticles (Co3O4NPs), owing to their distinctive redox properties and capacity to generate reactive oxygen species (ROS), have emerged as potential anticancer agents; however, their intrinsic effects against NSCLC cells and the underlying mechanisms remain poorly understood. This study therefore investigates the novel cytotoxic and pro-apoptotic effects of pristine Co3O4NPs in non-small A549 lung cancer cells using an integrated mechanistic approach. Cell viability was measured with the sulforhodamine B (SRB) assay, while genomic DNA damage, ROS generation, mitochondrial membrane potential, and apoptosis induction were evaluated using alkaline comet assay, 2′,7′-dichlorofluorescein diacetate, rhodamine-123 dye, and flow cytometry, respectively. Mechanistic insights were further supported by quantitative real-time PCR analysis of apoptosis- and mitochondria-related genes (p53, ND3, and Bcl-2). A 72-hour exposure to Co3O4NPs various concentrations significantly reduced A549 cell viability in a concentration-dependent manner with a markedly low IC50 value of 34.17 µg/ml. Moreover, treating A549 cells with the IC50 concentration (34.17 µg/ml) of Co3O4NPs caused excessive ROS production, severe genomic DNA damage, and dramatic mitochondrial depolarization. Gene expression analysis revealed remarkable upregulation of pro-apoptotic p53 and mitochondrial ND3 gene expression, alongside significant downregulation of anti-apoptotic Bcl-2 gene expression. Despite p53 activation the pronounced mitochondrial dysfunction and bioenergetic collapse favored necrotic cell death over classical apoptosis. Flow cytometric analysis confirmed this shift, showing a substantial increase in necrotic A549 cells alongside a marked reduction in apoptotic A549 populations following Co3O4NPs exposure. These findings collectively provide integrated evidence that pristine Co₃O₄NPs induce oxidative stress, genomic DNA damage, mitochondrial dysfunction, and cell death in A549 NSCLC cells. The predominance of necrotic cell death under the conditions examined suggests that severe oxidative and mitochondrial stress contributes to the observed cytotoxic response. Further studies using NSCLC models with different p53 statuses, appropriate normal-control cells, and in vivo systems are required to establish the selectivity, reproducibility, and broader biological relevance of these effects.