High-throughput transcriptomics: An insight on the pathways affected in HepG2 cells exposed to nickel oxide nanoparticles

Nickel oxide nanoparticles (NiO-NPs) have been used in several consumer goods, reported to demonstrate the hepatotoxic effects in vitro and in vivo test models. Nonetheless the molecular mechanism of hepatotoxicity is still missing. Hence, a toxicogenomic approach integrating microscopic techniques and high-throughput RNA sequencing (RNA-Seq) was applied to reveal hepatotoxicity in human hepatocellular carcinoma cells (HepG2). NiO-NPs induced a concentration dependent (5-100 μg/ml) cytotoxicity, with a No observed effect level (NOEL) of 5 μg/ml. Hypoxia-inducible transcription factor-1α (HIF-1α) and miR-210 microRNA were upregulated at 25 and 100 μg/ml, while significant alteration on transcriptome at mRNA and pathway level was observed at non-toxic level of NiO-NPs treatment. The treated cells also showed activation of glycolysis, glutathione, lysosomes and autophagy pathways by a pathway-driven analysis. Flow cytometric analysis affirmed the elevation in nitric oxide (NO), Ca⁺⁺ influx, esterase, and disruption of mitochondrial membrane potential (ΔΨm). Cell cycle dysregulation was affirmed by the appearance of 30.5% subG1 apoptotic peak in NiO-NPs (100 μg/ml) treated cells. The molecular responses were consistent with the microscopic observation that NiO-NPs induced subcellular alterations in HepG2 cells. We conclude that hypoxia stress played a pivotal role in NiO-NPs induced hepatoxicity in HepG2 cells. Concentration dependent effects on transcriptomics specify a powerful tool to evaluate the molecular mechanisms of nanoparticle induced cytotoxicity. Overall our study unequivocally affirmed the transcriptomic alterations in human cells, consequently the prevalent usage of NiO-NPs should be given subtle consideration owing to its effects on biological processes.