Co3O4 nanoparticles (NPs) are one kind of the important nanomaterials that have the application potential in catalyst, electrochromic devices, sensors, etc. However, their biological effect remains to be detailed. In this study, we investigated the effect of the as-synthesized Co3O4 NPs (15-30 nm) on the growth of mammalian cells, and found that the NPs severely inhibited cell growth at the sublethal concentrations from 12.5 to 200 mg/L Interestingly, the NPs did not cause obvious cell death and ROS accumulation, indicating that their inhibitory effect was not attributed to both apoptosis- or necrosis-related cell death and ROS accumulation. Transcription profiling analysis revealed that the NPs caused remarkable down regulation of the genes involved in mitochondria! functions. Transmission electron microscopy (TEM) and biochemical analysis further showed that the NPs might interact with the mitochondria, impairing the mitochondrial membrane potential (MMP) and ATP production. This study uncovers a mitochondrial respiratory chain-related and ROS-independent toxicity mechanism of Co3O4 NPs in eukaryotic cells. (C) 2018 Elsevier Inc. All rights reserved.