Graphene and graphene-based nanomaterials display novel and beneficial chemical, electrical, mechanical, and optical characteristics, which endow these nanomaterials with promising applications in a wide spectrum of areas such as electronics and biomedicine. However, its toxicity on health remains unknown and is of great concern. In the present study, we demonstrated that graphene oxide (GO) induced necrotic cell death to macrophages. This toxicity is mediated by activation of toll-like receptor 4 (TLR4) signaling and subsequently in part via autocrine TNF-α production. Inhibition of TLR4 signaling with a selective inhibitor prevented cell death nearly completely. Furthermore, TLR4-deficient bone marrow-derived macrophages were resistant to GO-triggered necrosis. Similarly, GO did not induce necrosis of HEK293T/TLR4-null cells. Macrophagic cell death upon GO treatment was partially attributed to RIP1-RIP3 complex-mediated programmed necrosis downstream of TNF-α induction. Additionally, upon uptake into macrophages, GO accumulated primarily in cytoplasm causing dramatic morphologic alterations and a significant reduction of the macrophagic ability in phagocytosis. However, macrophagic uptake of GO may not be required for induction of necrosis. GO exposure also caused a large increase of intracellular reactive oxygen species (ROS), which contributed to the cause of cell death. The combined data reveal that interaction of GO with TLR4 is the predominant molecular mechanism underlying GO-induced macrophagic necrosis; also, cytoskeletal damage and oxidative stress contribute to decreased viability and function of macrophages upon GO treatment.

The impact of silver nanoparticles (AgNPs) on the central nervous system is a topic with mounting interest and concern and the facts remain elusive. In the current study, the neurotoxicity of commercial AgNPs to rat cerebellum granule cells (CGCs) and the corresponding molecular mechanism are closely investigated. It is demonstrated that AgNPs induce significant cellular toxicity to CGCs in a dose-dependent manner without damaging the cell membrane. Flow cytometry analysis with the Annexin V/propidium iodide (PI) staining indicates that the apoptotic proportion of CGCs upon treatment with AgNPs is greatly increased compared to the negative control. Moreover, the activity of caspase-3 is largely elevated in AgNP-treated cells compared to the negative control. AgNPs are demonstrated to induce oxidative stress, reflected by the massive generation of reactive oxygen species (ROS), the depletion of antioxidant glutathione (GSH), and the increase of intracellular calcium. Histological examination suggests that AgNPs provoke destruction of the cerebellum granular layer in rats with concomitant activation of caspase-3, in parallel to the neurotoxicity of AgNPs observed in vitro. Taken together, it is demonstrated for the first time that AgNPs substantially impair the survival of primary neuronal cells through apoptosis coupled to oxidative stress, depending on the caspase activation-mediated signaling.

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