In Vitro Co-Exposure to CeO2 Nanomaterials from Diesel Engine Exhaust and Benzo(a)Pyrene Induces Additive DNA Damage in Sperm and Cumulus Cells but Not in Oocytes.

Benzo(a)pyrene (BaP) is a recognized reprotoxic compound and the most widely investigated polycyclic aromatic hydrocarbon in ambient air; it is widespread by the incomplete combustion of fossil fuels along with cerium dioxide nanomaterials (CeO2 NMs), which are used in nano-based diesel additives to decrease the emission of toxic compounds and to increase fuel economy. The toxicity of CeO2 NMs on reproductive organs and cells has also been shown. However, the effect of the combined interactions of BaP and CeO2 NMs on reproduction has not been investigated. Herein, human and rat gametes were exposed in vitro to combusted CeO2 NMs or BaP or CeO2 NMs and BaP in combination. CeO2 NMs were burned at 850 °C prior to mimicking their release after combustion in a diesel engine. We demonstrated significantly higher amounts of DNA damage after exposure to combusted CeO2 NMs (1 µg·L-1) or BaP (1.13 µmol·L-1) in all cell types considered compared to unexposed cells. Co-exposure to the CeO2 NMs-BaP mixture induced additive DNA damage in sperm and cumulus cells, whereas no additive effect was observed in rat oocytes. This result could be related to the structural protection of the oocyte by cumulus cells and to the oocyte's efficient system to repair DNA damage compared to that of cumulus and sperm cells.

CeO2 Nanomaterials from Diesel Engine Exhaust Induce DNA Damage and Oxidative Stress in Human and Rat Sperm In Vitro.

Cerium dioxide nanomaterials (CeO2 NMs) are widely used in nano-based diesel additives to decrease the emission of toxic compounds, but they have been shown to increase the emission of ultrafine particles as well as the amount of released Ce. The Organization for Economic Cooperation and Development included CeO2 NMs in the priority list of nanomaterials that require urgent evaluation, and the potential hazard of aged CeO2 NM exposure remains unexplored. Herein, human and rat sperm cells were exposed in vitro to a CeO2 NM-based diesel additive (called EnviroxTM), burned at 850 °C to mimic its release after combustion in a diesel engine. We demonstrated significant DNA damage after in vitro exposure to the lowest tested concentration (1 µg·L-1) using the alkaline comet assay (ACA). We also showed a significant increase in oxidative stress in human sperm after in vitro exposure to 1 µg·L-1 aged CeO2 NMs evaluated by the H2DCF-DA probe. Electron microscopy showed no internalization of aged CeO2 NMs in human sperm but an affinity for the head plasma membrane. The results obtained in this study provide some insight on the complex cellular mechanisms by which aged CeO2 NMs could exert in vitro biological effects on human spermatozoa and generate ROS.