Transcriptomic-based toxicological investigations of graphene oxide with modest cytotoxicity to human umbilical vein endothelial cells: changes of Toll-like receptor signaling pathways.

The wide uses of graphene oxide (GO) lead to the contact of GO with vascular systems, so it is necessary to investigate the toxicological effects of GO to endothelial cells. Recently, we reported that GO of small lateral size (<500 nm) was relatively biocompatible to human umbilical vein endothelial cells (HUVECs), but recent studies by using omics-techniques revealed that nanomaterials (NMs) even without acute cytotoxicity might induce other toxicological effects. This study investigated the effects of GO on HUVECs based on RNA-sequencing and bioinformatics analysis. Even after exposure to 100 μg/ml GO, the cellular viability of HUVECs was higher than 70%. Furthermore, 25 μg/ml GO was internalized but did not induce ultrastructural changes or intracellular superoxide. These results combined indicated GO's relatively high biocompatibility. However, by analyzing the most significantly altered Gene Ontology terms and Kyoto Encyclopedia of Gene and Genomes pathways, we found that 25 μg/ml GO altered pathways related to immune systems' functions and the responses to virus. We further verified that GO exposure significantly decreased Toll-like receptor 3 and interleukin 8 proteins, indicating an immune suppressive effect. However, THP-1 monocyte adhesion was induced by GO with or without the presence of inflammatory stimulus lipopolysaccharide. We concluded that GO might inhibit the immune responses to virus in endothelial cells at least partially mediated by the inhibition of TLR3. Our results also highlighted a need to investigate the toxicological effects of NMs even without acute cytotoxicity by omics-based techniques.