Lateral Size of Thin Graphene Oxide Sheets is a Critical Factor for Induction of Oxidative-Stress Mediated Cellular Responses

The interest for graphene and its translation into commercial products have been expanding at high pace during the last few years. In consideration of the previously described pulmonary health and safety concerns for carbon... [ view full abstract ]

The interest for graphene and its
translation into commercial products have been expanding at high pace during
the last few years. In consideration of the previously described pulmonary
health and safety concerns for carbon nanomaterials, there is a great need to
understand the critical parameters impacting interactions between graphene
based materials (GBMs) and the cells of the pulmonary system. With this regard,
the aim of present study was to determine the importance of two key parameters:
lateral dimensions of the material and coating with proteins from the serum in
relation to each other and their impact on human lung epithelial cell line
BEAS-2B. Secondly, we interrogated whether the cellular response to graphene oxide
(GO) could be explained and predicted using oxidative stress paradigm.

In order
to address these questions, we produced and thoroughly characterized
endotoxin-free material with two distinct lateral dimensions – large,
micrometer-sized GO (5 – 15 µm) and small, nanometer-sized GO (50 – 200 nm).
The role of protein adsorption in the initial phase of the interaction between
the GO flakes and cells was addressed by controlling the presence of serum
during the first 4 h of interaction. We exploited the intrinsic fluorescence of
GO to image the material without introducing additional surface modifications
or attaching fluorescent dyes. Using confocal live cell imaging, we were able
to show for the first time the behavior of the cells in response to the material
exposure. Toxicity was confirmed to be time and dose dependent, with large
material inducing higher levels of cellular death correlated with elevated ROS
production and increased expression of pro-inflammatory genes, which is in
agreement with oxidative stress paradigm. In addition, toxicity of small
material was completely alleviated at 24 h in the presence of FBS, while only
mitigated for large material, confirming the hypothesis that lateral dimensions
are a predominant factor of toxicity and inflammation in vitro.