In Vitro Assessment of the Genotoxic Potential of Pristine Graphene Platelets

Abstract

(1) Background: Graphene is a two-dimensional atomic structure with a wide range of uses, including for biomedical applications. However, knowledge of its hazards is still limited. This work brings new cytotoxic, cytostatic, genotoxic and immunotoxic data concerning the in vitro exposure of human cell line to two types of graphene platelets (GP). It also contributes to the formation of general conclusions about the health risks of GP exposure. (2) Methods: In vitro exposure of a THP-1 cell line to three concentrations of two GP over 40 h. The cytotoxic potential was assessed by the measurement of LDH and glutathione (ROS) and by a trypan blue exclusion assay (TBEA); the cytostatic and genotoxic potential were assessed by the cytokinesis-block micronucleus (CBMN) test; and the immunotoxic potential was assessed by the measurement of IL-6, IL-10 and TNF-α. (3) Results: We found a significant dose-dependent increase in DNA damage (CBMN). The lowest observed genotoxic effect levels (LOGEL) were 5 µg/mL (GP1) and 30 µg/mL (GP2). We found no significant leaking of LDH from cells, increase in dead cells (TBEA), induction of ROS, increased levels of cytostasis, or changes in IL-6, IL-10 and TNF-α levels. (4) Conclusions: The genotoxicity increased during the short-term in vitro exposure of THP-1 to two GP. No increase in cytotoxicity, immunotoxicity, or cytostasis was observed.

PLATOX: Integrated In Vitro/In Vivo Approach for Screening of Adverse Lung Effects of Graphene-Related 2D Nanomaterials

“…Notably, a proliferation-inhibiting effect of the GRNP materials was observed in MRC-5 cells using RICC and, thus, cell counts as an endpoint. The decrease in RICC in MRC-5 cells seemed to correlate with GRNP thickness, as P5, P3, P2 and P1 demonstrated the lowest BMD30 values (thickness between 0.6 and 1.2 nm), followed by P6 (1-3 nm), P4 (2-10 nm) and P7 (3 nm), and might indicate direct mechanical interference with the cytoskeleton and the mitotic apparatus of the cells, as discussed previously for induction of micronuclei by two types of pristine graphene nanoplatelets in THP-1 cells [45].…”

“…The induction of micronuclei by pristine graphene nanoplates and, thus, of fixed DNA damage was furthermore demonstrated in THP-1 cells, underlining the need to include genotoxicity testing in the screening of pristine graphene nanoplates [45]. The same robust endpoints could be mirrored and validated in the in vivo inhalation testing using cytotoxicity, differential cell count in lung lavage fluid as well as histopathological equivalents.…”

The Effect of Chronic Exposure of Graphene Nanoplates on the Viability and Motility of A549 Cells

“…The WST-1 assay was more sensitive than the alamarBlue™ assay, while direct counting of nuclei using highthroughput image cytometry did not reveal any toxicity of the GP1 nanoplates. The WST-1 assay tetrazolium salt is cleaved to a soluble formazan by a complex cellular mechanism that occurs primarily at the cell surface and was reported to yield false-positive results because of the optical interference between nanomaterials and tetrazolium salts and other suggested modes of interactions [31][32][33]. Resazurin in alamarBlue™ is reduced by enzymes located in cytoplasm and mitochondria, and its reduction may signify an impairment of cellular metabolism without necessarily implicating mitochondrial dysfunction [34].…”

Carbon-Based Nanomaterials Increase Reactivity of Primary Monocytes towards Various Bacteria and Modulate Their Differentiation into Macrophages

“…Studies focused on graphene oxide sheets showed the inhibition of the migration of A549 and HeLa cells by direct reaction with actin [64,65]. However, the possible disruption of cytoskeletal fibres by GPs seems to caused be their mechanical action [66].…”