No abstract

We showed previously that exposure of human lung cells (BEAS-2B) to TiO2 nanoparticles (nano-TiO2 ) produced micronuclei (MN) only when the final concentration of protein in the cell-culture medium was at least 1%. Nanoparticles localize in the liver; thus, we exposed human liver cells (HepG2) to nano-TiO2 and found the same requirement for MN induction. Nano-TiO2 also formed small agglomerates in medium containing as little as 1% protein and caused cellular interaction as measured by side scatter by flow cytometry and DNA damage (comet assay) in HepG2 cells. Nano-TiO2 also increased the activity of the inflammatory factor NFkB but not of AP1 in a reporter-gene HepG2 cell line. Suspension of nano-TiO2 in medium containing 0.1% protein was sufficient for induction of MN by the nanoparticles in either BEAS-2B or HepG2 cells as long the final concentration of protein in the cell-culture medium was at least 1%.

Nano-TiO2 and nano-CeO2 are among the most widely used engineered nanoparticles (NPs). We investigated a variety of endpoints to assess the toxicity of eight of these NPs to induce potentially adverse health effects in an In Vitro human respiratory epithelial cell model. These endpoints include cytotoxicity, reactive oxygen species (ROS)/reactive nitrogen species (RNS) production, 8-hydroxy-2_-deoxyguanosine (8-oxo-dG), endogenous DNA adducts, Apurinic/apyrimidinic (AP) sites, 4-Hrdoxynonenal (4-HNE) protein adducts, Malondialdehyde (MDA) protein adducts, and genomics analysis on altered signaling pathways. Our results indicated that cytotoxicity assays are relatively insensitive, and we detected changes in other endpoints at concentrations much lower than those inducing cytotoxicity. Among the ROS-related endpoints, 8-oxo-dG is relatively more sensitive than other assays, and nano-TiO2 induced more 8-oxo-dG formation than nano-CeO2. Finally, there are many signaling pathways changes at concentrations at which no cytotoxicity was observed. These alterations in signaling pathways correlated well with In Vitro toxicity that was observed at higher concentrations, and with in vivo adverse outcome pathways caused by nano-TiO2 and nano-CeO2 in experimental animals.

A series of six titanium dioxide and two cerium oxide engineered nanomaterials were assessed for their ability to induce cytotoxicity, reactive oxygen species (ROS), various types of DNA damage, and transcriptional changes in human respiratory BEAS-2B cells exposed in vitro at several concentrations for 72 hours. Only limited cytotoxicity was observed at concentrations up to 300 μg/ml for all of the nanomaterials. Small increases in 8-oxo-deoxyguanosine were induced by some of the nanomaterials, but did not achieve statistical significance. No increases in ethenoadenosine or ethenocytidine were detected by ELISA assays for any of the tested nanomaterials. Several of the nanomaterials exhibited concentration related increases in levels of apurinic/apyrimidinic sites, endogenous DNA adducts measured by 32P-postlabeling, lipid peroxidation, and ROS. Consistent with these findings, several of the nanomaterials also affected expression of genes involved in p53, ATM, and mismatch repair pathways. Integrin signaling pathways were also altered by a majority of the nanomaterials tested. There was general agreement between activity in DNA damage assays and extent of pathway transcriptional alteration. One out of the cerium oxide nanomaterials tested did not induce a high enough incidence of differentially expressed genes relative to controls to allow analysis at the pathway level, and also elicited the lowest response in multiple DNA damage assays. Taken together, these data are consistent with the contribution of DNA damage induced by reactive oxygen species as mediators of potentially adverse biological effects following exposure to engineered titanium and cerium oxide nanomaterials, and suggests the utility of short term in vitro tests to predict relative potencies of these particles. This is an abstract of a proposed presentation and does not necessarily reflect EPA policy. Citation Format: Garret B. Nelson, Sheau-Fung Y. Thai, Carlton P. Jones, Audrey Barbee, Micaela Killius, Jeffrey A. Ross. Comparative DNA damage and transcriptomic effects of engineered nanoparticles in human lung cells in vitro. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2814.