Hyun-Jeong Eom

Oxidative stress of CeO2 nanoparticles via p38-Nrf-2 signaling pathway in human bronchial epithelial cell, Beas-2B

To understand the molecular mechanism of previously observed cerium oxide (CeO(2)) nanoparticles-induced oxidative stress, an in vitro toxicity assay was conducted using human bronchial epithelial cell, Beas-2B, focusing on the involvement of the oxidative stress responding signal transduction pathway and transcription factors in the toxicity of CeO(2) nanoparticles. Extracellular signal-regulating kinase (ERK), p38 and c-Jun N-terminal kinase (JNK) signaling pathways, along with nuclear factor-kappaB (NF-kappaB) and nuclear factor-E2-related factor-2 (Nrf-2), were investigated as the upstream events of oxidative stress from exposure to CeO(2) nanoparticles. The overall results suggest that CeO(2) nanoparticles may exert their toxicity through oxidative stress, as they cause significant increases in the cellular reactive oxygen species (ROS) concentrations, subsequently leading to the strong induction of heme oxygenase-1 (HO-1) via the p38-Nrf-2 signaling pathway. Further studies on the mechanism by which CeO(2) nanoparticles induce the p38-Nrf-2 signaling pathway are warranted for a better understanding of the CeO(2) nanoparticles-induced oxidative stress; studies with other signaling pathways, with concentration-response and time course experiments would also be justified.

Oxidative stress of silica nanoparticles in human bronchial epithelial cell, Beas-2B

In this study, the potentially harmful effect of the exposure to fumed and porous silicon dioxide (silica) nanoparticles was investigated using human bronchial epithelial cell, Beas-2B, with a focus on the involvement of oxidative stress as the toxic mechanism. Silica nanoparticles-induced oxidative stress was assessed by examining the formation of reactive oxygen species (ROS) and induction of antioxidant enzymes, such as superoxide dismutase (SOD) and heme oxygenase-1 (HO-1). Subsequently, to understand the mechanism of nanoparticles-induced oxidative stress, the involvement of oxidative stress-responding transcription factors, such as, nuclear factor-kappaB (NF-kappaB) and nuclear factor-E2-related factor-2 (Nrf-2), as well as the mitogen-activated protein (MAP) kinase signal transduction pathway were investigated. From the overall results, silica nanoparticles exerted toxicity via oxidative stress, which lead to the induction of HO-1 via the Nrf-2-ERK MAP kinase signaling pathway; cells exposed to porous silica nanoparticles showed a more sensitive response than those exposed to fumed silica. Nevertheless, the parameters tested were rather limited in terms of gaining a full understanding of the oxidative stress and cellular response due to exposure to silica nanoparticles. Further studies on the mechanism by which silica nanoparticles induce the Nrf-2-ERK MAP kinase pathway, to more clearly elucidate the silica-induced oxidative stress, as well as on the relationship between the physico-chemical properties of nanoparticles and their cytotoxicity are warranted to gain an understanding of the phenomenon of different sensitivities between porous and fumed silica.

A systems toxicology approach to the surface functionality control of graphene–cell interactions

The raised considerable concerns about the possible environmental health and safety impacts of graphene nanomaterials and their derivatives originated from their potential widespread applications. We performed a comprehensive study about biological interaction of grapheme nanomaterials, specifically in regard to its differential surface functionalization (oxidation status), by using OMICS in graphene oxide (GO) and reduced graphene oxide (rGO) treated HepG2 cells. Differential surface chemistry (particularly, oxidation - O/C ratio) modulates hydrophobicity/philicity of GO/rGO which in turn governs their biological interaction potentiality. Similar toxic responses (cytotoxicity, DNA damage, oxidative stress) with differential dose dependency were observed for both GO and rGO but they exhibited distinct mechanism, such as, hydrophilic GO showed cellular uptake, NADPH oxidase dependent ROS formation, high deregulation of antioxidant/DNA repair/apoptosis related genes, conversely, hydrophobic rGO was found to mostly adsorbed at cell surface without internalization, ROS generation by physical interaction, poor gene regulation etc. Global gene expression and pathway analysis displayed that TGFβ1 mediated signaling played the central role in GO induced biological/toxicological effect whereas rGO might elicited host-pathogen (viral) interaction and innate immune response through TLR4-NFkB pathway. In brief, the distinct biological and molecular mechanisms of GO/rGO were attributed to their differential surface oxidation status.

Oxidative stress‐related PMK‐1 P38 MAPK activation as a mechanism for toxicity of silver nanoparticles to reproduction in the nematode Caenorhabditis elegans

In the present study, a toxic mechanism of silver nanoparticles (AgNPs) was investigated in the nematode, Caenorhabditis elegans, focusing on the involvement of oxidative stress in reproduction toxicity. Initially, AgNPs were tested as potential oxidative stress inducers, and increased formation of reactive oxygen species (ROS) was observed in AgNP-exposed C. elegans. Subsequently, the potential upstream signaling pathway activated in response to AgNP exposure was investigated, paying special attention to the C. elegans PMK-1 p38 mitogen-activated protein kinase (MAPK). Increased PMK-1 p38 MAPK gene and protein expressions were observed in C. elegans exposed to AgNPs. Expression of the p38-dependent transcription factor genes and glutathione S-transferase (GST) enzyme activity was also investigated in wildtype (N2) and pmk-1 mutant (km25) C. elegans exposed to AgNPs. The results indicated that AgNP exposure led to increased ROS formation, increased expression of PMK-1 p38 MAPK and hypoxia-inducible factor (HIF-1), GST enzyme activity, and decreased reproductive potential in wildtype (N2) C. elegans; whereas in the AgNP-exposed pmk-1 (km25) mutant, ROS formation and HIF-1 and GST activation were not observed, and decreased reproductive potential was rescued. These results suggest that oxidative stress is an important mechanism of AgNP-induced reproduction toxicity in C. elegans, and that PMK-1 p38 MAPK plays an important role in it. The results also suggest that GST and HIF-1 activation by AgNP exposure are PMK-1 p38 MAPK-dependent, and that both play an important role in the PMK-1 p38 MAPK-mediated defense pathway to AgNP exposure in C. elegans.

Comparative toxicity of silver nanoparticles on oxidative stress and DNA damage in the nematode, Caenorhabditis elegans.

This study examined the effects of polyvinylpyrrolidone (PVP) surface coating and size on the organismal and molecular toxicity of silver nanoparticles (AgNPs) on the nematode, Caenorhabditis elegans. The toxicity of bare AgNPs and 8 and 38 nm PVP-coated AgNPs (PVP8-AgNPs, PVP38-AgNPs) were compared. The toxicity of AgNO3 was also tested because ion dissolution and particle-specific effects are often important characteristics determining Ag nanotoxicity. Comparative toxicity across AgNO3 and the three different types of AgNPs was first evaluated using a C. elegans mortality test by a direct comparison of the LC50 values. Subsequently, mutant screening followed by oxidative stress, mitochondrial toxicity and DNA damage assays were carried out at equitoxic (LC10 and LC50) concentrations to further assess the toxicity mechanism of AgNO3 and AgNPs. AgNO3 and bare AgNPs had similar toxicities, whereas PVP coating reduced the toxicity of the AgNPs significantly. Of the PVP-AgNPs, the smaller NPs were more toxic. Different groups of mutants responded differently to AgNO3 and AgNPs, which indicates that their toxicity mechanism might be different. AgNO3 and bare AgNPs induced mitochondrial membrane damage. None of the silver materials tested caused detectable polymerase-inhibiting DNA lesions in either the nucleus or mitochondria as measured by a quantitative PCR assay, but AgNO3, bare AgNPs and PVP8-AgNPs induced oxidative DNA damage. These results show that coatings on the AgNPs surface and the particle size make a clear contribution to the toxicity of the AgNPs, and oxidative stress-related mitochondrial and DNA damage appear to be potential mechanisms of toxicity.

Integrated mRNA and micro RNA profiling reveals epigenetic mechanism of differential sensitivity of Jurkat T cells to AgNPs and Ag ions

In our previous in vitro study of the toxicity on silver nanoparticles (AgNPs), we observed a dramatically higher sensitivity of Jurkat T cells to AgNPs than to Ag ions, and DNA damage and apoptosis were found to be involved in that toxicity. In this study, to understand underlying mechanism of different sensitivity of Jurket T cells to AgNPs and Ag ions, mRNA microarray and micro RNA microarray were concomitantly conducted on AgNPs and Ag ions exposed Jurkat T cells. Surprisingly only a small number of genes were differentially expressed by exposure to each of the silver (15 altered mRNA by AgNPs exposure, whereas 4 altered mRNA by Ag ions exposure, as determined 1.5-fold change as the cut-off value). miRNA microarray revealed that the expression of 63 miRNAs was altered by AgNPs exposure, whereas that of 32 miRNAs was altered by Ag ions exposure. An integrated analysis of mRNA and miRNA expression revealed that the expression of hsa-miR-219-5p, was negatively correlated with the expression of metallothionein 1F (MT1F) and tribbles homolog 3 (TRIB3), in cells exposed to AgNPs; whereas, the expression of hsa-miR-654-3p was negatively correlated with the expression of mRNA, endonuclease G-like 1 (EDGL1) in cells exposed to Ag ions. Network analysis were further conducted on mRNA-miRNA pairs, which revealed that miR-219-5p-MT1F and -TRIB3 pairs by AgNPs are being involved in various cellular processes, such as, oxidative stress, cell cycle and apoptosis, whereas, miR-654-3p and ENDOGL1 pair by Ag ions generated a much simpler network. The putative target genes of AgNPs-induced miR-504, miR-33 and miR-302 identified by Tarbase 6.0 are also found to be involved in DNA damage and apoptosis. These results collectively suggest that distinct epigenetic regulation may be an underlying mechanism of different sensitivity of Jurkat T cells to AgNPs and Ag ion. Further identification of putative target genes of DE miRNA by AgNPs and Ag ions may provide additional clues for the mechanism of differential toxicity. Overall results suggest that epigenetic mechanism is involved in toxicity of AgNPs and Ag ions in Jurkat T cells.

Hypoxia inducible factor-1 (HIF-1)-flavin containing monooxygenase-2 (FMO-2) signaling acts in silver nanoparticles and silver ion toxicity in the nematode, Caenorhabditis elegans.

In the present study, nanotoxicity mechanism associated with silver nanoparticles (AgNPs) exposure was investigated on the nematode, Caenorhabditis elegans focusing on the hypoxia response pathway. In order to test whether AgNPs-induced hypoxia inducible factor-1 (HIF-1) activation was due to hypoxia or to oxidative stress, depletion of dissolved oxygen (DO) in the test media and a rescue effect using an antioxidant were investigated, respectively. The results suggested that oxidative stress was involved in activation of the HIF-1 pathway. We then investigated the toxicological implications of HIF-1 activation by examining the HIF-1 mediated transcriptional response. Of the genes tested, increased expression of the flavin containing monooxygenase-2 (FMO-2) gene was found to be the most significant as induced by AgNPs exposure. We found that AgNPs exposure induced FMO-2 activation in a HIF-1 and p38 MAPK PMK-1 dependent manner, and oxidative stress was involved in it. We conducted all experiments to include comparison of AgNPs and AgNO3 in order to evaluate whether any observed toxicity was due to dissolution or particle specific. The AgNPs and AgNO3 did not produce any qualitative differences in terms of exerting toxicity in the pathways observed in this study, however, considering equal amount of silver mass, in every endpoint tested the AgNPs were found to be more toxic than AgNO3. These results suggest that Ag nanotoxicity is dependent not only on dissolution of Ag ion but also on particle specific effects and HIF-1-FMO-2 pathway seems to be involved in it.

Involvement of Caenohabditis elegans MAPK Signaling Pathways in Oxidative Stress Response Induced by Silver Nanoparticles Exposure

In the present study, toxicity of silver nanoparticles (AgNPs) was investigated in the nematode, Caenohabditis elegans focusing on the upstream signaling pathway responsible for regulating oxidative stress, such as mitogen-activated protein kinase (MAPK) cascades. Formation of reactive oxygen species (ROS) was observed in AgNPs exposed C.elegans, suggesting oxidative stress as an important mechanism in the toxicity of AgNPs towards C. elegans. Expression of genes in MAPK signaling pathways increased by AgNPs exposure in less than 2-fold compared to the control in wildtype C.elegans, however, those were increased dramatically in sod-3 (gk235) mutant after 48 h exposure of AgNPs (i.e. 4-fold for jnk-1 and mpk-2; 6-fold for nsy-1, sek-1, and pmk-1, and 10-fold for jkk-1). These results on the expression of oxidative stress response genes suggest that sod-3 gene expression appears to be dependent on p38 MAPK activation. The high expressions of the pmk-1 gene 48 h exposure to AgNPs in the sod-3 (gk235) mutant can also be interpreted as compensatory mechanisms in the absence of important stress response genes. Overall results suggest that MAPK-based integrated stress signaling network seems to be involved in defense to AgNPs exposure in C.elegans.

Toxic potentiality of bio-oils, from biomass pyrolysis, in cultured cells and Caenorhabditis elegans

Bio‐oils, which are multicomponent mixtures, were produced from two different biomass (rice straw (rice oil) and sawdust of oak tree (oak oil)) by using the slow pyrolysis process, and chemical compositional screening with GC‐MS detected several hazardous compounds in both bio‐oil samples. The two bio‐oils vary in their chemical compositional nature and concentrations. To know the actual hazard potentialities of these bio‐oils, toxicological assessments were carried out in a comparative approach by using in vitro (Jurkat T and HepG2 cell) as well as in vivo (Caenorhabditis elegans) systems. A dose‐dependent increase in cytotoxicity, cell death (apoptosis), and genotoxicity were observed in cultured cell systems. Similarly, the in vivo system, C. elegans also displayed a dose‐dependent decrease in survival. It was found that in comparison with rice oil, oak oil displayed higher toxicity to all models systems, and the susceptibility order of the model systems were Jurkat T > HepG2 > C. elegans. Pursuing the study further toward the underlying mechanism by exploiting the C. elegans mutants screening assay, the bio‐oils seem to mediate toxicity through oxidative stress and impairment of immunity. Taken together, bio‐oils compositions mainly depend on the feedstock used and the pyrolysis conditions which in turn modulate their toxic potentiality.

Development of biomarker for detecting silver nanoparticles exposure using a GAL4 enhancer trap screening in Drosophila

Silver nanoparticles (AgNPs) have been widely used in commercial goods ranging from medical devices to home appliances. Their widespread application increase the risk related to their potential toxicity. Although several studies showed their acute hazardous effects on living animals, our understanding of chronic effects of AgNPs exposed by the environment we encounter in our everyday lives is still very limited. This is partly because of the lack of versatile animal model system for studying AgNPs effects on terrestrial animals including human. In this study, we used Drosophila model to study AgNPs toxicity in terrestrial animals, and found that long-term exposure of AgNPs, but not Ag ions, at low level (0.1 and 1μg/mL) significantly shortened the lifespan. By taking advantage of the power of Drosophila genetics, we also isolated a GAL4 enhancer trap line called M95, in which the expression of GAL4 is up-regulated in response to ingestion of AgNPs at concentrations as low as 0.1μg/mL. Interestingly M95 flies showed significantly increased tolerance to both AgNPs treatment and dry starvation probably due to up-regulation of JNK signaling. These findings suggest not only that M95 may be a very useful biomarker of AgNPs because of its high sensitivity and tolerance to AgNPs, but also that Drosophila may be a versatile terrestrial invertebrate model for studying the effects of AgNPs on human health.