Genotoxicity Response of Fibroblast Cells and Human Epithelial Adenocarcinoma In Vitro Model Exposed to Bare and Ozone-Treated Silica Microparticles.
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ABSTRACT:
SUBMITTER: Colafarina S
PROVIDER: S-EPMC8773945 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:We used mass spectrometry to profile changes in metabolites, proteins, and phosphorylation in silica-exposed BEAS-2B epithelial cells.
2022-07-28 | PXD035608 |
Project description:The toxicity of many xenobiotic compounds is believed to involve oxidative injury to cells. Direct assessment of mechanistic events involved in xenobiotic-induced oxidative stress is not easily achievable. Development of genetically encoded probes designed for monitoring intracellular redox changes represents a methodological advance with potential applications in toxicological studies.We tested the utility of redox-sensitive green fluorescent protein (roGFP)-based redox sensors for monitoring real-time intracellular redox changes induced by xenobiotics in toxicological studies.roGFP2, a reporter of the glutathione redox potential (E(GSH)), was used to monitor EGSH in cultured human airway epithelial cells (BEAS-2B cells) undergoing exposure to 0.15-1.0 ppm ozone (O(3)). Cells were imaged in real time using a custom-built O(3) exposure system coupled to a confocal microscope.O(3) exposure induced a dose- and time-dependent increase of the cytosolic EGSH. Additional experiments confirmed that roGFP2 is not directly oxidized, but properly equilibrates with the glutathione redox couple: Inhibition of endogenous glutaredoxin 1 (Grx1) disrupted roGFP2 responses to O(3), and a Grx1-roGFP2 fusion protein responded more rapidly to O(3) exposure. Selenite-induced up-regulation of GPx (glutathione peroxidase) expression-enhanced roGFP2 responsiveness to O(3), suggesting that (hydro)peroxides are intermediates linking O(3) exposure to glutathione oxidation.Exposure to O(3) induces a profound increase in the cytosolic E(GSH) of airway epithelial cells that is indicative of an oxidant-dependent impairment of glutathione redox homeostasis. These studies demonstrate the utility of using genetically encoded redox reporters in making reliable assessments of cells undergoing exposure to xenobiotics with strong oxidizing properties.
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Project description:Previous studies show variable ozone cytotoxicity and genotoxicity in cell cultures, laboratory animals and humans directly exposed to tropospheric ozone. The aim of this study was therefore to investigate and compare the cyto and genotoxic effects of ozone using adenocarcinoma human alveolar basal epithelial cells A549 and normal human fibroblasts Hs27. A cell culture chamber with controlled atmosphere (a simulation reactor) was built to inject a flow of 120 ppb of ozone, which is two times the threshold value for the protection of human health, fixed by the EU legislation. Cell proliferation was evaluated by a luminescent cell viability assay while we assessed the genotoxic potential of ozone by the induction of micronuclei as well as evaluating DNA strand breaks by the induction of micronuclei evaluated by means of the cytokinesis-block micronucleus (CBMN) assay as well as evaluating DNA strand breaks by Alkaline Comet Assay (CA) or Comet Assay. A549 cells viability decreases significantly at 24 hours treatment with 120 ppb of O3 while at 48 hours and 72 hours O3 treated cells viability doesn't differ in respect to the control. However a significative decrease of A549 viability is shown at 72 hours vs. 48 hours in both treated and not-treated cells. The viability trend in the Hs27 cells did not show any significant changes in treated samples compared to the control in all conditions. The two genotoxicity biomarkers, the micronucleus and the comet tests, showed in both the cell types exposed to ozone, a significant increase in the number of micronuclei and in the tail DNA % in respect to the control even if at different times/cell type. Moreover, we found that O3 provokes genotoxic effects more evident in A549 cancer cells than in normal fibroblasts Hs27 ones. We applied a cell growth simulation model referred to ozone treated or not cell lines to confirm that the ozone exposure causes a slackening in the cells replication.
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Project description:Epithelial tissues possess a barrier function that must be restored following injury. Repair often involves changes in cell fate, but it remains poorly understood how cell plasticity is induced and whether responses vary with distinct types of damage. Here, we reveal that genotoxic agents, but not mechanical damage, induce hyperplasia and lineage infidelity in the interfollicular epidermis, as well as in two epidermal subappendages: the mammary gland and hair follicle. DNA damage to the epidermis promotes stromal proliferation, and induces basal cell hyperplasia with formation of abnormal, multi-layered K14+/K10+ cells. Unexpectedly, this behavior is epithelial cell non-autonomous, but independent of immunity. Instead, dermal fibroblasts are both necessary and sufficient to induce the epithelial response. We identify a previously unrecognized fibroblast-specific NLRP3 inflammasome that drives this response via IL-1β. Thus, genotoxic agents, used chemotherapeutically to promote cancer cell death, can have the opposite effect on wild-type epithelia, paradoxically promoting hyperplasia and inducing both stemness and lineage infidelity via a non-autonomous, fibroblast-driven mechanism.
2020-12-16 | GSE139272 | GEO
Project description:Hollow microparticles are important materials, offering a larger surface area and lower density than their solid counterparts. Furthermore, their inner void space can be exploited for the encapsulation and release of guest species in a variety of applications. Herein, we present phosphazene-based silica hollow microparticles prepared via a surfactant-free sol-gel process through self-assembly of the alkoxysilyl-containing polymer in water-ethanol solution. Solely, a silane-derived polyphosphazene was used as the precursor for the microparticle formation, without additional classical silica sources. These novel hollow silica-based microparticles were prepared without surfactant, using a designed amphiphilic polyphosphazene for the particle formation made by two components, a hydrophilic unit consisting of 3-mercaptopropyl(trimethoxysilane), and a hydrophobic unit (dodecanethiol) attached to the double bonds from the poly(allylamine)phosphazene backbone via a thiol-ene photoreaction. Due to these two functionalities, a "vesicle"-like self-assembled structure was formed in the reaction medium, which could be then utilized for the microparticle preparation. The influence of NaOH during the synthesis was shown to affect the size and the wall thickness of the microparticles. This effect may enhance the possibilities to tailor such microparticles for drug delivery purposes or for future controlled release of other substances, such as drugs, fragrances, or anticorrosive pigments.
| S-EPMC9318875 | biostudies-literature
Project description:BackgroundDietary intake of the omega-3 family of polyunsaturated fatty acids (ω-3 FA) is associated with anti-inflammatory effects. However, unsaturated fatty acids are susceptible to oxidation, which produces pro-inflammatory mediators. Ozone (O3) is a tropospheric pollutant that reacts rapidly with unsaturated fatty acids to produce electrophilic and oxidative mediators of inflammation.ObjectiveDetermine whether supplementation with ω-3 FA alters O3-induced oxidative stress in human airway epithelial cells (HAEC).Methods16-HBE cells expressing a genetically encoded sensor of the reduced to oxidized glutathione ratio (GSH/GSSG, EGSH) were supplemented with saturated, monounsaturated, or ω-3 FA prior to exposure to 0, 0.08, 0.1, or 0.3 ppm O3. Lipid peroxidation was measured in cellular lipid extracts and intact cells following O3 exposure.ResultsRelative to cells incubated with the saturated or monounsaturated fatty acids, cells supplemented with ω-3 FA containing 5 or 6 double bonds showed a marked increase in EGSH during exposure to O3 concentrations as low as 0.08 ppm. Consistent with this finding, the concentration of lipid hydroperoxides produced following O3 exposure was significantly elevated in ω-3 FA supplemented cells.DiscussionSupplementation with polyunsaturated ω-3 FA potentiates oxidative responses, as indicated by EGSH, in HAEC exposed to environmentally relevant concentrations of O3. This effect is mediated by the increased formation of lipid hydroperoxides produced by the reaction of O3 with polyunsaturated fatty acids. Given the inflammatory activity of lipid hydroperoxides, these findings have implications for the potential role of ω-3 FA in increasing human susceptibility to the adverse health effects of O3 exposure.
| S-EPMC9131300 | biostudies-literature
Project description:To tailor the properties of colloidal materials, precise control over the self-assembly of their constituents is a prerequisite. Here, we govern the assembly of silica particles by functionalization with supramolecular moieties which interact with each other via directional and reversible hydrogen bonding. Through a generally applicable synthesis protocol, two different types of self-complementary hydrogen bonding moieties, BTA- and UPy-derivatives, are anchored to silica particles. Their self-assembly is initiated by the UV-induced removal of a photolabile protecting group, allowing the formation of hydrogen bonds between tethered molecules. The light-induced assembly of BTA- and UPy-decorated colloids in single-component dispersions and colloidal self-sorting in mixed dispersions is studied. Furthermore, we demonstrate that UPy-colloids can dissasemble upon addition of traces of a competitive binder (NaPy). This work provides further insight into the utility of supramolecular handles to orchestrate the assembly of micron-sized colloids via non-oligonucleotide hydrogen-bonding units.
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Project description:Diagnosing tumors at an early stage when they are easily curable and may not require systemic chemotherapy remains a challenge to clinicians. In order to improve early cancer detection, gas filled hollow boron-doped silica particles have been developed, which can be used for ultrasound-guided breast conservation therapy. The particles are synthesized using a polystyrene template and subsequently calcinated to create hollow, rigid nanoporous microspheres. The microshells are filled with perfluoropentane vapor. Studies were performed in phantoms to optimize particle concentration, injection dose, and the ultrasound settings such as pulse frequency and mechanical index. In vitro studies have shown that these particles can be continuously imaged by US up to 48 min and their signal lifetime persisted for 5 days. These particles could potentially be given by intravenous injection and, in conjunction with contrast-enhanced ultrasound, be utilized as a screening tool to detect smaller breast cancers before they are detectible by traditional mammography.
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Project description:BACKGROUND:The unique physicochemical properties of multi-walled carbon nanotubes (MWCNT) have led to many industrial applications. Due to their low density and small size, MWCNT are easily aerosolized in the workplace making respiratory exposures likely in workers. The International Agency for Research on Cancer designated the pristine Mitsui-7 MWCNT (MWCNT-7) as a Group 2B carcinogen, but there was insufficient data to classify all other MWCNT. Previously, MWCNT exposed to high temperature (MWCNT-HT) or synthesized with nitrogen (MWCNT-ND) have been found to elicit attenuated toxicity; however, their genotoxic and carcinogenic potential are not known. Our aim was to measure the genotoxicity of MWCNT-7 compared to these two physicochemically-altered MWCNTs in human lung epithelial cells (BEAS-2B & SAEC). RESULTS:Dose-dependent partitioning of individual nanotubes in the cell nuclei was observed for each MWCNT material and was greatest for MWCNT-7. Exposure to each MWCNT led to significantly increased mitotic aberrations with multi- and monopolar spindle morphologies and fragmented centrosomes. Quantitative analysis of the spindle pole demonstrated significantly increased centrosome fragmentation from 0.024-2.4 μg/mL of each MWCNT. Significant aneuploidy was measured in a dose-response from each MWCNT-7, HT, and ND; the highest dose of 24 μg/mL produced 67, 61, and 55%, respectively. Chromosome analysis demonstrated significantly increased centromere fragmentation and translocations from each MWCNT at each dose. Following 24 h of exposure to MWCNT-7, ND and/or HT in BEAS-2B a significant arrest in the G1/S phase in the cell cycle occurred, whereas the MWCNT-ND also induced a G2 arrest. Primary SAEC exposed for 24 h to each MWCNT elicited a significantly greater arrest in the G1 and G2 phases. However, SAEC arrested in the G1/S phase after 72 h of exposure. Lastly, a significant increase in clonal growth was observed one month after exposure to 0.024 μg/mL MWCNT-HT & ND. CONCLUSIONS:Although MWCNT-HT & ND cause a lower incidence of genotoxicity, all three MWCNTs cause the same type of mitotic and chromosomal disruptions. Chromosomal fragmentation and translocations have not been observed with other nanomaterials. Because in vitro genotoxicity is correlated with in vivo genotoxic response, these studies in primary human lung cells may predict the genotoxic potency in exposed human populations.
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Project description:BackgroundLoss of the genomic stability jeopardize genome stability and promote malignancies. A fraction of ovarian cancer (OvCa) arises from pathological mutations of DNA repair genes that result in highly mutagenic genomes. However, it remains elusive why the ovarian epithelial cells are particularly susceptible to the malfunction of genome surveillance system.MethodsTo explore the genotoxic responses in the unique context of microenvironment for ovarian epithelium that is periodically exposed to high-level steroid hormones, we examined estrogen-induced DNA damage by immunofluorescence in OvCa cell lines, animal and human samples.ResultsWe found that OvCa cells are burdened with high levels of endogenous DNA damage that is not correlated with genomic replication. The elevation of damage burden is attributable to the excessive concentration of bioactive estrogen instead of its chemomimetic derivative (tamoxifen). Induction of DNA lesions by estrogen is dependent on the expression of hormone receptors, and occurs in G1 and non-G1 phases of cell cycle. Moreover, depletion of homologous recombination (HR) genes (BRCA1 and BRCA2) exacerbated the genotoxicity of estrogen, highlighting the role of HR to counteract hormone-induced genome instability. Finally, the estrogen-induced DNA damage was reproduced in the epithelial compartments of both ovarian and fallopian tubes.ConclusionsTaken together, our study disclose that estrogen-induced genotoxicity and HR deficiency perturb the genome stability of ovarian and fallopian epithelial cells, representing microenvironmental and genetic risk factors, respectively.
| S-EPMC7579787 | biostudies-literature