Toxicology in Vitro最新文献

Background

Commensal bacteria colonizing oral mucosa and skin play an essential role in maintaining host-microbiome homeostasis. It is unknown whether cytotoxicity resulting from metal ions leaching from medical devices may be influenced by commensal microbes.

Objective

Determine whether the extent of apoptosis triggered by nickel or titanium ions is influenced by Streptococcus mitis and whether apoptosis occurs via the intrinsic or extrinsic apoptosis pathway.

Methods

Reconstructed Human Gingiva (RHG) and Skin (RHS) were topically exposed to titanium or nickel salts in the presence or absence of S. mitis. Cytotoxicity and apoptosis were assessed by histology, immunohistochemistry, TUNEL assay, and Western Blot.

Results

S. mitis alone resulted in negligible cytotoxicity. After metal exposure, localized apoptosis was observed in the epithelium and fibroblasts within the lamina propria hydrogel of both RHG and RHS. S. mitis enhanced metal-mediated apoptosis in gingiva but not in skin. Apoptosis was mediated via the extrinsic pathway caspase 8. Activation of the execution phase of apoptosis occurred via caspases 3 and 7, and PARP-1.

Conclusion

Our study supports the finding that metals have irritant, cytotoxic properties resulting in apoptosis when leaching into skin or gingiva. Particularly for gingiva, commensal microbes exaggerate this detrimental effect.

Studying percutaneous penetration of various cosmetic ingredients through intact and compromised skin can provide insight on quantitative exposure assessment for baby products intended for diapered skin. We developed an in vitro model (tape-stripped human skin) designed to achieve the Trans-Epidermal Water Loss values measured in babies with various degrees of diaper dermatitis. Six reference compounds showed the impact of physicochemical properties on absorption through this “diaper rash” skin model. Under simulated diaper conditions, dermal absorption of cosmetic ingredients (phenoxyethanol, sodium benzoate, benzyl alcohol, disodium EDTA, and propylene glycol) was different, but <100%. Additionally, the effect of diaper rash on dermal absorption of well-absorbed ingredients (phenoxyethanol, sodium benzoate, and benzyl alcohol) was limited (enhancement of 1.1–1.3), while the enhancement for moderately absorbed compounds (disodium EDTA and propylene glycol) was 1.8–3.3. Absorption via skin with “diaper rash” is specific to individual ingredients and exposure conditions, so a fixed uncertainty factor is not appropriate for safety assessment. The data support that the default 100% dermal absorption commonly used in first-tier risk assessments for diapered skin is conservative. This diaper rash skin model provides a practical tool of estimating absorption of various ingredients in baby products intended for diapered skin.

The thyroid gland, a vital component of the endocrine system, plays a pivotal role in regulating metabolic processes, growth, and development. To better characterize thyroid system disrupting chemicals (TSDC), we followed the next-generation risk assessment approach, which further considers the mechanistic profile of xenobiotics. We combined targeted in vitro testing with untargeted metabolomics. Four known TSDC, propyl-thiouracil (PTU), sodium perchlorate, triclosan, and 5-pregnen-3β-ol-20-one-16α‑carbonitrile (PCN) were investigated using rat in vitro models, including primary hepatocytes, PCCL3 cells, thyroid microsomes, and three-dimensional thyroid follicles. We confirmed each compound's mode of action, PTU inhibited thyroperoxidase activity and thyroid hormones secretion in thyroid cells model, sodium perchlorate induced a NIS-mediated iodide uptake decrease as triclosan to a lesser extent, and PCN activated expression and activity of hepatic enzymes (CYPs and UGTs) involved in thyroid hormones metabolism. In parallel, we characterized intracellular metabolites of interest. We identified disrupted basal metabolic pathways, but also metabolites directly linked to the compound's mode of action as tyrosine derivates for sodium perchlorate and triclosan, bile acids involved in beta-oxidation, and precursors of cytochrome P450 synthesis for PCN. This pilot study has provided metabolomic fingerprinting of dedicated TSDC exposures, which could be used to screen and differentiate specific modes of action.

Studies on in vitro-in vivo correlations of inflammatory and genotoxic responses are needed to advance new approach methodologies. Here, we assessed pro-inflammatory and genotoxic responses by 13 nanosized metal oxides (nMeOx) and quartz (DQ12) in alveolar epithelial cells (A549) and macrophages (THP-1a) exposed in submerged conditions, and in A549:THP-1a co-cultures in air-liquid interface (ALI) system. Soluble nMeOx produced the highest IL-8 expression in A549 and THP-1a cells in submerged conditions (≥2-fold, p < 0.05), whereas only CuO caused a strong response in co-cultures exposed in the ALI system (13-fold, p < 0.05). IL-8 expression in A549 cells with concentrations as nMeOx specific surface area (SSA) correlated with neutrophil influx in mice (r = 0.89–0.98, p < 0.05). Similarly, IL-8 expression in THP-1a cell with concentrations as mass and SSA (when excluding soluble nMeOx) correlated with neutrophil influx in mice (r = 0.81–0.84, p < 0.05). DNA strand breaks (SB) was measured by the comet assay. We used a scoring system that categorizes effects in standard deviation units for comparison of genotoxicity in different models. Concordant genotoxicity was observed between SB levels in vitro (A549 and co-culture) and in vivo (broncho-alveolar lavage fluid cells and lung tissue). In conclusion, this study shows in vitro-in vivo correlations of nMeOx-induced inflammatory and genotoxic responses.

The EU-ToxRisk project (2016–2021) was a large European project working towards shifting toxicological testing away from animal tests, towards a toxicological assessment based on comprehensive mechanistic understanding of cause-consequence relationships of chemical adverse effects. More than 40 partners from scientific institutions, industry and regulators coordinated their work towards this goal in a six-year long programme. The breadth and variety of data and knowledge generated, presented a challenging data management landscape.

Here, we describe our approach to data management as developed under EU-ToxRisk. The main building blocks of the data infrastructure are: 1) An easy-to-use, extensible data and metadata format; 2) A flexible system with protocols for data capture and sharing from the entire consortium; 3) A methods database for describing and reviewing data generation and processing protocols; 4) Data archiving using a sustainable resource; 5) Data transformation from the archive to the system that provides granular access; 6) Application Programming Interface (API) for access to individual data points; 7) Data exploration and analysis modules, based on a «web notebook» approach to executable data processing documentation; and 8) Knowledge portal that ties together all of the above and provides a collaboration space for information exchange across the consortium. This knowledge infrastructure is being extended and refined for the support of follow-up projects (RISK-HUNT3R, ASPIS cluster, European Open Science Cloud (2021–2026)).

Per- and Polyfluoroalkyl substances (PFAS) are a group of persistent long-lived chemicals with global environmental contamination. The published literature is rife with confusing and sometimes contradictory effects of PFAS on animal and cell models, as well as epidemiological studies. Cytotoxicity studies are often used as an early indicator to guide safety requirements, regulation, and further studies and thus can be useful to understand important toxicity differences by various PFAS. Recent studies have found that PFAS are not equivalently toxic on all cell types, and that not all cell types exhibit the same sensitivity to individual PFAS. However, immune cells have not been well studied. As immune cells are important for regulating responses to environmental toxins, infection, and cancer, we sought to discover the sensitivity of these cells to various PFAS, including legacy and replacement compounds. We assessed a range of concentrations and found that immune cells are generally more robust when exposed to PFAS, and that Jurkat T-cells were more sensitive than THP-1 monocytes. As monocytes are critical for coordinating inflammatory responses to external threats with cell death cascades, we further investigated these cells. We discovered that THP-1 cells do not undergo organized or programmed death, such as apoptosis or pyroptosis, and instead PFAS exposure results in a more necrotic/lytic and unorganized death, likely contributing to potential inflammatory effects downstream.

Hydroquinone (HQ) is one of benzene metabolites that can cause oxidative stress damage and Homologous recombination repair (HR). A good deal of reactive oxygen species (ROS) generated by oxidative stress can trigger apoptotic signaling pathways. The nuclear factor erythroid 2-related factor 2 (Nrf2) can regulate the cell response to oxidative stress damage. The aim of this study was to explore whether Nrf2 participate in HQ-induced apoptosis and its mechanism. The findings displayed that HQ triggered HR, promoted Nrf2 transfer into the cell nucleus and induced cell apoptosis, while Nrf2 deficient elevated cell apoptosis, attenuated the expression of PARP1 and RAD51. We also observed that Nrf2 deficient triggered Caspase-9. Thus, we speculated that Nrf2 might participate in HQ-induced cell apoptosis through Caspase-9 dependent pathways. Meanwhile, Nrf2 participated in HQ-induced DNA damage repair by regulating the level of PARP1 and RAD51.

Mimicking the microenvironment of seminiferous tubules plays an indispensable role in directing differentiation of stem cells toward germ cells in vitro. In this work, we fabricated electrospun gelatin (EG) mats (i.e., with diameter <500 nm) conditioned with Sertoli cells' extracellular matrix (ECM) to simulate both 3D structures and composition of normal testis tissue. Sertoli cells were isolated from mice testis and represented through immunocytochemistry (ICC) staining for expression of vimentin, a specific marker of Sertoli cells. The morphological characteristics of ECM-coated scaffold were investigated under scanning electron microscope (SEM). The efficient elimination of cells was confirmed by MTT assay. Furthermore, the cyto/biocompatibility of ECM-conditioned EG scaffold was determined for Sertoli cells and embryonic stem cells (ESCs), alone and as in co-culture. According to the results, the designed scaffold provided a mat for cell proliferation with negligible toxicity (almost 100% cell viability). SEM micrographs displayed cells with elongated shape and complete stretching morphology when compared with those cultured on scaffold without ECM. Moreover, an enhanced differentiation of ESCs toward sperm-generating cells was obtained through co-culturing of Sertoli cells and ESCs, where cell viability was found almost 100%. Our findings introduce the ECM-conditioned EG scaffold as a potentially influential engineered substrate for in vitro guidance of stem cells differentiation by mimicking the native microenvironment.

The application of carbon nanomaterials in diverse fields has substantially increased their demand for commercial usage. Within the earliest decade, the development of functional materials has further increased the significance of this element. Despite the advancements recorded, the potential harmful impacts of embracing carbon nanomaterials for biological applications must be balanced against their advantages. Interestingly, many studies have neglected the intriguing and dynamic cellular interaction of carbon nanomaterials and the mechanistic understanding of their property-driven behaviour, even though common toxicity profiles have been reported. Reiterating the toxicity issue, several researchers conclude that these materials have minimal toxicity and may be safe for contact with biological systems at certain dosages. Here, we aim to provide a report on the significance of some of the properties that influence their toxicity. After that, a description of the implication of nanotoxicology in humans and living systems, revealing piece by piece their exposure routes and possible risks, will be provided. Then, an extensive discussion of the mechanistic puzzle modulating the interface between various human cellular systems and carbon nanomaterials such as carbon nanotubes, carbon dots, graphene, fullerenes, and nanodiamonds will follow. Finally, this review also sheds light on the organization that handles the risk associated with nanomaterials.

Natural pyrethrins (NPs) are insecticidal compounds isolated and extracted from pyrethrum flowers and are primarily use to control sanitary pests. The lungs become the main target after exposure, and its use may pose potential hazards to respiratory health. Therefore, in this paper, the toxic effects of NPs on human lung cells A549 were investigated and the risk of respiratory toxicity of NPs was studied using zebrafish swim bladder as a model. The results showed that NPs induced cytotoxicity, caused oxidative DNA damage and triggered mitochondria-mediated apoptosis. In addition, exposure to NPs decreased zebrafish embryo survival, hatchability, and heartbeat, and may inhibit normal swim bladder development by disrupting Wnt and Hedgehog signaling pathways. In conclusion, our results suggest that NPs can induce cytotoxicity in A549 in vitro and developmental toxicity in zebrafish in vivo. This study provides a conceptual basis for understanding the mechanisms of toxicity of NPs and assessing respiratory health risks in humans.