In vitro studies of nanoparticle interactions within the human alveolar respiratory unit

Carbon nanotubes (CNTs) have attracted interest in many industries because of their novel and beneficial properties. Future exposure to public by inhalation is expected with the dramatic increase in manufacture and use of CNTs. Concurrently urban air pollution, including the contribution of diesel exhaust particles (DEPs), is already a heavy burden on public health on a global scale. The overarching hypothesis of the study was that the physicochemical properties of carbon-based nanoparticles would determine their bioreactivity and toxicity at the alveolar unit of the lung. As I was studying both CNTs and DEPs, I had two more specific hypotheses: 1) The cellular responses would be dependent on the source of exhaust emissions which determines the size and composition of the resultant DEPs. 2) The surface charge of multi-walled carbon nanotubes (MWCNTs) would determine the magnitude of the bioreactive responses in alveolar cells. Simple and complex in vitro models utilising the human alveolar type 1 cell line (TT1) and human primary cells: alveolar epithelial type 2 (AT2), alveolar macrophage (AM) and microvascular endothelial cells (HPMVEC) were used to test the hypotheses. The development and use of a complex in vitro cell model was a vital aspect of this study as it was imperative to understand the interaction not only between nanoparticles (NPs) and cells but also to gain an insight into the interaction between cells. Performing studies in simple and complex models comparatively allowed for greater awareness of the cell-cell interactions which inform toxicological outcomes following particle uptake including reactive oxygen species (ROS) formation and inflammation. Diesel particulate matter (PM) and carbon black induced moderate endothelial cell activation characterised by IL-6 release and a pro-thrombotic response. Recently generated automobile diesel PM and biodiesel PM were notably less reactive at the epithelium than forklift diesel PM and carbon black; however as all of the NPs were internalised by the epithelial type 1 cells and macrophages, the potential chronic and systemic effects of all of the particles should be considered. The conspicuous cytotoxicity of carbon black was likely a combination of its smaller size and surface activity. The MWCNTs were acutely reactive at the alveolar epithelium, inducing cytotoxicity, inflammation (including anti-protease production at even the lowest concentrations, 0.1 µg/ml) and ROS generation. All of the MWCNTs were internalised by the alveolar cells, by active and passive uptake mechanisms, however the positively charged CNT had distinctly more interaction with the alveolar cells and thus greater uptake, and even translocation in the epithelial type 1 cells. Of the MWCNTs, MAA and APTAC induced the smallest and greatest bioreactive responses respectively, in the cellular (simple and complex) models utilised, confirming the hypothesis. The association between nanoparticle exposure in humans to a number of adverse respiratory outcomes is well-established however the mechanisms by which these objects influence human health are still only partially understood. My study demonstrated that nanoparticle toxicity is dependent on their physicochemical properties, specifically charge in the case of MWCNTs, and generally size, as well as the cell models used. Understanding the toxicity of carbon nanomaterials, both anthropogenic and engineered, which are major contaminants of our environment and determinants of cardiopulmonary toxicity, is essential to generating safer nanomaterials and limiting airborne exposure to protect human health.Open Acces

Impaired expression of metallothioneins contributes to allergen-induced inflammation in patients with atopic dermatitis

Regulation of cutaneous immunity is severely compromised in inflammatory skin disease. To investigate the molecular crosstalk underpinning tolerance versus inflammation in atopic dermatitis, we utilise a human in vivo allergen challenge study, exposing atopic dermatitis patients to house dust mite. Here we analyse transcriptional programmes at the population and single cell levels in parallel with immunophenotyping of cutaneous immunocytes revealed a distinct dichotomy in atopic dermatitis patient responsiveness to house dust mite challenge. Our study shows that reactivity to house dust mite was associated with high basal levels of TNF-expressing cutaneous Th17 T cells, and documents the presence of hub structures where Langerhans cells and T cells co-localised. Mechanistically, we identify expression of metallothioneins and transcriptional programmes encoding antioxidant defences across all skin cell types, that appear to protect against allergen-induced inflammation. Furthermore, single nucleotide polymorphisms in the MTIX gene are associated with patients who did not react to house dust mite, opening up possibilities for therapeutic interventions modulating metallothionein expression in atopic dermatitis

Bioreactivity of a novel poly(epsilon-caprolactone) nanocapsule containing atrazine with human lung alveolar epithelial cells

Atrazine (ATZ), a commonly used agricultural herbicide, is potentially harmful to animals and humans. Nanoencapsulation of ATZ (NC–ATZ) within non-toxic, biodegradable poly(ε-caprolactone) (PCL) improves the herbicidal activity of ATZ 10-fold and moderates its environmental persistence, possibly reducing off-target ecological effects. These compounds may reach the pulmonary respiratory units following inhalation; thus, we investigated the effect of ATZ and NC–ATZ on an immortalised human lung alveolar type 1-like epithelial cell model (TT1 cells). The concentration-dependent effect of the compounds was analysed by assessment of viability, reactive oxygen species (ROS) production and inflammatory cytokine release. Confocal microscopy was used to visualise TT1 cell-nanoparticle interactions. NC–ATZ caused cellular effects not observed with ATZ or the PCL nanocapsule alone. NC–ATZ significantly increased lactate dehydrogenase (LDH) release at ≥1 μg ml−1 after 48 hours of exposure, peaking at 5 μg ml−1 (3-fold of the non-treated control, ***p < 0.001). Moreover, ≥1 μg ml−1 NC–ATZ was pro-inflammatory at 48 hours, peaking at 5 μg ml−1 (IL-6 release ∼125 pg ml−1; IL-8 release ∼46 pg ml−1). Confocal microscopy of fluorescently-labelled ATZ and NC–ATZ indicated high intensity fluorescence nanoparticle uptake into the cytoplasm and co-localisation in the Golgi, suggesting nanoparticle recycling within 24 hours. We provide evidence that nanoencapsulation of the pesticide ATZ alters bioreactivity, stimulating more necrosis and inflammation in human lung alveolar type 1 epithelial cells in comparison to ATZ or the PCL nanocapsule alone. However, nanoencapsulation improves the efficacy of pesticides, thus lower pesticide concentrations could be used, reducing environmental contamination. Further investigation, particularly with different exposure scenarios, is warranted in order to generate safer nanoencapsulated pesticides

Additional file 2: of Cerium dioxide nanoparticles exacerbate house dust mite induced type II airway inflammation

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