Development of lung tissue models and their applications.

The lungs are important organs that play a critical role in the development of specific diseases, as well as responding to the effects of drugs, chemicals, and environmental pollutants. Due to the ethical concerns around animal testing, alternative methods have been sought which are more time-effective, do not pose ethical issues for animals, do not involve species differences, and provide easy investigation of the pathobiology of lung diseases. Several national and international organizations are working to accelerate the development and implementation of structurally and functionally complex tissue models as alternatives to animal testing, particularly for the lung. Unfortunately, to date, there is no lung tissue model that has been accepted by regulatory agencies for use in inhalation toxicology. This review discusses the challenges involved in developing a relevant lung tissue model derived from human cells such as cell lines, primary cells, and pluripotent stem cells. It also introduces examples of two-dimensional (2D) air-liquid interface and monocultured and co-cultured three-dimensional (3D) culture techniques, particularly organoid culture and 3D bioprinting. Furthermore, it reviews development of the lung-on-a-chip model to mimic the microenvironment and physiological performance. The applications of lung tissue models in various studies, especially disease modeling, viral respiratory infection, and environmental toxicology will be also introduced. The development of a relevant lung tissue model is extremely important for standardizing and validation the in vitro models for inhalation toxicity and other studies in the future.

Airborne emissions from combustion of graphene nanoplatelet/epoxy composites and their cytotoxicity on lung cells via air-liquid interface cell exposure in vitro.

Graphene nanoplatelet (GNP) as a nanofiller improves the mechanical strength, electrical conductivity, and flame retardancy of the polymers significantly. With an increasing number of GNP-reinforced products, a careful safety assessment is needed to avoid social and economic setbacks. However, no study has addressed the effects of combustion-generated emissions from GNP-reinforced products in the lung, the most sensitive exposure route to airborne particles. Therefore, we studied the influence of GNP as a nanofiller on the emitted particles and polycyclic aromatic hydrocarbons (PAHs), and cytotoxicity of the emissions from the combustion of pure epoxy (EP) and GNP-reinforced epoxy (EP-GNP). GNP was not detected in the airborne emissions. PAHs were found in airborne particles of both emissions from EP and EP-GNP, with some differences in their concentrations. A first hazard assessment was performed on human alveolar epithelial cells exposed to the airborne emissions at air-liquid interface conditions. At 24 h and 96 h after the exposure, similar responses were observed between EP and EP-GNP except an acute transient decrease in mitochondrial activity after exposure to the emissions from EP-GNP. Both emissions from EP and EP-GNP had no acute effects on membrane integrity, cell morphology or expression of anti-oxidative stress markers (HMOX1 and SOD2 genes). Meanwhile, both emissions induced the activation of the aryl hydrocarbon receptor (CYP1A1 gene) and a transient (pro-) inflammatory response (MCP-1), but the effects between EP and EP-GNP were not significantly different.

Hazard assessment of abraded thermoplastic composites reinforced with reduced graphene oxide.

Graphene-related materials (GRMs) are subject to intensive investigations and considerable progress has been made in recent years in terms of safety assessment. However, limited information is available concerning the hazard potential of GRM-containing products such as graphene-reinforced composites. In the present study, we conducted a comprehensive investigation of the potential biological effects of particles released through an abrasion process from reduced graphene oxide (rGO)-reinforced composites of polyamide 6 (PA6), a widely used engineered thermoplastic polymer, in comparison to as-produced rGO. First, a panel of well-established in vitro models, representative of the immune system and possible target organs such as the lungs, the gut, and the skin, was applied. Limited responses to PA6-rGO exposure were found in the different in vitro models. Only as-produced rGO induced substantial adverse effects, in particular in macrophages. Since inhalation of airborne materials is a key occupational concern, we then sought to test whether the in vitro responses noted for these materials would translate into adverse effects in vivo. To this end, the response at 1, 7 and 28 days after a single pulmonary exposure was evaluated in mice. In agreement with the in vitro data, PA6-rGO induced a modest and transient pulmonary inflammation, resolved by day 28. In contrast, rGO induced a longer-lasting, albeit moderate inflammation that did not lead to tissue remodeling within 28 days. Taken together, the present study suggests a negligible impact on human health under acute exposure conditions of GRM fillers such as rGO when released from composites at doses expected at the workplace.

Effects of Combining Graphene Nanoplatelet and Phosphorous Flame Retardant as Additives on Mechanical Properties and Flame Retardancy of Epoxy Nanocomposite.

The effects of combining 0.1-5 wt % graphene nanoplatelet (GNP) and 3-30 wt % phosphorous flame retardant, 9,10- dihydro-9-oxa-10-phosphaphenanthrene-10-oxide (DOPO) as fillers in epoxy polymer on the mechanical, flame retardancy, and electrical properties of the epoxy nanocomposites was investigated. GNP was homogeneously dispersed into the epoxy matrix using a solvent-free three-roll milling process, while DOPO was incorporated into the epoxy resin by mechanical stirring at elevated temperature. The incorporation of DOPO reduced the crosslinking density of the epoxy resin. When using polyetheramine as a hardener, the structural rigidity effect of DOPO overshadowed the crosslinking effect and governed the flexural moduli of epoxy/DOPO resins. The flexural moduli of the nanocomposites were improved by adding GNP up to 5 wt % and DOPO up to 30 wt %, whereas the flexural strengths deteriorated when the GNP and DOPO loading were higher than 1 wt % and 10 wt %, respectively. Limited by the adverse effects on mechanical property, the loading combinations of GNP and DOPO within the range of 0-1 wt % and 0-10 wt %, respectively, in epoxy resin were further studied. Flame retardancy index (FRI), which depended on three parameters obtained from cone calorimetry, was considered to evaluate the flame retardancy of the epoxy composites. DOPO showed better performance than GNP as the flame retardant additive, while combining DOPO and GNP could further improve FRI to some extent. With the combination of 0.5 wt % GNP and 10 wt % DOPO, improvement in both mechanical properties and flame retardant efficiency of the nanocomposite was observed. Such a combination did not affect the electrical conductivity of the nanocomposites since the percolation threshold was at 1.6 wt % GNP. Our results enhance the understanding of the structure-property relationship of additive-filled epoxy resin composites and serve as a property constraining guidance for the composite manufacturing.

Release of graphene-related materials from epoxy-based composites: characterization, quantification and hazard assessment in vitro.

Due to their mechanical strength, thermal stability and electrical conductivity, graphene-related materials (GRMs) have been extensively explored for various applications. Moreover, GRMs have been studied and applied as fillers in polymer composite manufacturing to enhance the polymer performance. With the foreseen growth in GRM production, occupational and consumer exposure is inevitable, thus raising concerns for potential health risks. Therefore, this study aims (1) to characterize aerosol particles released after mechanical abrasion on GRM-reinforced epoxy composites, (2) to quantify the amounts of protruding and free-standing GRMs in the abraded particles and (3) to assess the potential effects of the pristine GRMs as well as the abraded particles on human macrophages differentiated from the THP-1 cell line in vitro. GRMs used in this study included graphene nanoplatelets (GNPs), graphene oxide (GO), and reduced graphene oxide (rGO). All types of pristine GRMs tested induced a dose-dependent increase in reactive oxygen species formation, but a decrease in cell viability was only detected for large GNPs at high concentrations (20 and 40 µg mL-1). The particle modes measured using a scanning mobility particle sizer (SMPS) were 300-400 nm and using an aerodynamic particle sizer (APS) were between 2-3 µm, indicating the release of respirable particles. A significant fraction (51% to 92%) of the GRMs embedded in the epoxy composites was released in the form of free-standing or protruding GRMs in the abraded particles. The abraded particles did not induce any acute cytotoxic effects.