Dry Generation of CeO2 Nanoparticles and Deposition onto a Co-Culture of A549 and THP-1 Cells in Air-Liquid Interface—Dosimetry Considerations and Comparison to Submerged Exposure

Abstract

Relevant in vitro assays that can simulate exposure to nanoparticles (NPs) via inhalation are urgently needed. Presently, the most common method employed is to expose lung cells under submerged conditions, but the cellular responses to NPs under such conditions might differ from those observed at the more physiological air-liquid interface (ALI). The aim of this study was to investigate the cytotoxic and inflammatory potential of CeO2 NPs (NM-212) in a co-culture of A549 lung epithelial cells and differentiated THP-1 cells in both ALI and submerged conditions. Cellular dose was examined quantitatively using inductively coupled plasma mass spectrometry (ICP-MS). The role of serum and LPS-priming for IL-1β release was further tested in THP-1 cells in submerged exposure. An aerosol of CeO2 NPs was generated by using the PreciseInhale® system, and NPs were deposited on the co-culture using XposeALI®. No or minor cytotoxicity and no increased release of inflammatory cytokines (IL-1β, IL-6, TNFα, MCP-1) were observed after exposure of the co-culture in ALI (max 5 µg/cm2) or submerged (max 22 µg/cm2) conditions. In contrast, CeO2 NPs cause clear IL-1β release in monocultures of macrophage-like THP-1, independent of the presence of serum and LPS-priming. This study demonstrates a useful approach for comparing effects at various in-vitro conditions.

Toxicological effects of zinc oxide nanoparticle exposure: an in vitro comparison between dry aerosol air-liquid interface and submerged exposure systems

“…Increased expression of multiple cytokines, for example IL-8, in hAELVi (human alveolar epithelial lentivirus immortalized) cells was seen in Mills-Goodlet et al, but where similar dose levels as in the current study were used. However, two other studies showed similar cellular responses in both ALI and submerged cultures, except for in the release of LDH, where higher levels were found in the ALI cultures (Cappellini et al 2020;Medina-Reyes et al 2020). These similarities were seen even though several study parameters varied between the two studies.…”

“…To ensure that the ALI exposure systems can be used for NP exposures, each system needs to be evaluated with different categories of NPs, and comparative studies between traditionally submerged and ALI exposure systems need to be performed. Multiple ALI systems have already been used in such comparative studies, for example MINUCELL (Lenz et al 2013), VITROCELL (Xie et al 2012;Panas et al 2014;Mihai et al 2015;Loret et al 2016;Hilton et al 2019;Medina-Reyes et al 2020;Mills-Goodlet et al 2020), XposeALI (Cappellini et al 2020), and ALICE (Lenz et al 2009;Stoehr et al 2015), and most of them have reported more pronounced effects with the ALI exposure systems than with the submerged at similar dose levels. However, a comparison between the NACIVT system and traditional submerged exposure systems has not previously been performed.…”

Air–Liquid Interface Exposure of Lung Epithelial Cells to Low Doses of Nanoparticles to Assess Pulmonary Adverse Effects

“…Meanwhile, there are a few published in vitro ALI studies which tested CeO 2 NPs [ 30 , 61 , 62 ]. Steiner et al [ 63 ] exposed a 3D coculture model prepared from A549 cells, human monocyte-derived macrophages and dendritic cells under ALI conditions to a CeO 2 NP suspension (0.75 µg/cm 2 ) employing a microsprayer and found increased expression of HO-1, but not of SOD1, TNF-α, IL-8 and no release of LDH.…”

“…These findings are in line with a later report employing an XposeALI system depositing CeO 2 NM-212 (max. 5 µg/cm 2 ) within 5–20 min onto a coculture of A549 and THP-1 where no major effects on cytotoxicity and cytokine release became evident [ 61 ]. Similarly, and also using a VITROCELL exposure system, adverse responses were investigated in A549, BEAS-2B (max.…”

Impact of mono-culture vs. Co-culture of keratinocytes and monocytes on cytokine responses induced by important skin sensitizers

“…The human monocytic THP-1 line was grown in RPMI-1640 supple-mented 10% FBS, 1% penicillin/streptomycin, and 1% L-glutamine. For the THP-1 mono-culture, cells were plated as described in Cappellini et al (2020) at 4 Â 10 4 cells/well (80 ml) in 96-well plates and treated with 80 ml of medium containing the test chemical or vehicle control (160 ml total volume/well).…”