Research Article: Atomic Layer Deposition Coating of Carbon Nanotubes with Aluminum Oxide Alters Pro-Fibrogenic Cytokine Expression by Human Mononuclear Phagocytes In Vitro and Reduces Lung Fibrosis in Mice In Vivo

Date Published: September 12, 2014

Publisher: Public Library of Science

Author(s): Alexia J. Taylor, Christina D. McClure, Kelly A. Shipkowski, Elizabeth A. Thompson, Salik Hussain, Stavros Garantziotis, Gregory N. Parsons, James C. Bonner, Jörn Coers.


Multi-walled carbon nanotubes (MWCNTs) pose a possible human health risk for lung disease as a result of inhalation exposure. Mice exposed to MWCNTs develop pulmonary fibrosis. Lung macrophages engulf MWCNTs and produce pro-fibrogenic cytokines including interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, and osteopontin (OPN). Atomic layer deposition (ALD) is a novel process used to enhance functional properties of MWCNTs, yet the consequence of ALD-modified MWCNTs on macrophage biology and fibrosis is unknown.

The purpose of this study was to determine whether ALD coating with aluminum oxide (Al2O3) would alter the fibrogenic response to MWCNTs and whether cytokine expression in human macrophage/monocytes exposed to MWCNTs in vitro would predict the severity of lung fibrosis in mice. Uncoated (U)-MWCNTs or ALD-coated (A)-MWCNTs were incubated with THP-1 macrophages or human peripheral blood mononuclear cells (PBMC) and cell supernatants assayed for cytokines by ELISA. C57BL6 mice were exposed to a single dose of A- or U-MWCNTs by oropharyngeal aspiration (4 mg/kg) followed by evaluation of histopathology, lung inflammatory cell counts, and cytokine levels at day 1 and 28 post-exposure.

ALD coating of MWCNTs with Al2O3 enhanced IL-1β secretion by THP-1 and PBMC in vitro, yet reduced protein levels of IL-6, TNF-α, and OPN production by THP-1 cells. Moreover, Al2O3 nanoparticles, but not carbon black NPs, increased IL-1β but decreased OPN and IL-6 in THP-1 and PBMC. Mice exposed to U-MWCNT had increased levels of all four cytokines assayed and developed pulmonary fibrosis by 28 days, whereas ALD-coating significantly reduced fibrosis and cytokine levels at the mRNA or protein level.

These findings indicate that ALD thin film coating of MWCNTs with Al2O3 reduces fibrosis in mice and that in vitro phagocyte expression of IL-6, TNF-α, and OPN, but not IL-1β, predict MWCNT-induced fibrosis in the lungs of mice in vivo.

Partial Text

Multi-walled carbon nanotubes (MWCNTs) are fiber-like, engineered graphene nanomaterials that have a wide range of applications in engineering, electronics, and medicine. They are currently being utilized for their superior mechanical strength, large surface area and electrical conducting properties in many consumer products and for industrial purposes [1]. MWCNTs also have potential for biomedical applications, including drug delivery and scaffolds for tissue regeneration [2], [3]. Human exposure to MWCNTs will be inevitable due to increased production and use in a variety of consumer products, so it is extremely important to better understand the potential risks of MWCNTs to human health in order to ensure safe design of materials containing MWCNTs [4].

An increasing number of studies demonstrate that multi-walled carbon nanotubes (MWCNTs) cause fibrosis when delivered to the lung of rodents, suggesting a human health risk. Moreover, a variety of post-synthesis modifications, including atomic layer deposition (ALD), are used to enhance the unique properties of MWCNTs. In this study we investigated whether aluminum oxide (Al2O3) surface coating applied to MWCNTs by ALD would alter the expression of pro-inflammatory or pro-fibrogenic cytokines in vitro using two human mononuclear cell models (THP-1 and PBMC) and whether ALD-coating would alter MWCNT-induced cytokine levels and fibrosis in vivo in the lungs of C57BL6 mice. MWCNTs coated with Al2O3 by ALD (A-MWCNT), when compared to uncoated (U)-MWCNTs, increased IL-1β secretion and yet decreased the production of IL-6, OPN, and TNF-α in cultured THP-1 cells and PBMCs in vitro. A-MWCNTs delivered to the lungs of mice by OPA had reduced mRNA or protein levels of IL-1β, IL-6, OPN and TNF-α relative to U-MWCNT at 1 or 28 days post-exposure and A-MWCNTs caused less lung fibrosis in mice relative to U-MWCNTs. These findings suggest that A-MWCNTs engineered by ALD with Al2O3 would reduce the risk of pulmonary fibrosis in humans compared to U-MWCNTs. A generalized scheme to depict the overall findings of this study is shown in Fig. 8.