Effect of silver nanoparticles and microbial ligands on the innate immune response of the human alveolar epithelium
File(s)
Author(s)
Zambianchi, Marta
Type
Thesis
Abstract
Over the past decade there has been a rapid increase in the development and use of nanoparticles (NPs, <100nm). Silver (Ag) NPs, due to their antimicrobial activity, are the most commercialised NPs in the world and are incorporated into several consumer goods. However, little is known about the potential adverse effects of AgNPs in humans. Approximately 50% of inhaled NPs preferentially deposit in the alveoli, where alveolar epithelial type-I cells (AT1) form 95% of the alveolar surface, thus constituting a significant target for deposition.
We hypothesised that (I) inhalation of AgNPs induces oxidative stress in AT1 cells, leading to a pro-inflammatory response, autophagy activation and DNA damage; (II) in the presence of a microbial infection, AgNP exposure potentiates the innate immune response, leading to increased inflammation and genotoxicity.
An immortalised human transformed alveolar type1-like cell line (TT1) was used to investigate these hypotheses. TT1 cells were exposed to AgNPs in the presence/absence of Poly I:C (TLR-3 ligand) and LPS (TLR-4 ligand) prior to assaying cell viability, inflammatory mediator release, oxidative stress, autophagy and DNA damage.
AgNPs induced oxidative stress in TT1 cells, as well as inflammation, autophagy activation and DNA damage. The AgNPs-Poly I:C combination further stimulated IL-6 release, DNA damage and delayed autophagic flux, whereas the AgNPs-LPS combination did not. We also discovered two different sources of AgNP-mediated oxidative stress, mitochondria and NADPH oxidase. Pre-treatment of TT1 cells with a TLR-4 inhibitor prevented AgNP-induced inflammation, autophagy activation and DNA damage. On the contrary, TLR-3 inhibition did not affect any of the pathways investigated. In conclusion, we showed that AgNPs induce oxidative stress and act via TLR-4, leading to a pro-inflammatory response, autophagy activation and DNA damage. These cellular pathways are potentiated by co-exposure with Poly I:C. This suggests that inhalation of AgNPs may exert deleterious effects on the lung and, in presence of a viral infection, a worsening of the pulmonary condition.
We hypothesised that (I) inhalation of AgNPs induces oxidative stress in AT1 cells, leading to a pro-inflammatory response, autophagy activation and DNA damage; (II) in the presence of a microbial infection, AgNP exposure potentiates the innate immune response, leading to increased inflammation and genotoxicity.
An immortalised human transformed alveolar type1-like cell line (TT1) was used to investigate these hypotheses. TT1 cells were exposed to AgNPs in the presence/absence of Poly I:C (TLR-3 ligand) and LPS (TLR-4 ligand) prior to assaying cell viability, inflammatory mediator release, oxidative stress, autophagy and DNA damage.
AgNPs induced oxidative stress in TT1 cells, as well as inflammation, autophagy activation and DNA damage. The AgNPs-Poly I:C combination further stimulated IL-6 release, DNA damage and delayed autophagic flux, whereas the AgNPs-LPS combination did not. We also discovered two different sources of AgNP-mediated oxidative stress, mitochondria and NADPH oxidase. Pre-treatment of TT1 cells with a TLR-4 inhibitor prevented AgNP-induced inflammation, autophagy activation and DNA damage. On the contrary, TLR-3 inhibition did not affect any of the pathways investigated. In conclusion, we showed that AgNPs induce oxidative stress and act via TLR-4, leading to a pro-inflammatory response, autophagy activation and DNA damage. These cellular pathways are potentiated by co-exposure with Poly I:C. This suggests that inhalation of AgNPs may exert deleterious effects on the lung and, in presence of a viral infection, a worsening of the pulmonary condition.
Version
Open Access
Date Issued
2014-08
Date Awarded
2014-01
Copyright Statement
Attribution NoDerivatives 4.0 International Licence (CC BY-ND)
Advisor
Thorley, Andrew
Sponsor
National Heart & Lung Institute
Imperial College London
Publisher Department
National Heart & Lung Institute
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)