Schlüsselwörter
(Deutsch)
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Nanoparticle exposure, EBC exhaled breath condensate, NTA nanoparticle tracking analysis, oxidative stress markers, passive smoking
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Kurzbeschreibung
(Deutsch)
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Die Studie soll abklären, ob sich die Nanoparticle Tracking Analysis (NTA) eignet, um in der kondensierten Ausatemluft (EBC) Nanopartikel nachzuweisen. Damit sind Rückschlüsse auf die effektive Nanopartikel-Belastung in der Lunge möglich. Der Nachweis der oxidativen Stressprodukte Wasserstoffperoxid und Malondialdehyd kann ebenfalls auf eine Partikelbelastung der Lunge hindeuten. Als Partikelquelle wird in der Studie Tabakrauch verwendet.
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Projektziele
(Deutsch)
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Introduction
A central hypothetical mechanism to explain the adverse effects of particulate matter, including
unintentionally or engineered nanoparticles (NP), is the ability of some components of such a
complex mixture to induce oxidative stress. Low levels of oxidative stress are associated with
the induction of antioxidant and detoxification enzymes. At higher levels of oxidative stress, this
protective response is overtaken by inflammation and cytotoxicity.
A fundamental principle of toxicology is that response is proportional to the concentration of the
affecter molecule at the site of action. Thus, the use of target tissue dose rather than less specific
measures of dose such as exposure or administered dose has been shown to improve the
dose-response correlations for drugs, chemicals and inhaled gases as well as particles
In the case of particle/nanoparticle exposure. the measurement of such a target tissue dose
leading to a biological effect (inflammation) is difficult to achieve. Many studies aimed at quantifying
airway inflammation have used either invasive (e.g. bronchoalveolar lavage fluids) or
semi-invasive techniques (e.g. sputum induction). Others have measured inflammatory mediators
in plasma or urine, which probably reflect systemic rather than organ specific inflammation.
By contrast, measurements of inflammatory markers in exhaled air or breath condensate are
completely non-invasive and are likely to reflect inflammation in the respiratory tract. As its
name indicates, exhaled breath condensate (EBC) corresponds to the liquid collected after condensing
exhaled air at low temperature. Although most of the condensate consists of water,
available evidences suggest that non-volatile substances in the lower respiratory tract can be
transported in the form of aerosols in exhaled breath. EBC is thus reported to reflect the composition
of the airway lining fluid. Proteins, eicosanoids and hydrogen peroxide (H202) among
others, have been found to be constituents of these aerosols. pH value of the EBC reflects the
airway homeostasis and is a simple and useful parameter to monitor. In addition, measurement
of exhaled nitric oxide (NO) has become an accepted method of monitoring inflammation in the
lung.
As lining fluid is the first media NP encounter when deposited on the lungs, it is expected that
oxidative stress and its subsequent release of end products will modify such media. Indeed different
markers of oxidative stress and/or inflammatory mediators have been detected in EBC
from healthy subjects exposed to particles. For example, cigarette smokers had a 5-fold higher
mean EBC H202 level than non-smokers. Malondialdehyde (MDA), a marker of oxidative reaction
on polyunsaturated fatty acids (lipoperoxidation), has been shown to increase in EBC of
asthmatic children exposed to polluted air, in workers exposed to cobalt and tungsten, in
healthy subjects after exercise with high PM, exposure, or in healthy subjects exposed to wood
smoke.
The number and the agglomerate size distribution of NP in a solution can be determined by a
technique called nanoparticle tracking analysis (NTA). It is necessary to have particle concentrations
in the order of 10'-109 particles/ml. The estimated particle concentration in EBC after an
8 hours exposure to a particle concentration typical for a polluted urban environment is _ 10'
particles/ml and is thus at the lower detection range of such a technique. Therefore it remains to
be demonstrated whether this technique proves to be adequate for NP detection in EBC.
Objectives
The objectives of the proposed pilot study are to:
A. Determine the feasibility of using NTA analysis on EBC solutions as a measure of particle
deposition in the lung.
B. Set up methodologies for determining oxidative stress markers (H2O2 and malondialdehyde)
in EBC.
The hypothesis which are underlying this pilot project are:
H1: Exhaled breath condensate (EBC) is representative of the lung lining fluid
H2: Particle concentration in the EBC reflects the deposited internal dose on the alveolar epithelial
cells (low or predictable condensation of the inhaled and un-deposited NP)
H3: H202 and malondialdehyde can be considered as biomarkers of effect due to the oxidative
stress generated by the lung deposited NP.
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Abstract
(Deutsch)
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Die Studie hat gezeigt, dass mit NTA Nanopartikel im EBC nachweisbar sind. Hingegen ist das verwendete Protokoll ungeeignet, um die Partikeldosis in der Lunge abzuschätzen, da nur die Zahl, nicht aber die Zusammensetzung und somit die Herkunft der in der Lunge deponierten Partikel bestimmt wird. Zu diesem Zweck müsste mit markierten Partikeln gearbeitet werden. Die Methoden für die Bestimmung der Marker für Entzündung und oxidativen Stress waren zwar empfindlich genug, um die entsprechenden Marker im EBC nachzuweisen, hingegen müssten mit methodologischen Verbesserungen mögliche Störfaktoren ausgeschaltet werden können. Die MDA-Messung wurde zudem als Biomarker auf individueller Ebene als ungeeignet betrachtet. Die Verwendung von Tabakrauch als Partikelquelle erwies sich aufgrund des Gehalts an reaktiven und möglicherweise interferierenden Substanzen als ungünstig gewählt, war jedoch die einzige Möglichkeit einer Partikelexposition, die aus ethischen Gründen akzeptiert werden kann.
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Umsetzung und Anwendungen
(Deutsch)
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Dieses Vorprojekt zeigte, dass Partikel und Entzündungs- oder Stressmarker im EBC grundsätzlich nachweisbar sind, dass die Methoden die Abschätzung der Partikeldosis in der Lunge noch nicht erlauben und weiter entwickelt werden müssten.
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