Visit our Expo - Redox and Inflammation signaling 2012

Session XIV : Transcriptional and translational control Poster XIV, 5
Gene expression modulation in A549 human lung cells in response to combustiongenerated
nano-sized particles.
Christa Baumstark-Khan, Andrea Arenz, Christine E. Hellweg and Horst-Henning
Grotheer.
Cellular Biodiagnostics, Department of Radiobiology, Institute of Aerospace Medicine,
German Aerospace Center, 51170 Köln, Germany. E-mail : christa.baumstarkkhan@dlr.de
Cell response to different kinds of environmental pollution is complex and involves the
participation of different classes of genes for different cellular outcomes (DNA repair, cell
cycle control, signal transduction, inflammation, apoptosis and oncogenesis). Ambient
particulate matter (PM) is a complex mixture of chemicals and particles that may be
compositionally diverse depending on geography and season. High levels of ambient air
pollution are associated with aggravation of asthma, respiratory morbidity, and
cardiopulmonary morbidity; long-term exposures to PM have been linked to possible
increases in lung cancer risk, chronic respiratory disease, and death rates. Nanometer-sized
particles (diameter 1.5 to 5 nm, mass numbers ranging from 1000 to 40.000 amu) which are
generated by combustion processes as soot precursors show surprising features, such as water
solubility and transparency, in contrast to the better known soot particles. The damaging
effects of nanoparticles are attributed to the fact that they are respirable and water soluble and
can penetrate lung mucosa. These particles are less readily cleared than fine particles, thereby
prolonging interaction with the lung epithelium and potentiating cellular damage. Particulate
matter has been shown to invoke inflammatory responses after exposure in animal models. Invitro
experiments have revealed that diesel exhaust and PM10 are capable to induce the
release of proinflammatory cytokines such as IL-6 and IL-8 from bronchial epithelial cells
mediated by the transcription factor NF-!B through a mechanism partially involving TNF- ".
Thus, an important aspect of our work was to determine whether nanoparticles cause
activation of NF-!B and expression of NF-!B dependent genes in pulmonary epithelial cells.
Recombinant A549 lung cells (A549-NF-!B-EGFP) were incubated with different
concentrations of condensation water samples collected from model flames (combustion of
gaseous fuels propane and ethylene under laboratory conditions). TNF- " treated cells were
used for comparison. RNA was extracted from exposed cells after various recovery times and
a real-time QRT-PCR assay was applied, which employs relative quantification of candidate
mRNA biomarkers. The expressions of different DNA damage inducible genes (GADD45#,
p21) and NF-kB dependent genes (NFkBIA, IL-6, “!B-EGFP”) were analysed. The results
show a reproducible up-regulation for the NF-kB dependent genes IL-6 (32x) and NF!BIA
(13x) with maximal values for 1 and 2 hrs treatment with TNF-", while the DNA damage
inducible genes are not induced. For nanoparticle treated cells, a down-regulation of NF-!B
dependent genes, especially for IL-6, could be shown for high particle concentrations
(Dilution ratios 1:8 to 1:20). For intermediate particle concentrations (Dilution ratio 1:100),
the DNA damage inducible gene GADD45 # is up-regulated (~5x) for up to 4 hrs and for
lower particle concentrations (Dilution ratio 1:200) the maximal induction value for
GADD45# is about 3. NF-!B dependent gene expression is down-regulated for the first hours
of nanoparticle incubation for NF!BIA (-3x, 4 hrs). For IL-6 a first down-regulation (-6.5, 30
min) is followed by a significant up-regulation for incubation times of 2 and 4 hrs.
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