Comparative analysis of fine and nanoparticles for cellular uptake, oxidative stress and genomic damage in human lung cells [Elektronische Ressource] / presented by Kunal Bhattacharya

Ht0162

Sujets

• Gesundheit

Informations

• Publié par: universitat_duisburg-essen
• Publié le: 01 janvier 2008
• Nombre de lectures: 29
• Langue: English
• Poids de l'ouvrage: 16 Mo

Extrait

Comparative analysis of fine and nanoparticles for cellular
uptake, oxidative stress and genomic damage in human
lung cells



Inaugural Dissertation for the Degree of
Doctor of Natural Science
Dr. rer. nat.



A Thesis Presented to
The Faculty of Biology and Geography
University of Duisburg-Essen
Germany




Presented by
Kunal Bhattacharya
from Lucknow, India

March, 2008





Die der folgenden Arbeit zugrunde liegenden Experimente wurden in der
Arbeitgruppe ‘In-vitro- und Molekulare Toxikologie’ am Institut für Hygiene und
Arbeitsmedizin der Universität Duisburg–Essen, durchgeführt.





1. Gutachter:

2. Gutachter:

3. Gutachter:



Vorsitzender des Prüfungsausschusses:


Tag der mündlichen Prüfung:






Dedicated to my Parents


The following publications were written during my doctoral tenure at the Institute of
Hygiene and Occupational Medicine, University of Duisburg–Essen, and are partially
attached to this thesis:

o Dopp, E., Yadav, S., Ansari, F.A., Bhattacharya, K., von Recklinghausen, U.,
Rauen, U., Rödelsperger, K., Geh, S., Rahman, Q. (2005). ROS-mediated
genotoxicity of asbestos-cement in mammalian lung cells in vitro. Part. Fibre
Toxicol. 2, 9.

o Bhattacharya, K., Dopp, E., Kakkar, P., Jaffery, F.N., Rahman, I., Jaurand, M.C.,
Rahman, Q. (2005). Biomarkers in risk assessment of asbestos exposure. Mutat.
Res. 579(1-2), 6-21.

o Bhattacharya, K., Alink, G.M., Dopp, E. (2007). Oxidative stress and changed
gene expression profiles in fiber-/particle-induced carcinogenesis. Int. J. Hum.
Gen. 7(1), 1-21.

o Bhattacharya, K., Cramer, H., Zimmermann, U., Yadav, S., Geh, S., Shi, T.,
Shokouhi, B., Schins, R., Rahman, Q., Rettenmeier, A.W., Dopp, E. (2008). Cyto-
and genotoxic effects of natural and surface-treated ultrafine titanium particles in
mammalian cells. J. Tox. Environ. Health. (In press)



Abbreviations

μg /cm Microgram per centimetre square
μm Micrometer
μM Micromolar
1
O Singlet oxygen 2
8-OHdG 8-hydroxyl–2-deoxyguanosine
AAS Atomic absorption spectroscopy
ANOVA Analysis of variance
Approx. Approximately
APS Aerodynamic particle sizer
As(III) Arsenic trivalent
As(V) Arsenic pentavalent
2+
Ca Calcium
cDNA Complementary desoxyribonucleic acid
CO Carbon dioxide 2
CPC Condensed particle counter
cRNA Complementary ribonucleic acid
Cu(II) Copper divalent
DCF 2’,7’-dichlorofluorescein
DEP Diesel exhaust particles
DFG Deutsche forschungsgemeinschaft
D-KSFM Defined keratinocyte serum free medium
DMA Differential mobility analyzer
DMPO 5,5-dimethyl-pyrroline-N-oxide
DNA Desoxyribonucleic acid
EDX Electron dispersive X-ray analysis
ELISA Enzyme linked immunosorbent assay
EMEM Earl’s modified Eagle medium
EPR Electron paramagnetic resonance
ER Endoplasmic reticulum
FACS Fluorescence-assisted cell sorting
Fe(II) Iron divalent
Fe O Ferric oxide 2 3
FeAsS / FeAs S / FeAs S Natural arsenopyrite mineral 0.9 1.1 1.1 0.9
Fig Figure
G Grams
GO Gene ontology consortium
h Hours
H DCFDA 2',7’-Dichlorodihydrofluorescein diacetate 2
H O Water 2

²
μg /cm Microgram per centimetre square
H O Hydrogen peroxide 2 2
H SO Sulphuric acid 2 4
HBSS Hank’s buffer saline solution
HOCL Hypochlorous acid
HPLC High pressure liquid chromatography
ICP-MS Inductively coupled plasma mass spectrometry
kDa Kilodalton
keV Kiloelectron volts
l/min Litres per minute
LMPA Low melting Agarose
M Molar
2
M /g Meter square per grams
3
mg/m Milligram per cubic meter
+
Mg Magnesium
mins Minutes
mM Millimolar
mRNA Messenger ribonucleic acid
NADPH Nicotinamide adenine dinucleotide phosphate-oxidase
NfкB Nuclear factor kappa B
ng Nanograms
nm Nanometer
O Oxygen
O Molecular dioxygen 2
•-
O Superoxide radicals 2
O Ozone 3
o
C Degree centigrade
•
OH Hydroxyl radicals
•
ONOO Peroxynitrite
OSHA Occupational safety and health administration
OTM Olive tail moment
p Probability Value
PAH Polycyclic aromatic hydrocarbons
PBS Phosphate buffer saline
PEL Permissible exposure limits
PHEM Piperazine-N-bis
PI Propidium iodide
PM Particulate matter
PM Mass of particles ≤ 10 μm mass median aerodynamic diameter 10
PM Mass of particles ≤ 2.5 μm mass median aerodynamic diameter 2.5
ppm Parts per million

²
μg /cm Microgram per centimetre square
PSLT Poorly soluble, low-toxicity
RF Radiofrequency
•
RO Alkoxyl radicals
•
RO Peroxyl radicals 2
rpm Rotation per minute
RT Room temperature
S Sulphur
sec Seconds
SEM Scanning electron microscope
Si Silica
SMPS Scanning mobility particle sizer
SO Sulphur dioxide 2
TEM Transmission electron microscopy
TiO Titanium dioxide 2
TLV Threshold limit value
TNS Trypsin neutralizing solution
Tris - EDTA Tris–ethylenediaminetetraacetic acid
TWA Time weighted average
UV Ultra violet
W Watt
w/v Weight by volume
WCE Whole cell extract
WTC World trade centre
x g Time multiplied by gravity

²
CONTENT
1 INTRODUCTION................................................................................................................................ 10
1.1 Particle-inflicted health risks............................................................................................................ 10
1.2 Ambient air particulate matter characteristics ................................................................................. 12
1.3 Particle-induced health effects based on size distribution .............................................................. 15
1.4 Mechanism of reactive oxygen species (ROS)-induced damage inside the lung tissue ................ 18
1.5 Intrinsic DNA damage repair mechanisms...................................................................................... 23
1.6 Particles of interest.......................................................................................................................... 26
1.6.1 Hematite (fine and nanoparticles) ........................................................................................................... 26
1.6.2 Arsenopyrite ash particles....................................................................................................................... 28
1.6.3 Titanium dioxide nanoparticles................................................................................................................ 30
1.6.4 Quartz (DQ12) fine particles ................................................................................................................... 32
Aim of this study .................................................................................................................................... 34
2. MATERIALS AND METHODS.......................................................................................................... 36
2.1 Particle source................................................................................................................................. 36
2.4 Scanning electron microscopy (SEM) and electron dispersive X-ray analysis (EDX) .................... 38
2.5 Inductively coupled plasma mass spectrometry (ICP-MS).............................................................. 39
2.6 Determination of total iron content in the particles .......................................................................... 41
2.7 Transmission electron microscopy (TEM)....................................................................................... 43
2.8 Trypan blue assay ........................................................................................................................... 44
2.9 Fluorescence assisted cell sorting (FACS) ..................................................................................... 44
2.10 Alkaline comet assay..................................................................................................................... 47
2.12 2',7’- Dichlorodihydrofluorescein diacetate (H DCFDA)................................................................ 51 2
2.14 Microarray analysis........................................................................................................................ 55
3. RESULTS.......................................................................................................................................... 58
3.1 Chemical and physical analysis of the particles.............................................................................. 58
3.1.1 Particle size analysis............................................................................................................................... 58
3.1.2 Morphological and elemental analysis of the particles by SEM and EDX ..............