Detecting the low dose ionizing radiation

induced effects bioinformatics and

microarray data.

Christina Vasileiou

School of Applied Mathematics and Physical Sciences

National and Technical University of Athens
 Main Concern: Low doses
e.g. Diagnostic medical procedures

CT-scan

Hall and A.J. Giaccia, Radiobiology for the Radiologist, Seventh Ed.,
Lippincott, Williams and Wilkins, 2012

X-ray-scan
Relatively low dose < 0.5 Gy
Low dose < 0.1 Gy

The use of Ionizing Radiation (IR) is currently one of the most

common approaches in the diagnosis and treatment of many
kinds of disease, including most types of cancer.
 DNA Microarrays
 A DNA microarray is a two-dimensional array of microscopic spots immobilized with
nanotechnology methods on a solid surface. Each spot contains an amount (picomoles) of a
specific DNA sequence known as probe.
DNA microarray technology is used for parallel gene expression analysis for thousands
of genes of known and unknown functions

The core principle behind microarrays is hybridization

between two DNA strands
 DNA Microarray -
Experimental design
A typical Microarray experiment is
composed by the following steps:
 Selection of our biological query.

 Total mRNA extraction.

 Hybridization of an oligo-dT
oligonucleotide to the polyA tail of the
mRNA
 Reverse transcription into cDNA and

labeling with a fluorescent substance

 Coating on the tile and hybridization

 Optical Laser Scanning and

Measurement of Probe Intensity

 GEO
Gene Expression Omnibus (GEO) is a database repository of high throughput
gene expression data and hybridization arrays, chips, microarrays.

 7 series of studies
 (GSE20629, GSE23901, GSE29344, GSE12435, GSE8917, GSE52918, GSE59861)
 Chips: Affymetrix, Illumina, Agilent

Type of Radiation Dose range Samples

lowLET p, Blood, keratinocytes EPI-
a-particles, 200,fetal lung fibroblasts
X-rays, 0.005<D<0.5 IMR90
Cs-137, Co-60 Dermal fibroblasts AG1522,
prostate fibroblasts.
 Bioinformatic approach
R programming language
RStudio : is a free and open-source integrated development environment
(IDE) for R, a programming language for statistical computing and graphics

Bioconductor
(Limma, Database
lumi, RMA..)
Microarrays

Log2 -transformation

Background correction
Quantile normalization

Check with boxplots

Database Microarrays
 Meta-analysis
Complex set of data  statistical analysis  Increase
of statistical significance
Meta-analysis
Control-Low: 218
STATA :Data Analysis and Statistical Software statistically significant
Random effect model differentially
expressed genes

P-value for every comparison (C-L) for each gene

 Terms of representation-WebGestalt
Hypergeometric distribution
FDR-multiple test correction

- GO analysis : terms of gene ontology corresponding to either BP, CC or MF

- KEGG pathway analysis
- WikiPathways analysis Biological pathways

- Pathway commons analysis

 Genes Analysis
cellular response to GO
CDKN1A CRY1 SNAI2
radiation
cellular response to GO Biological
CDKN1A SNAI2 Processes
ionizing radiation
FZD10 RAB23 GO
G1/G0 transition FOXG1 NFIB

cyclin binding GAK CDKN1A GO

heat shock protein GAK NASP DLST Molecular

GO Functions
binding TOMM34
oxidoreductase
activity, acting on the
CH-CH group of DHDH BLVRB GO
donors, NAD or NADP
as acceptor
 Genes Analysis

DNA damage CDKN1A FRAT1

Wiki
response (only ATM LDLR
dependent)
GAB1 signalosome CDKN1A CSK PC

Pyrimidine
DTYMK POLE3 KEGG
Metabolism
DPYS PRIM1

Pathway Analysis
<<G1->G0 transition>>:

• cell cycle arrest during phase G1, then the cell

enters in phase G0 or rest phase
• G1 phase is a phase cell restraint where the cell cycle
can be controlled

<<GAB1 Signalosome>>: signaling activation through AKT

• Threonine/Serine Kinase AKT: proto-oncogene

• Regulates cellular functions such as metabolism,
development and protein synthesis
• It is related with carcinogenesis
• Heat shock
• Expression of proteins which protect from
decomposition
• Indication of thermal stress

• Oxidoreductase activity with

NAD, NADP receptors
• Indication of oxidative stress
CDKN1A – p21 Cell cycle arrest G1/S

• It is induced by the p53

tumor suppressor protein

• CDK-inhibitor

• Cell cycle arrest

• Phase G1 / S
• DNA Damage
• Oxidative stress
• Active Oncogenes
 Conclusions/Future Plans
• Response and Repair mechanisms seem to appear but
its difficult to have a clear and well defined result

• Design our microarray experiment in order to confirm

our results in the lab

• ‘Cell lines’ response to radiation is related to cell cycle

• Further experiments analysis is neccessary

 Acknowledgements
- Theodora Daphne Michalettou NTUA,

- Dr Ioannis Michalopoulos BRFFA,

- Associate Professor Aleksandros Georgakilas NTUA,

-Associate Professor Pantelis Bagos, University of Thessaly

-Dr Panagiota Kontou, University of Thessaly

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