Transcript Diapositiva 1

The THz-BRIDGE Project: Tera-Hertz radiation in Biological Research,
Investigation on Diagnostics and study on potential Genotoxic Effects
An R&D project funded by the European Commission - 5th Framework Programme:
Quality of Life and Management of Living Resources - Key Action 4-Environment and Health
Contract QLK4-CT2000-00129
The aim of this research project is to investigate the interaction of Terahertz (THz) radiation with biological systems. Present and rapidly increasing future applications of radiation in this spectral range, necessitate preventive rather than reactive
research. The project follows a streamline of increasing complexity from bio-molecules to cells, e.g. membranes, chromosomal and DNA integrity. The objectives of the project are to analyze the physical mechanisms of interaction, to assess risk of
potential damage to biological activity, both functional and morphological, and to guide and accompany the development of microscopic imaging at THz frequencies for biological and biomedical applications.
The project will bridge the existing gap of knowledge regarding the effects of Terahertz (THz) radiation on biological systems. THz radiation covers the frequency range
between 100 GHz and 20 THz (i.e. a wavelength between 3 mm and 15 µm), which spans the spectral interval between the microwave- and the infrared regions of the
electromagnetic spectrum.
RESEARCH OBJECTIVES OF THz-BRIDGE PROJECT
To provide a spectroscopic database for selected enzymes, proteins, biological membranes and cells in the frequency range from 100 GHz to
20 THz under irradiation conditions that preserve the integrity and functionality of the biological samples. (Workpackage 1)
THz-BRIDGE PROJECT KEY FACTS

Project start date: February 1, 2001
Duration: 36 months
Total cost : 2 MEuro - EU Contribution : 1.4 MEuro
To identify critical frequencies, which might induce damages on biological systems, and to determine the spectral regions for optimal contrast in
imaging applications. (Workpackage 2)

To assess potential risks due to the exposure of membranes, cells, and DNA to pulsed and CW THz radiation and to define exposure standards
for THz biomedical imaging applications; to monitor and recommend THz exposure conditions at specific occupational sites. (Workpackage 3)

Interdisciplinary consortium: 9 groups, 5 countries
ENEA-Frascati (coordinator)
 Forschungszentrum Rossendorf-Dresden
(I)
(D)

Tel-Aviv University - Dept. Physiology and Pharmacology
(IL)

Stuttgart University - I. Physics Institute
(D)

J.W. Goethe University - Biophysics Institute –Frankfurt
(D)

IREA -Naples, IMS-Rome, DIE-University of Rome - (ICEmB)
(I)


National Hellenic Research Foundation – Athens
(EL)

Teraview Limited – Cambridge
(UK)

University of Nottingham - Biomedical School
(UK)
Identification of critical frequencies
Liposomes, blood & skin-cell cultures
Interface with
International Projects
Dissemination of results
WP-1
WP-2
Spectroscopy of proteins, enzymes
membranes and cells
Evaluation of biological effects
in vitro
Project Management
Evaluation of results:
Spectral database & Reps.
Risk Assessment
Safety Issues
Definition of exposure levels
Spectral information on
contrast in THz Imaging
WP-3
Safety Issue at specific
Occupational sites
Expected achievements and
preliminary results
Role of T-rays in the biomedical field
Biological applications are based on the specific spectroscopic
fingerprints of biological matter in the THz spectral regions.
The different values of the
absorption
coefficient
and
index of refraction between
water and tissue carbonated
proteins at such frequencies,
provide a unique contrast
mechanism for biomedical
imaging applications.
Spectral database
on biological systems
Spectroscopic measurements are used to identify the frequency intervals
where THz radiation is absorbed by a specific system.
First spectroscopic investigations have been carried
out on human blood, human hemoglobin, melanin and
uric acid in the THz region.
An exposure set-up for the irradiation of human
lymphocytes has been designed and constructed.
First irradiation experiments on whole blood have
been performed at a frequency of 120 GHz.
Following irradiation, the Micronucleus assay was
performed by ICEmB on human lymphocytes
cultures.
The preliminary
results
obtained
indicate that, under the adopted
experimental conditions, THz radiation
1
does not affect micronucleus frequency
0,8
and cell cycle kinetics in peripheral
control
0,6
sham
blood human lymphocytes, as shown by
0,4
exposed
0,2
comparing sham exposed cultures with
0
exposed ones. Preliminary exposure of
Treatments
human skin cells to THz radiation has
also been undertaken.


Auston Switch
BIAS 5 mm



10 mm
An ultrafast laser pulse is used to “switch”
on a circuit (a coplanar transmission line
antenna). A current transient is
generated, producing a time-dependent
dipole moment that emits radiation. The
short duration of the pulse allows THz
emission.
Evaluation of Biological Effects In Vitro DNA Bases and Human Lymphocytes:
ICEmB, TAU, NHRF, ENEA, FZR-Dresden
Evaluation of Biological Effects In Vitro: Membranes and epithelial-cell cultures:
ICEmB, ENEA, UNOTT, FZR-Dresden, TVL


EXAMPLES OF SPECTROSCOPY MEASUREMENTS
1,0000
Transmission of 120 GHz ENEA-FEL radiation
was measured in saline solution, culture
medium (not shown in the graph), serum and
whole blood. Results were compared to water
absorption
Phys. Sol.
I/I0
Serum
Whole blood
0,1000
0,0100
Culture medium a = 83 cm-1
Saline Solution: a = 79 cm-1
Whole blood: a = 75 cm-1
Serum: a = 71 cm-1
Water (non reported on the graph): = 80 cm-1
0,0010
0,0001
0,00
0,05
0,10
0,15
nominal thickness (cm)

Study of the exposition of human blood to THz radiation as a function of:
frequency, average and peak power, modulation conditions
Genotoxicity Test on human lymphocytes: Micronucleus, Comet, FISH
Irradiation of DNA bases and measurement of the presence of fragments
by mass spectroscopy techniques
Study of the exposure of liposomes to THz radiation: on-line evaluation of
the carbonic anhydrase (CA) activity under THz exposure as a function of
frequency, average and peak power, and of modulation conditions
Investigations on epithelial cells: keratinocytes cell lines (NHK) and
corneal cells (HCE-T) (Alamar-Blue cell cell viability assay) effects on the
adhesion molecules of epithelial tissues (Fluorescein Leakage assay on
tight junctions)
Modelling of the interaction mechanism
In a FREE ELECTRON LASER
radiation is produced by a relativistic
electron beam traveling through a
magnetic structure (undulator). FELs
can produce very short – high power
THz laser pulse.
Safety issues at specific occupational sites: TVL, FZR-Dresden, UNOTT, TAU



Survey of exposure conditions of technical personnel at specific
occupational sites, where THz sources are employed or developed.
Distribution of a questionnaire to collect information
on the main radiation parameters of THz sources currently in
use: (e.g. frequency range, power level, modulation)
on biological and potential biomedical applications of such
sources
on the exposure conditions (if any) of technical personnel
on the safety measurements or precautions currently adopted
Recommendations on safe exposure conditions.
EXAMPLE OF GENOTOXICITY TEST
THE MICRONUCLEI ASSAI
Whole blood samples are exposed to THz
radiation under controlled conditions
3
2.0

The non-linear properties of selected semiconductors
are utilized to produce optical rectification of
ultrashort laser pulses. The non linear polarization so
generated travels in the medium with a lower phase
velocity, due to the lattice vibration contribution. The
result is an emission in a typical Cerenkov cone
configuration.The rapid oscillations of the laser E field
are rectified and only the envelop remains.
Workplan – WP 3
Spectroscopy of Proteins, Enzymes, Biological Membranes and Cells
(Human Lymphocytes) : UFRANK, FZR-Dresden, USTUTT, ENEA, TVL

Free Electron Laser
P|E0(z,t)|2
E0(z,t)
Workplan – WP 2

Optical Rectification
hu
30 mm
% MN

Coherent radiation from Free Electron Lasers, Gas Lasers and Solid State Sources is available in a wide spectral
range at the partner sites and at collaborating European FEL facilities.
Features: Wide tunability and power ranges, amplitude modulation
Soft X-ray microscopy, SEM, Bio-labs available at ENEA, TAU and FZR
Workplan – WP1
Development of biological samples, basic spectroscopic investigations in
the spectral range 100 GHz - 20 THz
Measurements of Reaction Induced Differential Spectra (RIDS)
Time-resolved Spectroscopy at FEL Facilities
“Time Domain Spectroscopy” (TDS) by solid state THz sources
Assignment of absorption spectra to molecular bonds and functions
Assessment of THz
radiation effects
THE THz SOURCES

In the above picture, the THz image of basal cell carcinoma (BCC)
obtained by Terahertz Pulse Imaging (TPITM) is shown on the left.
The red areas indicate areas of cancer. On the right is the visible
image showing no obvious signs of cancer growth.

Recommendations on
THz exposure
After preparation of cultures,
lymphocytes are induced to divide
Uric Acid
47.6 cm
-1
Human hemoglobin
1.5
O
H
-1
1.0
O
N
N
40.3 cm
-1
2
C
N
H
0.5
H
O
N
H
absorption
absorption
79.1 cm
1
0.0
0
-0.5
0.0
0.5
1.0
1.5
2.0
frequency / THz
2.5
3.0
0.0
0.5
1.0
1.5
2.0
2.5
3.0
frequency / THz
After 44 h of growth Cytocalasin-B is added to stop
cell division at nuclear level
Survey on the use of THz radiation on the THz-BRIDGE web site
The THz absorption spectra of uric acid and human hemoglobin (Teraview Ltd.)
1
Significant progress has been made on the design of spectroscopic cells
and on the choice of suitable materials transparent in the frequency
range of interest.
% MN
0,8
control
0,6
sham
0,4
exposed
0,2
FURTHER INFORMATION
0
Treatments
Slide preparation and observation of
Micronuclei frequency
Statistical analysis of
experimental results
EFFECTS ON MEMBRANE MODEL SYSTEM
A good model for the membrane
system is composed by carbonicanydrase loaded liposomes vesicles.
With such a model it is possible to
evaluate the effects of THz radiation
on the membrane permeability
Polystyrene exhibits excellent optical properties in the wavelength range
between 200 mm and 3 mm (frequency range 1.5 THz to 100 GHz).
Other materials like ZnTe can be used at shorter wavelengths.




Typical Time Domain Spectroscopy layout
The substrate has a very low self-diffusion rate across intact liposome bilayer.
The substrate can be hydrolyzed by CA inside the liposomes only if it permeates
across the bilayer.
Any radiation induced increase of hydrolysis rate of substrate catalyzed by CA
accounts for the effect on membrane permeability.
The kinetic measurement is made following for 2-3 minutes the appearance of
reaction product p-nitrophenolate anion at its peak absorbance (=400 nm) on an
Cary 50 Scan spectrophotometer
THz-BRIDGE Web site
http://www.frascati.enea.it/THz-BRIDGE
FREE ACCESS AREA

Project Description

Progress reports

Official document

Contacts
Project Coordinator
Dr. G.P. Gallerano – ENEA Frascati
E-mail: [email protected]
Tel: +39-06-94005223
fax: +39-06-94005334
EU Scientific Officer
Minna Wilkki - EUROPEAN COMMISSION
E-mail: [email protected]
Tel: +32-2-299 5573
fax: +32-2-296 4322
This work has been carried out with financial support from the Commission of the European Communities,
specific RTD programme “Quality of Life and Management of Living Resources”, Key Action 4
“Environment and Health” - contract QLK4-2000-00129.