The molecular mechanisms involved in the biological response to environmental radiation are still little known and need to be investigated. Important information can be acquired by analyzing possible differences between parallel biological systems, one kept in a reference radiation environment (RRE) and another one in a low radiation environment (LRE). To this purpose, the underground Gran Sasso National Laboratory (LNGS-INFN) represents a unique opportunity being cosmic radiation almost absent and neutron flux reduced by a 103 factor.
The Pulex experiments, carried out on cells of different origin (yeast, rodent and human), have shown that environmental radiation might act as a trigger of defence mechanisms against endogenous damage: cells grown under reference conditions were more resistant to damage than those grown in extremely low radiation environment. The Cosmic Silence Project aims to deepen the investigation of the molecular mechanisms involved in the biological response to environmental radiation of in-vitro and in-vivo model systems with different levels of phylogenetic complexity.
Life evolved over billions of years under environmental conditions that include exposure to radiation from both the space and the Earth mantle. Different studies have assessed the influence of environmental radiation on living matter and showed that low-radiation environmental conditions affect cell metabolism. In 1995, Satta and co-workers taking advantage of the opportunity given by the underground Gran Sasso National Laboratory (LNGS), carried out experiments aimed at investigating the possible modulation of the mutagenic potential of chemical agents in yeasts kept for 120 generations in an extremely low-radiation environment (LRE). Their work showed that the permanence in LRE impairs the biological defence of the yeast Saccharomyces cerevisiae against chemical radiomimetic agents.
Since then, in the framework of a wider collaboration, the underground biological research at LNGS has continued. The Pulex experiment (so called in contrast to the MACRO experiment that was running in those years at LNGS) further investigated the influence of the radiation environment on the metabolism and the stress response mechanisms of cultured rodent and human cells maintained in parallel, for a similar number of generations as yeasts, under different environmental radiation conditions. The overall in-vitro data showed that, similarly to yeasts, mammalian cells maintained in an extremely LRE exhibit a different biochemical behaviour as compared to cells maintained in a reference radiation environment (RRE). Specifically, cells cultured in the LNGS underground laboratory are less protected from DNA damage induced by chemical and physical agents, and show a lower Reactive Oxygen Species (ROS) scavenging power than cells cultured in a ground laboratory, e.g. at the Istituto Superiore di Sanità, Rome.
More recently, the Cosmic Silence Project started with the general aim to better understand the role played by background radiation in biological systems and, in particular, to deepen the investigation of the molecular mechanisms involved in the biological response by using sensitive in-vitro (A11 hybridoma cells derived from transgenic pKZ1 mice) and in-vivo models (Drosophila melanogaster and, eventually, pKZ1 transgenic mice) with different levels of phylogenetic complexity.
Data on A11 cells (kindly donated by Prof. Pamela Sykes, Flinders University, Adelaide, Australia) obtained after 1 month of culture in LRE and RRE corroborate the hypothesis that environmental radiation contributes to the development and maintenance of defence mechanisms against oxidative stress. Moreover, experiments aimed at investigating the expression of some stress response genes in LRE showed no difference between cells grown in the presence and absence of a 5-cm Fe shield (able to reduce the gamma component of the radiation spectrum by a factor of about 10). This finding indicates that a 10-fold increase in the gamma component of the environmental radiation does not significantly influence the biological response.
To gain further insight into this aspect, experiments are being conducted in collaboration with J.B. Smith and co-workers (Mexico State University) aimed at increasing the gamma background at the LRE using KCL salt as radiation source. Using mammalian and bacterial cells grown under reduced radiation environmental conditions at the Waste Isolation Pilot Plant (WIPP), USA, this group have recently obtained results consistent with our findings.
For the interpretation of the results obtained in the framework of the Pulex-Cosmic Silence projects it is mandatory to characterise in detail the radiation field in the environments where the in-vitro and in-vivo experiments are carried out. To this purpose measurements are ongoing in the different sites of interest, namely in the underground laboratories of Gran Sasso (INFN) and in the ground laboratories at L’Aquila University and at ISS. Thermoluminescence dosimeters (LiF), optically stimulated luminescent dosimeters (Al2O3), a large dimension BF3 detector, high pressure ionization chambers (Reuter-Stokes) have been chosen to measure the dose, dose rate due to terrestrial and cosmic radiation. Radon activity concentration in air is measured and continuously monitored with an Alfaguard instrument during the biological experiments. We have also planned to integrate the experimental measurements with GEANT4 simulations (collaboration with Milano-Bicocca University), in order to implement the modelling of the Cosmic Silence installation for predicting the dose to the target, starting from particle fluxes.
The overall in-vitro data obtained so far corroborate the hypothesis that environmental radiation contributes to the development and maintenance of defence mechanisms in organisms living today. However, it is well known that not all the damage has a local origin, and that part of the biological response is due to non-cell autonomous physiological mechanisms that point to studies at the organism level. To verify whether extremely low doses of environmental ionizing radiation, including cosmic radiation, cause effects in-vivo, a new facility for housing living organisms of different phylogenetic complexity has been designed, and is presently under construction next to the PULEX cell culture laboratory. It will be provided with temperature, humidity and light control systems as well as with an independent ventilation system. Once it is ready, the first planned experiments will be done with Drosophila melanogaster. In the future, after authorization, it will be possible to perform experiments with mice. To this end, the facility has been designed to host a 60-cage mouse rack.
The importance of reducing the uncertainties in the evaluation of damage due to low chronic radiation doses is a crucial issue (Horizon 2020-EURATOM Foster Radiation Protection programme). An approach is recommended that integrates epidemiological studies with in-vitro and in-vivo experimentation. In this context, radiation infrastructures are crucial tools and underground facilities represent a unique opportunity to investigate biological effects, and their underlying mechanisms, in radiation environments below the average radiation background.
Maria Antonella Tabocchini (Istituto Superiore di Sanità, INFN-Gr.coll.Sanità, Centro Fermi, Rome, Italy)
Istituto Superiore di Sanità and Istituto Nazionale di Fisica Nucleare (INFN), Roma1-Gr. coll.Sanità: M.Belli, E.Bortolin, C.DeAngelis, G.Esposito, P.Fattibene, C.Nuccetelli, M.A.Tabocchini
Radon laboratory of Istituto Nazionale di Metrologia delle Radiazioni Ionizzanti (INMRI) of Ente nazionale per le nuove tecnologie, l’energia e lo sviluppo economico sostenibile (ENEA): F.Cardellini
Laboratori Nazionali di Frascati, LNF-INFN, Unità funzionale di Fisica Sanitaria: M.Chiti, A.Esposito
L’Aquila University: E.Alesse, A.Tessitore, F.Zazzeroni, R.Iorio
“La Sapienza” University of Rome, Department of Biology & Biotechnology “C. Darwin”, Section of Genetics: G.Cenci
LNGS-INFN, Servizio di Chimica ed Impianti Chimici: M.Balata, L.Ioannucci
Centro Fermi: E.Fratini, F.Fischietti, L.Satta, G.Simone, M.A.Tabocchini
Università di Milano “La Bicocca”: M. Pavan
- Flinders University, Adelaide, Australia: P.Sykes, R.Omrsby
- New Mexico State University and WIPP Facility, underground repository, New Mexico, USA: G.Smith, H.Castillo
- ASCR, Prague, Czech Republic: M. Davidkova
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