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Philippe Baert, Geert Meesen, Dr. Patrick Van Oostveldt, Dr. Andre Poffijn Ghent University Dept. of Molecular Biotechnology Coupure Links 653 , 9000 Gent Dept. of Subatomic & Radiation Physics Proeftuinstraat 86, 9000 Gent. RAMIROS. Experiment name

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ramiros
Philippe Baert, Geert Meesen,

Dr. Patrick Van Oostveldt, Dr. Andre Poffijn

Ghent University

Dept. of Molecular Biotechnology

Coupure Links 653, 9000 Gent

Dept. of Subatomic & Radiation Physics

Proeftuinstraat 86, 9000 Gent

RAMIROS
lesson outline contents
Experiment name

Experiment Team/University/Sponsoring Agency

Scientific Background

Proposed Experiment

Experiment Hardware

Experiment Protocol

Benefits to Science/Mankind

Lesson outline / Contents
experiment name
“RAMIROS”

Radiation And Microgravity Related Oxidative Stress

Experiment Name
experiment team university sponsoring agency
Dr. P. Van Oostveldt, Ph. Baert

Dept. of Molecular BiotechnologyCoupure Links 653B-9000 GentTel: +32 9 264 59 69Fax: +32 9 264 62 19E-Mail: Patrick.VanOostveldt@rug.ac.be, Philippe.Baert@rug.ac.be

Dr. A. Poffijn, G. Meesen

Dept. of Subatomic and Radiation Physics (Radiation and Environmental Physics group)Proeftuinstraat 86B-9000 GentTel: +32 9 264 65 40Fax: +32 9 264 66 97E-Mail : Andre.Poffijn@rug.ac.be, Geert.Meesen@rug.ac.be

Contract grant sponsor: ESA

Experiment Team/University/Sponsoring Agency
scientific background
Cosmic radiation

Primary radiation :

Trapped particle radiation (TR): protons and electrons

Galactic cosmic radiation (GCR):85% protons14% He

1% HZE’s (Charged Particles with Atomic Number > 2; peak for 26Fe)

Solar particle events (SPE)protons, electrons

Secondary radiation :

Brehmstralung radiation

Neutrons

Scientific Background
scientific background1
Very little human radio-epidemiology data on bio-effects of high-energy charged particle radiation

Radiation induces DNA damage through direct ionisation or indirectly through the generation of reactive oxygen species

Acute and long term effects of radiation;

DNA damage is a starting point for:

senescence, ageing

cancer

Scientific Background
proposed experiment
AIM:

To analyse the biological effects of heavy particle (HZE) radiation on primary mammalian tissue in space, in order to understand how single cells and their environment deal with HZE impact, with a contribution to radiation safety guidelines for human space activities in mind.

Proposed Experiment
proposed experiment1
Confluent murine bone marrow culture on top of HZE track detector stack

Co-localisation of tracks and hit cells

Analysis of tracks (HZE characterisation) with corresponding effect/response both at DNA and protein level in the single cell level and its microenvironment

Proposed Experiment
proposed experiment physics
Use of high contrast two photon/confocal fluorescence microscopy enabling 3D imaging of bothtrack detector and biology

Irradiation

Etching

Damage trail

(Latent track)

SSNTD

HZE particle

Proposed Experiment - Physics
proposed experiment biology
Use of high contrast two photon/confocal fluorescence microscopy enabling 3D imaging of both track detector and biology

DNA adducts and localization/expression of protein targets

Immunochemical methods for detecting:

DNA single and double strand breaks,

modified DNA bases,

DNA-repair machinery

Proposed Experiment - Biology
slide13

Proposed Experiment - Biology

  • Irradiation induced DNA lesions
  • single or double strand breaks
  • base modifications
  • destruction of sugars
  • base dimer formation
  • need for efficient DNA repair, if not:
      • less efficient gene reading, senescence (ageing) features
      • DNA mutations
      • Cancerous transformation (late effect)
proposed experiment biology1
Track position linked with hit cells and immediate surroundings called bystander effects

To be linked with information of HZE (SSNTD’s) and bulk radiation (TLD’s)

~25 HZE hits/day randomly distributed throughout the 800 mm² cell culture plane

Proposed Experiment - Biology
proposed experiment2
Semi automated image acquisition and processing

co-localization of tracks and cells

image enhancement (deconvolution)

DNA damage and protein expression profiles

track parameter extraction :

length

direction

Proposed Experiment

Dedicated software routines

experiment hardware
Experiment Hardware
  • CCM based concept for plunger activation
  • wet and dry compartment for biology and
  • physics respectively
  • 3 plungers/plungerbox
  • 2 fluids: -MEM, 0.5% Formaldehyde
experiment hardware2
1st level of containment: plunger module

2nd level of containment: Type II/E or CIS container (flushed with CO2)

Scenario: 4 plungerbox units in 2 Type II/E containers (excluding electrical activation unit)

- mass: 200 g x 2 x 2 = 0.8 kg

- volume: 595 ml x 2 = 1.190 dm³

3th level of containment?

Experiment Hardware
experiment protocol
Prior to operations:

passive (if ambient T° is close to 20°C) or

Aquarius CTA (< 27 W)

Transfer Aquarius CTA from Soyuz to ISS

At ISS (T0): cosmonaut should activate the time line for plunger activation and set 37°C condition for Aquarius (<27 W)

first plunger activation: T0 +2.5 days

second (fixative) and third plunger (detector shift) activation: T0 +5 days

set 5°C

Experiment Protocol
slide20

= Crew activity

Experiment Protocol

t0 =start of exp. timeline

Set CTA

temperature

to +37°C

Activate

4 plungers

Fixation and

Detector shift

(2x4 plungers)

Transfer CTA

from Soyuz to ISS

1

2

3

4

22°C 0xg 22°C >> 37°C, 60 h

37°C 60-84 h

RAMIROS 1

RAMIROS 2

CTR

CTR

2 EC’s Type II/E stored at +22°C in CTA for 3 days (tbc)

Activator unit

Activator unit

1

Chemicals used inside the plunger boxes and Type II/E containers:

1) Activation: αMEM medium

2) Fixation: 0.5% Formaldehyde

experiment protocol1
Post-operations:

transfer 2 type II/E containers to CTR

Transfer CTR from ISS to Soyuz

Experiment Protocol
benefits to science mankind
This research will help :

1) to refine the knowledge about the space radiation environment

2) to better understand fundamental biological processes such as the occurrence of different DNA lesions (hallmarks of cancer and ageing) under particular conditions

3) to better understand the relation between radiation and biological damage (long duration space flights, heavy particle radiation therapy)

Benefits to Science/Mankind
summary
Single cell analysis procedure for HZE impact

Experiment could be self-supporting

Summary