PRINCIPLES OF DETECTION OF RADIATION INJURES

Accidental dosimetry PHYSICAL DOSIMETR Y BIOLOGICAL DOSIMETRY CLINICAL DOSIMETRY DOSE RECONSTRUCTION, Personal Dosimeters CYTOGENETIC DOSIMETRY Dicentrics, FISH, PCC, MNA NAUSEA, VOMITING, BLOOD CELLS COUNTS, SKIN REACTIONS... OTHER BIOINDICATOR S

Physical dosimetry

Instruments for detecting and measuring radiation  Survey meters Geiger-Mueller (GM) instruments Geiger-Mueller (GM) instruments Ionization chamber instruments Ionization chamber instruments Scintilation instruments Scintilation instruments  Laboratory counters  Personnel dosimeters Photographic film dosimeters Photographic film dosimeters Thermoluminescent dosimeters Thermoluminescent dosimeters Pocket dosimeters Pocket dosimeters

Primary use of radiation instrument Level of radioactive contamination Level of radioactive contamination Radiation dose rate in area Radiation dose rate in area Identity and quantity of radioactive material Identity and quantity of radioactive material Accumulated dose to individuals in area Accumulated dose to individuals in area Survey meters Laboratory counters Personnel dosimeters

TLD Film badge Electronic dosimeter

Photographic film dosimeters Advantages Advantages Permanent record Permanent record Energy and nature of exposure Energy and nature of exposure Cost Cost Disadvantages Energy dependence Fading Size

Thermoluminescent dosimeters

Pocket dosimeters

Digital pocket dosimeter

Clinical dosimetry

Clinical and laboratory sings of acute radiation syndrome  Prodromal clinical effects Time of onset Time of onset Degree of symptoms Degree of symptoms  Haematological changes Lymphocyte counts Lymphocyte counts Leukocytes counts Leukocytes counts Biological dosimetry Biological dosimetry

Clinical dosimetry at radiation vomiting Crude estimate of absorbed dose obtainable from clinical presentation  Vomiting  Onset: 2 h after exposure or later  Onset: 1-2 h after exposure or later  Onset: earlier than 1 h after exposure  Onset: earlier than 30 min after exposure MILD ARS (1-2 Gy) MODERATE ARS (2-4 Gy) SEVERE ARS (4-6 Gy) VERY SEVERE ARS (6-8 Gy)

Radiation dose under 5 Gy   No immediate life- threatening hazard exists   Prodromal symptoms of moderate severity Onset > 1 hour Duration < 24 hours

Fatal radiation   Nausea and vomiting within minutes (during the first hour)  Within hours (on the first day): Explosive bloody diarrhoea Explosive bloody diarrhoea Hyperthermia Hyperthermia Hypotension Hypotension Erythema Erythema Neurological signs Neurological signs

Triage categories of radiation injuries according to early symptoms

Guide for management of radiation injuries on the basis of early symptoms No vomiting Vomiting 2-3 h after exposure Vomiting 1-2 h after exposure Vomiting earlier than 1 h, other severe symptoms, like hypotension hyperthermia, diarrhea, oedema, erythema, CNS symptoms < 1 Gy 1-2 Gy 2-4 Gy > 4 Gy Outpatient with 5-week surveillance Surveillance in a general hospital (or outpatient for 3 weeks) followed by hospitalization Hospitalization in a hematological department Hospitalization in a well equipped hematological or surgical department with transfer to a specialized centre for radiopathology

Clinical signs of skin injury depending on dose of radiation exposure

Laboratory dosimetry using early changes in lymphocyte counts

Change of lymphocytes counts depending on dose of acute whole body exposure Degree of ARS Dose (Gy) Lymphocyte counts (cells/  L) 2 days after first exposure Preclinical phase MildModerateSevere Very severe Lethal >

Laboratory dosimetry using granulocyte counts

Cytogenetic dosimetry

Analysis of chromosomal aberrations in peripheral blood lymphocytes - widely used biological dosimetry method for assessing radiation dose, especially useful  in persons not wearing dosimeters while exposed to radiation  in cases of claims for compensation for radiation injuries not supported by unequivocal dosimetric evidence  for validation of occupational radioprotection cases involving suspected low-dose exposures

Biophysical background to chromosome damage ***************************** High LET * * * * * * * * Low LET

Classification of chromosomal aberrations Inversion Symmetrical (STABLE) Breaks Intrachange Asymmetrical (UNSTABLE) Centric Ring Interchange TranslocationDicentric

Biological dose assessment using standard dicentric analysis  Introduced by M. Bender in 1964  Isolated lymphocytes stimulated by phytohaemagglutin (PHA) into mitosis  Arrest of metaphase using colchicine  Scoring of dicentric chromosome aberrations in metaphase spreads

Dicentric chromosome aberrations in metaphase spreads dic f f f f

Dose curves at high LET and low LET radiation Y = A+  D +  D 2

Gamma rays, X-rays acute exposure (Low LET) Gamma rays X-rays chronic exposure (Low LET)  particles Fast neutrons (High LET) Dose Dicentric yield Y = c +  D +  D 2 Y = c +  D Dose curves at acute and chronic exposure Effect

Dose estimation of a partial body radiation exposure (non-uniform irradiation)

Dicentric assay  Most accurate method for dose estimation with sensitivity threshold of about 0.1 Gy for whole body low LET radiation  Especially useful in cases where dosimeter not used, e.g. radiation accident to support physical dosimetry results in radiation protection and safety practice to determine partial body exposure not detected by locally placed dosimeter

Limitations of dicentric analysis for dose estimation  Dicentrics are unstable and lymphocytes carrying aberration elimininated with time (average lifetime days, depending on dose), hence can underestimate magnitude of dose  Method useful only within few months of irradiation

Translocation assay  In retrospective dosimetry and chronic exposure reciprocal translocations used for dose assessment  Translocations considered stable in cell division so yield should not fall with time  Typically detected using specific whole chromosome DNA hybridization probes and FISH methodology

Stable chromosome aberration analysis with G-banding A normal G banded male karyotype An idiogram showing the banding patterns of individual chromosomes by fluorescent and Giemsa staining

Stable chromosome aberration analysis with FISH Translocation Deletion

Applicability of stable chromosome aberration analysis for biological dosimetry Method based on scoring stable chromosome aberrations (translocations and insertions) detected with fluorescent in-situ hybridization of whole chromosomes Requires complex procedures and technical equipment May be use decades after exposure Sensitivity threshold a few cGy but method not feasible for doses less than 0.2 Gy because of expense and time needed for analysis Spontaneous level of stable chromosome aberrations not well established

Premature chromosome condensation (PCC) assay  Initially introduced by Johnson and Rao (1970)  Mitotic-inducer cells (i.e. CHO) isolated using chemical (colcemid) and physical (rapid shaking of flask) technique  Test cells (i.e. human lymphocytes) fused with CHO cells using polyethylene glycol (PEG)  Interphase DNA of test cells condense into chromatid/chromosome-like structures (46 for non- irradiated human cells)

PCC technique PERIPHERAL BLOOD FICOL SEPARATION FUSE IN PEG LYMPHOCYTES CHO CHINESE HAMSTER OVARY (CHO) CELLS (Grown in BrdU) COLCEMID MITOTIC SHAKE OFF (METAPHASE CELLS) PCC Incubate 1 h (Medium+PHA+Colcemid)

PCCs and FISH Irradiated cells with excess break Unirradiated control

Estimation of irradiated body fractions

Applicability of PCC assay for biological dosimetry  Dose estimates obtainable within 48 hours of receipt of blood in laboratory  Radiation induced mitotic delay does not interfere with assay since performed on interphase nuclei and does not require cell division  Method envisioned applicable after partial- body / supra-lethal exposure and improves detection level of lower doses

Micronucleus assay Cytochalasin B

Micronucleus and nucleoplasmic bridges in binucleated cells A B

Micronucleus assay with pancentromeric probe A B centromere negative centromere positive

Application of micronucleus assay for biological dosimetry  Micronucleus not specific to radiation exposure  Discrimination between total and partial body exposure more difficult  High doses of radiation interfere with cell division  High baseline frequency and age dependency make reliability of assay questionable

Glycophorin A (GPA) somatic cell mutation assay  Performed by two-color immunofluorescence flow cytometry on peripheral blood erythrocytes  Based of measuring N/0 variants of erythrocytes, which display phenotype consistent with loss of expression of GPA (M) allele  Can be performed only on individuals heterozygous at this locus that codes for the N/M blood group antigens (approximately half of population)  Prompt but requires complex and expensive equipment  Sensitivity threshold about Gy

Application of GPA assay for biological dosimetry Relationship between glycophorin A mutant frequency in red blood cells and radiation dose for about 1200 A-bomb survivors

Biophysical assays - ESR (electron spin resonance) l Persistent free radicals formed in solid matrix biomaterial (e.g. dental enamel, nail clippings, hair) from accidentally exposed victim can be detected via ESR l Measurements provide reliable biophysical dose estimates and partial body exposure information l In some circumstances, certain clothing material, particularly hard plastics and buttons, may be measured and absorbed dose estimated

Characterization of biological dosimetry methods

Summary of lecture In radiation accidents, important to estimate the absorbed doses in victims to plan appropriate medical treatment In most accidents, physical dosimetry of absorbed dose is not possible. Even where possible, important to confirm the estimates by other methods Most commonly used method cytogenetic analysis of chromosomal aberration in peripheral blood lymphocytes using dicentrics, translocations, PCC and micronuclei assays

Lecture is ended THANKS FOR ATTENTION In lecture materials of the International Atomic Energy Agency (IAEA), kindly given by doctor Elena Buglova, were used