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MEDRADSC 2X03 MIDTERM EXAM STUDY GUIDE Fall 2018 find more resources at oneclass.com 2X03 MIDTERM NOTES: Radiobiology → study of action of ionizing radiation on living things Safety in use of radiation → controlling quantity received, body irradiated, conditions, minimizing detriment and maximizing...

MEDRADSC 2X03 MIDTERM EXAM STUDY GUIDE Fall 2018 find more resources at oneclass.com 2X03 MIDTERM NOTES: Radiobiology → study of action of ionizing radiation on living things Safety in use of radiation → controlling quantity received, body irradiated, conditions, minimizing detriment and maximizing benefit to us. Contributions from man-made radiation sources → fallout from weapon testing, extra cosmic rays from air travel, dental x-rays, nuclear power plans, luminous wristwatches, buried radioacti e aste, TV’s a d co puter o itors a d edical x-rays [radiography and NucMed & therapeutic procedures like radiation therapy and radiopharmaceutical therapy & medical devices like pacemakers and arterial stents] ALARA principle balances → benefit of use of radiation, likelihood of detriment from radiation and cost of minimization of exposure. X-ray interactions with matter: Exponential function Depends on: o Mass attenuation coefficient/linear attenuation coefficient o Density of the material o Thickness of the material o Incident x-ray intensity For diagnostic xrays (range of 10-150 kEV) → o Photoelectric effect ▪ Dependent on the x-ray energy and the Z of the material. find more resources at oneclass.com find more resources at oneclass.com Removal of inner shell electron – orbital electrons rearrange them selves producing either a characteristic or an auger electron. o , Coherent scatter ▪ No change in energy, uses low energy (less than 10 kev), has a forward scatter and has little contribution to the image (since photon is so small with respect to energy when it interacts with the atom it has nothing left) o Compton scatter. ▪ Uses an outer shell electron, uses high energy, the xray deflects at an angle with reduced energy (could be 0-80% less) but still free to interact with others Radiation therapy ranges from 1.02 meV the predominant types of interactions are: Pair production o Incident photon approaches nucleus then it disappears and the energy is transformed to produce 2 particles – negative and positive charged electron Photodisintegration o Occurs at 5-10 mev o Incident photon penetrates nucleus and deposits energy – excess energy in nucleus results in a particle being ejected. High energy Neutrons: Produced as a by product of x-ray and electron beams in linear accelerators operating at 10 meV Experience multiple scatterings within room (walls, floors, ceiling) Small fraction reaches room entrance Elastic Scattering: o In tissue, usually proton of hydrogen nucleus set in motion – total EK conserved. Inelastic scattering: o Some transferred energy used in overcoming binding forces (final total EK < initial EK), nuclear fragments are particulate radiations Direct Ionizing Radiation: Collisional interaction of electrostatic fields causes ionization → high energy charged particles are directly ionizing. Ex. Protons, alpha particles Indirectly Ionizing Radiation: Energy transfer causes released of energetic charged particles (recoil electrons), which then are directly ionizing → photons and uncharged particles are indirectly ionizing. Ex. Gamma rays, x-rays, neutrons ▪ IONIZATIONS → BROKEN CHEMICAL BONDS → CHANGED MOLECULAR STRUCTURE → CHANGED CHEMICAL BEHAVIOUR Radiation Interaction in the Cell: Effect mainly due to damage to biologic macromolecule → dna/rna/protein which are considered critical targets find more resources at oneclass.com find more resources at oneclass.com The targets are damaged by radiation (direct) or products of radiation interactions (indirect) Direction action: Energy is absorbed by macromolecule, ionization/excitation results, which cause a chemical bond to break leaving a chemical behaviour change of macromolecule. Indirect Action: Energy is absorbed by a non critical molecule → H20 Cytotoxic products then migrate through cell to damage critical target. (most radiation is indirect) Free Radical: Atom or molecule with unpaired electron in outermost orbit Highly reactive Able to migrate through cell to distant site May cause bond breakage of another molecule to obtain second electron to achieve stability. find more resources at oneclass.com find more resources at oneclass.com REFER TO LECTURE 3 FOR RADIOLYSIS PARTS Oxygen Effect: Extends lifespan of free radicals by reducing likelihood of radicals recombining with H20, with this increased lifespan it will be more likely for the radical to reach and interact with a critical target → increased damage for same radiation quantity. Macromolecules: Proteins → determine cell characteristics/functions o Structural proteins, enzymes, hormones, antibodies. Mechanism of Radiation Injury: Ionization/excitation o If io izatio it is either i dire t/dire t…if i dire t it’s a free radi al. o If ionization happens to critical target → chemical bond repair OR chemical bond breakage. If there is a breakage there is enzymatic repair (no permanent damage) If cell affected by DNA damage → either cell dies or there is a DNA mutation → cell survives but is abnormal/germ cells/somatic cells → cancer REFER TO LECTURE 4 FOR MITOSIS REVIEW Damage to DNA bc of radiation: Base damage → change or loss of base (u instead of t) Cross-link → abnormal bond across/between molecules [adding relationships] Single-strand break → one chain broken [one side of dna helix] – only happen at point mutations, indirect radiation Double strand break → both chains broken [much harder to repair – direct radiation], if breaks are further apart 2 repair activities can occur at the same time – no lasting damage as long as repair faculties are working If base changes occur in a filler gene area it wont matter, but if a base change occurs in a tumour suppressor region that can have an effect of the entire cell Consequences of double strand breaks: Restitution → broken ends rejoin – chromosome restored Deletion → loss of part of chromosome/chromatic Rearrangement of broken ends → distortion of chromosome – shape change or rearrangement of genetic material Chromosome Aberrations: Occur when radiation damage occurs BEFORE dna replication (before s phase, in G1): o During synthesis, aberration is replication – at mitosis both chromatids exhibit the break – both daughters at telophase exhibit damaged chromosome. Chromatid Aberrations: Occurs when radiation damage occurs AFTER dna replication (g2 post synthesis): o Only one daughter cell will be affected – if not repaired prior to dna synthesis both daughters will exhibit the damage. find more resources at oneclass.com find more resources at oneclass.com Dicentric fragment aberration: Are from breaks on one chromatid of each of two chromosomes Dicentric always results in cell death Translocation: Alternate outcome of creation of dicentric/acentric fragments Unstable aberration Similar to base changes Ring Aberration: Caused by one break on each arm of same chromosome – two sticky ends attack – replication cannot occur therefore unstable Lethal Aberrations: Ring, dicentric, anaphase bridge [2 arms of separate chromosomes are attached] Result in cell reproductive death – inability of cell to divide Non-lethal aberrations: Symmetrical translocations, inversions, small deletions Cell survives but chromosome content has changed Cell reproduces with the mutation Associated with malignancies (mutation into oncogene or loss of tumour suppressor gene) Chromosome 9 and 22 → if end up being brought together form a fusion gene that leads to chronic mylogenesis leukemia and its called BCR ABL. Symmetric Translocation Mutation: Formation of a fusion gene that is associated with over 90% of chronic myelogenous leukemia Dose Estimation using Lymphocyte aberrations: Useful for whole body exposure to radiation Can use either lethal or non lethal aberrations Steps: o Lymphocytes of blood sample are stimulated to divide – chromosomes become visible during metaphase – rate of aberrations is compared to standards – effect on time on number of lethal aberrations is seen. Apoptosis: Aka interphase death Programmed cell death Can occur spontaneously without injury to cell/can be induced Chromatin condenses – nucleus fragments – cell condenses and breaks into pieces – fragments are phagocytosed by surrounding cells Mitotic Death: Death while attempting to divide If radiatio does ’t kill ell the first ti e it ould e the 2 nd or the 3rd. m/c type of cell death Cell survival: find more resources at oneclass.com find more resources at oneclass.com Has ability to undergo repeated cell divisions and are called clonogenic and this can produce large colonies of cells. Plating Efficiency: # Colonies counted/ # cells seeded Survival Fraction: # colonies counted/[cells seeded X (PE/100)] Multi-target Multi-hit model: Assumes cell must sustain multiple hits to critical targets within it for cell death to occur. Does ’t elieve o e is e ough. o Shoulder portion of curve (beginning) DQ qn ▪ Initial slope ▪ Results from accumulation of single – event damage o Straight line portion (end of curve) Do, ▪ 2nd interaction → cell death ▪ Final/terminal slope Linear energy transfer affect shape of curve: o Low energy transfer (LET): ▪ The shoulder in the curve at lower doses indicates cells ability to repair some damage → tissues should be able to repair itself o High energy transfer (LET): ▪ Typically has no shoulder, indicating little or no repair – if they deposit a lot of energy it can cause a double strand break. Extrapolation Number (n): o Value on y-axis where extrapolated line of linear portion would intersect o Lower the n value → more damage seen to particular cell o Higher n value → shoulder to curve and extends higher. o Is ’t a real u er → just determines how radiosensitive a cell is Quasithreshold dose (Dq): o Dose at which extrapolated line of linear portion would have a Survival fraction of unity (1). find more resources at oneclass.com find more resources at oneclass.com D0 – ea lethal dose Quantity of dose that delivers, on average, one inactivating event per cell o Some cells will be hit once, others more, and others not at all, but on average radiation hits once per cell. Logen = Dq/Do Limitation of math expression of curve: Expression addresses straight line portion only, does not tell exact shape of initial shoulder since all curves start at SF = 1. Does indicate where shoulder meets straight line portion Linear Quadratic Model of Cell Survival: 2 parallel components to cell killing o One is proportional to dose o Other is proportional to square of dose Recognizes that chromosome damage could be due to: o Single radiation (ex. Electronic passes through one DNA molecule) causing one DSB or point mutation o Two radiations causing two separate breaks resulting in a lethal aberration Fractionation → leads to better tumour control. Disadvantages → space out too long you can increase tumour cell proliferation. Under 2Gys tend to spare late reacting normal tissues which allows prolonged treatment. Allows for estimation of radtherapy schedule find more resources at oneclass.com find more resources at oneclass.com Alpha and beta constants are tissue specific. find more resources at oneclass.com find more resources at oneclass.com Cell survival may be expressed as a single fraction (equation above) Dose where alpha and beta are equal is our alpha beta ratio → same amount of damaging result of two interactions vs. single interactions Alpha – single, beta – double. Clinical Use of equation D= a/B: Dose- response curves are dif for two categories of responders: o Problem: multiple organs are irradiated during treatment, not all respond the same o Used in determining treatment plan that balances early vs late effects and tumour control Large alpha beta rations 10-20, early or acute reacting tissues Small alpha beta ratios 2, late reacting tissues find more resources at oneclass.com find more resources at oneclass.com Early responders → mucosa, skin, GI → reproducing all the time, typically frequent cell cycles. Larger alpha beta ratio → alpha outweighs beta, which means more single hit kinetics. Anything that divides is more sensitive → initially show a bit steeper slope bc more affected by first radiation. Alpha component dominates at lower doses. [Steeper initial slope until get to higher doses] Takes about 10 Gys for components of cells to reach late responders Late responders → lung tissue and spinal cord → pretty spe ified, do t usually divide. Less of an alpha component than a beta component. Smaller alpha beta ratio (3Gys Take more activating event. Beta component has influence at low doses [where you see line curve] Limitations of linear quadratic: Continuously bending curve, never straightens Fits experimental in vitro data for first few generations only Initial region of curve correlates best with clinical data o Later generations show straight-line relationship not bending curve. Comparison of Survival Curves: Radiosensitivty of the cell find more resources at oneclass.com find more resources at oneclass.com Relative effectiveness of radiation types on a given cell type Dose needed to produce a specific effect. Limitations of Use of In-Vitro cell survival curves: Cells reproductive integrity is not only the clinically relevant end point For tumour cells from biopsy specimens, poor plating efficiency, difficult to determine accurate survive fraction to dose of 2Gy Each tumour type has a wide range of radio sensitivities recorded, pt varies The bystander effect: Inductions of biological effects in cells that are not directly traversed by a charged particle but are in close proximity to those that are. Experimental data: o Narrow beam radiation to portion of cell culture o Addition of irradiates cells to non-irradiated cells o Both show increase cell kill to non-irradiated cells that are in prox. To irradiated cells. o Indicates that the target for radiation damage may be larger than the cell itself Importance primarily at low doses, significant to therapy and protection practices. Cell Cycle Time (Tc): Time required to complete one cell cycle Mitotic cycle time Variation in Tc for mammalian cells is mainly due to dif. in duration of G1 phase o Mitosis completed in approx. 1 hour o Dna synthesis within 15 hr In vivo cells: Normally asynchronous Cells in all 4 phases (G1, S, G2, M) are present – proportional in each stable over time Mitotic index – fraction of cells in mitosis In Vitro Cells: Normally asynchronous Synchronized population can be produced in lab In vitro testing: o Synchronized population obtained using mitotic harvest and drug induced G1 hold Test procedure: o Synchronized cells are irradiated at various times in cycle o Surviving fractions are compared Division Delay: Can occur in both non-lethal and lethal damage Disturbs mitotic index of asynchronous population of cells o Initial decrease following radiation and mitotic overshoot follows find more resources at oneclass.com find more resources at oneclass.com Graph of mitotic index (% of cells still in mitosis) as function of time post-irradiation Timing of division delay: o Occurs at 2 points in interphase ▪ G2 phase – delays cells about to enter mitosis – due to check point gene ▪ S phase – at start of dna synthesis (smaller contributor) Effect on dose: o Dose affects the: magnitude of decrease of mitotic index (depth of curve drop), length of delay (width of curve drop) and size of overshoot o At relatively low doses, mitotic index returns to normal (non lethal) o At high doses, delay occurs but lethal damage Mitotic Overshoot: Increase in number of actively dividing cells following mitotic delay Due to synchronizing effect of radiation Increased number of cells entering mitosis due to: o Recovery of delayed cells plus normal progression of unaffected cells Comparison of Survival Curve shapes: find more resources at oneclass.com find more resources at oneclass.com For M and G2: no shoulder, steep slope – repair unlikely – many lethal events For S (especially early) – noticeable shoulder, slope less steep – repair of sub lethal damage especially at lower doses. Radio sensitivity General Patterns: Least sensitive: o Late S → due to homologous recombination repair using sister chromatids find more resources at oneclass.com find more resources at oneclass.com Most sensitive → mitosis and late G2 G1 sensitivity changes over time Radio sensitivity is highest in cells that: actively dividing, undifferentiated and have long future of dividing. Effect of radiation type on age-related response of cells: Main effect is magnitude of dif in sensitivity. o High LET radiation → less variation across cell cycle o Low LET radiation → higher proportion of single vs double strand breaks ▪ Resultant damage more easily repaired Differentiated Cells: ▪ Functionally/structurally specialized – mature/end cell in population Undifferentiated cells: ▪ Few functional/structural characteristics, immature cell/primary function is to divide → precursor or stem cells in population Stem Cells: ▪ Sole purpose to divide and produce more cells ▪ Basal cells in epidermis Transit Cells: ▪ Intermediate cell on way from stem to end cell compartment ▪ Reticulocyte in bone marrow Static population: ▪ Fully differentiated, little/no mitotic activity ▪ Adult nervous tissue, muscle Casarett’s Classification of Cell Radiosensitivity: ▪ Based on early histological signs of cell death not loss of reproductive potential ▪ Defined 5 categories of cell populations in terms of radio sensitivity.[highest to lowest] 1. Vegetative Intermitotic Cells (VIM) a. Rapidly dividing, undifferentiated, short life b. Type A Spermatogonia 2. Differentiating Intermitotic Cells (DIM) a. Produced by division of VIM, more differentiated but still actively mitotic. b. Intermediate spermatogonia 3. Multipotential Connective Tissue Cells: a. Supporting structures, divide regularly, more differentiated then VIM/DIM b. Endothelial 4. Reverting Post-Mitotic Cells (RPM): a. Do t or ally u dergo itosis ut still apa le of dividing, more differentiated, longer life → mature lymphocytes 5. Fixed Postmitotic Cells (FPM): a. Do not divide, highly differentiated → erythrocytes find more resources at oneclass.com find more resources at oneclass.com Lymphocytes→ exception to the rules: Cell is structurally RPM but is one of the most radiosensitive cells – post irradiation they undergo induced apoptosis. find more resources at oneclass.com