cellular radiation effects

1. Cellular Radiation Effects Cell membrane - Alteration in permeability Cellular organelles - Functional Aberrations Nuclear membrane - Altered permeability & Function DNA - Chromosomes - Functional aberrations

3. DNA Structure Double stranded helix (twisted ladder millions of rungs long) with side rails of ladder composed of Sugar molecules bound together by a phosphate Rungs are composed of the nitrogenous bases Adenine, Thymine, Guanine and Cytosine. Adenine and Thymine combine to make up one type of rung and Guanine and Cytosine combine to make up another type. A given base may be on either side of the helix

4. DNA Structure DNA is a very large molecule. There are about 2 x 109 base pairs in the mammalian genome distributed across 15-100 chromosomes. The stearic configuration (shape) of the molecule changes constantly and is important to function. DNA is replicated at cell division

5. DNA Structure

6. DNA Structure

7. Mechanism of radiation Injury Direct ionization of a portion of the DNA molecule. Indirect injury by free radicals in the DNA environment. H+, 0H-, H202-, etc.

8. Mechanism of radiation Injury

9. DNA Radiation Injuries Base pair deletion Cross-linking injuies Single Strand Break Double Strand Break Multiple (complex) lesions

10. DNA Radiation Injuries

11. DNA Radiation Injuries

12. DNA Replication DNA is replicated during S Phase prior to the onset of mitosis The original DNA is used as a template for the building of the new DNA. Quite rapid process, requires less than 15 hours.

13. DNA Replication

14. Cell Division Mitosis Multistep process DNA organizes into identifiable chromosomes (Prophase ) DNA aligns with centromeres on equatorial plate (Metaphase) DNA Separates and moves to opposite ends of cell (Anaphase) Cell cytoplasm divides at equatorial plate (Telophase)

15. Cell Division

16. Mitosis Cell resumes normal functional operations (interphase) Through this process radiation induced aberrations in the DNA may result in significant loss of DNA to one or both of the daughter cells. Only requires about one hour

17. Radiation Induced Chromosomal Aberations Chromatid exchanges. Sister Unions Acentric Fragments Rings Dicentric Unions

18. Radiation Induced Chromosomal Aberations

19. Radiation Induced Chromosomal Aberations

20. Cell Cycle Tissues grow and are maintained through cell replication (regeneration) Some cells never divide once adulthood is reached. There are a specific set of steps involved G1 (G0) Gap Phase 1 Functional cell S Synthesis DNA synthesis G2 Gap phase 2 Rest M Mitosis Cell Division

21. Cell Cycle

22. Repair of Radiation Injury Cellular mechanisms are in place which can repair most if not all types of radiation injury to the DNA. Repair is a time sensitive process Repair is a cell cycle dependent process Repair is a dose rate dependent process Repair is dose dependent Repair is radiation type dependent

23. Cellular Mechanisms of Repair Base Excision Repair Damaged bases must be repaired The complementary base on the opposite strand serves as a template. This type of repair is quite efficient Loss of this repair mechanism increases the incidence of mutations.

24. Cellular Mechanisms of Repair Nucleotide Excision Repair (NER) Repairs DNA damage due to pyrimidine dimer adducts added to the DNA by injury. - Enzymatic removal of lesion and associated backbone. - Lesion is then sealed by DNA polyemerase and ligase. - Defective mechanism increases sensitivity to UV light

25. Cellular Mechanisms of Repair Double Strand Break Repair Nonhomologous End Joining Occurs primarily in S phase when no sister chromatid is present. In some instances the base pair sequence is filled in by repair processes without a template. Complex process with multiple pathways Because it is an error prone process it tends to promote development of mutations.

26. Cellular Mechanisms of Repair Double Strand Break repair Homologus Recombination repair Uses sister chromatid as a template to faithfully recreate the damage section and join the ends together properly Occurs in G1 phase when sister chromatids present Error free process Loss of ability increase radiation sensitivity and mutation rate.

27. Cellular Mechanisms of Repair Single strand break repair Occurs via similar pathway to Base Excision Repair. Efficiently done and vast majority of lesions are repaired.

28. Cellular Mechanisms of Repair Because of the efficiency of repair mechanisms for all but double strand breaks the majority of the cell killing occurring at low doses is due to double strand breaks which are not repaired. At high doses accumulated DNA injury due to many single strand breaks and base pair deletions becomes more important.

29. Types of DNA Damage Lethal Damage Irreversible and irreparable � fatal to cell Potentially Lethal Damage (PLD Damage which is lethal unless modified by post irradiation events Sublethal Damage (SLD) Repairable injury to the DNA

30. Lethal Damage Non repairable injury associated with double strand breaks Increases with LET up to a point Increases with higher doses

31. Potentially Lethal Damage Not repaired and lethal under normal circumstances. Repair increased by conditions which are suboptimal to the division of the cell Reduced temperature Hypoxia Low pH Others Increased capability = radioresistance

32. Sublethal Damage Repair (SLD) Refers to DNA damage that is repaired Splitting radiation dose increases survival Occurs in 1-6 hours after irradiation Affected by phase of cell cycle Affected by cell cycle time Long cycle usually increases repair Indicated by shoulder on survival curve

33. Repair is a time sensitive process Repair of DNA injury of all types is essentially complete by 6 hours post irradiation. External factors that affect cellular metabolic rate may delay or accelerate it Foundation of modern radiotherapy

34. Repair is a cell cycle dependent process Different phases have different repair capabilities Mitosis has the least repair capability G2 G1/G0 S phase has the most repair capability Capability varies in G1 and S

35. S-phase Radiation Resistance Likely due to Homologous Recombination Can result in cell population synchrony S G2 blockade and increased survival in S More important in rapidly dividing cells May be important in some tumor lines

36. Reassortment Cells in G2 & M are preferentially killed Cells in S are preferentially spared. Alters proportion of cells in each phase Cell population tends to reestablish normal proportions within 2-3 cycles. Killed cells replaced by cells from G1 Moves cells to more sensitive G2 & S

37. Repair - dose rate dependency Dose rate decreased by two mechanisms Splitting dose into smaller fractions w/ time between the fractions Smaller fractions increase time if spacing constant Reducing the actually rate at which dose is delivered Repair between ongoing during doses Repopulation may occur

40. Repair is dose rate dependent At very low dose rates repair of SLD can keep up with radiation damage. SLD predominate type of injury. Repopulation can account for LD and SLD Dependent on cycling cell population Cell cycle time short relative to dose rate Affected by radiation quality Mutation rates may be increased

41. Repair - dose rate dependency

42. Repair - dose rate dependency

43. Repair is dose dependent Lethal Damage increase with dose PLD increases with dose Accumulation of SLD increase with dose Survival curve is continuously bending Some repair always present Various forms of damage interact

45. Repair is radiation type dependent Low LET radiation is repaired Little repair of High LET radiation injury Dense ionization track Double strand breaks more likely Energy deposition curve dependent Sublethal damage less important Single strand breaks, base pair deletion, etc.

46. LET vrs. Survival