Transcript Tissue Effects of Radiation at the Cellular Level Jeffrey Bryan, DVM, MS, PhD, DACVIM(Oncology) [email protected].

Tissue Effects of Radiation at the
Cellular Level
Jeffrey Bryan, DVM, MS, PhD, DACVIM(Oncology)
[email protected]
Cellular Radiation Effects
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Cell membrane - Alteration in permeability
Cellular organelles - Functional Aberrations
Nuclear membrane - Altered permeability & Function
DNA - Chromosomes - Functional aberrations
DNA (Chromosomes)
• The DNA makes up the chromosomes of the cell and carries all of the
functional encoding information of the cell or organism
• All of the chromosomes together make up the genome
• The genome is composed of many genes (60,000 in humans)
• The individual genes are composed of sequences of nitrogenous bases
attached to the molecular backbone. These sequences encode for protein
functions etc. which control all cell functions
• Large areas of a DNA strand may not be expressed in individual cells
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
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
DNA Structure
DNA Structure
DNA Size
DNA Radiation Injuries
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.
Mechanism of radiation Injury
DNA Radiation Injuries
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Base pair deletion
Cross-linking injuries
Single Strand Break
Double Strand Break
Multiple (complex) lesions
Radiation Induced Chromosomal
Aberations
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Chromatid exchanges.
Sister Unions
Acentric Fragments
Rings
Dicentric Unions
Radiation Induced Chromosomal
Aberrations
Radiation Induced Chromosomal
Aberrations
Radiation Induced Chromosomal
Aberrations
Radiation Induced Chromosomal
Aberrations
http://www.geneticarchaeology.com/research/DNA_Damage_To_Nuclear_Test_Vets_Prompts_Call_For_Study_Of_Children.asp
Radiation Induced Chromosomal
Aberrations
Chromosome 5 pair
Comet Assay
Radiation Induced Chromosomal
Aberrations
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.
DNA Replication
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)
Cell Division
Mitosis
• Cell resumes normal functional operations
(interphase)
• Only requires about one hour
• Through this process radiation induced
aberrations in the DNA may result in
significant loss of DNA to one or both of the
daughter cells.
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)
–S
– G2
–M
Gap Phase 1 Functional cell
Synthesis
DNA synthesis
Gap phase 2 Rest
Mitosis
Cell Division
Cell Cycle
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
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.
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
Cellular Mechanisms of Repair
• Double Strand Break Repair
– Non-homologous End Joining
• Occurs primarily in G1 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.
Non-homologous End Joining
Cellular Mechanisms of Repair
• Double Strand Break repair
– Homologous Recombination repair
• Uses sister chromatid as a template to faithfully
recreate the damage section and join the ends together
properly
• Occurs in S phase when sister chromatids present
• Error free process
• Loss of ability increases radiation sensitivity and
mutation rate.
Homologous Recombination repair
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.
• Predominately error free process
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.
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
Lethal Damage
• Non repairable injury associated with double
strand breaks
• Increases with LET up to a point
• Increases with higher doses
Potentially Lethal Damage
• Not repaired and is 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
Sublethal Damage Repair (SLD)
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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
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
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
Next Time
• Cell Cycle and Differentiation Effects