Oxidative damage 02/04 Flashcards
List the 3 free radical ROS
Free radicals have an unpaired electron.
- *Hydroxyl OH**- (Most reactive, very short half life, does not travel far so is retained within cell)
- *O2-** (Superoxide, highly reactive)
- *LOO-** (Lipid peroxyl)
Why do mitochondria become damaged with age?
Mitochondria contain Mt-DNA coding for enzyme transport chain (ETC) proteins (Except complex II). Mt-DNA is circular with few introns. It resides unprotected in the matrix, lacks histones and has relatively few repair enzymes. It is therefore prone to mutations.
What are the consequences of Mt-DNA damage?
- Cells accumulate mitochondria with mutations
- These mitochondria produce more ROS
- ETC complex activity declines
- ROS are transferred to neighbouring cells
- Cells die by necrosis
Why do damaged mitochondria accumulate within cells?
They increase in size and can no longer be degraded by autophagy as normal
How do apoptosis and necrosis differ?
Apoptosis is an organised, controlled cell death. It begins with the contraction and shrinkage of the cell. Organelles are partitioned off and destroyed. This is the preferable cell death mechanisms. In necrosis, the cell begins to swell, and bulge, spewing out lysozymes and proteasomes, causing local inflammation. ATP deficiency triggers cells to necrose rather than apoptose.
Evidence that mitochondrial damage is a cause of aging, not just a consequence
- SOD2 KO mice (lacking mitochondrial SOD) show reduced lifespan and reduced complex I and II activity - Polg proof reads mitochondrial DNA for errors -/-mice show increased errors and reduced survival. However -/+ mice do not show reduced survival.
reduction/oxidation mnemonic
OILRIG oxidation is loss, reduction is gain
Where are the bulk reactive oxygen species (ROS) produced?
They are produced in the mitochondria in the ETC. Electrons leak at complex III (or complex I in ischemic states) and form ROS.
How are mitochondrial ROS produced?
Electron leakage at complex I and III results in electrons binding to 0.1-0.2% of oxygen being reduced and forming superoxide.
Highly dependent on concrentraton of oxygen - more oxygen = higher ROS
This occurs when an unstable intermediate (Semi-quinone) is formed as part of the electron transferral process).
What are the other sources of RNS/ROS
Nitric oxide synthase produces NO- for vasodilation. However when the reaction is ‘uncoupled’ O2- and ONOO- forms.
Uncoupling occurs as a result of reduced BH4 or L-arginine availability. NADPH oxidase (NOX) also produces ROS. It is used by neutrophils to produce superoxide to kill pathogens. It is also expressed by vascular cells and endothelial cells.
Neurotransmitter degradation.
Cell signalling e.g. insulin receptor kinase
How is mitochondrial ROS removed?
mitochondrial ROS scavenging systems
What are some basal antioxidant enzymes?
Superoxide dismutase forms less reactive H2O2 from O2-.
Catalase produces water from H2O2.
Glutathione. GSH turns oxidised proteins to reduced proteins.
How are inducible antioxidant systems switched on?
They are inducible because they are switched on by changes to their environment. After eating a fatty meal, lipid and cholesterol levels increase, and your body produces lipid peroxides. These generate ROS.
Inducible antioxidant systems remove these lipid peroxide. The Nuclear factor E2 defence pathway is activated, which is the ‘master regulator’ of inducible defenses.
Nrf2 is constitively transcribed and translated. In an unstressed environment, keap-1 sequesters NRf2 for degradation - nrf2 has a half life of 15 min.
When a large fatty meal is eaten, keap-1-NFR2 complex becomes stabilised. Any de novo Nrf2 therefore does not bind to keap-1. Nrf2 can then activate expression of downstream antioxidant defence genes by binding to antioxidant response elements
What is the oxidative damage theory of aging?
In health there is an equilibrium of ROS/RNS creation and removal -> redox homeostasis. In ageing this is disrupted and cell constituents become more oxidised leading to DNA damage and cellular dysfunction.
In health we have an equilibrium between ROS/RNS generation and their remove by endogenous defences. In health, cells exist in a partially reduced state.
in ageing this redox balance is disrupted and cell constituents become more oxidized leading to DNA damage & cellular dysfunction. cells therefore shift to a more oxidised status
5 pieces of correlatory evidence for oxidative damage theory of ageing?
- Cold blooded animals (with slower metabolic rate) live longer - less mitochondrial acitivty, less production of ROS
- Lifespan in mammals is inverse to metabolic rate
- Calorie restriction results in reduced core body temperature, reduced DNA damage and a longer lifespan.
- Mean lifespan correlates with SOD activity
- Mitochondrial damage markers increase with age and antioxidant systems decline in function
How does the mitochondrial ETC produce ATP?
- NADH donates electrons to complex 1.
- These are passesd to c3 via c2 by the q enzyme (along with e- from kreb cycle.) - Q cycle
- complex III to cytochrome C to complex IV.
- electrons released to reduce oxygen to water
All electron transfer used to drive H+ across its [gradient]
- ATP produced by ATP synathase as H+ flows back down gradient.
