Antioxidant System in body

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Antioxidant System in body

• H. Singh
• NTRS 525

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DNA damage

• Over 74,000 damage incidences occur in DNA per
  cell per day, mostly by oxidation, hydrolysis,
  alkylation, radiation or toxic chemicals that can
  either directly damage one of the 3 billion bases
  contained in DNA or create breaks in the
  phosphodiester backbone that the bases sit on.
• The result can be mutations in genes which are
  transferred the gene product (protein). If these
  mutations are in genes that normally control cell
  proliferation or suppress tumor growth, the cells
  may start to grow uncontrollably. Cells have
  therefore developed mechanisms to repair DNA
  damage but when they stop working efficiently,
  the number of mutations in our genome increases
  and cancer can develop.

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Agents that Damage DNA

• Certain wavelengths of radiation
• ionizing radiation such as gamma rays and x-rays
• ultraviolet rays, especially the UV-C rays (260
  nm) that are absorbed strongly by DNA but also
  the longer-wavelength UV-B that penetrates the
  ozone shield .
• Highly-reactive oxygen radicals produced during
  normal cellular respiration as well as by other
  biochemical pathways.

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Who is responsible?

• Major cause
• Reactive Oxygen Species (ROS)
• Reactive Nitrogen Species (RNS)

Oxidants are also generated by different types of
radiation, with X-irradiation generating the
hydroxyl radical and irradiation with ultraviolet
light generating electronically excited
states with subsequent radical formation.
Ultrasound and microwave radiation can also
generate reactive oxygen species. Even shear
stress, e.g. in homogenization, is known to
generate radicals.
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Free radicals
Molecular oxygen can be reduced to water. The
intermediate steps of' oxygen reduction are the
formation of- the superoxide anion radical,
hydrogen peroxide and the hydroxyl
radical, corresponding to the steps of' reduction
by one, two and three electrons, respectively.
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Types of DNA Damage

• All four of the bases in DNA (A, T, C, G) can be
  covalently modified at various positions.
• One of the most frequent is the loss of an amino
  group ("deamination") resulting, for example,
  in a C being converted to a U.
• Mismatches of the normal bases because of a
  failure of proofreading during DNA replication.
• Common example incorporation of the pyrimidine U
  (normally found only in RNA) instead of T.

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http//www.benbest.com/lifeext/aging.htmlradical
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Other damages

• Breaks in the backbone.
• Can be limited to one of the two strands (a
  single-stranded break, SSB) or
• on both strands (a double-stranded break (DSB).
• Ionizing radiation is a frequent cause, but some
  chemicals produce breaks as well.
• Crosslinks -- Covalent linkages can be formed
  between bases
• on the same DNA strand ("intrastrand") or
• on the opposite strand ("interstrand").
• Several chemotherapeutic drugs used against
  cancers crosslink DNA.

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How?

• Mutagenesis by ROS/RON contribute to cancer due
  to
• 1.Cause structural changes e.g. base pair
  modification, rearrangement, deletion, insertion
  and sequence amplifications
• 2.Affect cytoplasmic and nuclear signal
  transduction
• 3.Modulate the activity of the proteins and genes
  that respond to stress and which act to regulate
  the genes that are related to cell
  proliferation, differentiation and apoptosis

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• Fig. 1. Reactive oxygen species (ROS) can play a
  role in cell signaling. Oxidative stress can
  activate numerous intracellular signaling
  pathways via ROS-mediated modulation of various
  enzymes and critical transcription factors. In
  one scenario, transcription factors activated in
  response to an increase in ROS or oxidative
  damage travel from the cytoplasm to the nucleus
  within a cell and bind to promoter regions of
  particular genes. As a result, these
  stress-activated pathways can have a significant
  impact on gene expression, which will ultimately
  affect the fate of a cell (e.g., apoptosis,
  proliferation, cytokines). The balance between
  ROS production, cellular antioxidant defenses,
  activation of stress-related signaling pathways,
  and the production of various gene products, as
  well as the effect of aging on these processes,
  will determine whether a cell exposed to an
  increase in ROS will be destined for survival or
  death.

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Where?

• Mitochondria more than nuclear DNA
• Intracellular source is Mitochondrial electron
  transport may generate radicals ROS
• Prevented by low calorie intake and free radical
  inhibitors
• Fe and Cu are associated with ROS

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Why different types of antioxidants?

• The half-lives of the major reactive oxygen
  species are vastly different, underscoring the
  necessity for different types of defense
  mechanisms
• Highest rate constants for the reaction with
  target molecules are found for the hydroxyl
  radical its reactions are diffusion limited,
  i.e. they take place practically at the site of
  generation.
• In contrast, some peroxyl radicals are relatively
  stable, with half-lives in the range of seconds.
  Such molecules may diffuse away from their site
  of generation and thus transport the radical or
  oxidant function to other target sites.

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Oxidative Stress

• An imbalance between oxidants and antioxidants in
  favor of the oxidants, potentially leading to
  damage, is termed 'oxidative stress
• Antioxidant defense involves several strategies,
  
• enzymatic and
• non-enzymatic.

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Natures Strategy Prevention, Interception and
Repair

• Prevention
• Cytochrome oxidase, which carries out most of the
  cellular oxygen reduction, does not release
  superoxide or other radicals, even though it
  contains iron and copper ions.
• Likewise, the three-dimensional structure of the
  enzyme Ribonucleotide reductase keeps the radical
  character of the tyrosyl function in subunit B
  from spreading to the environment by forming an
  appropriate 'cage'.
• Metal chelation is a major means of controlling
  lipid peroxidation and DNA fragmentation. Thus,
  the metal-binding proteins ferritin, transferrin,
  coeruloplasmin and others, e.g. metallothionein,
  are of central importance in the control

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Prevention (contd. )

• Protection of cells from incident radiation may
  occur through specialized pigments, e.g. the
  melanins for ultraviolet radiation or the
  carotenoids for electronically excited states
  such as singlet oxygen. However, these and other
  strategies are not completely preventative,
  because they operate by decreasing the yield of a
  given challenging agent with less than 100
  efficiency.
• The intestinal mucosal cells --- These cells are
  exposed to a variety of reactive intermediates
  and xenobiotics, and the rate of accumulation of
  products of oxidative damage in these cells is
  high. The turnover and elimination of whole cells
  prevents further spread of the challenging
  species.

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2. Interception

• Antioxidant defense involves several strategies,
  
• enzymatic and
• non-enzymatic.
• In general, this means transferring the oxidizing
  equivalents from the hydrophobic phases into the
  aqueous phases, e.g. from the membrane to the
  cytosol or from lipoproteins to the aqueous phase
  of the plasma.
• Such intercepting chain-breaking antioxidants are
  often phenolic compounds. (R,R,R)-a-Tocopherol is
  probably the most efficient compound in the lipid
  phase
• A prerequisite for efficient interception by the
  phenolic antioxidants is that the lifetime of the
  radical to be intercepted must not be too short.

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Non-enzymatic

• In the lipid phase, tocopherols and carotenes as
  well as oxy-carotenoids are of' interest, (as are
  vitamin A and ubiquinols.
• In the aqueous phase, there are ascorbate,
  glutathione and other compounds.
• In addition to the cytosol, the nuclear and
  mitochondrial matrices and extracellular fluids
  are protected.

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Supplements

• Antioxidants from our diet appear to be of great
  importance in controlling damage by free
  radicals.
• it is not clear if supplements should be taken
  and, if so, how much. Once thought to be
  harmless, we now know that consuming mega-doses
  of antioxidants can be harmful due to their
  potential toxicity and interactions with
  medications. Remember -- antioxidants themselves
  may act as pro-oxidants at high levels.

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Enzymatic

• Overall, these low molecular mass (like Vit. E)
  antioxidant molecules add significantly to the
  defense provided by the enzymes
• superoxide dismutase,
• catalase and
• glutathione peroxidases

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Antioxidant Enzymes

• The antioxidant enzymes are proteins with
  antioxidant properties. There are three known
  classes of antioxidant enzymes

• Superoxide dismutases
• Catalases
• Glutathione peroxidases

There are many forms of each class of protein. In
general, cancer cells have low levels of these
enzymes, when compared to an appropriate normal
cell control.
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Primary Antioxidant Enzyme System
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http//lpi.oregonstate.edu/f-w97/reactive.html
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SOD

• FeSOD which contains an iron ion and is generally
  found in some prokaryotes, and CuZnSOD, which is
  active in the cytoplasm of eukaryotic cells.
• CuZnSOD occurs as a dimer of identical 16 KDa
  subunits. Each subunit is 151 amino acids long,
  and the total protein weighs 32 KDa
• Absence of Zn
• Alzheimer Disease
• SOD act as pro-oxidant than Antioxidant

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SOD

• Superoxide dismutase is categorized as an
  oxidoreductase class of enzyme, and has the
  Enzyme Commission identifier of EC1.15.1.1. SOD
  is a metalloenzyme, meaning that in addition to
  amino acids, it contains metal ions.
• Cu2 O2- ? Cu O2
• Cu O2- 2H ? Cu2 H2O2
• 2 O2- 2H ? H2O2 O2

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Repair

• Since prevention and interception processes are
  not completely effective, products of damage are
  continuously formed in low yields and hence may
  accumulate. there are multiple enzyme systems
  involved in
• DNA repair and lipolytic as well as proteolytic
  enzymes capable of serving the functions of
  restitution or replenishment.

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Peroxynitrile (RON)

• which is formed from nitric oxide and superoxide.
  It was observed that a seleno-organic compound
  reacts very efficiently with peroxynitrile

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Biomarkers-1

• Lipid Peroxidation The appearance of
  8-epi-prostaglandin PGF2a (8-epi-PGF2a) in plasma
  or urine has been suggested by a number of
  investigators as a reliable index of in vivo free
  radical generation and oxidative lipid formation.
  There is very strong evidence from animal studies
  that 8-epi-PGF2a increase in plasma and urine as
  a result of oxidative stress, and in human, this
  product is elevated in smokers. Comparison with
  other measures of lipid peroxidation, 8-epi-PGF2a
  is specific product of lipid peroxidation, and is
  very stable. In addition, its formation is
  modulated by antioxidant status, and its level is
  not affected by lipid content of the diet.

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Biomarkers-2

• Protein Oxidation Oxidative damage can affect
  proteins giving rise among others to protein
  carbonyl derivatives, via a variety of mechanisms
  that include fragmentation and amino acid
  oxidation. Protein oxidation has major
  deleterious effects on normal functioning of
  organism. 2-Oxohistine and nitrotyrosine are
  thought to be an indicator of protein oxidation
  induced by peroxyl radical and peroxynitrite,
  respectively.

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Biomarkers-3

• DNA damage One of the major products of oxygen
  radical attack is 8-hydroxy-2'-deoxyguanosine.
  Numerous publications reported that there is an
  age-dependent increase in the level of this
  adduct in human brain tissue. Brunswick
  Laboratories has developed a LC/MS method to
  measure 8-hydroxy-2'-deoxyguanosine in urine.