Extension of lifespan by overexpression of superoxide dismutase in drosophila melanogaster
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Extension of Lifespan by Overexpression of Superoxide Dismutase in Drosophila melanogaster. Orr and Sohal. Background. Hypothesis is that oxygen free radicals/reactive oxygen species (ROS) cause of aging

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Extension of lifespan by overexpression of superoxide dismutase in drosophila melanogaster l.jpg

Extension of Lifespan by Overexpression of Superoxide Dismutase in Drosophila melanogaster

Orr and Sohal


Background l.jpg
Background Dismutase in

  • Hypothesis is that oxygen free radicals/reactive oxygen species (ROS) cause of aging

  • Main assumption of this theory is that normal antioxidant defense levels are not sufficient, so that some ROS escape elimination.

  • ROS cause molecular damage, some of which is irreparable, accumulates with age


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Methods l.jpg
Methods established.

  • Created transgenic flies that had two copies of the SOD and catalase genes

  • Compared lifespan to controls

  • Compared metabolism, activity to controls


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Results established.

  • Flies that overexpressed SOD and catalase

    • Lived ~ 30% longer than controls (median and maximum lifespan)

    • Had lower levels of damage due to ROS

    • Had higher metabolic rates at older ages

    • Had delayed loss of motor ability


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  • The results were different from an earlier study by the authors, in which only SOD or only catalase were overexpressed.

  • In that study, average lifespan was extended up to 10%, but maximum lifespan was unchanged.

  • The combination of SOD and catalase overexpression is critical.


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Does Over-expression of SOD1 Extend Lifespan in Drosophila? authors, in which only SOD or only catalase were overexpressed.

Orr, Sohal


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  • This paper discusses the shortcomings of previous experiments (including those of the authors).

  • Lifespan has been extended in short-lived flies.

  • This was achieved by bolstering antioxidant defenses.

  • But is this possible only with flies with compromised genetic backgrounds?

  • We don’t really know the effect of SOD1 on robust backgrounds.


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  • So we must infer that: hypothesis.

    • cells normally exist under a certain level of oxidative stress

    • oxidative stress increases with age

    • accumulation of molecular damage is major factor in senescence.


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Over-expression of SOD1 in flies lifespan.

  • Three groups (including Orr and Sohal) report positive findings.

  • But did not prove SOD1 had a beneficial effect on a robust genetic background.


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Transgenic effects on longevity only obvious in short-lived strains.

  • One study found 48% extension of lifespan in one strain.

  • This strain lives about 37 days.

  • The control group lived 25 days.

  • Even normal lab flies live 50-70 days.


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Extension of Drosophila Lifespan by Over-expression of Human SOD1 in Motorneurons

Parkes, Elia, Dickinson, Hilliker et al.


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Hypotheses SOD1 in Motorneurons

  • 1. Chronic and unrepaired oxidative damage to motor neurons may be a factor in aging.

  • 2. Sod mutants can be rescued by restoring Sod activity in motorneurons alone.

  • 3. Lifespan extension involves the catalytic activity of SOD in motorneurons.

  • 4. Lower metabolic rate contributes to extended lifespan.


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Materials SOD1 in Motorneurons

  • Expression of Human Sod1 transgene (HS) in fruit flies achieved by using yeast GAL4/UAS system.

  • D42-GAL4 activator used here is:

    • expressed broadly in embryogenesis

    • but restricted to motorneurons & interneurons in the adult CNS.


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Method for each hypothesis SOD1 in Motorneurons

  • 1. Chronic and unrepaired oxidative damage to motor neurons may be a factor in aging.

    • Overexpress HS on motorneurons.

  • 2. Sod mutants can be rescued by restoring Sod activity in motorneurons alone.

    • Selectively overexpress Sod in Sod mutants.


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Results in motorneurons.

  • Targeting motorneurons causes dramatic life extension.

    • HS increased lifespan 40%

    • doubles survival (95%) between 27-50 days of age.

  • Note: Previous studies show increased Sod levels in many tissues had little effect unless combined with an increase in catalase.

  • Note: In situ hybridization shows that expression of HS is limited to adult motorneurons. (including those controlling flight muscles)


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Conclusions lifespan in a dose dependent manner in Sod mutant flies.

  • Overexpressing human SOD1 in adult motorneurons:

    • extends lifespan 40%

    • rescues lifespan of short lived Sod null mutants

    • elevates resistance to oxidative stress.


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Targeted Neuronal Gene Expression and Longevity in Drosophila

Phillips, Parkes, Hilliker


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  • Most organisms have two (or more) types of superoxide dismutase (SOD):

    • SOD1 throughout the cell (CuZnSOD)

    • SOD2 in the mitochondria (MnSOD)

  • SOD1 in motorneurons previously shown to extend lifespan by 140%.

  • SOD1 activated in motorneurons also rescues the lifespan of Sod1 null mutants


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  • Expression of SOD2 in motorneurons: dismutase (SOD):

  • wildtype showed lifespan increase of 30%

  • SOD1 null mutants partially rescues adult lifespan

  • Expression of SOD1 in skeletal muscle showed no effect on lifespan.


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Expression of catalase in motorneurons dismutase (SOD):

  • catalase null mutants show massive mortality in first 2-3 days of adulthood

  • the few that survive have nearly normal lifespan

  • restoration of catalase reduces mortality and leads to normal lifespan

  • wildtype showed no change in lifespan with catalase overexpression


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Co-expression of SOD1 & CAT in motorneurons dismutase (SOD):

  • SOD1 alone extends lifespan by 40%

  • Combination of catalase with SOD1 negates effect of SOD1; normal lifespan

  • elevated CAT activity probably offset the effect of SOD1


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Conclusions dismutase (SOD):

  • Motorneurons limit normal lifespan of fruit flies.

  • ROS part of the mechanism.


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Extension of Lifespan with Superoxide Dismutase/Catalase Mimetics in Worms

Melov, Ravenscroft, Malik et al.


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Background Mimetics in Worms

  • If ROS contributes to aging, then aging can be slowed by reducing the effects of ROS.

  • This can be done in 2 ways:

    • reduce the amount of ROS generated

    • increase the amount of antioxidant repair activities.

  • Genetic mutations and manipulations that resist oxidation also extend lifespan.


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Hypothesis Mimetics in Worms

  • Synthetic superoxide dismutase/catalase mimetics can :

    • Extend lifespan in wildtype worms

    • Restore lifespan in short-lived worm mutants that lack mitochondrial SOD.


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Materials Mimetics in Worms

  • 2 mimetics were tested:

    • EUK-8 (has SOD & catalase-like activity)

    • EUK-134 (an analog of EUK-8 with more catalase activity).

  • Adult worms


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  • Divide worms into several groups: Mimetics in Worms

    • untreated wildtype control

    • worms treated with EUK drugs

    • short-lived mutants (lacking mitochondrial SOD) .

  • Introduce varying concentrations of mimetics into the medium.

  • Mimetics entered worms by ingestion


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Results: wildtype worms Mimetics in Worms

  • SOD/catalase mimetic increased lifespan of wildtype 54%.

    • no overall dose response observed

  • aging worms eat less, mimetic levels decline

  • fertility unchanged

  • body size unchanged


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Results: mutant worms Mimetics in Worms

  • restored normal lifespan (up 67%)


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Conclusions Mimetics in Worms

  • Findings are consistent with amelioration of chronic endogenous oxidative stress.

  • Mimetics extend lifespan by bolstering natural antioxidant defenses.


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Reversal of Age-related Learning Deficits and Brain Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics

Liu, Liu, Bi, Thompson


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Background Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics

  • Loss of learning and memory function from 8-11 months in aging mice is associated with increases in markers of brain oxidative stress.

  • Contextual fear learning and levels of protein oxidation in brain show strong negative correlation.


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Hypothesis Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics

  • Clinical application of synthetic catalytic scavengers of ROS are beneficial in reversing age-related learning deficits.


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Treat mice with SOD mimetics Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics

  • SOD mimetics from Eukarion: EUK-189 and EUK-207.

  • Female mice at 8 months old randomly assigned to 6 groups (16-18 per group)

    • control

    • untreated control

    • low dose EUK-189

    • high dose EUK-189

    • low dose EUK-207

    • high doseEUK-207


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  • Minipumps Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics implanted in anesthetized mice.

  • drug delivered for 28 days

  • low rate ~9nmol/day

  • high rate ~ 0.09 mol/day

  • Pumps replaced twice of over 3 months of treatment.


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Behavioral testing after 3 months Oxidative Stress in Mice with Superoxide Dismutase/catalase Mimetics

  • Plexiglass cages.

  • Videocam to record freezing behaviour (index of fear conditioning).

  • Computer to control events.

  • Mice placed alone in clean chamber.


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  • Day 1: 3 minutes elapse, 3 tones sound, footshock, 1 minute later mouse removed.

  • Day 2: testing for conditioning to context, no sound, no footshock, 8 minutes later mouse removed

  • Day 3: testing for conditioning to tone, mice placed in different chamber,1 minute elapses, 1 tone sounds, 7 minutes later mouse removed


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Behavioral Analysis later mouse removed.

  • Fear conditioning measured as % of time mice exhibited a freezing response (absence of all movement except breathing)

  • Measured for auditory (startle threshold) and visual functions.

  • Measured for nociception ( onset latency to tail flick at 51C hotplate)


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Effects of SOD/catalase mimetic on fear conditioning learning

  • control mice show low levels of freezing for both tone and context (indicates impaired learning and memory at middle age)

  • SOD mimetic mice show increased tone and context freezing at both doses

  • Compared EUK-207 mice at 11 months old to control mice at 8 months old: drug appears to reverse impairment of older mice


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Effects of SOD/catalase mimetic on brain oxidative stress learning

  • Mimetics decreased age-related free-radical damage:

    • decreased lipid peroxidation

    • decreased protein oxidation


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