© tnd 2005 oxidative stress and inflammation in neurodegenerative diseases george perry and mark a....
TRANSCRIPT
© TND 2005
Oxidative Stress and Inflammation in Neurodegenerative Diseases
George Perry and Mark A. Smith
Case Western Reserve University
Cleveland, Ohio
© TND 2005
I. Alzheimer’s Disease
Background© TND 2004
© TND 2005
Alzheimer’s Disease
4 million Americans have Alzheimer’s Disease; by 2050, 14 million will have AD.1
AD is one of the top 10 leading causes of death in Americans over 65 years of age.2
AD is the third most costly disease after heart disease and cancer.1
Federal funding for AD is 4 to 7 times lower than for heart disease, cancer or AIDS.1
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PET (positron emission tomography) scans show differences in brain activity between a normal brain and a brain affected by Alzheimer’s disease. Blue and black in the images above denote inactive areas.
Normal Alzheimer
Metabolism is the primary source of oxidants.
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Brain Inflammation in Alzheimer’s Disease
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Proposed Etiologies of Alzheimer Disease
• Aluminum or other toxins
• Infectious agents
• Synaptic abnormalities
• Vascular/immune abnormalities
• Lack of growth factor
• Lipoproteins
• Protein abnormalities
• Oxidative stress
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0
10
20
30
40
50
60
65-69 70-74 75-79 80-84 85-89 90-94 95+
Age
Per
cent
Pre
vale
nce
A leading hypothesis of the biological basis of aging is oxidative stress.
The prevalence of AD is strictly age-dependent
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Proposed Chronology of Changes in Proposed Chronology of Changes in ADAD
Tauist and BAPtistTauist and BAPtist
Amyloid- Deposition[Senile Plaque]
Tau Phosphorylation[Neurofibrillary Tangles]
Neu
ronal d
eath/D
ysfun
ction:
Dem
entia
Tau Phosphorylation[Neurofibrillary Tangles]
Amyloid- Deposition[Senile Plaque]
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Proposed Chronology of Changes in ADProposed Chronology of Changes in ADTauist and BAPtistTauist and BAPtist
Amyloid- Deposition[Senile Plaque]
Tau Phosphorylation[Neurofibrillary tangles]
Neuro
nal d
eath
:D
em
entia
Tau phosphorylation[Neurofibrillary tangles]
Amyloid-b deposition[Senile plaque]
Causes Consequences
Age-RelatedNeuronal dysfunction
Senile plaques&
neurofibrillary tangles
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II. Oxidative Stress
Role in AD
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Oxidative Stress
1. Classic definition: The production of reactive oxygen in excess of antioxidant mechanisms
2. Modern definition: Altered homeostatic balance resulting from oxidant insult.
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Oxidative Modifications Affect All Cellular Macromolecules
ControlAlzheimer
Lipid Peroxidation/Protein Adduction(4-HNE)
Protein Oxidation (Free Carbonyl Groups)
Nucleic Acids (8-OH-Guanosine)
Alzheimer
Alzheimer
Control
Control
Alzheimer Control
Glycoxidation (Carboxymethyllysine)
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Is oxidative stress an early event in AD? In the proposed sequence of degenerative events, it occurs earlier than cytoskeletal alterations.
? .....
.
Glycation
NormalNeuron
? Pre-NFT I-NFT E-NFT
FREE CARBONYLS
80HG
HNE
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Causes of Reactive Oxygen Species Generation in Alzheimer’s Disease
Active microglia
Redox active metalsAmyloid-
Advanced glycation endproducts
Mitochondria
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Partial reduction of oxygen generates ROS
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Consequences
Oxidation Inflammation
Superoxide dismutase
NSAIDS confer protection
Complement pathway activation
Microglial activation and association with amyloid plaques
Heme-oxygenase- 1 induction
Induction of “adaptive” gene responses and repair enzymes
Apoptosis pathway
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III. Mitochondrial and microtubule abnormalities are found in Alzheimer’s Disease.
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Mitochondrial DNA is increased in Alzheimer’s pyramidal neurons.
*
*
*
*
Deleted
Wild type
Alzheimer Control
mtDNA
Wild type probe
Chimera probe
4977 bp deletion
Deleted mtDNANormal mtDNA
8482 10897
8454 10941
1347513475
8454
8482/1346013460
ACACAAACTACCACCTACCTCCCTCACCATTGGCAGCCTA GCATT
CAACAACCTATTTAGCTGTTCCCCAACCTTTTCCTCCGACCCCCT
16,569 bp 11,592 bp
OHOH
4977 bp deletion contains ATPase subunit 8, subunit 6, cytochrome-c oxidase subunit III, and NADH-coenzyme Q oxidoreductase subunit 3, 4 and 5.
In situ hybridization of mtDNA
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mtDNA
8OHG
Nitrotyrosine
The distribution of increased neuronal levels of mtDNA (A), 8OHG (B) and nitrotryosine (C) in Alzheimer’s Disease completely overlaps.
The same neurons in adjacent serial sections are numbered.
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Mitochondria components are in autophagosomes.
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Normal
Alzheimer
Could the mitochondrial problem be related to microtubules?
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Study of biopsy samples shows microtubules are reduced specifically in AD pyramidal neurons.
Pyramidal neurons
Non-pyramidal neurons
p=0.000004
p=0.90
R2 = 0.9841p=0.016
0
2
4
6
8
10
12
60 65 70 75 80
Age (yr)
Num
ber
or m
icro
tubu
les/
m²
Numbers of microtubules decrease
with normal aging
Microtubule Density
0
1
2
3
4
No
. o
f p
yra
mid
al n
eu
ron
s
0
1
2
3
4
No. of microtubules/mm²
No
. o
f n
on
-pyra
mid
al n
eu
ron
s
0
1
2
3
4
No. o
f pyr
amid
al n
euro
ns
0
1
2
3
4
Length (mm x 10²) of microtubules/mm²N
o. o
f n
on
-pyra
mid
al n
eu
ron
s
Microtubule Density Microtubule Length Control AD -PHF AD +PHF
Control AD -PHF AD +PHF
Control AD -PHF AD +PHF
Control AD -PHF AD +PHF
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Microtubules (arrowheads) remain intact even in close proximity to paired helical filaments (*).
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IV. Interplay of pathological lesions and oxidative stress
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Accumulation is Associated with a Reduction in Oxidative Stress
/80HG HO-1
Heme Oxygenase Alz50
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Overexpression of HO-1 Induces tausuppression
1 2 3 4
HO-1
HO-2
Tau
Ferritin
1 Cep2 Antisense3 HO-14 HO-1 + inhibitor
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Increased amyloid - (brown) correlates with decreasedoxidative damage (8OHG, blue).
Down Syndrome
17 yr.
61 yr.
31 yr.
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How do lesions protect?
• Oxidatively damaged
• Metal binding sites
• Reduce oxidative stress
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Is amyloid- protective against a cauldron of oxidative stressors in Alzheimer’s Disease?
Are there signs of established antioxidant responses in AD?
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V. Phosphorylation of Cytoskeletal Proteins Drives Oxidative Modifications.
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p-ERK p-JNK/SAPK p-p38
Stress response kinases are induced
tau assembly induced by HNE is dependent on phosphorylation
Effect of HNE on tau assembly.
Phospho-tau polymers following 1mM HNE.
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0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1
0 500 1000 1500 2000
HNE (mM)
Rela
tiv
e d
en
sit
y
B Unt
C -PO4
0 30 125 500 2000
HNE (mM)
A
NFH
NFH-PO4
In vitro modification of NFH and NFM by HNE is dependent upon
lysine residues…P h o s p h o r y l a t e d N F H
0
5 0
1 0 0
1 5 0
2 0 0
2 5 0
0 5 1 0 1 5 2 0 2 5 3 0 3 5
A g e i n m o n t h s
Relativ
e Dens
ity
C 5 7 B L 6 J
B 6 C 3 F 1
H N E - M i c h a e l
01 02 03 04 05 06 07 0
0 5 1 0 1 5 2 0 2 5 3 0 3 5
A g e i n m o n t h s
Relativ
e Dens
ity
H N E - P y r r o l e
05
1 01 52 02 53 03 54 0
0 5 1 0 1 5 2 0 2 5 3 0 3 5
A g e i n m o n t h s
Relativ
e Dens
ity
A
B
C
Levels of HNE do not accumulate with age in the
mouse’s sciatic nerve
…and phosphorylation state.
NFHNFM
A B C D
Antibody recognition of NFH and NFM (A) is abolished by HNE-lysine (B) but not cyteine (C) or histidine(D).
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VI. Therapeutics
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Progression/Incidence
EstrogenAcetylsalicylicacid (Aspirin) (-) Deprenyl
(selegiline)
Ibuprofen
Dapsone
Acetyl-L-Carnitine(ALCAR)
Vitamin E
Tenilsetam
Antioxidants and anti-inflammatories are protective
DietLipoic Acid
Fruits and vegetables
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Nutrients per 1000 kilocalories Alzheimer Disease Cases Controls p Value n=104 n=223
Vitamin A (RE) 855 983 0.001 Carotene (mcg) 294 389 0.001 Carotene (mcg) 1921 2370 0.003Pro-A Carotene (mcg) 2231 2809 0.001Lutein (mcg) 972 1214 0.015Lycopene (mcg) 666 927 0.001Vitamin C (mg) 74.6 86.7 0.007Vitamin E (a TE) 5.6 5.9 NS
Servings per dayYellow/green vegetables 2.0 2.3 0.022Vitamin C fruits, vegetables 2.4 2.6 NS
Antioxidant diet is protective
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VII. Summary
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Primary etiology
Cellular response
Oxidative stress and inflammation
Aand
AD phenotype
Current therapeutic
targets
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MitochondriaNitric Oxide
Mutated genes (SOD, PP, synuclein)
Phospholipid metabolismProteolysis
Redox Active Metals Advanced Glycation
EndproductsMicroglia
ProteinsLipids
Nucleic AcidApoptosis
Alzheimer DiseaseParkinson Disease
ALSStroke
Multiple Sclerosis
Vitamin ELipoic Acid
Metal Chelation
STOP
Amyloid Neurofilament
proteintau
GlutathioneHemeOxygenase
-1Superoxide dismutase
Classic Cellular Protein-based
STOP
STOP
STOP
© TND 2005
Conclusions
• Metal catalyzed oxidative damage to all categories of macromolecules is increased.
• Antioxidant pathways and inflammatory responses are induced.
• “Pathological changes” may be compensations that are critical to maintaining oxidative homeostasis.