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Assessment of Oxidative Stress

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Title: Assessment of Oxidative Stress


1
Assessment of Oxidative Stress
2
Oxidative stress involves in various diseases
Aging
Cancer
Inflammation
Arteriosclerosis
Mucosa of the Gut Disease
Hepatic Disease
Ischemic Reperfusion
Diabetes
Cardiac Disease (cardiac infarct etc.)
Respiratory Disorder
Renal Disease
Cranial Nerves Disease
Skin Disease
Multiple Organ Failure
Ocular Disease (Cataract)
Dr. T Yoshikawa
3
Balance of Oxidative Stress and Anti-oxidative
System
O2-
SOD
Vitamin E
OH
GSH-Px
Carotenoids
Uric Acid
H2O2
1O2
Catalase
Flavonoids
Vitamin C
LOO
L
LO
Other Antioxidant Enzymes
Other Antioxidants
Antioxidative System
Reactive Oxygen Species Lipid Peroxide
Balance is Important
OXIDATIVE STRESS
Life Style-Related Diseases Parkinson
disease High blood pressure Arterial
sclerosis Myocardial infarction Dementia,
Cataract etc.
Immune Disorder
Cancer
Aging
4
(No Transcript)
5
Oxidative Stress Markers
DNA Oxidation
Lipid Oxidation
8-OHdG Thymidine Glycol
Acrolein Crotonaldehyde Hexanoyl-Lysine 4-hydroxy
-2-hexenal 4-hydroxy-2-nonenal 7-ketocholesterol M
alondialdehyde Methylglyoxal
Protein Oxidation
Dibromo-Tyrosine Dithyrosine
6
Formation of 8-OHdG
Cell
Anti-Oxidative System
Reactive Oxygen Species
Repair System
Urine
Oxidative Stress
DNA (dG)
Damaged DNA (8-OHdG)
Normal DNA (dG)
Blood
Damaged Base (8-OHdG)
Normal Base (dG)
7
Anti-8-OHdG Monoclonal Antibody (N45.1) 8-OHdG
ELISA kit
Assessment of Oxidative Stress inside the body.
a) Highly specific for 8-OHdG. b) Applications
urine, serum, tissue, cultured cells, saliva c)
Assay time 3.5 hours (New 8-OHdG Check) d)
Number of samples (N3 18 samples / kit) e)
Risk management of health and aging. f)
Monitoring the effect of chemicals, UV ,
radiation etc g) Assessment the effect of
anti-oxidants.
Concentration of Urinary 8-OHdG (250 healthy
subjects)
Patent No. 3091974 Patent No. 2850128 Prized
Grand invention award (1999 Japan)
Director-general of the science
and technology agency premium (2000 Japan)
Number of Person
Urinary 8-OHdG (ng/mg CRE)
8
Relationship between 8-OHdG and diseases (1)
1) Diabetes (8-OHdG in urine)
Diabetic patients(macro albuminuria) 19.2
/- 16.8 ug/24h (24 h urine) Normal
controls 8.1
/- 1.8 ug/24h Reference Study of urinary
8-hydroxydeoxyguanosine as a biomarker of
oxidative DNA damage in diabetic nephropathy
patients. J Pharm Biomed Anal. 2004 Sep
2136(1)101-4. Xu GW, et.al.
2) Cancer (8-OHdG in urine) Normal
controls 1.11 /-
0.62 umol/mol CRE Cancer patients (before
treatment) 1.84 /- 1.12 umol/mol CRE

? After initiation of therapy 2.18
/- 1.44 umol/mol CRE.
Irradiation and chemical treatment result in
higher 8-OHdG formation. Reference
Determination of urinary 8-hydroxydeoxyguanosine
by automated coupled-column high performance
liquid chromatography a powerful technique for
assaying in vivo oxidative DNA damage in cancer
patients. Eur J Cancer. 1995 Jun31A(6)934-40.
Tagesson C, et.al.
3) Dialysis (8-OHdG in serum) Serum 8-OHdG
concentration from dialysis patient is higher
than that from normal controls. Reference
Blood 8-hydroxy-2'-deoxyguanosine is associated
with erythropoietin resistance in haemodialysis
patients. Nephrol Dial Transplant. 2003
May18(5)931-6. Kato A, Odamaki M, Hishida A.
9
Relationship between 8-OHdG and diseases (2)
4) Cystic fibrosis (8-OHdG in urine)
Cystic fibrosis patients 2.78 /- 1.21 nmol/mol
CRE Normal controls 1.51 /-
0.38 nmol/mol CRE Reference
Oxidative damage to DNA in patients with cystic
fibrosis. Free Radic Biol Med. 1995
Apr18(4)801-6. Brown RK, et. al.
5) Alzheimer's disease (8-OHdG in tissue and
cerebrospinal fluid) 1. 8-OHdG levels in the
intact DNA from cerebrospinal fluid with AD (9
patients) is higher than normal controls.
Reference Increased DNA oxidation and
decreased levels of repair products in
Alzheimer's disease ventricular CSF. J
Neurochem. 1999 Feb72(2)771-6. Lovell MA,
Gabbita SP, Markesbery WR. 2. 8-OHdG levels in
brain tissue with AD became high.
Reference Oxidative alterations in
Alzheimer's disease. Brain Pathol. 1999
Jan9(1)133-46. Markesbery WR, Carney JM.
6) Administration AZT to HIV Patients (8-OHdG in
urine) HIV patients (Administration AZT)
355 /- 100 pmol/kg/day HIV patients
(Avoid AZT) 182 /- 29
pmol/kg/day Reference AZT
treatment induces molecular and ultrastructural
oxidative damage to muscle mitochondria.
Prevention by antioxidant vitamins. J
Clin Invest. 1998 Jul 1102(1)4-9. de la
Asuncion JG, et.al.
10
Relationship between 8-OHdG and diseases (3)
7) Childhood Bacterial Meningitis (8-OHdG in
cerebrospinal fluid) Bacterial
meningitis patients cerebrospinal fluid 8-OHdG
levels became high significantly.
Reference Oxidative stress in childhood
meningitis measurement of 8-hydroxy-2'-deoxyguano
sine concentration in cerebrospinal fluid.
Redox Rep. 20005(5)295-8. Tsukahara H,et.al.
8) Periodontitis (8-OHdG in Saliva)
Untreated patients with periodontitis (78
patients) 4.28 /- 0.1ng/mL Normal
controls (17
patients) 1.56 /- 0.1ng/mL
Reference New biomarker evidence of
oxidative DNA damage in whole saliva from
clinically healthy and periodontally
diseased individuals. J Periodontol. 2002
May73(5)551-4. M Takane, et.al.
11
Chemical Industry Workers Chemical Industry
Workers Urinary 8-OHdG levels became High.
Reference Increased urinary excretion of the
oxidative DNA adduct, 8-hydroxydeoxyguanosine, as
a possible early indicator of occupational
cancer hazards in the asbestos, rubber, and
azo- dye industries. Pol J Occup Med Environ
Health, 1993, 64, 357-68
12
Oxidative stress by inhalation of asbestos
Reference Levels of 8-hydroxy-2'-deoxyguanosin
e in DNA of white blood cells from workers highly
exposed to asbestos in Germany. Mutat Res, 2000
Jul, 4682, 195-202
13
Oxidative stress caused by benzene Benzene cause
oxidative damage to DNA in peripheral blood
cells.
Reference The study of DNA oxidative damage
in benzene-exposed workers. Mutat Res, 1996 Oct,
3703-4, 145-50
14
Influence of psychological stress
8-OHdG in peripheral blood cells from women under
higher psychological stress are higher.
Reference Relationships between perceived
workload, stress and oxidative DNA damage. Int
Arch Occup Environ Health 2001 Mar74(2)153-7
15
Sperm DNA from smokers
Reference Detection of oxidative DNA damage
in human sperm and the association with cigarette
smoking. Reprod Toxicol, 1997 Sep, 115, 675-80
16
Vegetable juice(Green Juice) reduce urinary
8-OHdG
17
Effect of lemon flavonoids to diabetic rats
18
Oxidative Stress Markers
DNA Oxidation
Lipid Oxidation
8-OHdG Thymidine Glycol
Acrolein Crotonaldehyde Hexanoyl-Lysine 4-hydroxy
-2-hexenal 4-hydroxy-2-nonenal 7-ketocholesterol M
alondialdehyde Methylglyoxal
Protein Oxidation
Dibromo-Tyrosine Dithyrosine
19
Anti Hexanoyl-Lysine(HEL) Monoclonal Antibody
HEL ELISA kit
Assessment of Oxidative Stress inside the body.
a) Highly specific for HEL. b) Applications
urine, serum, tissue, cultured cells, saliva c)
Assay time overnight and 2 hours d) Number of
samples (N3 18 samples / kit) e) Risk
management of health and aging. f) Monitoring the
effect of chemicals, UV , radiation etc g)
Assessment the effect of anti-oxidants.
What is HEXANOYL-LYSINE ?
20
Relationship between HEL and diseases
1) Dry eyes (HEL in cornea) Reference
Selenoprotein p controls oxidative stress in
cornea. PLoS One. 2010 Mar 295(3)e9911, Higuchi
et. al.
2) Cerebral Infarction (HEL in serum)
Reference Effect of prophylactically
administered edaravone during antegrade cerebral
perfusion in a canine model of old cerebral
infarction. J Thorac Cardiovasc Surg. 2007
Mar133(3)710-6, Suzuki et. al.
3) Atherosclerosis (HEL in tissue) Reference
Immunohistochemical detection of oxidative
stress biomarkers, dityrosine and
N(epsilon)-(hexanoyl)lysine, and C-reactive
protein in rabbit atherosclerotic lesions. J
Atheroscler Thromb. 2008 Aug15(4)185-92,
Fukuchi et. al.
21
Oxidative Stress Markers
DNA Oxidation
Lipid Oxidation
8-OHdG Thymidine Glycol
Acrolein Crotonaldehyde Hexanoyl-Lysine 4-hydroxy
-2-hexenal 4-hydroxy-2-nonenal 7-ketocholesterol M
alondialdehyde Methylglyoxal
Protein Oxidation
Dibromo-Tyrosine Dithyrosine
22
Anti 4-hydroxy-2-hexenal (4-HNE) Monoclonal
Antibody (HNEJ-2)
Assessment of Oxidative Stress inside the body.
Specifications a) Highly specific for 4-HNE. b)
Clone HNEJ-2 c) Immunogen 4-HNE-modified
Keyhole Limpet Hemocyanin d) Subclass Mouse
IgG1(kappa) e) Form Lyophilized Powder
(containing 20 ug or 100 ug of IgG) c)
Applications Immunohistochemistry and western
blotting.
Structure of 4-hydroxy-2-nonenal
23
Relationship between 4-HNE and diseases
1) Sjögren's syndrome (4-HNE in salivary gland)
Reference Thioredoxin may exert a
protective effect against tissue damage caused by
oxidative stress in salivary glands of patients
with Sjogren's syndrome, J Rheumatol. 2007
Oct34(10)2035-43, Kurimoto et. al.
2) Active liver disease (4-HNE in liver tissue)
Reference M-30 and 4HNE are sequestered
in different aggresomes in the same hepatocytes.,
Exp Mol Pathol. 2007 Dec83(3)296-300., Amidi
et. al.
3) Chronic obstructive pulmonary disease
(COPD)(4-HNE in lung tissue)
Reference Activation of nuclear factor-kappaB
in airway epithelial cells in patients with
chronic obstructive pulmonary disease.,
Respiration. 200673(5)610-6, Yagi et.al.
24
Antioxidant Assay
Antioxidant assay by Test kit for antioxidant
(PAO)
a) Applications Human and animal serum samples,
foods and beverage samples. b) Assay time 3
minutes c) Number of samples (N3 26 samples /
kit) d) Monitoring the effect of chemicals, UV ,
radiation etc
Principle of this assay
25
Relationship between antioxidants and diseases
1) Alzheimer disease (Antioxidants in plasma)
ReferenceOxidative imbalance and
cathepsin D changes as peripheral blood
biomarkers of Alzheimer disease a pilot study.,
FEBS Lett. 2005 May 23579(13)2759-66, Straface
et. al.
2) Coronary artery disease (Antioxidants in
plasma) ReferenceOxidative stress and
its association with coronary artery disease and
different atherogenic risk factors., J Intern
Med. 2004 Oct256(4)308-15, Vassalle et. al.
26
Conclusion
1) Oxidative stress markers involves in various
diseases. 2) 8-OHdG, HEL, 4-HNE are detected in
various biological samples such as urine, serum,
tissue, cerebrospinal fluid, saliva and cultured
cells. 3) For antioxidant assay, PAO is useful.
PAO is applicable for serum, foods and beverage
samples. 4) Oxidative stress markers and
antioxidant assay may be useful for pathological
analysis, development of medicines, anti-aging
research, or evaluation of functional foods.
27
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