new emerging team on fasd: oxidative stress, biomarkers & antioxidant therapy
DESCRIPTION
New Emerging Team on FASD: Oxidative Stress, Biomarkers & Antioxidant Therapy. Leader: James F. Brien, Queen’s University. Canadian Institutes of Health Research. CIHR NEW EMERGING TEAM ON FASD. Antioxidant Therapy. C. A. B. Biomarkers. Oxidative Stress. Members of NET on FASD. - PowerPoint PPT PresentationTRANSCRIPT
New Emerging Team on FASD:Oxidative Stress, Biomarkers & Antioxidant Therapy
New Emerging Team on FASD:Oxidative Stress, Biomarkers & Antioxidant Therapy
Leader: James F. Brien, Queen’s UniversityLeader: James F. Brien, Queen’s University
Canadian Institutes of Health ResearchCanadian Institutes of Health Research
Biomarkers
B
Antioxidant Therapy
AOxidative Stress
C
CIHR NEW EMERGING TEAM ON FASDCIHR NEW EMERGING TEAM ON FASD
Members of NET on FASDMembers of NET on FASDAlan D. Bocking, obstetrics and maternal-fetal physiology,
University of Toronto;
James F. Brien, basic developmental pharmacology & toxicology, Queen’s University;
Gideon Koren, pediatrics and clinical pharmacology & toxicology, Hospital for Sick Children, Toronto;
Stephen G. Matthews, developmental neuro-endocrinology, University of Toronto;
James N. Reynolds, developmental neuroscience, Queen’s University;
Joanne Rovet, developmental neuropsychology, Hospital for Sick Children;
Wendy J. Ungar, health economics and population health, Hospital for Sick Children.
RESEARCH OBJECTIVESRESEARCH OBJECTIVES
A. To test the hypothesis that oxidative stress is an important mechanism of the brain injury of FASD;
B. To identify and validate reliable biomarkers for fetal ethanol exposure at critical periods of vulnerability during gestation and for the magnitude of fetal ethanol exposure;
C. To discover and develop innovative antioxidant treatment strategies for preventing or attenuating ethanol-induced oxidative stress in fetal life and decreasing its impact on brain function in postnatal life.
Coronal Section of the BrainCoronal Section of the Brain
RESEARCH OBJECTIVESRESEARCH OBJECTIVES
A. To test the hypothesis that oxidative stress is an important mechanism of the brain injury of FASD;
B. To identify and validate reliable biomarkers for fetal ethanol exposure at critical periods of vulnerability during gestation and for the magnitude of fetal ethanol exposure;
C. To discover and develop innovative antioxidant treatment strategies for preventing or attenuating ethanol-induced oxidative stress in fetal life and decreasing its impact on brain function in postnatal life.
OBJECTIVE AOBJECTIVE ATo determine whether oxidative stress is a
mechanism of the brain injury of FASD.
Definition of Oxidative StressDefinition of Oxidative StressOxygen radicals: highly reactive molecules
generated during cell metabolism.
Cell production
of O2 radicals
Cell degradation
of O2 radicals
Overabundance of O2 radicals/Oxidative Stress
Proposed Mechanism of Brain Injury of FASD
Proposed Mechanism of Brain Injury of FASD
Maternal Ingestion of Ethanol
Fetal Brain Exposure to Ethanol
Damage to Key Cell Molecules(DNA, Proteins, Membrane Phospholipids)
Neuronal Cell Death
Oxidative Stress / Increased Reactive Oxygen Species
H2O2 O2– OH
Brain Injury of FASD
Measures of Oxidative StressMeasures of Oxidative Stress
1. Glutathione (GSH):• Intracellular GSH localized primarily in mitochondria
and cytoplasm.
2. F2-Isoprostanes:• Prostaglandin F2-like compounds. • Formed in vivo by nonenzymatic free radical-induced peroxidation of
arachidonic acid.• Specific and stable products of lipid peroxidation.
8-iso-Prostaglandin F2
HIPPOCAMPUS
Ethanol Offspring
Control Offspring
Experimental Animal Study DesignExperimental Animal Study DesignTimed Pregnant Guinea Pigs
Ethanol Isocaloric-Sucrose/ Water(4g/kg MBW/day) Pair-Feeding
Term Fetus (GD 65)
[GSH] in mitochondria and cytoplasm
[8-iso-PGF2] in homogenate
Hippocampus
Fetal HippocampusFetal Hippocampus
MITOCHONDRIA
Ethanol Sucrose Water0.0
2.5
5.0
a a
b
TREATMENT
[GS
H]
(mm
ol/
mg
pro
tein
)
CYTOSOL
Ethanol Sucrose Water0
10
20
30
40
50
60
TREATMENT
[GS
H]
(mm
ol/
mg
pro
tein
)
GLUTATHIONE
Ethanol Sucrose Water0
50
100
150
200
250GD 65PD 0
TREATMENT
[8-i
so-P
GF
2
](p
g/m
g p
rote
in)
Fetal HippocampusFetal Hippocampus
8-iso-PGF2
G. Weaver, University of Colorado at Denver
Modified from J.E. Dawson and L.M Winn
Apoptosis (Programmed Cell Death) and Caspase-3
Apoptosis (Programmed Cell Death) and Caspase-3
CYTOCHROME CYTOCHROME CC (FETAL HIPPOCAMPUS) (FETAL HIPPOCAMPUS)
Ethanol Sucrose
Ethanol Sucrose Water0
10
20
30 *
TREATMENT
AC
TIV
AT
ED
CA
SP
AS
E-3
IMM
UN
OR
EA
CT
IVIT
Y(%
of
all
Ce
lls
)
ETHANOL SUCROSE WATER
ACTIVATED CASPASE-3 (FETAL HIPPOCAMPUS)
SUMMARYSUMMARY
Chronic ethanol exposure produces in the fetal hippocampus:
• depletion of mitochondrial [GSH];• mitochondrial cytochrome c leakage into cytoplasm;• increase in caspase-3 enzymatic activity; • no change in [8-iso-PGF2].
Disruption of the mitochondria and consequent apoptosis play key roles in the mechanism of the brain injury of FASD involving the hippocampus.
CONCLUSIONCONCLUSION
RESEARCH OBJECTIVESRESEARCH OBJECTIVES
A. To test the hypothesis that oxidative stress is an important mechanism of the brain injury of FASD;
B. To identify and validate reliable biomarkers for fetal ethanol exposure at critical periods of vulnerability during gestation and for the magnitude of fetal ethanol exposure;
C. To discover and develop innovative antioxidant treatment strategies for preventing or attenuating ethanol-induced oxidative stress in fetal life and decreasing its impact on brain function in postnatal life.
EthanolSucrose
INDIVIDUAL FAEE
3.0
2.0
1.0
0ME
CO
NIU
M I
ND
IVID
UA
L [
FA
EE
]
(
nm
ol/
g)
Individual [FAEE] in meconium of term fetal offspring of the ethanol and isocaloric-sucrose/pair-fed groups
Ethanol Sucrose Water
*4.0
3.0
2.0
1.0
0ME
CO
NIU
M T
OT
AL
[F
AE
Es]
(n
mo
l/g
)
TREATMENT
Total [FAEEs] in meconium of term fetal offspring of the ethanol, isocaloric-sucrose/pair-fed and water groups
RESEARCH OBJECTIVESRESEARCH OBJECTIVES
A. To test the hypothesis that oxidative stress is an important mechanism of the brain injury of FASD;
B. To identify and validate reliable biomarkers for fetal ethanol exposure at critical periods of vulnerability during gestation and for the magnitude of fetal ethanol exposure;
C. To discover and develop innovative antioxidant treatment strategies for preventing or attenuating ethanol-induced oxidative stress in fetal life and decreasing its impact on brain function in postnatal life.
Rationale for Vitamin C + Vitamin E StudyRationale for Vitamin C + Vitamin E Study
In women at increased risk of pre-eclampsia, pharmacological doses of vitamin C (1000 mg/day) and vitamin E (400 IU natural-source/day) starting at 16-22 weeks’ gestation and continued throughout the second half of pregnancy:
• decreased the occurrence of pre-eclampsia.
• were apparently safe with no obvious adverse fetal effects.
L.C. Chappell et al., Lancet (1999).
Chronic Treatment Regimen:Daily oral administration of vitamins C (250mg) + E (100mg) OR vehicle (milk/cream).
Two hours later,Oral administration of 4g ethanol/kg maternal body weight OR isocaloric-sucrose/pair-feeding for five consecutive days, followed by no treatment for two days, each week.
PD 45: Morris water-maze task for spatial learning and memory.
Ethanol
Vitamins C + E Vehicle Vitamins C + E Vehicle
Nutritional ControlEthanol
Vitamins C + E Vehicle Vitamins C + E Vehicle
Nutritional Control
MORRIS WATER MAZE
PD0: Brain and hippocampal weights
Ethanol decreased brain weight compared with control; Vitamin C plus E treatment protected hippocampal weight in ethanol offspring.
E +Vit E + Veh S + Vit S + Veh0.0
0.5
1.0
1.5
2.0
2.5
a ab b
Treatment
Bra
in W
eig
ht
(g)
E + Vit E + Veh S + Vit S + Veh0.000
0.025
0.050
0.075
0.100
0.125
ab b b
Treatment
Hip
po
cam
pu
s W
eig
ht/
Bra
in W
eig
ht
Rat
io
E = Ethanol; S = Sucrose Control; Vit = Vitamins C + E; Veh = Vehicle
PD 45: Morris water maze
Vitamins C + E protected against the ethanol-induced deficit in retention of new memory (Old Locations).
Vitamins C + E produced deficits in both acquisition (New Locations) and retention (Old Locations) of new memory in control offspring.
"New Locations" of Platform
E + Vit E + Veh S + Vit S + Veh0
10
20
30
ba
aa
TREATMENT
AV
ER
AG
E T
IME
RE
QU
IRE
D T
OL
OC
AT
E H
IDD
EN
PL
AT
FO
RM
(s) "Old Locations" of Platform
E + Vit E + Veh S + Vit S + Veh0
5
10
15
20
25
ad
b,ca,c
TREATMENT
AV
ER
AG
E T
IME
RE
QU
IRE
D T
OL
OC
AT
E H
IDD
EN
PL
AT
FO
RM
(S
)
E = Ethanol; S = Sucrose Control; Vit = Vitamins C + E; Veh = Vehicle
Chronic maternal ethanol administration:
• decreased brain weight in the neonate.• impaired offspring performance in the Morris water-maze
task, resulting in deficits in the acquisition and retention of new memory.
Maternal administration of vitamins C + E:
• protected hippocampal weight in ethanol-exposed offspring at birth.
• protected ethanol-exposed offspring from deficit in retention of new memory.
• produced deficits in acquisition and retention of new memory in control offspring.
SUMMARYSUMMARY
Vitamins C + E dose studies are being conducted to determine optimal antioxidant vitamin therapy for the brain injury of FASD.
CONCLUSIONCONCLUSION
Biomarkers
B
Antioxidant Therapy
Training
D
AOxidative Stress
C
CIHR NEW EMERGING TEAM ON FASDCIHR NEW EMERGING TEAM ON FASD
PhD
MSc
R. Cohen-Kerem & G. Koren, Neurotoxicol. Teratol. (2003).
Ethanol-induced Oxidative Stress MechanismEthanol-induced Oxidative Stress Mechanism
HIPPOCAMPAL CA1 PYRAMIDAL CELL LOSSHIPPOCAMPAL CA1 PYRAMIDAL CELL LOSS
GD 62: No cell loss
PD 1: 25% Cell Loss
PD 5: 30% Cell Loss
PD 12: 30% Cell Loss
McGoey et al., 2003
Ethanol Sucrose Water0
10
20
30 *
TREATMENT
CL
EA
VE
D P
AR
PIM
MU
NO
RE
AC
TIV
ITY
(% o
f a
ll C
ells
)
ETHANOL SUCROSE WATER
CLEAVED PARP (FETAL HIPPOCAMPUS)
PD 45: Morris water maze
Ethanol and/or vitamins C + E treatment did not affect swim speed in locating the hidden platform.
"New Locations" of Platform
E + Vit E + Veh S + Vit S + Veh0
50
100
150
200
250
TREATMENT
SW
IM S
PE
ED
IN
LO
CA
TIN
GH
IDD
EN
PL
AT
FO
RM
(cm
/s)
"Old Locations" of Platform
E + Vit E + Veh S + Vit S + Veh0
50
100
150
200
250
TREATMENTS
WIM
SP
EE
D I
N L
OC
AT
ING
HID
DE
N P
LA
TF
OR
M (
cm/s
)
E = Ethanol; S = Sucrose Control; Vit = Vitamins C + E; Veh = Vehicle