the possibility of a dementia-free future: fantasy or reality?
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THE POSSIBILITY OF A DEMENTIA-FREE FUTURE: FANTASY OR REALITY?. 3. 2. 1. Developing a Mind…. Newborn 3months 15months 2years. Connections Give Ever Deeper MEANING over time…. - PowerPoint PPT PresentationTRANSCRIPT
THE POSSIBILITY OF A DEMENTIA-FREE FUTURE: FANTASY OR REALITY?
Developing a Mind…
Newborn 3months 15months 2years
Connections Give Ever Deeper MEANING over
time…
THE BIOLOGICAL BASIS OF THE
MIND IS THE
PERSONALISATION OF THE
BRAIN THROUGH UNIQUE
DYNAMIC CONFIGURATIONS OF
NEURONAL CONNECTIONS,
DRIVEN BY UNIQUE
EXPERIENCES
Developing And Losing One’s Mind…
…Reflected In Brain Cell Branching (Connections)
CURRENT APPROACHES
CURRENT APPROACHES
INVASIVE
APPROACHES
CURRENT APPROACHES
INVASIVE
APPROACHES
NON-INVASIVE
APPROACHES
CURRENT APPROACHES
INVASIVE
APPROACHES
NON-INVASIVE
APPROACHES
THE ‘ANTICIPATORY’
APPROACH
But degeneration can start 20-30 years before the onset of visible symptoms!
THE ANTICIPATORY APPROACH
(A) Marker for in blood detects degeneration before symptoms appear
+(B) Medication for arresting further
neuronal death
=PERMANENT PREVENTION OF
SYMPTOM ONSET!
AN EVENTUAL ‘CURE’?
What is the Basic Mechanism of Neurodegeneration?
Clue 1: Co-pathology: Alzheimer's, Parkinson’s, and ALS
In search of the Basic Mechanism
of Neurodegeneration
‘Global Neurons’
SERIAL Primary sensory,
relays, cerebellum, thalamus, hippocampus, cortex
Amino acids Alar plate Electrically silent Lose plasticity Insensitive to trophic
factors
GLOBAL Locus C., Raphe N.,
A9, A10, basal forebrain motorneurons
Ach, DA, NA, 5-HT Basal plate Spontaneously
active Robust plasticity Sensitive to trophic
factors
‘SERIAL VS GLOBAL’ NEURONS (Woolf, 1996)
SERIAL Primary sensory,
relays, cerebellum, thalamus, hippocampus, cortex
Amino acids Alar plate Electrically silent Lose plasticity Insensitive to trophic
factors
GLOBAL Locus C., Raphe N.,
A9, A10, basal forebrain motorneurons
Ach, DA, NA, 5-HT Basal plate Spontaneously
active Robust plasticity Sensitive to trophic
factors
‘SERIAL VS GLOBAL’ NEURONS (Woolf, 1996)
Cholinergic Striatal Interneurons
Cholinergic Basal Forebrain
Histamine Hypothalamus
Dopaminergic Substantia Nigra
Cholinergic Pontomesencephalon
Noradrenergic Cerulear Region
Serotonergic Raphe
Cholinergic Motor Neurons
Woolf 1996
The ‘Isodendritic Core’: Rossor 1981
NORADRENALINE ACETYLCHOLINESTERASE (AChE)
THE LOCUS COERULEUS Degree of cell death correlates with disease
duration (Albanese & Butcher 1980)
Clue 1: Co-pathology: Alzheimer's, Parkinson’s, and ALS
Clue 2: Presence of key protein (AChE) in all Global neurons/Isodendritic
core
Clue 3: AChE is released
In search of the Basic Mechanism
of Neurodegeneration
Release of AChE from the Substantia Nigra:
(Llinás & Greenfield 1987)
Clue 1: Co-pathology: Alzheimer's, Parkinson’s, and ALS
Clue 2: Presence of key protein (AChE) in all Global neurons/Isodendritic
core
Clue 3: AChE is released
Clue 4: AChE promotes cell growth
In search of the Basic Mechanism
of Neurodegeneration
Trophic action of AChE in Hippocampus
(Day & Greenfield 2002)
Control
AChE
Trophic-Toxic Consequences of Increasing Levels of Calcium:
(Dickie et al., 1996)
Trophic or Toxic? Another Key Factor:
AGE!
Neonatal
Growth factors (e.g. AChE)
Brain develops normal structure
and function
Adult
AChE cells retain plasticity, sensitivity to growth factors and regenerative capacity
Cell Death
Attempt to repopulate in response to growth factors,
harmful ‘developmental’ response
Neurodegenerative Disorders
Non-AChE cells stabilise, lose plasticity and proliferative responses
Cell Death
Compensation by extant cells, no
proliferation
Adult
INSULT
Neurodegeneration: An Aberrant Form of
Development
We have identified the molecular fragment of
AChE that can operate
independent of enzyme function:
‘AChE-Peptide’
AChE C-Terminal Peptides
APP ..VKMDAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVVIA.. AChE ..RQWKAEFHRWSSYMVHWKNQFDHYSKQDRCSDL
T14
T30
AB42
Control AChE Peptide
Administration of AChE-Peptide can be Toxic
(Greenfield et al 2004)
NEURODEGENERATION IS AN ABERRANT
FORM OF DEVELOPMENT:
AChE-PEPTIDE IS THE PIVOTAL TOXIC
MOLECULEI. BiomarkerII. In Vitro and In Vivo
ModelsIII. Therapeutic Intervention
NEURODEGENERATION IS AN ABERRANT
FORM OF DEVELOPMENT:
AChE-PEPTIDE IS THE PIVOTAL TOXIC
MOLECULEI. Biomarker
Discovery of Biomarkers:Mass Spectrometry
Sampl
e selection
• Selection of Alzheimer patients sample (Diagnosed for 1 to 3 years)
• Selection of age matched control patients
Sampl
e processing
• Filter the proteins with a 10kDa cut-off membrane• Desalting samples for mass spectrometry analysis
Sampl
e analysis
• All samples are analysed by Liquid chromatography coupled with a mass spectrometer
• The data are then manually processed
The AD-index:
Higher AD-index at the early stage of the disease
83% of the values are above the median of the control
50% of the values are above the upper quartile of the control
60 samples (30 AD and 30 control)R2 = 0.1297; P = 0.0040
Time after diagnosis (years)
AD
-ind
ex
NEXT STEPS
• Determine how early the AD-index can diagnose Alzheimer’s disease
• Confirm the specificity of this index (comparison with non-neurodegenerative disorders)
• Produce a diagnostic feature for Alzheimer’s disease (ELISA kit, etc.)
• Establish the relationship between the AChE-peptide and the current biomarker peptides
• Identify the proteases involved in the cleavage of AChE, and subsequent metabolites of AChE-peptide in current samples
NEURODEGENERATION IS AN ABERRANT
FORM OF DEVELOPMENT:
AChE-PEPTIDE IS THE PIVOTAL TOXIC
MOLECULEI. BiomarkerII. In Vitro and In Vivo
Models
(A) CHROMAFFIN/PC12 CELLS:
(1) Lewy body inclusions (Averback 1983)
(2) Neurofibrilliary tangles & paired helical filaments: (Izumiyama et al., 1990)
(3) Expression of APP (Takeda et al., 1994)
(4) Aberrant secretion of AChE (Appleyard & MacDonald 1991)
(5) A ‘window on the brain’ (Bornstein et al., 2012)
(A) CHROMAFFIN/PC12 CELLS:
AChE-Peptide (T30) is Toxic in PC12 cells....
30 % Reduction on Cell Viability (1 h)
A Specific Compensatory Response?
Could A and T30 peptide share a
final common path of action?
Synergy of A and T30 peptide after 1 hour
Cell viability AChE activity in perfusate
Advantages: Highly reductionistEasy and quick to cultureImplicated already in ADRelease AChE
Disadvantages: Not brain cells
(A) CHROMAFFIN/PC12 CELLS:
(B) OPTICAL IMAGING: BRAIN SLICES
Badin et al., 2013
(B) OPTICAL IMAGING: BRAIN SLICES
Advantages:Brain cellsFunctional circuitry retained (>10m
cells)Time resolution of action potentialsDifferential spatial and temporal
dynamics
Disadvantages: In Vitro
(C) IN VIVO RAT
The Cylinder Test:Sensitive indicator of neurological
damage (Schallert, 2000)
Control
T30 1μM
T30 100μM
7 days post treatmentGFAP IR IN THE BASAL FOREBRAIN
(C) IN VIVO RAT
Advantages: Functional effectsHistological effects
Disadvantages: Indirect actionBasic mechanism not revealed
(A) CHROMAFFIN/PC12 CELLS(B) OPTICAL IMAGING: BRAIN SLICES(C) IN VIVO RAT
In Vitro and In Vivo Models
NEURODEGENERATION IS AN ABERRANT
FORM OF DEVELOPMENT:
AChE-PEPTIDE IS THE PIVOTAL TOXIC
MOLECULEI. BiomarkerII. In Vitro and In Vivo
ModelsIII. Therapeutic Intervention
The 7 Receptor: The Target for AChE-Peptide?
The 7 Nicotinic Receptor• Coexpresses with AChE in developmental brain
(Broide et al,1996)
• Calcium permeability even greater than NMDA
(Séguéla et al,1993)
• Transgenic AD mouse (elevated Ab1-42): upregulation
(Svedverg et al, 2002)
• Binds amyloid peptide (Aβ) (Wang et al, 2000)
• Implicated in AD in conjunction of Aβ (Dineley,2001; Nagele, 2002)
B
10 μM BuChE peptide
10 μM AChE peptide
10 nM AChE peptide
10 μM BuChE peptide
10 μM AChE peptide
120 s
200 nA
α7
α4/β2
A
Log[AChE peptide] M
Per
cen
tag
e A
Ch
res
po
nse
B
C
-14 -12 -8 -10 -6 -4
50
100
150
AChE-Peptides act via 7-nAChR(Greenfield et al., 2004)
Compared with Control (A) & T15 peptide (D) Chronic Treatment with T14 (B) and T30 (C)
Increases α7-nAChR: A FEEDFORWARD MECHANISM!
Bond et al 2009
Normal AChE-Peptide Alpha-7 Blocker
Allosteric Modulator Site
Intracellular Pathways
Ranya Bechara
Cytotoxicity Assays
SRB MTT LDH
Mitochondria
Collagen
MTT
Cell Adhesion to collagen
Mitochondrial Function
Membrane Permeability
Measuring Cytotoxicity
Formazan
Comparison of T30 and Aβ
Cell Adhesion to collagen
Mitochondrial Function
Membrane Permeability
Ca2 +Calpain
Mitochondria
Spectrin
Nucleus
α7nAChR
Caspases
GSK
PFactor
s
Selective Intracellular Signalling Mechanisms of the
AChE Peptide
Ca2 +Calpain
Mitochondria
Spectrin
Nucleus
α7nAChR
Caspases
GSK
P
Factors
Selective Intracellular Signalling Mechanisms of the
AChE Peptide
(A) Marker for AChE PEPTIDE in blood detects degeneration before symptoms appear
+(B) Medication for arresting further
neuronal death by BLOCKING PEPTIDE ACTION
=PERMANENT PREVENTION OF
SYMPTOM ONSET!
Adding the Two Scenarios: An Eventual ‘Cure’?