Expanding the Window of Opportunity: Intransal Delivery of Wnt 3a and Whisker Stimulation in the Ischemic Stroke Mouse Model Público
Sharpe, Michka G. (2012)
Published
Abstract
Abstract
Expanding the Window of Opportunity: Intransal Delivery of Wnt 3a
and Whisker
Stimulation in the Ischemic Stroke Mouse Model
Each year, one out of six individuals will suffer a stroke. Despite
extensive basic and clinical
investigations, stroke remains to be the leading threat to human
life and health in the US and
worldwide. Up to now, there are very few effective treatments for
acute stroke patients. Novel
neuroprotective therapy and regenerative therapy are urgently
needed for clinical applications.
The purpose of this investigation was to test a combination therapy
of a canonical Wnt signaling
protein involved in neurodevelopment, Wnt 3a and whisker
stimulation in the enhancement of
neurogenesis and angiogenesis following focal ischemic stroke in
Nestin-GFP mice. In an
ischemic barrel cortex stroke mouse model induced by ligations of
distal branches of the middle
cerebral artery (MCA) in Nestin-GFP mice, Wnt 3a was administrated
intranasally to stimulate
the Wnt signaling pathway while peripheral whisker stimulation was
applied as a central specific
rehabilitation treatment after stroke. The combination therapy
showed increased expression of
brain-derived neurotrophic factor (BDNF) and vascular endothelial
growth factor (VEGF) in the
penumbra region 14 days after stroke. Co-labeling of
5-Bromo-2-deoxyuridine (BrdU) and NeuN
positive cells in the penumbra demonstrated augmented neurogenesis,
while increased co-
labeling of BrdU and Collagen IV suggested increased angiogenesis
in these mice. Wnt 3a paired
with whisker stimulation also increased local blood flow compared
to the saline control. In
addition, the SDF-1 receptor CXCR4 expression increased when Wnt 3a
was paired with
peripheral stimulation, suggesting enhanced sensitivity to
chemoattractant directed migration that
may promote homing of neuroblasts from the subventricular zone to
the infarct site. Cellular
migration was confirmed with positive Doublecortin (DCX) and BrdU
co-labeling.
These data support that the combination therapy targeting the
Wnt pathway paired with peripheral signals
may be beneficial for long-term tissue repair and functional
recovery after ischemic stroke.
Table of Contents
Table of Contents
Terminology and Abbreviations ...2
Abstract ...3 Introduction ...4i. Stroke
ii. Ischemic Cascade
iii. Penumbra as a therapeutic target region in stroke
iv. Limitations of Current Stroke Therapy
v. Peripheral Whisker Stimulation
vi. Wnt 3a and Canonical Wnt Signaling
vii. The Adult Brain and Neurogenesis
Specific Aims and Hypotheses ...14 Materials and Methods ...15i. Focal Ischemia in Animals
ii. Wnt 3a Intranasal Delivery
iii. BrdU Injections
iv. Whisker Stimulation
v. Measurement of Cerebral Blood Flow
vi. Immunohistochemistry
vii. Western Blot Analysis
viii. Sterological Cell Countingix. Statistical Analysis
Results ...20 Figures ...25Figure 1. Wnt 3a+Stimulation enhances blood flow to the penumbra 14 days after stroke.
Figure 2. Stimulation and not Wnt 3a+Stimulation enhances cerebral blood flow 21 days after stoke.
Figure 3. Immunohistochemistry for vessels, neuronal nuclei and newly proliferating cells.
Figure 4. Combination therapy further enhances neurogenesis in the penumbra 14 days after stroke.
Figure 5. Increased BDNF expression with combination of Wnt 3a and whisker stimulation.
Figure 6. Combination treatment of Wnt 3a+Stimulation further enhances angiogenesis 14 days after ischemic stroke.
Figure 7A. Wnt 3a+Whisker stimulation demonstrates cell migration towards the penumbra 14 days after ischemic stroke.
Figure 7B. Wnt 3a+Stimulation has increased CXCR4 expression compared to Wnt 3a and saline groups but not stimulation only group.
Figure 8. Increased expression of VEGF when stimulation is paired with Wnt 3a.
Figure 9. XAV939 Mechanism of Action.
Figure 10A. Ischemic stroke in saline control C57/B6 mouse.
Figure 10B. XAV939 Wnt antagonist truncates de novo neurogenesis 14 days following ischemic stroke.
Figure 11. XAV939 blocks Wnt 3a activity in vivo.
Discussion ...36 References ...45About this Honors Thesis
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