Early-Onset Formation of Parenchymal Plaque Amyloid Abrogates Cerebral Microvascular Amyloid Accumulation in Transgenic Mice.

Article (PDF Available)inJournal of Biological Chemistry 289(25) · May 2014with52 Reads
DOI: 10.1074/jbc.M113.536565 · Source: PubMed
Abstract
The fibrillar assembly and deposition of amyloid β-protein (Aβ), a key pathology of Alzheimers disease (AD), can occur in the form of parenchymal amyloid plaques and cerebral amyloid angiopathy (CAA). Familial forms of CAA exist in the absence of appreciable parenchymal amyloid pathology. The molecular interplay between parenchymal amyloid plaques and CAA is unclear. Here we investigated how early-onset parenchymal amyloid plaques impact the development of microvascular amyloid in transgenic mice. Tg-5xFAD mice, which produce non-mutated human Aβ and develop early-onset parenchymal amyloid plaques, were bred to Tg-SwDI mice, which produce familial CAA mutant human Aβ and develop cerebral microvascular amyloid. The bigenic mice presented with elevated accumulation of Aβ and fibrillar amyloid in brain compared to either single transgenic line. Tg-SwDI/Tg-5xFAD mice were devoid of microvascular amyloid, the prominent pathology of Tg-SwDI mice, but exhibited larger parenchymal amyloid plaques compared to Tg-5xFAD mice. The larger parenchymal amyloid deposits were associated with a higher loss of cortical neurons and elevated activated microglia in the bigenic Tg-SwDI/Tg-5xFAD mice. The periphery of parenchymal amyloid plaques was largely composed of CAA mutant Aβ. Non-mutated Aβ fibril seeds promoted CAA mutant Aβ fibril formation in vitro. Further, intrahippocampal administration of biotin-labeled CAA mutant Aβ peptide accumulated on and adjacent to pre-existing parenchymal amyloid plaques in Tg-5xFAD mice. These findings indicate that early-onset parenchymal amyloid plaques can serve as a scaffold to capture CAA mutant Aβ peptides and prevent their accumulation in cerebral microvessels.
1
EARLY-ONSET FORMATION OF PARENCHYMAL PLAQUE AMYLOID ABROGATES
CEREBRAL MICROVASCULAR AMYLOID ACCUMULATION IN TRANSGENIC MICE*
Feng Xu
1
, AnnMarie E. Kotarba
1
, Ziao Fu
2
, Judianne Davis
1
, Steven O. Smith
2
,
and William E. Van Nostrand
1
Departments of Neurosurgery & Medicine
1
and Biochemistry & Cell Biology
2
,
Stony Brook University, Stony Brook, NY 11794-8122
Running Head: Parenchymal amyloid inhibits vascular amyloid formation in mice
Address correspondence to Dr. W.E. Van Nostrand, Department of Neurosurgery, HSC T-12/089, Stony
Brook University, Stony Brook, NY 11794-8122. Tel. No. 631-444-1661; FAX No. 631-444-2560; E-
Mail: William.VanNostrand@sbumed.org
Keywords: Protein misfolding; amyloid; Alzheimers disease; transgenic mice; pathology, cerebral
vascular; plaque
Background: Mis-folded amyloid proteins deposit
in plaques and blood vessels in Alzheimers disease
brain.
Results: Early formation of amyloid plaques
impedes subsequent amyloid accumulation in
brain blood vessels.
Conclusion: Amyloid deposition in one
compartment can impact amyloid accumulation in
another compartment in brain.
Significance: Learning how amyloid proteins can
spread or block further formation is important for
understanding the progression of pathology in
neurodegenerative diseases.
ABSTRACT
The aggregation and deposition of amyloid
ß-protein (Aß), a key pathology of Alzheimers
disease (AD), can occur in the form of
parenchymal amyloid plaques and cerebral
amyloid angiopathy (CAA). Familial forms of
CAA exist in the absence of appreciable
parenchymal amyloid pathology. The
molecular interplay between parenchymal
amyloid plaques and CAA is unclear. Here we
investigated how early-onset parenchymal
amyloid plaques impact the development of
microvascular amyloid in transgenic mice. Tg-
5xFAD mice, which produce normal, wild-type
human and develop early-onset
parenchymal amyloid plaques, were bred to
Tg-SwDI mice, which produce familial CAA
mutant human and develop cerebral
microvascular amyloid. The bigenic mice
presented with elevated accumulation of
and fibrillar amyloid in brain compared to
either single transgenic line. Tg-SwDI/Tg-
5xFAD mice were devoid of microvascular
amyloid, the prominent pathology of Tg-SwDI
mice, but exhibited larger parenchymal
amyloid plaques compared to Tg-5xFAD mice.
The larger parenchymal amyloid deposits were
associated with a higher loss of cortical neurons
and elevated activated microglia in the bigenic
Tg-SwDI/Tg-5xFAD mice. The periphery of
parenchymal amyloid plaques was largely
composed of CAA mutant Aß. Wild-type Aß
fibril seeds promoted CAA mutant Aß fibril
formation in vitro. Further, intrahippocampal
administration of biotin-labeled CAA mutant
peptide accumulated on and adjacent to
pre-existing parenchymal amyloid plaques in
Tg-5xFAD mice. These findings indicate that
early-onset parenchymal amyloid plaques can
serve as a scaffold to capture CAA mutant
peptides and prevent their accumulation in
cerebral microvessels.
The molecular seeding and deposition of mis-
folded proteins is a common process of various
neurodegenerative disorders (1,2). In Alzheimers
disease (AD
1
) and related disorders the
extracellular deposition of mis-folded amyloid ß-
protein (Aß) in brain is a prominent pathological
feature (3,4). is a 39-43 amino acid peptide
that exhibits a high propensity to self-assemble
into ß sheet-containing oligomeric forms and
fibrils (5,6). peptides are proteolytically
derived from a large type I integral membrane
precursor protein, termed the amyloid ß-protein
precursor (AßPP), which is encoded by a gene
located on chromosome 21 (7-9). The
2
amyloidogenic processing of AßPP initially
involves a cleavage at the amino terminus of
peptide sequence by ß-secretase, an aspartyl
proteinase named BACE (10,11). Subsequent
cleavage of the remaining amyloidogenic
membrane spanning PP carboxyl terminal
fragment by g-secretase liberates the predominant
Aß40 or Aß42 residue peptides. Presenilin (PS)
proteins contain the proteolytic active site as part
of a multi-protein γ-secretase complex (12,13).
In addition to plaques in the AD brain
parenchyma another prominent site of
extracellular Aß deposition is within and along
primarily small and medium-sized arteries and
arterioles of the cerebral cortex and leptomeninges
and in the cerebral microvasculature, a condition
known as cerebral amyloid angiopathy (CAA)
(14,15) There are two prominent forms of CAA
known as CAA type-1 and CAA type-2 (16). In
CAA-type 2 the amyloid deposition is restricted
within the vessel wall of the cortical and
meningeal arterioles and arteries and does not
promote surrounding neuroinflammation (15-17).
Accumulation of fibrillar in CAA type-2 has
been shown to cause degeneration and cell death
of smooth muscle cells in the affected larger
cerebral vessels and trigger hemorrhage (15-18).
On the other hand, CAA type-1 involves amyloid
deposition along the brain capillaries (16). In
contrast to CAA type-2 where the amyloid is
confined within the vessel wall, CAA type-1
results in penetrance of the fibrillar amyloid
deposits into the surrounding brain parenchyma
promoting a robust localized neuroinflammatory
response characterized by strong perivascular
microglial activation, accumulation of
hyperphosphorylated tau protein, and neuronal
degeneration (17,19-21). Furthermore, an
increasing number of studies have implicated
cerebral microvascular Aß deposition in
promoting neuroinflammation and dementia in AD
and in familial and sporadic cases of CAA
(17,22,23). In particular, cerebral microvascular,
but not parenchymal, amyloid deposition is more
often correlated with dementia in individuals
afflicted with AD and spontaneous CAA disorders
as well as in mouse models (24-26).
In addition to the prominent CAA that is
found in AD and in spontaneous cases of this
condition, several monogenic, familial forms of
CAA exist that result from mutations that reside
within the Aß peptide sequence of AßPP gene (27-
31). The most recognized example of familial
CAA is the Dutch-type E22Q substitution in
that causes early and severe cerebral vascular
amyloid deposition leading to recurrent, and often
fatal, intracerebral hemorrhages at mid-life
(27,28,32-34). Similarly, the Iowa-type D23N
substitution in Aß also causes early and severe
cerebral vascular amyloid deposition (31). In
contrast to the Dutch-type disease, Iowa-type
CAA appears more localized to the cerebral
microvasculature. In contrast to AD, in both
Dutch-type and Iowa-type familial CAA there is a
striking absence of appreciable parenchymal
amyloid plaque pathology despite the highly
fibrillogenic nature of Dutch and Iowa CAA
mutant peptides (35-37). In any case, the impact of
parenchymal amyloid plaques on the development
of CAA is not well understood.
We previously developed a unique transgenic
mouse model, designated Tg-SwDI, that expresses
low levels of familial Dutch/Iowa CAA mutant
human AßPP in brain and develops early-onset
and progressive accumulation of microvascular
CAA (38,39). Tg-SwDI mice do not develop
parenchymal fibrillar Aß plaque deposits
consistent with the pathology observed in Dutch-
type and Iowa-type patients. In close association
with cerebral microvascular fibrillar amyloid there
is a robust neuroinflammatory response in Tg-
SwDI mice characterized by highly increased
numbers of activated microglia and elevated levels
of pro-inflammatory cytokines (39,40). Moreover,
Tg-SwDI mice exhibit behavioral deficits that
coincide with the development of the early-onset
cerebral microvascular amyloid and
neuroinflammation (40,41). On the other hand,
Tg-5xFAD, a commonly used transgenic mouse
model that expresses high levels of human AßPP
in brain, produces wild-type human peptides
and develops progressive parenchymal fibrillar
deposits (42).
Cerebral vascular fibrillar amyloid can occur
in the presence or absence of parenchymal fibrillar
amyloid pathology, as in AD or in familial CAA,
respectively. Although parenchymal and cerebral
vascular fibrillar amyloid exists in distinct
compartments of the brain the molecular interplay
between these two pathologies is poorly
understood. Accordingly, in the present study we
generated bigenic Tg-SwDI/Tg-5xFAD mice to
3
investigate the impact of early-onset parenchymal
plaque formation on the development of
microvascular CAA. Here we show that in the
bigenic mice there is a conspicuous absence of
microvascular CAA, the prominent pathology in
Tg-SwDI mice, and a shift to larger parenchymal
fibrillar amyloid plaques compared to Tg-5xFAD
mice. This alteration in fibrillar amyloid pathology
resulted in more severe neuronal loss and
microglial activation in the Tg-SwDI/Tg-5xFAD
mice. Wild-type Aß42 amyloid fibril seeds
promoted the fibrillar assembly of Dutch/Iowa
CAA mutant Aß in vitro. Lastly, intrahippocampal
injection of biotin-labeled Dutch/Iowa CAA
mutant Aß40 strongly deposited on and adjacent to
pre-existing parenchymal fibrillar amyloid plaques
in Tg-5xFAD mice. Together, these findings show
that early-onset parenchymal fibrillar amyloid
plaques primarily composed of wild-type Aß42
can serve as a scaffold to recruit the co-deposition
of CAA mutant Aß peptides and prevent the
development of microvascular CAA. This suggests
that in brain mis-folded protein deposition in the
parenchymal compartment can significantly
impact mis-folded protein deposition in the
vascular compartment.
EXPERIMENTAL PROCEDURES
Reagents and Chemicals - peptides were
synthesized at the Keck Peptide Synthesis facility
at Yale University using tBOC-chemistry.
Hydrofluoric acid was used for cleavage and
deprotection. The peptides were purified by
reverse phase HPLC, using linear water-
acetonitrile gradients containing 0.1%
trifluoroacetic acid. The purity was estimated at
>90-95% on the basis of analytical RP-HPLC and
matrix-assisted laser desorption ionization mass
spectrometry. peptides were initially prepared
in 1,1,1,3,3,3-hexafluoro-2-propanol, flash frozen,
lyophilized to remove solvent, and resuspended in
pure DMSO. Thioflavin-T was purchased from
Sigma-Aldrich (St. Louis, MO).
AnimalsAll work with animals was approved by
the Stony Brook University Institutional Animal
Care and Use Committee and in compliance with
the guidelines established by the Public Health
Service Guide for the Care and Use of Laboratory
Animals. Generation and characterization of Tg-
SwDI mice were previously described (36,37).
Tg-5xFAD mice were generated as described by
Oakley et al. (42) and obtained from the Jackson
Laboratories. Heterozyous Tg-SwDI mice on a
pure C57/Bl6 background were bred with
heterozygous Tg-5xFAD mice on a mixed
C57/Bl6/Sjl background to obtain heterozygous
Tg-SwDI, heterozygous Tg-5xFAD, and bigenic
Tg-SwDI/Tg-5xFAD mice all on the same mixed
background. Eight to ten mice of each genotype
were examined at 3, 6 and 9 months of age for
quantitative immunochemical and pathological
studies.
Tissue Preparation Mice were sacrificed at
designated time points, the brains were
immediately removed and bisected in the mid-
sagittal plane. One hemisphere was snap-frozen
and used for the protein analyses. The other
hemisphere was placed in 70% ethanol, followed
by xylene treatment and embedding in paraffin.
Sagittal sections were cut at 10 µm thickness using
a microtome, placed in a flotation water bath at 45
o
C, and then mounted on Colorfrost/Plus slides
(Fisher Scientific, Houston, TX) for
immunohistochemical and histological analyses.
ELISA Measurement of Peptides Soluble
pools of Aß40 and Aß42 were determined by
using specific ELISAs on carbonate extracted
mouse forebrain tissue and subsequently the
insoluble Aß40 and Aß42 levels were determined
by ELISA of guanidine lysates of the insoluble
pellets resulting from the carbonate extracted brain
tissue as described (43,44). Total Aß40 and Aß42
levels were determined by combining the soluble
and insoluble pools of each species. In the
sandwich ELISAs Aß40 and Aß42 were captured
using their respective carboxyl-terminal specific
antibodies m2G3 and m21F12 and biotinylated
m3D6, specific for human Aß, was used for
detection (43). Each brain lysate was measured in
triplicate and compared to linear standard curves
generated with known concentrations of human
using a Spectramax M2 plate reader
(Molecular Devices, Sunnyvale, CA). Total Aß40
and Aß42 levels were determined by combining
the soluble and insoluble levels of each form.
Immunohistochemical Analysis Procedures were
performed as previously described (38,39).
4
Briefly, sections were cut in the sagittal plane at
10 µm thickness using a microtome,
deparaffinated and rehydrated. Antigen retrieval
was performed by treatment with proteinase K (0.2
mg/ml) for 10 min at 22
o
C for Aß, neuronal and
collagen labeling, and by 10 mM sodium citrate
solution (pH 9.0) for 30 min at 90
o
C in a water-
bath for activated microglia labeling. The
following antibodies were used for
immunolabeling analysis: rabbit polyclonal
antibody directed towards residues 15 of human
Aß (1:200, 45), mouse monoclonal 4G8, which
recognizes an epitope between residues 17-24 of
wild-type (1:200; Covance, Princeton, NJ),
mouse monoclonal antibody NeuN to identify
neurons (1:500; Chemicon, Temecula, CA), rabbit
polyclonal antibody to collagen type IV to identify
cerebral microvessels (1:100; Research
Diagnostics Inc., Flanders, NJ), mouse
monoclonal antibody 5D4 to identify activated
microglia (1:1000; Seikagaku Corp., Tokyo,
Japan). Primary antibodies were detected with
horseradish peroxidase-conjugated secondary
antibodies (1:1000; Vector Labs, Burlingame, CA)
and visualized with a stable diaminobenzidine
solution (Invitrogen, Carlsbad, CA) as substrate.
Alternatively, deposited fibrillar amyloid was
detected with thioflavin-S staining and the primary
antibodies to collagen type IV to visualize cerebral
microvessels or mAb5D4 for activated microglia
were detected with an Alexa Fluor 594-conjugated
secondary antibody (1:1000; Invitrogen, Carlsbad,
CA).
Quantitative Analysis of Regional Parenchymal
and Microvascular Amyloid DepositionTotal Aß
and fibrillar amyloid burden in the regions of the
cortex, thalamus and subiculum was quantified on
the same set of systematically sampled
immunostained or thioflavin-S stained sections,
respectively, using NIH image J 1.32 software.
The percentage of thioflavin-S labeled
microvessels in the regions of the fronto-temporal
cortex, thalamus and subiculum was respectively
quantified on the same set of systematically
sampled thioflavin-S stained sections using
stereological principles as described (46).
Quantitative Analysis of Cortical Neuronal
Denisities Total numbers of neurons in cortical
layer V were estimated using a computerized
stereology system (Stereologer, Systems Planning
and Analysis, Alexandria, VA). Every tenth
section was selected and generated 1015 sections
per reference space in a systematic-random
manner. Immuno-positive cells were counted
using the optical fractionator method with the
dissector principle and unbiased counting rules
(46).
Thioflavin T Measurements All peptides
were solubilized in pure DMSO and diluted in 10
mM phosphate buffer to a concentration of 20 µM
at a pH of 7.4. For seeding experiments, wild-type
42 seeds were grown by incubating the peptide
at 37 °C to form mature fibrils. Fibril formation
was confirmed by transmission electron
microscopy. 42 seeds were sonicated in a glass
vial for 10 min before use. Fresh solutions of the
Dutch/Iowa CAA mutant 40 peptide were
prepared as above and Aß42 seeds were added to
make a final peptide solution that was 20 µM
Dutch/Iowa Aß40 and either 10 or 20 µM wild-
type Aß42 (as seeds). Thioflavin T fluorescence
was monitored with a SpectraMax M2 microplate
reader every 15 s with excitation, emission and
automatic cutoff wavelengths of 446, 490 and 475
nm, respectively. Each experimental point is the
mean of the fluorescence signal of three wells
containing aliquots of the same solution.
Intrahippocampal Injection of Biotinylated-
Aß40DI Peptide in Tg-5xFAD Mice Six months
old Tg-5xFAD mice were anesthetized with
Avertin (250 mg/kg), then secured in a stereotaxic
frame (David Kopf Instruments, Tujunga, CA). A
sagittal incision was made caudal to rostral
allowing the scalp to be retracted and held in place
with micro-clips to expose the skull surface. To
insert the Hamilton syringe (30 gauge), a small
burr hole was drilled in the parietal bone. Two µl
of freshly prepared biotin-labeled 40DI peptide,
dissolved in sterile saline at concentration of 0.5
mg/ml was stereotactically injected into the
hippocampus (bregma -2.00 mm anterior, lateral
1.50 mm, 1.80 mm deep) at a rate of 0.3 µl/min
using a micro-injection unit. The needle was left in
place for 5 min post-injection to minimize reflux
before being slowly removed. The scalp was
closed under sterile conditions using 4-0 nylon
sutures and the animals were placed in a cage
warmed with a heating pad and observed until it is
5
alert and mobile. Twenty four hours later the mice
were sacrificed, the brains removed and processed
as described above.
Statistical Analysis The data were analyzed by
one-way ANOVAs for each measure at each brain
region. Significant ANOVAs (p < 0.05) were
followed by Fisher’s post-hoc tests the results of
which are reported in the corresponding figure
legends.
RESULTS
Bigenic Tg-SwDI/Tg-5xFAD Mice Accumulate
Elevated Levels of Cerebral Peptides In this
study we sought to determine the influence of
early-onset parenchymal amyloid plaque
formation on the development of cerebral
microvascular amyloid accumulation in transgenic
mice. Tg-5xFAD mice, which produce human
wild-type and develop early-onset
parenchymal amyloid plaques, were bred to Tg-
SwDI mice, which produce human Dutch/Iowa
CAA mutant and develop cerebral
microvascular amyloid. Comparisons were
performed of the bigenic mice generated from this
cross with the single transgenic animals. After
aging three to nine months, quantitative ELISAs
were performed to measure human Aß in forebrain
homogenates prepared from each line of mice. As
shown in Fig. 1, all mice showed a progressive
accumulation of in brain. At all ages Tg-
5xFAD mice accumulated much higher levels of
cerebral than Tg-SwDI mice consistent with
earlier studies (26). However, the bigenic Tg-
SwDI/Tg-5xFAD mice accumulated much higher
levels of than each single transgenic line. In
particular, at three and six months of age there was
a significant 44% (p < 0.05) and 67% (p < 0.01)
respective increase in total cerebral Aß in the
bigenic mice compared to the additive amounts of
cerebral Aß present in each single transgenic line.
We next performed immunolabeling studies to
characterize the deposition of human Aß in the
brains of the different mice. As shown in Fig. 2, at
three months of age Tg-SwDI mice showed early
stage deposition most prominent in the
subiculum region (Fig. 2A and 2J), whereas the
Tg-5xFAD mice and bigenic Tg-SwDI/Tg-5xFAD
mice exhibited more widespread deposition of Aß,
but was also highest in the subiculum region (Fig.
2D, 2G and 2J). As all three lines of mice
continued to age to six and nine months the
amount of deposited markedly increased and
was most robust in the bigenic Tg-SwDI/Tg-
5xFAD mice reflecting the ELISA results
obtained in Fig. 1.
Bigenic Tg-SwDI/Tg-5xFAD Mice Exhibit Altered
Fibrillar Amyloid Deposition To evaluate
fibrillar amyloid deposition in brain we performed
thioflavin S staining. Overview images revealed
little fibrillar amyloid was observed at three
months of age in Tg-SwDI mice (Fig. 3A) but an
increase in amyloid was observed as the mice aged
from six to nine months, primarily in the
subiculum region (Fig. 3B, 3C and 3J). Although
Tg-SwDI mice deposit appreciable in the brain
(Fig. 2A-C) this is mostly in diffuse form with
fibrillar amyloid primarily restricted to small
microvascular deposits (38,39). In contrast, both
Tg-5xFAD mice (Fig. 3D-F) and Tg-SwDI/Tg-
5xFAD mice (Fig. 3G-I) showed more widespread
fibrillar amyloid deposition that was generally
higher in the bigenic animals (Fig. 3J).
Subsequent examination of the brain tissues at
higher magnification showed that over the course
of three to nine months the Tg-SwDI mice
exhibited the typical progressive pattern of fibrillar
amyloid accumulation primarily in the form of
small cerebral microvascular deposits (Fig. 4A-C).
On the other hand, Tg-5xFAD mice showed
progressive accumulation of parenchymal fibrillar
amyloid plaques (Fig. 4G-I). Interestingly, the
bigenic Tg-SwDI/Tg-5xFAD mice (Fig. 4D-F)
exhibited striking differences compared to each
single transgenic line. Remarkably, in the bigenic
animals there was essentially a complete loss of
cerebral microvascular amyloid deposition as
characteristically observed in Tg-SwDI mice.
Quantitative analysis showed very high levels of
cerebral microvascular amyloid in the subiculum
region of nine months old Tg-SwDI mice that
were essentially absent in the Tg-5xFAD mice and
bigenic Tg-SwDI/Tg-5xFAD mice (Fig. 4J). The
other notable feature was the tendency for larger
fibrillar amyloid plaques in the bigenic mice
compared to Tg-5xFAD mice. The size
distribution of fibrillar amyloid deposits was
measured in the subiculum region of the different
lines of mice. As shown in Fig. 4K, 90% of the
fibrillar amyloid deposits in Tg-SwDI mice were
6
<50 µm
2
presented primarily as small
microvascular deposits (Fig. 4A-C). In contrast,
Tg-5xFAD mice present a range of fibrillar
amyloid deposits, in this case parenchymal
plaques with the majority between 50-250 µm
2
.
However, in the bigenic Tg-SwDI/Tg-5xFAD
mice the size distribution of fibrillar plaques
markedly changed. For example, there was a 32%
reduction in fibrillar deposits <50 µm
2
(p < 0.01)
whereas there was a 13-fold increase in the
percentage of very large fibrillar plaques of >500
µm
2
(p < 0.0001). Together, these results indicate
that in bigenic Tg-SwDI/Tg-5xFAD mice there is
a complete loss of microvascular amyloid and
growth of larger parenchymal amyloid plaques.
Increased Parenchymal Fibrillar Amyloid
Deposits in Bigenic Tg-SwDI/Tg-5xFAD Mice
Enhances Neuronal Loss and Microglial
Activation It was previously reported that Tg-
5xFAD mice exhibit neuronal loss, specifically in
cortical layer V, with increasing age and
pathology (42,47). Therefore, we examined
neuronal loss in the different transgenic lines with
respect to Aß deposition, fibrillar amyloid and
microglial activation. Although six months old Tg-
SwDI mice show accumulation of considerable
parenchymal diffuse Aß in the cortex they have
only minimal amounts of microvascular fibrillar
amyloid and sparse microglial activation resulting
in no neuronal loss in cortical layer V (Fig. 4E-
H,Q). In contrast, at the same age Tg-5xFAD mice
have substantial parenchymal deposition,
fibrillar amyloid plaques and accompanying
microglial activation that coincide with an 11%
loss in cortical layer V neurons (Fig. 4I-L,Q).
Moreover, the bigenic Tg-SwDI/Tg-5xFAD mice
have even higher amounts of parenchymal
deposition, fibrillar amyloid plaques and increased
microglial activation resulting in a near doubling
of the amount of neuronal loss in cortical layer V
(Fig. 4M-P,Q). As the mice aged to nine months
the loss of cortical layer V neurons further
increased in Tg-5xFAD mice and Tg-SwDI/Tg-
5xFAD mice while no changes occurred in Tg-
SwDI mice (data not shown).
Since the parenchymal fibrillar amyloid
plaques tended to be larger, were associated with
more extensive microglial activation and neuronal
loss we next sought to determine the distribution
of wild-type and CAA mutant in the
plaques of bigenic Tg-SwDI/Tg5xFAD mice. To
discriminate between human wild-type and
human Dutch/Iowa CAA mutant peptides in
the plaques we used the monoclonal antibody 4G8,
which recognizes a mid-region epitope on human
(48). The presence of the E22Q Dutch and
D23N Iowa mutations abolishes the 4G8 epitope
in human . Fig. 6A shows dot blot analysis
confirming that the N-terminal human Aß rabbit
polyclonal antibody (pAb-Aß) recognizes both
wild-type and Dutch/Iowa CAA mutant
whereas mAb4G8 only recognizes wild-type Aß.
As shown in Fig. 6B-D, deposits in Tg-SwDI
mice are recognized by pAb-Aß, but not mAb4G8.
On the other hand, parenchymal deposits in
Tg-5xFAD mice are recognized by both pAb-
and mAb4G8 (Fig. 6E-G). Although the core of
the plaques in bigenic Tg-SwDI/Tg-5xFAD mice
were labeled with both pAb-and mAb4G8 the
periphery of the plaques contained a halo of small
deposits that were labeled only with pAb-
suggesting that these satellite plaques were
composed primarily of Dutch/Iowa CAA mutant
Aß (Fig. 6J).
Wild-type Aß42 Fibrils Seed Dutch/Iowa CAA
Mutant Aß Fibril Formation and Deposition The
above findings suggest that early-onset
parenchymal fibrillar plaques, primarily composed
of wild-type Aß42, can act as a scaffold to recruit
the co-deposition of Dutch/Iowa CAA mutant
in the bigenic Tg-SwDI/Tg-5xFAD mice. To
directly test if wild-type Aß42 fibrils could
promote Dutch/Iowa CAA mutant Aß40 assembly,
we measured the kinetics of fibril formation of the
latter in the presence or absence of wild-type Aß42
fibril seeds. Fibril seeds were generated by
incubating Aß42 at 37 °C to form mature fibrils
followed by bath sonication to break the fibrils
into short segments. The addition of seeds to a
population of wild-type Aß42 monomers
eliminates the lag phase associated with formation
of a fibril nucleus (data not shown). As shown in
Fig. 7, the Dutch/Iowa CAA mutant Aß40 when
incubated at a monomer concentration of 20 µM
exhibits a lag phase of 2 h prior to an increase of
thioflavin T fluorescence, which is characteristic
of fibril formation. The addition of monomeric
Dutch/Iowa CAA mutant Aß40 to wild-type 42
fibril seeds at concentrations of either 10 µM or 20
µM eliminated the lag phase and resulted in a
7
rapid rise of thioflavin T fluorescence. The
increase in fluorescence is attributed to the
addition of the Dutch/Iowa CAA mutant Aß40 to
fibril seeds. The wild-type Aß42 seeds without
added CAA mutant Aß40 monomer do not result
in a change of fluorescence as expected.
We next tested this phenomenon in vivo by
injecting biotin-labeled Dutch/Iowa CAA mutant
Aß40 into the hippocampal region of six months
old Tg-5xFAD mice, which exhibit prominent
parenchymal plaque fibrillar amyloid deposition
(see Fig. 3H). The injected biotin-labeled
Dutch/Iowa CAA mutant Aß40 showed strong
accumulation on pre-existing parenchymal fibrillar
amyloid plaques (Fig. 8A-D). No accumulations
were found when biotin alone was injected into
similarly aged Tg-5xFAD mice (Fig. 8E-H).
Interestingly, higher magnification of the
parenchymal plaques (Fig. 8D) showed that biotin-
labeled Dutch/Iowa CAA mutant accumulated
around the periphery of the plaques forming a halo
of deposits highly reminiscent of its deposition in
the bigenic Tg-SwDI/Tg-5xFAD mice (Fig. 6J).
Together, these results indicate that wild-type
Aß42 fibrils can seed rapid fibrillar assembly and
co-deposition of Dutch/Iowa CAA mutant Aß onto
parenchymal amyloid plaques.
DISCUSSION
The seeding, spreading and deposition of mis-
folded proteins in brain, first associated with prion
disorders, is now recognized as a common
pathological process of many neurodegenerative
diseases including AD, Parkinson’s disease,
amyotrophic lateral sclerosis and fronto-temporal
dementia (1,2). In the case of AD, the
accumulation of fibrillar occurs in two distinct
compartments: in the brain parenchyma in the
form of plaques and in cerebral blood vessels as
vascular amyloid (5,14,15). Although the
pathology at both sites involves Aß deposition the
crosstalk between the parenchymal and cerebral
vascular amyloid remains unclear. Here, we
addressed this issue by crossing Tg-5xFAD mice,
a model of early-onset parenchymal plaque
amyloid, with Tg-SwDI mice, a model of cerebral
microvascular amyloid, to determine the influence
on the amyloid pathology in each compartment.
We show that in the bigenic Tg-SwDI/Tg-5xFAD
mice there is an overall loss of microvascular
amyloid and general shift to larger sized
parenchymal amyloid plaques accompanied with
more severe pathological consequences.
In AD patients and mouse models of
pathology the emergence of CAA is generally a
secondary event and exists in the presence of
abundant parenchymal fibrillar amyloid plaques
(3,4). Previous studies using transgenic rodent
models have shown that primarily parenchymal,
and some cerebral vascular, pathology can be
seeded and spread with exogenous intracerebral
administration of amyloid material in a cis manner
using fibrillar seeds (49-52). Alternatively, in
certain cases pathology could be seeded in a
trans manner using distinct amyloid material such
as prion proteins (53).
In contrast to AD, in familial forms of CAA
involving mutated forms of Aß and in
corresponding mouse models the cerebral vascular
amyloid is more extensive and occurs in the
absence of appreciable parenchymal fibrillar
amyloid deposition (31-34). However, familial
CAA patients generally produce a mixture of wild-
type and CAA mutant Aß peptides in brain. In
earlier efforts to replicate this scenario, transgenic
mouse models of relatively late parenchymal
amyloid deposition were bred with familial CAA
transgenic mouse models that exclusively develop
cerebral vascular amyloid (54,55). In these studies,
the bigenic mice exhibited a marked increase in
cerebral vascular amyloid composed of large
amounts of wild-type suggesting that familial
CAA mutant vascular amyloid deposits can
capture and co-deposit wild-type peptides. On
the other hand, the influence of early-onset
parenchymal fibrillar amyloid plaques on
subsequent cerebral vascular amyloid formation is
less clear. To address this issue we chose the Tg-
5xFAD mouse model that aggressively develops
amyloid plaque pathology starting at about two
months of age (26,42). When crossed with Tg-
SwDI mice, the bigenic Tg-SwDI/Tg-5xFAD mice
accumulated higher amounts of Aß and fibrillar
amyloid than either single transgenic line.
However, the compartmental distribution of
fibrillar amyloid was significantly altered in the
bigenic Tg-SwDI/Tg-5xFAD mice. Notably, there
was a complete loss of cerebral microvascular
amyloid and a general increase in the size
8
distribution of parenchymal amyloid deposits (Fig.
4).
The composition of the enhanced amyloid
plaques in bigenic Tg-SwDI/Tg-5xFAD mice is
intriguing. The use of differential antibody
recognition indicates that the core of the bigenic
amyloid plaques is likely composed of wild-type
and perhaps CAA mutant human Aß.
However, the plaque periphery appears to be
composed primarily of CAA mutant Aß, since it
lacks recognition by the mAb4G8 antibody (Fig.
6). This finding suggests that in the bigenic mice
parenchymal fibrillar amyloid plaques can act as a
scaffold to capture Dutch/Iowa CAA mutant
and promote its local assembly and deposition into
hybrid plaques, thus precluding microvascular
amyloid formation. This notion is supported by
both our in vitro data demonstrating that wild-type
Aß42 fibrillar seeds can increase the rate of
Dutch/Iowa CAA mutant fibril assembly and
in vivo results showing that exogenously
administered Dutch/Iowa CAA mutant
strongly co-deposits on and adjacent to fibrillar
amyloid plaques in Tg-5xFAD mice. The
core/periphery composition of related, but distinct,
peptides in the parenchymal amyloid plaques
of the bigenic Tg-SwDI/Tg-5xFAD mice is highly
similar to previous findings in mice where one
strain of prion protein was shown to form an initial
prion plaque core that could act as a scaffold for
the aggregation and deposition of another strain of
prion protein along the periphery to form hybrid
plaques (56). The formation of these small,
peripheral satellite plaque deposits of Dutch/Iowa
CAA mutant may further cluster to grow the
size of the initial plaque deposit as shown recently
in APP/PS1 mice (57).
In addition to their increase in size, the
parenchymal amyloid plaque deposits in the
bigenic Tg-SwDI/Tg-5xFAD mice appear to have
enhanced pathological consequences. For
example, there appears to be a more robust
activated microglial response around the amyloid
plaques in the bigenic mice compared to the Tg-
5xFAD mice (Fig. 5). Previously, we showed that
the small cerebral microvascular amyloid deposits
in Tg-SwDI mice, composed of Dutch/Iowa CAA
mutant Aß, promote a strong neuroinflammatory
response and activation of microglia around the
affected capillaries (39,40). Thus, the
accumulation of Dutch/Iowa CAA mutant
deposits on the periphery of the parenchymal
plaques in the bigenic animals may similarly lead
to enhanced microglial activation. In addition, the
presence of Dutch/Iowa CAA mutant Aß in
parenchymal plaques results in further loss of
cortical neurons in the bigenic mice compared to
Tg-5xFAD mice. This result is consistent with
recent studies showing that a marked shift from
microvascular amyloid to parenchymal fibrillar
amyloid deposits of Dutch/Iowa CAA mutant
in bigenic Tg-SwDI/hApoE mice led to cortical
neuronal loss (58). Together, these findings
suggest that shifting CAA mutant fibrillar amyloid
from the cerebral vascular compartment to the
parenchymal compartment in the form of plaques
can enhance the surrounding pathology and
neuronal loss.
In both Tg-5xFAD mice and Tg-SwDI mice
the source of wild-type and CAA mutant human
peptides is through neuronal expression and
processing of transgene human PP (38,42). A
normal clearance route for neuronally-derived
peptides in transgenic mice, as well as in humans,
is migration through the brain via interstitial fluid
to the cerebral capillaries where it is either
transported across the blood-brain barrier into the
blood or continues through perivascular drainage
pathways into the CSF (59-62). In Tg-5xFAD
mice the wild-type Aß can assemble and deposit as
parenchymal amyloid plaques whereas in Tg-
SwDI mice the CAA mutant Aß does not, but
instead migrates to the brain capillaries where
rather than be transported into the blood it
assembles and deposits as microvascular fibrillar
amyloid.
It remains uncertain why the Dutch E22Q and
Iowa D23N mutations in lead to preferential
accumulation of fibrillar amyloid in the cerebral
vessels whereas parenchymal fibrillar amyloid
deposits are largely absent is uncertain. There are,
however, several properties of CAA mutant
peptides that may contribute to their affinity for
vascular assembly and deposition relative to the
brain parenchyma. For example, both the Dutch
E22Q and Iowa D23N mutations result in loss of a
negative charge at their respective adjacent sites.
Earlier studies have demonstrated that these
substitutions increase the fibrillogenic and
cytotoxic properties of CAA mutant Aß compared
with wild-type Aß (35-37,63-65). Furthermore,
the presence of these substitutions in CAA mutant
9
forms of may induce conformational changes
in the monomeric peptide and/or oligomeric
assemblies that suppress or enhance further
assembly and deposition in the parenchyma or
cerebral vasculature, respectively. Consistent with
this idea, it was previously shown that the GM3
ganglioside, which is present in cultured
cerebrovascular cells and in cerebral vessels,
preferentially enhances fibrillar assembly of CAA
mutant (66,67). Lastly, CAA mutant
peptides show significantly impaired clearance
from brain via transport across the blood-brain
barrier into the circulatory system and perivascular
drainage to the cerebrospinal fluid (60,68). This
reduced clearance could allow for abnormal
accumulation of CAA mutant at the cerebral
vessels. These markedly altered properties of CAA
mutant forms of likely provide for its
preferential and excessive accumulation as fibrillar
amyloid in the cerebral vasculature.
Familial CAA patients and the bigenic Tg-
SwDI/Tg-5xFAD mice produce both wild-type
and CAA mutant Aß in brain. However, in familial
CAA patients the fibrillar amyloid is primarily
restricted to the cerebral vasculature whereas in
the bigenic mice parenchymal amyloid pathology
increased while cerebral microvascular amyloid
pathology essentially vanished. This difference
may result from the relative amounts of wild-type
Aß and CAA mutant Aß in human and the bigenic
mouse brain. Familial CAA patients generally
harbor one CAA mutant and one wild-type AßPP
allele thus yielding similar amounts of the
corresponding Aß peptide. One the other hand,
Tg-SwDI mice express human AßPP and produce
CAA mutant Aß at physiological levels (38,60)
whereas Tg-5xFAD mice vastly over-express
human AßPP and produce very high levels of
wild-type (42) which is reflected in the
disparate amounts of in the brains of each
transgenic line (Fig. 1). Further, the appearance of
fibrillar amyloid plaques is very aggressive in Tg-
5xFAD mice with an onset at about two months of
age (42). Consequently, the very early appearance
of parenchymal fibrillar amyloid plaques in the
bigenic Tg-SwDI/Tg-5xFAD mice appears
sufficient to serve as an effective scaffold to trap
and deposit neuronally-derived CAA mutant
preventing its normal migration to the cerebral
microvasculature. In conclusion, although fibrillar
amyloid can seed and spread subsequent
pathologies, our findings indicate that in certain
situations fibrillar amyloid can act as an efficient
sink to enhance amyloid accumulation in the
parenchymal compartment and forego amyloid
deposition in the vascular compartment of the
brain.
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14
FOOTNOTES
*This work was supported by National Institutes of Health grants AG033209 (WEVN) and AG027317
(SOS), Alzheimer’s Association grant IIRG-09-15254, and a gift from the Cowles Charitable Trust. The
ELISA antibody reagents were generously provided by Lilly Research Laboratories. The costs of
publication of this article were defrayed in part by the payment of page charges. This article must
therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate
this fact.
1
The abbreviations used are: AD, Alzheimer's disease; Aß, amyloid ß-protein; AßPP, amyloid ß-protein
precursor; CAA, cerebral amyloid angiopathy; 40DI, Dutch/Iowa CAA mutant 40 peptide;, ThS,
thioflavin S; mAb, monoclonal antibody; pAb, polyclonal antibody.
15
FIGURE LEGENDS
Fig. 1. ELISA measurement of levels in brains of Tg-SwDI, Tg5xFAD and bigenic Tg-
SwDI/Tg5xFAD mice. The total levels of were determined in mouse forebrain extracts of three to
nine month old Tg-SwDI mice (black bars), Tg-5xFAD mice (blue bars) and bigenic Tg-SwDI/Tg-
5xFAD mice (red bars) by ELISA analysis as described in Methods. The data presented are the mean ±
S.D. of eight to ten mice per timepoint. *p < 0.05; **p < 0.01.
Fig. 2. Progressive accumulation of Aß deposition in Tg-SwDI, Tg5xFAD and bigenic Tg-SwDI/Tg-
5xFAD mouse brain. Representative images of three to nine months old transgenic mouse brain sections
were immunostained for as described in Methods. A-C, Tg-SwDI mice; D-F, bigenic Tg-SwDI/Tg-
5xFAD mice; and G-I, Tg5xFAD mice. Scale bar = 1 mm. J, Regional cerebral Aß deposition in Tg-
SwDI mice (black bars), Tg-5xFAD mice (blue bars) and bigenic Tg-SwDI/Tg-5xFAD mice (red bars)
was determined by image analysis of immunostaining. The data presented are the mean ± S.D. of
eight mice of each genotype at each age.
Fig. 3. Progressive accumulation of fibrillar amyloid deposition in Tg-SwDI, Tg-5xFAD and bigenic
Tg-SwDI/Tg-5xFAD mouse brain. Three to nine months old transgenic mouse brain sections were
stained for fibrillar amyloid using thioflavin S (green) as described in Methods. A-C, Tg-SwDI mice; D-
F, bigenic Tg-SwDI/Tg-5xFAD mice; and G-I, Tg5x-FAD mice. Scale bar = 1 mm. J, Regional fibrillar
amyloid deposition in Tg-SwDI mice (black bars), Tg-5xFAD mice (blue bars) and bigenic Tg-SwDI/Tg-
5xFAD mice (red bars) was determined by image analysis of thioflavin S staining. The data presented are
the mean ± S.D. of eight mice of each genotype at each age.
Fig. 4. Altered fibrillar amyloid deposition in bigenic Tg-SwDI/Tg-5xFAD mouse brain. Three to
nine months old transgenic mouse brain sections were stained for fibrillar amyloid using thioflavin S
(green) and immunolabeled for cerebral blood vessels using an antibody to collagen IV (red) as described
in Methods. A-C, Tg-SwDI mice; D-F, bigenic Tg-SwDI/Tg5xFAD mice; and G-I, Tg-5xFAD mice.
Scale bar = 50 µm. J, the amount of cerebral microvascular and K, the size distribution of fibrillar
amyloid deposits in Tg-SwDI mice (black bars), Tg-5xFAD mice (blue bars) and bigenic Tg-SwDI/Tg-
5xFAD mice (red bars) was determined in the subiculum region of each transgenic mouse line at nine
months of age using stereological principles as described in Methods. The data presented are the mean ±
S.D. of eight mice per group. **p < 0.01; ***p < 0.0001.
Fig. 5. Increased neuronal loss and microglial activation in bigenic Tg-SwDI/Tg-5xFAD mouse
brain. Six months old transgenic mouse brain sections were immunolabeled for neurons using an
antibody to NeuN, for Aß using polyclonal antibody to human Aß, for activated microglia using mAb5D4
and stained for fibrillar amyloid using thioflavin S. A-D, representative images of neuronal staining in
cortical layers II-VI. Scale bars = 100 µm. E-H, representative images of neuronal staining in cortical
layer V. Scale bars = 50 µm. I-L, representative images of immunostaining in cortical layer V. Scale
bars = 50 µm. M-P, representative images of thioflavin S staining for fibrillar amyloid (green) and
immunolabeling for activated microglia (red). Scale bars = 50 µm. Q, the numbers of neurons in cortical
layer V in wild-type mice (gray bars), Tg-SwDI mice (black bars), Tg-5xFAD mice (blue bars) and
bigenic Tg-SwDI/Tg-5xFAD mice (red bars) were determined using stereological principles as described
in Methods. The data presented are the mean ± S.D. of seven mice per group. **p < 0.01; ***p < 0.0001.
16
Fig. 6. Analysis of wild-type and Dutch/Iowa CAA mutant accumulation in Tg-SwDI, Tg-
5xFAD and bigenic Tg-SwDI/Tg-5xFAD mouse brain. A, Dot blot analysis was performed to
demonstrate that rabbit polyclonal antibody to human (pAb-Aß) recognizes both wild-type and
Dutch/Iowa CAA mutant whereas mAb4G8 only recognizes wild-type . Brain sections obtained
from the different transgenic mouse lines at nine months of age were immunolabeled with pAb-Aß in red
(B,E,H), mAb4G8 in green (C,F,I) and merged (D,G,J). Dutch/Iowa CAA mutant Aß deposits in Tg-
SwDI mice are immunolabeled with pAb- (B), but not mAb4G8 (C). Wild-type Aß deposits are
immunolabeled with both pAb-Aß (E) and mAb4G8 (F). plaque core deposits in bigenic Tg-
SwDI/Tg-5xFAD mice are immunolabeled with both pAb-Aß (H) and mAb4G8 (I). Merged image shows
a halo of Aß deposits around the periphery of plaques that are not immunolabeled with mAb4G8
suggesting that they are composed of Dutch/Iowa CAA mutant Aß (J). Scale bars = 50 µm.
Fig. 7. Wild-type 42 amyloid fibril seeds promote the assembly of Dutch/Iowa CAA mutant
Aß40 fibrils. Freshly prepared solutions of Dutch/Iowa CAA mutant Aß40 (20 µM) were incubated in
the absence (----) or presence of 10 µM (----) or 20 µM (----) of wild-type Aß42 fibrillar seeds. The rate
and extent of fibril formation was assessed by thioflavin T fluorescence measurements. The fluorescence
curves of the wild-type Aß42 fibril seeds with added Dutch/Iowa CAA mutant Aß40 exhibit two
components. The rapid component is attributed to the growth of CAA mutant Aß40 fibrils on the wild-
type Aß42 seeds. The slower component is attributed to fibril formation of CAA mutant Aß40 following
nucleation of the monomeric peptide. Wild-type Aß42 fibril seeds in the absence of added monomeric
CAA mutant peptide do not exhibit a change in fluorescence (----). The data shown are the mean ±
S.D. of triplicate determinations.
Fig. 8. Wild-type parenchymal amyloid in Tg-5xFAD mice promotes the accumulation of injected
biotin-labeled Dutch/Iowa CAA mutant Aß. Biotin-labeled Dutch/Iowa CAA mutant Aß40 (A-D) and
biotin (E-H) were injected into the hippocampal region of six months old Tg-5xFAD mice. Brain sections
were prepared, fibrillar amyloid was visualized by staining with thioflavin S (green) and biotin-labeled
Dutch/Iowa CAA mutant Aß40 or biotin alone was detected using Texas red-labeled streptavidin (red).
A-C and E-G, scale bars = 50 µm. D,H, scale bars = 12.5 µ m.
17
FIGURE 1
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FIGURE 2
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FIGURE 3
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FIGURE 4
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FIGURE 5
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FIGURE 6
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FIGURE 7
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FIGURE 8
    • "In vivo 2-photon microscopy showed progressive CAA in 5xFAD mice, characterized by perivascular amyloid deposition, possibly corresponding to the tunica media/adventitia, on large caliber vessels. The latter was previously unreported (Xu et al., 2014) potentially due to the limitations associated with conventional immunohistochemistry as compared to longitudinal 2-photon preparations. Our results are relevant as AD patients frequently show CAA (Kovari et al., 2015) (Thal et al., 2008). "
    [Show abstract] [Hide abstract] ABSTRACT: Clinical and experimental evidence point to a possible role of cerebrovascular dysfunction in Alzheimer's disease (AD). The 5xFAD mouse model of AD expresses human amyloid precursor protein and presenilin genes with mutations found in AD patients. It remains unknown whether amyloid deposition driven by these mutations is associated with cerebrovascular changes. 5xFAD and wild type mice (2 to 12months old; M2 to M12) were used. Thinned skull in vivo 2-photon microscopy was used to determine Aβ accumulation on leptomeningeal or superficial cortical vessels over time. Parenchymal microvascular damage was assessed using FITC-microangiography. Collagen-IV and CD31 were used to stain basal lamina and endothelial cells. Methoxy-XO4, Thioflavin-S or 6E10 were used to visualize Aβ accumulation in living mice or in fixed brain tissues. Positioning of reactive IBA1 microglia and GFAP astrocytes at the vasculature was rendered using confocal microscopy. Platelet-derived growth factor receptor beta (PDGFRβ) staining was used to visualize perivascular pericytes. In vivo 2-photon microscopy revealed Methoxy-XO4(+) amyloid perivascular deposits on leptomeningeal and penetrating cortical vessels in 5xFAD mice, typical of cerebral amyloid angiopathy (CAA). Amyloid deposits were visible in vivo at M3 and aggravated over time. Progressive microvascular damage was concomitant to parenchymal Aβ plaque accumulation in 5xFAD mice. Microvascular inflammation in 5xFAD mice presented with sporadic FITC-albumin leakages at M4 becoming more prevalent at M9 and M12. 3D colocalization showed inflammatory IBA1(+) microglia proximal to microvascular FITC-albumin leaks. The number of perivascular PDGFRβ(+) pericytes was significantly decreased at M4 in the fronto-parietal cortices, with a trend decrease observed in the other structures. At M9-M12, PDGFRβ(+) pericytes displayed hypertrophic perivascular ramifications contiguous to reactive microglia. Cerebral amyloid angiopathy and microvascular inflammation occur in 5xFAD mice concomitantly to parenchymal plaque deposition. The prospect of cerebrovascular pharmacology in AD is discussed.
    Full-text · Article · Jan 2016
    • "Failure in the clearance of Aβ peptides may therefore result in inflammation and neurotoxicity (Giri et al., 2000; Li et al., 2009) and CAA is likely to play a key part in brain damage and loss of cognitive skills typical of AD (Lee et al., 2014). In accordance to this hypothesis, human studies and animal models of AD document that cerebrovascular dysfunction precedes the development of cognitive decline and AD pathology (de la Torre, 2004b; Bell and Zlokovic, 2009; Xu et al., 2014). Several biochemical alterations of endothelial cell physiology have been identified in response to exposure to amyloidogenic peptides (Figure 2). "
    [Show abstract] [Hide abstract] ABSTRACT: Alzheimer’s disease (AD) is the most common neurodegenerative cause of dementia in the elderly. AD is accompanied by the accumulation of amyloid peptides in the brain parenchyma and in the cerebral vessels. The sporadic form of AD accounts for about 95% of all cases. It is characterized by a late onset, typically after the age of 65, with a complex and still poorly understood aetiology. Several observations point towards a central role of cerebrovascular dysfunction in the onset of sporadic AD (SAD). According to the “vascular hypothesis”, AD may be initiated by vascular dysfunctions that precede and promote the neurodegenerative process. In accordance to this, AD patients show increased hemorrhagic or ischemic stroke risks. It is now clear that multiple bidirectional connections exist between AD and cerebrovascular disease, and in this new scenario, the effect of amyloid peptides on vascular cells and blood platelets appear to be central to AD. In this review, we analyze the effect of amyloid peptides on vascular function and platelet activation and its contribution to the cerebrovascular pathology associated with AD and the progression of this disease.
    Full-text · Article · Mar 2015
  • Full-text · Article · Jan 2016
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