disorders, as well as Traumatic Brain Injury (TBI). TBI induces
cognitive changes affecting millions of people, as well as increased
incidence of age-related neurodegeneration. Evidence from our lab
and others suggests tau that forms on route to NFT formation—
oligomers—are the most toxic tau species. We have shown increased
levels of tau oligomers in neurodegenerative disease brains, as well as
in TBI models. Using immunoprecipitation, we isolated tau oligo-
mers from controlled cortical impact injured mice, fluid percussion
injured and blast injured rat brains. Oligomers were characterized
biochemically and by atomic force microscopy and were injected
bilaterally in the hippocampi of mice overexpressing human tau
(Htau mice). Mice were cognitively evaluated using novel object
recognition and Y-maze tasks and brains were collected following
testing. We found that tau oligomers form as a result of brain injury
in three different rodent models of TBI and accelerated the onset of
cognitive deficits, absent increased levels of cell death, in Htau mice.
As we have seen previously in other diseases, oligomers collected
from TBI can seed the aggregation of tau monomer
in vitro.
Moreover, mice injected with oligomers exhibited elevated levels of
oligomeric tau in multiple brain regions, supporting the prion-like
seeding and propagation capability of TBI-derived oligomeric tau.
Our results suggest that tau oligomers play an important role in the
toxicity underlying TBI, making them a viable therapeutic target for
TBI and in preventing the increased acquisition of neurodegenera-
tive disease later in life. These studies were completed as part of an
interdisciplinary research team funded by The Moody Project for
Translational Traumatic Brain Injury Research.
Keywords: TBI, Tau, Protein aggregation, Tau oligomers, Tau
pathology
B2-09
ACUTE REGION OF INTEREST CHANGES IN KEY BRAIN
INJURYMARKERS FOLLOWINGPENETRATINGBALLISTIC-
LIKE BRAIN INJURY
Casandra Cartagena
, Ying Deng-Bryant, Hye Hwang, Frank
Tortella, Angela Boutte
WRAIR, BTNN, Silver Spring, USA
Penetrating brain injury (PBI) is associated with high mortality and
morbidity. PBI involves accelerated infiltration of peripheral immune
cells and increased edema. However, molecular mechanisms under-
lying PBI are not widely studied. Here we evaluated changes in key
markers of PBI across regions of interest (ROIs) including frontal
cortex (FCx), striatum (St), hippocampus (Hc), and residual midbrain
(RMb), using the model of penetrating ballistic-like brain injury
(PBBI). Injury was induced in anesthetized rats by inserting a probe
through the right FCx and St followed by rapid balloon inflation
causing a temporary cavity. Key injury markers were evaluated 24h
post-PBBI to determine molecular effects in ROIs proximal to the
injury tract (FCx, St) versus surrounding areas (Hc, rMb). Targeted
proteins were glial fibrillary acidic protein (GFAP), GFAP breakdown
products (GFAP-BDPs), spectrin, spectrin breakdown products
(SBDPs), B-cell lymphoma-2 (BCL2), BCL2-associated protein X
(BAX), amyloid associated protein (APP), APP
a
- and
b
- C-terminal
fragments (APP
a
CTF, APP
b
CTF), and
tau
protein. PBBI ROIs were
compared to comparable Sham ROIs (n
=
10/group). GFAP levels
increased 60% in FCx (p
<
0.05) and 53% in St (P
<
0.05). GFAP-
BDPs levels increased 262% in FCx (p
<
0.05) and 1342% in St
(p
<
0.01). Spectrinlevels decreased 75% in FCx (p
<
0.01) and 53% in
St (p
<
0.05). SBDPs levels increased 251% in FCx (p
<
0.01), 431% in
St (p
<
0.01) and 156% in RMb (p
<
0.01). BCL2 levels decreased 63%
in FCx (p
<
0.0001). BAX levels increased 1764% in FCx
(p
<
0.0001), 222% in Hc (p
<
0.05), 5869% in St (p
<
0.0001), and
387% in RMb (p
<
0.01). No significant changes were detected in APP
or APP
a
CTF levels. However, APP
b
CTF levels increased 330% in
FCx (p
<
0.001) and 1089% in St (p
<
0.001). Total
tau
levels de-
creased 30% (p
<
0.01) in FCx. These results indicate that acute (24 h)
alterations in astrogliosis and markers of neurodegeneration are lim-
ited to ROIs proximal to the injury tract. However, structural protein
abnormalities and pro-apoptotic mechanisms expand into surrounding
ROIs. Ongoing studies will evaluate these markers at subacute and
chronic time-points post-PBBI.
Keywords: Amyloid, Tau, GFAP, Spectrin, apoptosis
B2-10
BRAIN CATHEPSIN B IS ELEVATED IN BOTH MILD-
CLOSED AND SEVERE-PENETRATING TRAUMATIC
BRAIN INJURY MODELS
Angela Boutte
1
, Brittany Abbatiello
1
, Shonnette Grant
1
, Gregory
Hook
2
, Vivian Hook
3
, Frank Tortella
1
, Deborah Shear
1
1
Walter Reed Army Institute of Research, Brain Trauma Neuropro-
tection and Neurorestoration Branch, Silver Spring, USA
2
American Life Science Pharmaceuticals, Inc., Research and Devel-
opment, San Diego, USA
3
University of California, San Diego, Skaggs School of Pharmacy and
Pharmaceutical Sciences, Dept. of Neurosciences, La Jolla, USA
Comprehensive analysis of key mediators involved in traumatic brain
injury (TBI) is tantamount to understanding mechanisms involved in
injury progression. Cathepsin B is a cysteine protease implicated in
several neurodegeneration and TBI models, such as controlled cortical
impact. This preliminary study determined if brain cathepsin B was
up-regulated in penetrating ballistic-like brain injury (PBBI) or re-
peated projectile concussive impact (rPCI). For PBBI and sham/
craniotomy controls, coronal brain tissue sections were isolated at
various time-points post-injury. Repeated (r)PCI was conducted once
daily for 4 consecutive days (d). Control groups received anesthesia
alone. Righting-reflex (RR) was determined immediately after injury.
Select PCI brain tissue regions were collected 1d after the last con-
cussion. Both pro-(
*
37–43 kDa) and mature (
*
20–25 kDa) Cathe-
psin B protein levels were determined by western blotting and
densitometry (mean
+
/
-
SEM arbitrary units (AU)). Enzymatic ac-
tivity was determined by generation of amino-methyl coumarin
(AMC). Comparisons between injured and control groups are dis-
cussed (2-tailed, t-Test, p
£
0.05); correlative analysis is indicated
(1-way, Pearson
r
). Pro-cathepsin B upregulation in brain slices was
monophasic and peaked 2–3 d after PBBI (13.3
+
/
-
1.2 and
15.2
+
/
-
2.3 AU) compared to Sham (1.2
+
/
-
0.1 to 3.1
+
/
-
1.3
AU). Interestingly, mature cathepsin B was maximally increased 7d
after PBBI (384.1
+
/
-
39.7 AU), versus Sham (174.2
+
/
-
25.4 AU).
In rPCI, pro-cathepsin B was increased to (1.7
+
/
-
0.5 AU) in the
prefrontal cortex (not detectable in sham/anesthesia controls). In this
brain region, mature cathepsin B was
*
7-fold greater after rPCI
(14.2
+
/
-
3.6 AU) compared to controls (2.2
+
/
-
1.5 AU); proteo-
lytic activity was marginally increased. Surprisingly, cerebellar pro-
teolytic activity increased nearly 3-fold after rPCI (3.0
+
/
-
0.6
l
moles) compared to anesthesia alone (1.4
+
/
-
0.2
l
moles), and
positively associated with RR (
r
= +
0.65, p
=
0.12). Conversely, de-
creased activity in this region was negatively correlated with RR
(
r
= -
0.98, p
=
0.008) among anesthesia controls. These findings
suggest that brain cathepsin B has a role in multiple TBI models and is
linked to neurological deficits.
Keywords: Cathepsin B, Penetrating Ballistic-like Brain Injury,
Projectile Concussive Impact
A-49