Surgery for Aortic Disease
Differential Protein Kinase C Isoform Abundance in
Ascending Aortic Aneurysms From Patients With Bicuspid
Versus Tricuspid Aortic Valves
Jeffrey A. Jones, PhD; Robert E. Stroud, MS; Brooke S. Kaplan, BS; Allyson M. Leone, BS;
Joseph E. Bavaria, MD; Joseph H. Gorman, III, MD;
Robert C. Gorman, MD; John S. Ikonomidis, MD, PhD
Downloaded from http://circ.ahajournals.org/ by guest on June 18, 2017
Background—It is recognized that different events contribute to the initiation of ascending thoracic aortic aneurysms
(ATAAs) in patients with bicuspid aortic valves (BAV) versus patients with tricuspid aortic valves (TAV), but the
molecular signaling pathways driving aneurysm formation remain unclear. Protein kinase C (PKC) is a superfamily of
kinases which differentially mediate signaling events that lead to altered gene expression and cellular function, and may
regulate downstream mediators of vascular remodeling. The present study tested the hypothesis that ATAA development
in patients with BAV versus TAV proceeds by independent signaling pathways involving differential PKC signaling.
Methods and Results—ATAA samples were collected from BAV (n⫽57) and TAV (n⫽55) patients and assessed for 10
different PKC isoforms by immunoblotting. Results were expressed as a percent change in abundance (mean⫾SEM)
from a nonaneurysmal control group (100%, n⫽21). Correlation analysis was performed, and relationships between
PKC and matrix metalloproteinase abundance were reported. In the BAV group, classic and novel PKC isoforms
(PKC-␣, ␤⌱, ␥, ␧, ␪) were increased, whereas PKC-␩ and atypical PKC-␨ were decreased. In the TAV group, classic
and novel isoforms were decreased and atypical PKC-␨ was elevated. Positive correlations between PKC and matrix
metalloproteinase abundance were identified.
Conclusions—Differential PKC isoform abundance was observed in ATAA samples from patients with BAV versus TAV,
suggesting independent molecular signaling pathways may be operative. Induction of independent transcriptional
programs may result and may provide a mechanistic foundation for developing selective diagnostic/therapeutic
strategies for patients with ATAAs secondary to BAV or TAV. (Circulation. 2007;116[suppl I]:I-144–I-149.)
Key Words: valves 䡲 aneurysm 䡲 aorta 䡲 PKC 䡲 metalloproteinases
T
he congenital bicuspid aortic valve (BAV) is the most
common cardiac malformation, occurring in 1% to 2% of
the population.1–3 Approximately 33% of patients with a
BAV will develop serious complications that require treatment. A common consequence of BAV is aortic dilatation
leading to the formation of ascending thoracic aortic aneurysms (ATAAs).2,4,5 There have been 2 predominant theories
put forth to explain the increased incidence of ATAA in
patients with BAV: (1) genetic abnormalities which may
include defects in the neural crest-origin cells and alterations
in fibrillin-1 function, and (2) enhanced hemodynamic stress
on the ascending aortic wall as a result of turbulent blood
flow over the malformed valve. ATAA formation is a
multifactorial process that involves both cellular and extracellular mechanisms that converge on multiple signaling
pathways and result in the maladaptive remodeling of the
vascular extracellular matrix. Studies from this laboratory and
others have demonstrated differential matrix metalloproteinase (MMP) profiles in ATAA samples from patients with a
BAV versus patients with idiopathic medial degenerative
disease and a tricuspid aortic valve (TAV).6 –10 These differential profiles suggest that aneurysm formation in each valve
group may proceed by unique mechanisms that require
discrete signaling pathways and different intermediate effectors. However, although there are several studies comparing
the dysregulation of MMP expression secondary to BAV
versus TAV, little information is known regarding the upstream signaling events that orchestrate these changes in each
valve group.
Protein kinase C (PKC) is a family of lipid regulated
serine/threonine kinases that mediate cellular responses and
cellular function.11–13 There are at least 10 different isoenzymes, encoded by 9 different genes, classified into 3 groups
that are differentiated by their activation requirements and
From the Cardiothoracic Surgical Research (J.A.J., R.E.S., B.S.K., A.M.L., J.S.I.), Division of Cardiothoracic Surgery, Medical University of South
Carolina, Charleston; and Division of Cardiothoracic Surgery (J.E.B., J.H.G., R.C.G.), University of Pennsylvania, Philadelphia.
Presented at the American Heart Association Scientific Sessions, Chicago, IL November 12–15, 2006.
Correspondence to John S. Ikonomidis, MD, PhD, Associate Professor of Surgery, Division of Cardiothoracic Surgery, Department of Surgery, Medical
University of South Carolina, 96 Jonathan Lucas St, Suite 409 CSB, Charleston, SC 29425. E-mail [email protected]
© 2007 American Heart Association, Inc.
Circulation is available at http://circ.ahajournals.org
DOI: 10.1161/CIRCULATIONAHA.106.681361
I-144
Jones et al
PKC Signaling in BAV vs TAV Aortic Aneurysms
I-145
Patient Demographics
Reference Control
BAV
TAV
Count
21
57
55
Gender, %male⫾SEM
76⫾4
75⫾7
62⫾6
Age, y⫾SEM
48⫾3
57⫾2*
66⫾2*†
Aortic diameter, cm⫾SEM
2.8⫾0.1
5.3⫾0.1*
5.4⫾0.2*
Aortic insufficiency/regurgitation, %
4.8
47.9
45.8
Aortic stenosis, %
0.0
25.0
2.1
42.9
56.3
54.2
Hypertension, %
*P⬍0.05 vs reference control; †P⬍0.05 vs BAV
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downstream targets. The classic isoforms (cPKC; ␣, ␤⌱, ␤⌱⌱,
and ␥) require both diacylglycerol and calcium for activation,
whereas the novel isoforms (nPKC; ␦, ␧, ␩, and ␪) require
only diacylglycerol for activation and are calcium independent. The atypical isoforms (aPKC; ␫ and ␨) are both calcium
and diacylglycerol-independent but are regulated by other
glycero- and sphingo- lipids.11
Because the activation of various PKC isoforms has been
implicated in the regulation of downstream effectors of
vascular remodeling,14 –19 we hypothesized that independent
signaling pathways involving differential PKC signaling contribute to the unique etiologies of ATAA in patients with
BAV versus TAV.
Methods
Study Population
The study population consisted of aortic tissue samples collected
from ATAA patients at time of surgical resection that had either a
BAV (n⫽57), or a TAV (n⫽55). All patients underwent aortic root
replacement, and all tissue specimens were collected from the
ascending aorta as opposed to the sinus segment or proximal arch.
Results were compared with nonaneurysmal aortic samples collected
from the ascending aorta of 21 reference control patients (8 heart
donors, 12 heart transplant recipients, and 1 coronary artery bypass
graft patient). Thus, in all cases, reference control and aneurysmal
tissue specimens were collected from the same region of the
ascending aorta for biochemical comparison. There were no gender
differences between groups (control: 76⫾4% male, BAV: 75⫾7%
male, TAV: 62⫾6% male; P⫽0.231 ␹2); however, the mean ages
between groups were different (control: 48⫾3 years; BAV: 57⫾2
years, P⫽0.002 from control; and TAV 66⫾2 years, P⬍0.001 from
control, and P⫽0.002 from BAV). Aortic diameters were not
different between ATAA subgroups, but both were different from the
reference control group (control: 2.8⫾0.1 cm; BAV: 5.3⫾0.1 cm,
P⬍0.001; TAV: 5.4⫾0.2 cm, P⬍0.001). Significant proportions of
the BAV (47.9%) and TAV (45.8%) patients had documented aortic
insufficiency or regurgitation, and 25% of the BAV patients had
documented aortic stenosis that was moderate to critical. Approximately half of the patients in all 3 groups had hypertension (Control
42.9%, BAV 56.3%, and TAV 54.2%). All patient demographics are
summarized in the Table. No patient experienced giant cell arteritis,
had an aortic dissection, or had an established connective tissue
disorder. This study was approved by the Institutional Review
Boards of both the Medical University of South Carolina and the
University of Pennsylvania, and informed consent was obtained from
all patients.
Sample Preparation
Resected aortic tissue specimens were homogenized in cold acidic
extraction buffer to prevent protease activation during the extraction
process.20 The aortic homogenates were then centrifuged (4°C, 10
minutes, 1200g) and the final protein concentration was determined
(BCA Protein Assay).
Immunoblotting
The relative abundance of 10 different PKC isoforms were determined using quantitative immunoblotting techniques.20 Briefly, 10
␮g of each aortic homogenate was fractionated on a 4% to 12%
Bis-Tris gradient polyacrylamide gel (Invitrogen Corp). The proteins
were then transferred to nitrocellulose membrane (0.45 ␮m; BioRad) and blocked with 5% nonfat dry milk in phosphate buffered
saline (PBS) for 1 hour at room temperature. The membrane was
incubated with antiserum (0.4 ␮g/mL in 5% nonfat dry milk/PBS)
containing specific antibodies for each of the PKC isoforms tested
(␣⫽alpha, ␤⌱⫽beta-⌱, ␤⌱⌱⫽beta⌱⌱, ␥⫽gamma, ␦⫽delta, ␧⫽epsilon,
␪⫽theta, ␩⫽eta, ␫⫽iota, and ␨⫽zeta; Santa Cruz Biotechnology).
Following incubation with primary antibody, the membrane was
washed extensively (3⫻10 minutes, 25 mL PBS) to reduce nonspecific antibody interactions. A secondary peroxidase-conjugated antibody (species dependent on the primary antibody used) was applied
(1:5000 dilution in 5% nonfat dry milk/PBS) and allowed to incubate
for 1 hour at room temperature. The membrane was again washed
extensively (4⫻15 minutes, 25 mL PBS). Positive immunoreactivity
was identified by briefly incubating the membranes with a chemiluminescent substrate (Western Lighting Chemiluminescence Reagent
Plus; Perkin Elmer) and exposing the blot to film (Hyperfilm;
GE Healthcare).
Data Analysis
Immunoblots were digitized and quantitative densitometric image
analysis was performed (Gel Pro Analyzer; Media Cybernetics).
PKC abundance in ATAA specimens was expressed as a percentage
increase or decrease as compared with reference control values,
which were set at 100%.
All statistical procedures were carried out using the Stata statistical package (Intercooled Stata v8.2; StataCorp LP). Patient demographics were compared by ␹2 analysis or 1-way ANOVA using
Tukey wsd post hoc analysis. One-sample mean comparison tests
were performed to determine statistical differences in PKC abundance. Pairwise correlation analysis was performed to reveal relationships between PKC abundance and MMP/tissue inhibitor of
metalloproteinase (TIMP) expression (previously reported8) or ascending aortic diameter. Significant correlations were reported with
regression values and probability values. Statistical analyses were
also performed to compare PKC isoform abundance to age, presence
of hypertension, aortic valve pathology (insufficiency/regurgitation
versus stenosis), and aortic diameter indexed to body surface area.
All data are presented as a mean⫾SEM, and values of P⬍0.05 were
considered statistically significant.
The authors had full access to the data and take responsibility for
its integrity. All authors have read and agree to the manuscript as
written.
Results
Analysis of PKC Abundance
The aortic specimens were assayed by immunoblotting for 10
different PKC isoforms that were classified into 3 different
I-146
Circulation
September 11, 2007
(54)
βΙ
(55)
βΙΙΙΙ
(54)
#
(52)
*
(52)
#
(53)
γ
(53)
δ
(54)
ε
(55)
θ
(54)
η
(52)
ι
(56)
ζ
(55)
(51)
TAV
*
#
α
βΙ
βΙΙΙΙ γ
δ ε θ η
ι ζ
Novel
Atypical
BAV
TAV
Classical
*
*
*
Figure 2. Summary of the significant changes in PKC isoform
abundance in ATAA specimens from BAV versus TAV patients.
(52)
*
(52)
(53)
Downloaded from http://circ.ahajournals.org/ by guest on June 18, 2017
0
*
#*
50
100
150
200
Percent Change from Reference Control
Figure 1. Comparison of classic, novel, and atypical PKC isoform abundance in BAV and TAV aneurysm samples (BAV [dark
gray], TAV [light gray]; representing the mean⫾SEM). A representative immunoblot of each PKC isoform is shown on the
right. The number of observations are shown in parentheses on
each bar; *P⬍0.05 versus reference control, #P⬍0.05 versus
BAV.
activation groups (Figure 1). Of the classic PKC isoforms
(presented as integrated optical density ratio, percent change
from reference control, probability value) PKC-␣ (479.1/
293.1, 163.5⫾19.3%, P⫽0.002), - ␤ ⌱ (230.0/165.9,
138.7⫾16.2%, P⫽0.020), and - ␥ (269.1/165.9,
162.2⫾21.7%, P⫽0.006) were elevated in the BAV specimens, whereas PKC-␤⌱⌱ remained unchanged. In the TAV
samples, PKC-␤⌱⌱ (617.9/947.9, 65.2⫾8.1%, P⬍0.001) and
-␥ (80.6/165.9, 48.6⫾7.7%, P⬍0.001) were decreased from
control values, whereas PKC-␣ and -␤⌱ remained unchanged.
Of the novel PKC isoforms PKC-␧ (168.3/105.1,
160.0⫾24.9%, P⫽0.020) and -␪ (308.8/194.7, 158.6⫾20.3%,
P⫽0.006) were elevated in the BAV specimens, whereas
PKC-␩ (3973.9/5397.8, 73.6⫾9.9%, P⫽0.010) was decreased and PKC-␦ remained unchanged. In the TAV specimens, PKC-␧ (80.0/105.1, 76.1⫾11.5%, P⫽0.043) and -␩
(2766.0/5397.8, 51.2⫾8.1%, P⬍0.001) were decreased from
control values and PKC-␦ and -␪ remained unchanged. Of the
atypical PKC isoforms, PKC-␨ (968.0/1180.1, 82.0⫾5.2%,
P⫽0.001) was decreased in the BAV specimens, whereas
PKC-␫ remained unchanged. In the TAV specimens, PKC-␨
(1443.7/1180.1, 122.3⫾7.0%, P⫽0.002) was elevated from
control values, whereas PKC-␫ remained unchanged. The
abundance of the following PKC isoforms was significantly
different between BAV and TAV specimens: PKC-␣
(P⫽0.034), -␤⌱⌱ (P⫽0.001), -␥ (P⬍0.001), -␧ (P⫽0.006), -␪
(P⫽0.011), and -␨ (P⬍0.001). These results are summarized
in Figure 2.
Pairwise Correlation Analysis
To examine the relationships between PKC abundance and
potential downstream regulators of vascular remodeling,
A
350
MMP-2 Abundance
*
pairwise correlation analysis was performed. The percent
change from reference control of each PKC isoform was
compared with the abundance of each MMP and TIMP
species, as previously reported, for each patient sample.8
Significant correlations were identified between PKC-␪
(170⫾23%, P⬍0.05 versus reference control) and MMP-2
(138⫾7%, P⬍0.05 versus reference control) in the BAV
samples (r⫽0.4995, P⫽0.0002; Figure 3A), and PKC-␨
(122⫾7%, P⬍0.05 versus reference control) and MMP-7
(300⫾82%, P⬍0.05 versus reference control) in the TAV
samples (r⫽0.4076, P⫽0.0025; Figure 3B). No significant
correlations were identified between PKC isoform abundance
and ascending aortic diameter in either BAV or TAV patients.
Additional analyses examining the affect of age, aortic valve
pathology (aortic insufficiency/regurgitation, stenosis), and
aortic size indexed to body surface area on PKC isoform
abundance failed to reveal any significant relationships.
Comparisons of PKC isoform abundance in ATAA specimens from BAV and TAV patients with and without hypertension, revealed the PKC-␣ (P⫽0.007), -␧ (P⫽0.014), -␪
(percent of reference control)
#
(53)
(51)
*
*
*
300
250
200
150
100
50
(r=0.4995, p=0.0002)
0
0
200
400
600
800
PKC-θ
θ Abundance
(percent of reference control)
B
1000
(r=0.4076, p=0.0025)
MMP-7 Abundance
#
(52)
(percent of reference control)
Atypical
Novel
Classical
CON BAV
α
800
600
400
200
0
0
50
100
150
200
250
300
PKC-ζ
ζ Abundance
(percent of reference control)
Figure 3. Correlation analysis between PKC and MMP abundance. A, PKC-␪ versus MMP-2 in BAV samples; B, PKC-␨ versus MMP-7 in TAV samples. The r value and P value for the
correlation are shown. Gray lines indicate the 95% CI.
Jones et al
PKC Signaling in BAV vs TAV Aortic Aneurysms
(P⫽0.018), and -␫ (P⫽0.034) were elevated in ATAA specimens from BAV patients with hypertension as compared
with those without.
Discussion
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Although it is recognized that different natural histories and
pathophysiological events contribute to the initiation of
ATAA formation in patients with a BAV versus patients with
a TAV, the molecular signaling pathways that drive aneurysm
formation in these two valve groups remain unclear. Because
PKC has been directly implicated in regulating downstream
effectors of extracellular matrix remodeling in other disease
states, the present study measured the abundance of 10
different PKC isoenzymes in ATAA samples resected from
BAV and TAV patients. When compared with nonaneurysmal aortic specimens collected from a reference control
group, the unique findings of this study demonstrate that
differential PKC isoenzyme expression profiles exist in
ATAAs from BAV versus TAV patients (Figures 1 and 2). In
the BAV samples, increased abundance of the diacylglycerolsensitive classic and novel PKC isoforms was observed along
with decreased novel PKC-␩ and atypical PKC-␨. Alternatively, in the TAV samples, the classic and novel isoforms
were decreased from control, whereas atypical PKC-␨ was
increased. Moreover, when pairwise comparisons were performed between the abundance of PKC isoenzyme species
and the MMPs and their endogenous inhibitors (as previously
reported8), significant correlations were identified (Figure 3).
Together, these data suggest that independent transcriptional
programs mediated by differential PKC signaling may account for the disparate proteolytic processes driving ATAA
formation in BAV versus TAV patients. Accordingly, this
differential expression of PKC isoforms may hold future
diagnostic and prognostic implications.
PKC functions as a critical signaling mediator, transducing
intracellular and extracellular signals, by regulating kinase
cascades that ultimately influence a diverse number of cellular mechanisms including mitogenic and apoptotic pathways,
vesicular transport, lipid metabolism, and gene expression.11,21–23 Importantly, PKC has been shown to be operative
in pathways regulating the expression of MMPs.15 The MMPs
are a family of at least 27 distinct zinc-dependent proteases
capable of degrading all extracellular matrix constituents, and
thus have been established as critical mediators of extracellular matrix remodeling.24 –30 Recently, this laboratory has
reported that the MMPs and their endogenous inhibitors are
differentially expressed in aneurysmal tissue from patients
with BAV versus TAV.8 Because regulation of MMP expression has been suggested to occur primarily at the transcriptional level,15 the differential MMP profiles identified in these
valve groups likely arose from the activation of different
transcriptional programs during aneurysm development. The
present study results suggest that PKC signaling, affected
through differential isoenzyme expression, may contribute to
the disparate transcriptional programs regulating the distinct
MMP expression profiles demonstrated in each ATAA valve
group.
Previous studies have implicated PKC as an upstream
mediator of MMP production and release. Park and col-
I-147
leagues demonstrated in glioblastoma cells that phorbol ester
treatment could induce MMP-9 secretion, MMP-2 activation,
and the translocation of MT1-MMP to the plasma membrane.18 These observations were shown to be dependent on
PKC-mediated p38MAP kinase activation, and were accompanied by the down regulation of TIMP-1 and TIMP-2.17
PKC-mediated MMP production was also confirmed by Chu
and coworkers who demonstrated that inflammatory cytokines could induce the production and secretion of MMP-2
and MMP-9 by a PKC-dependent mechanism in osteoarthritis
cell cultures.16 Furthermore, MMP release from neutrophils
was shown to be dependent on PKC activity by Chakrabarti
et al, who demonstrated that 85% of MMP-9 specific granule
release could be blocked by a pan-specific PKC inhibitor.14
Together these data strongly support the hypothesis that the
expression and release of MMPs is at least in part dependent
on a common fundamental signaling pathway involving PKC.
In the present study, the increased abundance of the
diacylglycerol-sensitive classic and novel PKC isoforms -␣,
-␤⌱, -␥, -␧, and -␪ in BAV samples suggests that aneurysm
formation may be driven by a process that results in the
sustained activation of upstream mediators of diacylglycerol
production; such as phospholipase C (PLC). PLC is a family
of lipid metabolic enzymes that function to remove the
phospo-head group of several specific phospholipid species,
resulting in the formation of diacylglycerol. Schütze and
coworkers demonstrated that tumor necrosis factor-␣ could
stimulate diacylglycerol levels in U937 cells by activating a
phosphatidylcholine-specific PLC. The increase in diacylglycerol was linked to the potent activation of PKC, as PKC
activation could be inhibited with a specific PLC inhibitor.31
Although there is a paucity of information regarding the
active signaling pathways in aortic aneurysm formation,
previous studies of intracranial aneurysms have identified
increased PLC activity in both experimental models and
patient samples, and associated the increase in PLC activity
with increased PKC activation.32–34 Thus, the persistent
activation of lipid metabolic enzymes such as PLC could
contribute to aneurysm formation through the activation of
the diacylglycerol-sensitive PKC isoforms and the subsequent induction of PKC-mediated MMP expression.
With regard to the TAV samples, a general decrease was
observed in the diacylglycerol-sensitive isoforms concomitant
with an increase in atypical PKC-␨. PKC-␨, although insensitive
to calcium and diacylglycerol, requires specific interactions with
phospholipids or sphingolipids for activation.35 Among these
lipids, the generation of phosphotidylinositol-3,4,5-trisphosphate
by phosphatidylinositol-3 kinase has been identified as a potent
regulator of PKC-␨ activation.36 Interestingly, PKC-␨ has been
shown to be a potent activator of nuclear factor ␬-B, a transcription factor with target binding sites in several MMP promoter
regions. Esteve et al demonstrated that interleukin-1 and tumor
necrosis factor-␣ could induce MMP-9 expression by an nuclear
factor ␬-B– dependent mechanism that required the activation of
PKC-␨.37 This work confirmed the observations of Hussain and
coworkers who demonstrated that PKC-␨ activity was essential
for cytokine-dependent induction of MMP-1, -3, and -9 through
a mechanism that involved the activation of nuclear factor ␬-B.38
Thus, activation of atypical PKC in ATAA samples from TAV
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September 11, 2007
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patients may also result in the production of MMPs through the
stimulation of a potentially distinct transcriptional program.
In an effort to establish a relationship between the MMP/
TIMP profiles and PKC isoenzyme expression in BAV versus
TAV aneurysms, pairwise correlation analysis was performed
comparing MMP or TIMP abundance with PKC isoform
abundance. Several statistically significant interactions were
uncovered. Of greatest interest was a strong positive relationship (r⫽0.4995, P⫽0.0002) between MMP-2 abundance and
PKC-␪ in the BAV samples. Both MMP-2 and PKC-␪ were
elevated as compared with reference control values. Previous
studies from several investigators have described elevated
MMP-2 in valvular and aneurysmal aortic tissues from BAV
patients, representing the most consistent finding in BAV
specimens across laboratories.6 –10 Interestingly, Xie and coworkers, through a series of in vitro studies in adult rat
cardiac fibroblasts, have demonstrated that activation of
PKC-␪ plays a critical role in interleukin-1␤–mediated induction of MMP-2 expression and activity.39 Thus, these data
support the present study findings and may suggest that
PKC-␪ activation is required for MMP-2 induction during
ATAA formation in patients with BAV.
In the TAV samples, MMP-7 and PKC-␨ were both
elevated compared with reference control values and likewise
displayed a positive relationship (r⫽0.4076, P⫽0.0025) by
correlation analysis. Although no literature precedence has
been established directly linking PKC-␨ activation with
MMP-7 expression, indirect evidence suggests they may be
linked. Activator protein-1 (AP-1) is a heterodimeric leucine
zipper transcription factor comprised of the Fos and Jun
family of transcriptional regulators. MMP-7, as well as
several other MMPs, have AP-1 consensus binding sites
within their promoter regions.40 In a study by Ways et al,
overexpression of PKC-␨ in U937 cells induced differentiation which was characterized by increased levels of the c-jun
proto-oncogene and increased AP-1 DNA binding activity;
implicating PKC-␨ as an upstream activator of AP-1.41 More
recently, lipopolysaccharide induced activation of AP-1 was
likewise shown to require PKC-␨ activation.42 Thus, PKC-␨
could potentially activate AP-1-mediated transcription of
MMP-7 during ATAA formation in patients with TAV.
Because this study was performed on resected aortic
specimens collected at the time of surgical intervention, there
are some inherent limitations to this data. First, it is unclear
whether the differential PKC profiles identified in each valve
group are representative of specific phases of aneurysm
progression or summary of all phases. Furthermore, other
confounding factors such as patient exposure to cyclo-oxygenase inhibitors, steroids, or statins may also affect aneurysm formation and progression. Until noninvasive methods
for following specific targets over time are identified, this
limitation cannot be addressed in human patients. This study,
however, is well suited for testing in an animal model of
thoracic aortic aneurysm in which temporal PKC profiles can
be determined and experimentally modified. Second, care
was taken to be sure that all tissue specimens were harvested
from the same region of the ascending aorta, as regional
aortic heterogeneity may exist such that PKC isoform expression may be different in the descending aorta or the aortic
arch. Furthermore, regional expression differences may exist
within each specific aortic segment. Hence, additional studies
will be required to determine the topographic expression
differences of the PKC isoforms, and how those expression
differences affect aneurysm formation. Third, altered PKC
abundance in these specimens does not necessarily imply that
PKC isoform activity has been altered. A more direct measure
of specific activity will be required to further establish the
role of PKC activation in aneurysm formation. Last, it is also
important to note that potential upstream activators of PKC
(diacylglycerol, phospholipase C, phosphatidylinositol-3 kinase) were not measured in this study. Identification of the
operative signaling intermediates that function upstream of
PKC in each valve group will need to be addressed in future
experimentation.
Nevertheless, the results of the present study imply that
differential PKC signaling may be operative during aneurysm
formation in ATAA samples from BAV versus TAV patients.
This differential signaling may result in the activation of
distinct transcriptional programs that could explain the differential proteolytic profiles previously described in these
ATAA subgroups. Furthermore, the altered abundance of the
different classes of PKC isoforms in BAV (classic and novel)
versus TAV (atypical) suggest that the upstream stimuli are
different and add to the data further describing the divergent
operational pathobiology in ATAA formation secondary to
BAV versus TAV. Continued definition of the roles played
by upstream signaling pathways in these pathological conditions may eventually allow identification of the critical
stimuli which initiate aneurysm formation with obvious
therapeutic implications.
Sources of Funding
This work was supported by NIH/NHLBI R01 HL075488-04.
Disclosures
None.
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Differential Protein Kinase C Isoform Abundance in Ascending Aortic Aneurysms From
Patients With Bicuspid Versus Tricuspid Aortic Valves
Jeffrey A. Jones, Robert E. Stroud, Brooke S. Kaplan, Allyson M. Leone, Joseph E. Bavaria,
Joseph H. Gorman III, Robert C. Gorman and John S. Ikonomidis
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Circulation. 2007;116:I-144-I-149
doi: 10.1161/CIRCULATIONAHA.106.681361
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