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Abstracts e155
Investigación Biomédica Sant Pau (IIB Sant Pau), Barcelona, SPAIN,
6
Epithe-
lial Systems Biology Laboratory, National Heart Lung and Blood Institute (NIH),
Bethesda, USA,
7
Laboratori de Proteòmica CSIC/UAB, Instituto de Investigacio-
nes Biomédicas de Barcelona (IIBB-CSIC-IDIBAPS), Barcelona, SPAIN,
8
Anax-
omics Biotech SL, Barcelona, SPAIN
Objective: Diabetic nephropathy (DN) is the major cause of end-stage renal dis-
ease. Renin-angiotensin system (RAS) inhibition is the preferred treatment to
slow its progression. We have studied the urinary proteomes of patients with DN
(high albuminuria) to investigate the pathophysiology of renal disease and iden-
tify disease markers and predictors of clinical outcome.
Design and method: We included diabetic men with (n = 9) and without DN (n
= 12) (control cohort). Data collection included clinical and laboratory evalua-
tion of blood and urine at baseline (control cohort and DN-basal), and in patients
with DN after 3 months of losartan treatment (DN-treated). Urinary proteome was
analyzed and quantified by Tandem Mass Tag (TMT) labeling on a LTQ-Orbitrap
mass spectrometer.
Results: Patients enrolled in the study showed no differences regarding basic
clinical parameters. Urinary proteome analysis have identified 166 differentially
excreted proteins when comparing the proteomes of controls and DN patients,
27 comparing DN-treated and DN-basal patients, and 182 among patients DN-
treated and controls. Systems biology approach comprising functional proteomic
networks and artificial neural networks (TPMS technology) have identified 80 key
proteins involved in the pathophysiology of DN and 15 key proteins involved in
the efficacy of losartan. There are 7 proteins identified in the urine proteome that
are essential in both DN pathophysiology and treatment efficacy. Vascular cell
adhesion molecule-1 (VCAM-1) and the angiotensin-metabolizing neutral endo-
peptidase neprilysin (NEP) stand out from the other identified proteins because
they are the only ones that are DN effectors. They are differentially expressed in
the urinary proteome and are also key proteins in both DN pathophysiology and
RAS inhibition efficacy.
Conclusions: NEP is a membrane-bound zinc-containing metalloproteinase showing
great abundance in the brush border of proximal renal tubular cells. NEP is responsible
for the processing and catabolism of several vasoactive peptides including angiotensin
II and endothelin which may explain its pathogenic role in the development of DN.
PP.07.07 CIRCULATING MICROVESICLES (C-MVS) LEVELS
AND ENDOTHELIAL FUNCTION IN PATIENTS WITH
MULTIFOCAL FIBROMUSCULAR DYSPLASIA (FMD):
A CROSS-SECTIONAL STUDY
X. Loyer
1
, H. Kettab
2
, A. Lorthioir
2
, M. Frank
2
, R. Niarra
2
, J.M. Renard
1
,
Y. Chambon
2
, X. Jeunemaitre
1
, P.F. Plouin
2
, L. Amar
2
, P. Boutouyrie
2
,
C.M. Boulanger
1
, M. Azizi
2
.
1
INSERM U970, Paris Cardiovascular Research
Center, PARCC, Paris, FRANCE,
2
APHP, Clinical Investigation Center,
Departments of Genetics, Pharmacology and Hypertension Unit, HEGP,
Paris, FRANCE
Objective: FMD is an idiopathic, segmental, non-atherosclerotic non-inflam-
matory arterial disease of unknown origin which occurs mostly in middle-aged
women and affects medium-sized arteries (renal and carotid arteries in particu-
lar). The objective of the study was to identify new hemodynamic and biological
biomarkers of the pathology. We investigated i) the flow-mediated dilation (FD)
and endothelium-independent dilation (ED) of the brachial artery (BA); ii) c-MVs
from different vascular cell origins.
Design and method: We conducted a cross sectional study with 50 patients with
multifocal FMD, 50 essential hypertensive (EH) patients matched for age, sex,
ethnicity and BP and 50 healthy subjects (HS) matched for age, sex and ethnic-
ity. Exclusion criteria were: tobacco consumption, hypercholesterolemia, diabe-
tes, aspirin or statin treatment. We measured blind to the phenotype: 1) changes
in BA diameter after release of hand ischemia (FD) and glyceryl trinitrate (ED)
by high-resolution radiofrequency-based echotracking system; and 2) endothelial
and smooth-cell derived MVs plasma concentrations by flow cytometry analysis
of human platelet free plasma.
Results: FMD, EH and HS were well matched. FMD and EH had signifi-
cantly higher SBP than HS despite antihypertensive treatments. Circulat-
ing levels of total MVs (annexinV+MVs), endothelial MVs (CD144+MVs,
CD62E+MVs and CD31+CD41-MVs), CD11a+MVs and smooth muscle
derived MVs (SMA (smooth muscle actin)+ MVs) displayed large between-
subject variability within each group and did not significantly differ between
groups. FD or ED changes in BA diameter did not significantly differ be-
tween groups.
Conclusions: In conclusion, we could not identify specific changes in c-MVs
levels of endothelial or smooth muscle origin in patients with FMD when com-
pared with age-, sex-, and ethnicity-matched patients with EH or HS. This result is
consistent with the similar acute vasodilatory responses to flow and glyceryl trini-
trate observed in FMD patients compared to EH and HS. Taken all together, these
results demonstrate that endothelial function is not affected in patients with FMD.
PP.07.08 METABOLOMIC STUDY OF PATHWAYS
UNDERLYING HEXADECANEDIOATE INDUCED
BLOOD PRESSURE ELEVATION
N. Alharbi, M.W. McBride, D. Graham, S. Padmanabhan. Institute of Cardiovas-
cular and Medical Sciences, College of Medical, Veterinary and Life Sciences,
University of Glasgow, Glasgow, UNITED KINGDOM
Objective: A putative novel pathway for blood pressure regulation involving the
dicarboxylic acid hexadecanedioate was identified in a study associating blood
pressure and mortality outcomes with fasting blood metabolites. The functional
role of hexadecanedioate on blood pressure elevation and vascular reactivity was
confirmed by oral administration of hexadecanedioic acid to Wistar Kyoto (WKY)
rats. This study aimed to characterize the metabolic effects of hexadecanedioic
acid administration in WKY rats to identify metabolic pathways underlying hexa-
decanedioate-induced blood pressure elevation.
Design and method: Eleven-week-old male WKY were treated orally with hexa-
decanedioic acid (250 mg/kg per day; n = 5) or vehicle (n = 5) for 3 weeks. At
sacrifice tissues (aorta, heart, brain, adipose, kidney, and liver) were harvested
and global metabolic profiles analysed by UPLC-MS/MS (Metabolon). Data was
analysed using random forest classification and the top rank metabolites were
highlighted for further investigation.
Results: Treatment with hexadecanedioic acid increased hexadecanedioate levels in
all tissues tested except the brain, potentially reflecting poor diffusion through the
blood-brain barrier. Random Forest (RF) classification of metabolites gave a predic-
tive accuracy of 80% for heart and 100% for kidney. The RF Importance Plot of
the 30 top-ranking metabolites indicates key differences in lipid, carbohydrate and
amino acid metabolism. In heart, increased fatty acids (e.g. acylcarnitines, complex
lipids, and lysolipids) in treated rats suggest impairment of fatty acid beta-oxidation.
Heart also demonstrated elevated levels of glycogen breakdown products indicat-
ing increased glucose demand. In contrast, kidneys demonstrated an increase in
long-chain fatty acids and ketone body beta-hydroxybutyric acid, and a decrease in
maloylcarnitine suggesting increased beta-oxidative use. In adipose tissue dicarbox-
ylate fatty acids (tetradecanedioate and octadecanedioate) were elevated suggesting
hexadecandioic acid caused a decrease in beta-oxidation and a shift towards use
of peroxisomal omega-oxidation. In liver, hexadecanedioic acid increased cysteine-
glutathione disulfide levels suggesting an enhanced oxidising environment.
Conclusions: Exogenous administration of hexadecanedioic acid in addition
to increasing blood pressure impacts a number of metabolic readouts including
changes related to lipid and glucose metabolism and redox homeostasis. These
pathways are putative targets for further research to elucidate the mechanism by
which hexadecanedioate affects blood pressure.
PP.07.09 GENETIC RISK SCORE AND CARDIOVASCULAR
COMPLICATIONS IN PATIENTS WITH
HYPERTENSION
M. Wirtwein
1
, O. Melander
2
, M. Sjogren
2
, M. Hoffmann
3
, K. Narkiewicz
3
,
M. Gruchala
4
, W. Sobiczewski.
1
Medical University of Gdansk, Department of
Pharmacology, Gdansk, POLAND,
2
Lund University, Department of Clinical