bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 465: Evolution of Cardiovascular Lifestyle Behaviours Occurring in Hypercholesterolemic Patients Concomitantly to the Intake of Phytosterol-supplemented Yoghurt.
Authors: Bruckert Eric , Masama L , Descamp O. , Bosi E. , Allaert Francois
Published: March 2022

Objective:
to assess the evolution of cardiovascular lifestyle behaviours in hypercholesterolemic patients occurring concomitantly with changes in their daily intake of phytosterol-supplemented yoghurt (Phyto-SY).

METHODS:
Nationwide prospective observational study conducted in Spanish and French general practice Each GP proposed lifestyle modification to 5 consecutive patients presenting with hypercholesterolemia, (taking or not hypocholesterolemic drugs) and recommended daily consumption of Phyto-SY. Study design included an inclusion visit, a patient’s self-monitoring assessment after 1 month and a final end-visit after four months. Main evaluation criterion : nutritional lifestyle score. Secondary criteria: Total, LDL and HDL cholesterol, waist circumference, daily walking time.

Results:
2376 hypercholesterolemic patients, 56.2 years old ± 11.9 (women 54.8%) were included. Nutritional lifestyle score decreased from 15.4 ± 5.4 to 8.7± 4.0 (p<0,0001), either in never-treated patients (15.3 ± 5.4 to 8.6 ± 3.9; p<0,0001) or in patients treated with hypocholesterolemic drugs (15.5 ± 5.4 to 8.9 ± 4.1; p<0,0001). Total cholesterol decreased 10.6% (<0.0001), HDL-C increased 8.0% (<0.0001), and LDL-C decreased 12.7% (<0.0001). Frequency of walking (>30 min) increased from 59.3% to 78.3% (p<0.0001); by contrast overweight frequency decreased from 22.8% to 17.5% (p<0.0001) and waist circumference from 94.6 ± 13.3 cm to 93.0 ± 12.8 (p<0.0001).

Conclusion:
Improvement in nutritional lifestyle score, involving regular consumption of Phyto-SY over 4 months was significantly linked to a healthier lifestyle and to beneficial modification of atherogenic lipid profile, thereby reflecting patient empowerment in a “real life” context.
Abstract 47: Angiotensin Receptor Agonistic Autoantibody Mediated Vascular Impairment via sFlt-1 Contributes to Decreased Aldosterone Production in Preeclampsia
Authors: Siddiqui Athar H , Irani Roxanna , Zhang Weiru , Wang Wei , Blackwell Sean C , Ramin Susan M , Kellems Rodney E , Xia Yang
Published: March 2022

Preeclampsia (PE) is a multi-system disorder of pregnancy characterized by high blood pressure and proteinuria. Despite high blood pressure, a decreased circulating level of aldosterone is associated with PE. The molecular mechanisms underlying reduced aldosterone production in PE are unknown. In the present study we demonstrate for the first time that reduced aldosterone levels in the sera of preeclamptic women could be associated with the presence of angiotensin II type 1 receptor agonistic autoantibody (AT1-AA) and elevated soluble Fms-like tyrosine kinase-1 (sFlt-1), two prominent factors related to the pathophysiology of PE. Using a well established adoptive transfer animal model of PE, we provide in vivo evidence that the injection of IgG from women with PE, but not IgG from normotensive pregnant women (NT-IgG), resulted in hypertension, proteinuria and a reduction in aldosterone production. The aldosterone level decreased from 1377±272 pg/ml to 544±92 pg/ml (p<0.05) in pregnant mice. These features were prevented by co-injection with an epitope peptide that blocks the antibody-mediated AT1 receptor (AT1R) activation. In contrast, injection of IgG from preeclamptic women into non-pregnant mice resulted in an increase in the adolsterone levels from 213±24 pg/ml to 615±48 pg/ml (p<0.05). These results indicate that maternal circulating autoantibody in preeclamptic women is a detrimental factor for decreased aldosterone production via AT1R activation in a pregnancy-dependent manner. Next, we found that circulating sFlt-1 was increased more in the autoantibody-injected pregnant mice compared to the NT-IgG injected mice (56.1±4.6 vs 21.87±6.4 ng/ml, p<0.05). Furthermore upon infusion of VEGF121, we saw a recovery in the circulating levels of aldosterone (from 544±92 to 1110±269 pg/ml, p<0.05). Collectively, these results suggest that AT1-AA-induced sFlt-1 elevation is a novel pathogenic mechanism that promotes vascular impairment in adrenal glands leading to decreased aldosterone production in PE. Overall, our findings reveal novel factors and signaling cascades involved in decreased aldosterone production that could reveal possible therapeutic interventions in the timely management of PE.
Abstract 472: Effect of Acetylcholinesterase Inhibition with Pyridostigmine on Hemodynamics, Autonomic and Cardiac Function in Heart Failure C57Bl/6J Mice.
Authors: Durand Marina T , Oliveira Mauro , Prado Cibele M , Rossi Marcos A , Fazan Rubens , Salgado Helio C
Published: March 2022

We investigated the effects of pyridostigmine (PYR) on hemodynamics, autonomic and cardiac function in heart failure (HF) mice. Telemetry transmitters (DSI) were implanted into the carotid artery under isofluorane anesthesia. After recovery, basal arterial pressure (AP, mmHg) and heart rate (HR, bpm) were recorded and parasympathetic and sympathetic tone (Δbpm) was evaluated by means of methyl atropine and propranolol. After the basal recording, the mice were submitted to left coronary ligation (LCL) and mini pumps implantation (PYR: 3mg/kg/day). Control mice were implanted with empty mini pumps. One and four weeks after LCL, the recordings were repeated. LCL promoted a myocardial infarction of 10-20%. HF mice presented decreased systolic AP (SAP) (113±2 vs 120±2 basal) and increased HR (548±12 vs 489±20 basal) only in the 1st week (n=8). In addition, LCL decreased vagal tone in the 1st week (121±14 vs 177±9 basal), increased sympathetic tone in the 1st and 4th week (-41±6 and -49±10 vs -23±2 basal). PYR prevented the increase of HR (505±10 vs 495±7 basal) but did not affect the decrease of the SAP (113±3 vs 123±1 basal) in the 1st week (n=3). PYR prevented the increase in the sympathetic tone (-23±15 and -23±6 vs -17±10 basal), but did not affect the parasympathetic tone (218±30 vs 199±37 basal) throughout the study. Cardiac function was assessed in separated mice (n=23) under isofluorane anesthesia. HF mice exhibited a decreased ejection fraction (28±4 vs 45±1% sham LCL) and increased diastolic and systolic volume (31±2 and 24±3 vs 18±2 and 12±1μl sham LCL). PYR increased the ejection fraction (44±4%) and attenuated the increase of both volumes (24±4 and 14±3μl). In conclusion, PYR prevented the tachycardia, decreased sympathetic tone and protected the cardiac function during the development (4 weeks) of HF in mice.
Abstract 48: Hypoxia-induced Heparanse Regulates sFlt-1 Release from Placental Chorionic Villi
Authors: George Eric M
Published: March 2022

Preeclampsia is one of the most pervasive and serious complications of pregnancy in the US and worldwide, which currently has no effective pharmacological intervention. While the underlying cause of the disorder remains unclear, it is generally accepted that placental hypoperfusion leads to placental ischemia. In response pathogenic factors are produced by the placenta and secreted into the maternal circulation, leading to the maternal symptomatic phase of the disorder. One of the factors which has received a great deal of attention is the soluble VEGF receptor/antagonist sFlt-1, which is sufficient to produce maternal hypertension and is believed to be a major contributor to maternal endothelial dysfunction. The factors which regulate sFlt-1 secretion from the placenta are unclear, however recent evidence has shown that in normal pregnancy, localization of sFlt-1 (i.e. retained in the placenta vs. secretion into the circulation) is controlled by binding to extracellular heparan in the placenta. Here we have examined the effect of placental ischemia on the production and activity of heparanase, which can cleave extracellular heparan and release the locally retained sFlt-1. In response to placental ischemia, placental heparanase expression is increased ∼50% (1±0.03 vs 1.49±0.07 normalized expression, p<0.05) a result which is supported by in vitro experiments demonstrating a ∼2-fold increase (1±0.07 vs 2.02±0.3 normalized units, p<0.05) in secreted heparanase by chorionic villus explants cultured under hypoxic conditions. Continuous infusion of heparin, which can mimic heparanase by displacing sFlt-1 bound to extracellular heparan, displaces sFlt-1 from the placenta (913±33 vs 762±48 pg/ml, p<0.05), decreasing the bioavailable VEGF in the maternal circulation (1174±62 vs 1174±38 pg/ml, p<0.05). This is accompanied by a significant (10mmHg, p<0.05) increase in mean arterial pressure. Together, these results suggest an important role for heparan binding and heparanase expression in the control of sFlt-1 secretion from the ischemic placenta, and suggest a potential new therapeutic approach for the management of hypertension in preeclampsia.
Abstract 480: Dopamine d1 Receptor Requires Both gQ/11? and gI for Downstream Signaling in Human Kidney 2 Cells
Authors: Pokkunuri Indira , Asghar Mohammad
Published: March 2022

Renal dopamine D1 receptor (D1R) plays pivotal role in maintaining sodium homeostasis and blood pressure (BP). It does so by either coupling to G
s
or G
q
proteins and engaging downstream signaling molecules. Renal proximal tubular cells from human kidney, HK-2 cells, are important
ex vivo
tool to study mechanistic aspect of D1R signaling cascade involved in BP regulation. Recently, however, it is reported that D1R is uncoupled from G
s
protein suggesting attenuation of the receptor’s downstream signaling in HK-2 cells.
In this study we wanted to determine whether D1R is coupled to Gq proteins in HK-2 cells
. To test this, we measured the activity of a downstream surrogate marker of G
q
stimulation, protein kinase C (PKC), by undertaking following experimental manipulations and comparing them with a known activator of G
q
, angiotensin II (Ang II). Fenoldopam (FD, 1μM, 10 min), a D1R agonist, increased PKC activity to the levels of a direct activator (PMA, a phorbol ester, 1μM, 15 min) of PKC activity (in ng/μl: Control vs FD vs PMA: 0.31 + 0.04 vs 1.08 + 0.05 vs 1.02 + 0.07). FD-mediated PKC stimulation was attenuated in the presence of D1R antagonist (
SCH23390
, 10μM), PKC inhibitor (chelerythrin chloride, 1μM), G
q
depletion (siRNA, 200 nM) and pertussis toxin (PTX, an inhibitor of G
i
protein, 100 ng/ml). Similarly, Ang II (1μM, 10 min) increased PKC activity (in ng/μl: Control vs Ang II: 0.31 + 0.04 vs 1.05 + 0.15) which was attenuated in the presence of Ang II AT1 receptor blocker (candesartan, 1μM), chelerythrin chloride, G
q
siRNA and PTX. Furthermore, FD increased
35
S-GTPγS binding, an index of D1R function (cpm/μg protein: Control vs FD: 414 + 104 vs 1188 + 43), and abundance of G
q
(Density units: Control vs FD: 30 + 8 vs 47 + 12) in the cellular membranes. Our results suggest that (i) D1R is functional; (ii) not only D1R but also AT1R downstream signaling is intact, which requires (iii) both G
q
and G
i
G-proteins in HK-2 cells.
Abstract 487: AT1a Receptor-Mediated Uptake of Extracellular Angiotensin II and NHE3 Expression in Mouse Proximal Tubule Cells partly Involve the Multi-ligand Endocytic Receptor Megalin
Authors: Li Xiao C , Hopfer Ulrich , Zhuo Jia L
Published: March 2022

Megalin, the multi-ligand endocytic receptor abundantly expressed in apical (AP) membranes of proximal tubules, plays a crucial role in mediating the uptake of low molecular weight (LMW) proteins in the kidney. Deletion of megalin leads to the development of LMW proteinuria in mice. In the present study, we tested the hypothesis that megalin in AP membranes mediates the uptake (or endocytosis) of angiotensin II (ANG II) in mouse proximal tubule cells (mPCT) by interacting with AT
1a
receptors. Semi-confluent, polarized monolayers of mPCT cells of wild-type (WT) and AT
1a
-deficient (AT
1a
-KO) mice grown on transwell permeable supports or 6-well plates were treated from the AP surface with vehicle, losartan (10 μM), PD123319 (10 μM), a selective megalin-siRNA or a scrambled siRNA for 48 h. The time-dependent uptake of fluorescein (FITC)-labeled ANG II (10 nM, 37
o
C) was then determined by fluorescence imaging. As expected, AT
1a
receptor and megalin proteins were abundantly expressed in AP membranes of WT mPCT cells, whereas AT
1a
receptors were absent in AT
1a
-KO mPCT cells (
p
<0.01 vs. WT cells). In WT mPCT cells, the uptake of FITC-ANG II was peaked at 30 min and visualized in the nuclei by 1 h. These responses were blocked by losartan (
p
<0.01), whereas PD123319 had no effect. Furthermore, AT
1a
receptor-mediated FITC-ANG II uptake was largely blocked in AT
1a
-KO mPCT cells (
p
<0.01 vs. WT cells). In both WT and AT
1a
-KO mPCT cells, the specific megalin-siRNA knocked down 86.3±5.2% of megalin protein expression (
p
<0.01), but it inhibited only 32.8±3.1% of FlTC-ANG II uptake in WT mPCT cells (
p
<0.01 vs.losartan). Megalin-siRNA had no effect on FITC-ANG II uptake in AT
1a
-KO mPCT cells. The AT
1a
receptor-mediated uptake of FITC-ANG II was associated with 3-fold increases in phosphorylated MAP kinases ERK1/2 proteins (control: 0.36±0.06 vs. ANG II: 1.07±0.10,
p
<0.001) and 1-fold increase in p-NHE3 proteins (control: 0.18±0.03 vs. ANG II: 0.37±0.04,
p
<0.01) in WT mPCT cells, which was blocked by losartan (0.17±0.07,
p
<0.01 vs. ANG II) and megalin-siRNA (0.19±0.02
p
<0.01 vs. ANG II), respectively. These results suggest that AT
1a
receptor-mediated uptake of extracellular Ang II and NHE3 expression in mPCT cells may partly involve the multi-ligand endocytic receptor megalin.
Abstract 492: Experimental Hyperleptinemia Induces Changes on Renal Function and Morphology in Rats
Authors: Thieme Karina , Oliveira-Souza Maria
Published: March 2022

Aim:
To evaluate the effect of experimental hyperleptinemia for 7 days and the participation of the Renin-Angiotensin II System (RAS) on renal function and morphology.

Methods:
40-days male Wistar rats were divided into 4 groups (n = 5 - 6 per group): sham (
Sh
), Leptin (
Lep
; s.c. infusion 0.5mg/kg/day, osmotic pump, Alzet), Losartan (
Los
; s.c. 10 mg/kg/day), Leptin plus Losartan (
LL
). Systolic blood pressure (SBP) was measured by tail-cuff method. On 8
th
day, rats were anesthetized and prepared to renal clearance experiments. Renal plasmatic flow (RPF) and glomerular filtration rate (GFR) were determined by p-Aminohippuric acid sodium and inulin clearance. Urinary NH
4
+
, pH, Na
+
, K
+
and osmolality were analyzed. Renal tissue was submitted to histological studies.

Results:
7-days Leptin treatment increased SBP (
Sh
: 105,8 ± 0,7
vs
Lep
: 118,0 ± 1,5 mmHg; *p<0.0001) and
LL
group reversed this parameter (
LL
: 103,4 ± 0,7 mmHg; *p<0.0001). Our preliminary results showed that Leptin and/or Losartan treatment does not affect renal hemodynamic (RPF -
Sh
: 20,48 ± 0,77
vs
Lep
: 15,79 ± 1,02
vs
Los
: 17,96 ± 2,21
vs
LL
: 18,99 ± 1,54 ml/min/kg and GFR -
Sh
: 7,94 ± 0,46
vs
Lep
: 9,19 ± 0,59
vs
Los
: 7,46 ± 0,60
vs
LL
: 6,97 ± 0,52 ml/min/kg). However, filtration fraction increased significantly in
Lep
group (
Sh
: 40,40 ± 1,72
vs
Lep
: 59,79 ± 2,46 %*; *p < 0.0001) and was corrected by Losartan (
LL
34,42 ± 3,06 %
#
;
#
p< 0.0001). Urinary flux increased in
Lep
rats and was also corrected by Losartan (
Sh
: 0,18 ± 0,01
vs
Lep
: 0,25 ± 0,02; *p<0.001
vs
LL
: 0,18 ± 0,008
#
;
#
p< 0.0001 ml/min/kg). Na
+
excretion, which had an 337% increase in
Lep
group, was normalized in
LL
. Urinary excretion of NH
4
+
and K
+
and urinary osmolality was not different between the studied groups. All parameters of
Los
group were not statistically different from
Sh
rats.
Lep
rats showed an increase in total glomerular area, which was also corrected in
LL
rats (
Sh
: 6708 ± 62
vs
Lep
: 7796 ± 130
vs
Los
: 6779 ± 23
vs
LL
6822 ± 20 μm
2
).

Discussion:
Lep
rats showed higher SBP, but not hypertension. Although there were no significant changes in renal hemodynamic, Leptin treatment caused glomerular hypertrophy. As many parameters were corrected by co-treatment with Losartan, it seems that RAS has a critical role in hyperleptinemic induced renal changes.
Abstract 5: Angiotensin II-induced Alterations of T Cell Receptor Vβ Chain Usage in Kidney and Mesenteric Vasculature
Authors: Trott Daniel W , Harrison David G
Published: March 2022

We have previously shown that T cells play a critical role in the genesis of hypertension, and that activated T cells infiltrate the vasculature and the kidney to promote vasoconstriction and sodium retention. Stimulation of the T cell receptor (TCR) with a specific antigen leads to clonal expansion of cells with identical TCRs. In contrast, if multiple antigens are presented, T cells with multiple TCRs proliferate. We sought to determine if hypertension promotes a mono- or polyclonal T cell response. TCR Vβ chain gene transcript length can be used to examine T cell clonality. We produced hypertension in C57Bl/6 mice by chronic (2 week, 490 ng/kg/min) infusion of angiotensin II and used vehicle infused mice as controls. PCR was employed to amplify the 24 TCR Vβ chain subfamilies in the kidney and mesenteric vasculature. PCR products were subjected to high-resolution capillary electrophoresis fragment analysis to determine TCR Vβ chain transcript length. Eleven of the 24 Vβ length profiles exhibited greater than 20% skewing with angiotensin II infusion in kidney and 6 of 24 exhibited greater than 20% skewing in mesentery. Dominant transcript lengths were present with angiotensin II infusion for Vβ 1 (30 ± 5 vs. 51 ± 7), 3 (29 ± 3 vs. 51 ± 12) and 20 (0 ± 0 vs. 41 ± 18) in kidney and 1 (30 ± 2 vs. 43 ± 4), 4 (27 ± 4 vs. 45 ± 3) and 20 (15 ± 3 vs. 46 ± 6) in mesentery (data are expressed as % ± S.E.M. of TCR transcripts of a given length within each Vβ family, sham vs. angiotensin II, n = 5 per group). These results suggest that hypertension results in oligoclonal T cell expansion in the kidney and mesentery. Alterations in Vβ 1 and 20 in both kidney and mesentery suggest a common TCR response between tissues.
Abstract 50: Characterization of the Gwas-nominated Agtrap-plod1 Locus Using Zfn-mutated Rats
Authors: Flister Michael J , Tsaih Shirng-Wern , Endres Bradley , Sarkis Allison B , Geurts Aron M , Lazar Jozef , Moreno Carol , Dwinell Melinda R , Jacob Howard J
Published: March 2022

Background:
Genome-wide association studies (GWAS) are frequently used to nominate candidate genes for complex diseases, but are largely unable to identify the causative variant(s). One example: the AGTRAP-PLOD1 locus contains 6 genes in close proximity that are all associated with blood pressure (BP), making it difficult to delineate specific allele(s) that underlie human hypertension at this locus. Here we present a novel rat model that was developed to rapidly dissect GWAS loci using high-throughput gene mutation on a single hypertensive background.

METHODS:
Agtrap, Mthfr, Clcn6, Nppa, Nppb, and Plod1 were individually mutated in the Dahl SS rat by zinc finger nuclease (ZFN) injections. ZFN-mutated and wild type (WT) control SS rats (n=8-25 per group) were assessed for BP and renal damage after 10 days on a 4% NaCl diet. Temporal gene expression during development of hypertension was assessed by qRT-PCR and confirmed by RNA-seq.

Results:
Expression of all AGTRAP-PLOD1 transcripts changed significantly in response to elevated BP, indicating possible roles of the entire AGTRAP-PLOD1 locus in salt-sensitive hypertension. However, compared with WT, only NPPA mutation further increased mean BP (Δ+27mmHg, P<0.001) and heart weight (1.62±0.03 vs. 1.13±0.03g, P<0.001) in response to 4% NaCl diet. In contrast, CLCN6 mutation significantly decreased diastolic BP (Δ-22mmHg, P<0.001) but had no effect on systolic BP compared to WT. Following salt challenge, proteinuria was elevated by mutation of PLOD1 (147±12 mg/day, P<0.001) and MTHFR (132±23 mg/day, P<0.05) compared with WT littermates (91±5 mg/day), whereas AGTRAP mutation decreased protein excretion (54±9 mg/day, P<0.05). Analysis of haplotype blocks that are associated with gene expression and human hypertension confirmed that NPPA, CLCN6, and MTHFR are the primary mediators of BP changes linked to the AGTRAP-PLOD1 locus.

Conclusions:
Combined with human GWAS, our data show for the first time that NPPA and CLCN6 are divergent mediators of BP at the AGTRAP-PLOD1 locus, while MTHFR mutation directly increases susceptibility to end-stage renal disease. These novel mechanistic data provide rationale for developing haplotype-specific therapies for treating hypertension.
Abstract 502: Salt-Excess Induced Exacerbated Hypertension Prevention by Telmisartan in Spontaneously Hypertensive Rats
Authors: Susic Dinko , Fares Hassan , Frohlich Edward D
Published: March 2022

The effects of angiotensin receptor blocker (AGA), diuretic, a calcium antagonist (CA), and their combination were evaluated on the progression of cardiovascular damage in spontaneously hypertensive rats (SHR) given salt-excess. To this end, 8-week male SHR rats were divided into 7 groups. The control group (C) received regular NaCl (0.6%) diet. All other groups were given 8% NaCl rat chow. In addition, Group 2 (HS) was given placebo, Group 3 (T) the AGA telmisartan (10 mg/kg/day), Group 4 (D) received the chlorothiazide (80 mg/kg/day), Group 5 (T+D) telmisartan plus diuretic, Group 6 (A) was given the CA amlodipine (10 mg/kg/day), and Group 7 (T+A) telmisartan plus amlodipine. All treatments lasted 8 weeks. Compared with controls, mean arterial pressure (MAP), renal blood flow, coronary flow reserve (CFR), minimal coronary vascular resistance, diastolic time constant, and maximal rate of ventricular pressure fall (-dP/dT) were all adversely affected by salt-loading (Table). Increased left ventricular weight (LVWI) with marked cardiac fibrosis was also found with salt overload. Telmisartan normalized all indices except MAP, whereas diuretic and amlodipine only partially restored cardiac function and mass. Combination therapy with telmisartan and either diuretic or amlodipine also normalized all indices including arterial pressure (Table). These data show that: (1) cardiovascular and renal damage induced by salt-excess in SHR were not pressure dependent; and (2) compared with the CA and diuretic, AGA was more effective in this model suggesting that local renin in target organs participated in salt-induced hypertension.
Abstract 511: Nitric Oxide Attenuates Matrix Metalloproteinase-9 Production by Endothelial Cells
Authors: Meschiari Cesar A , Izidoro-Toledo Tatiane C , Gerlach Raquel F , Tanus-Santos José
Published: March 2022

Impaired nitric oxide (NO) bioavailability and imbalanced matrix metalloproteinase (MMP) activity are key pathogenetic mechanisms involved in cardiovascular diseases. However, there is little evidence supporting a direct link between these mechanisms, although NO is known to interfere with nuclear factor kappa B (NFB) activity, an important modulator of MMP-9 expression. Moreover, it is not known whether the possible effects of NO on MMPs is dependent on cyclic GMP formation.

Objective:
We examined the effect of NO donors on MMP-9 production by endothelial cells, and if this effect is dependent on cGMP formation or NFB activation.

Methods:
Human umbilical vein endothelial cells were cultured in appropriate medium and treated for 24 hours with 10 nM phorbol myristate acetate (PMA; a MMP-9 inducer) and other drugs: NO donors (S-nitroso-N-acetylpenicillamine; SNAP, or DetaNONOate), 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; a soluble guanylyl cyclase inhibitor), BAY 11-7082 (a NFB inhibitor), or their vehicles. Conditioned media were analyzed by gelatin zimography.

Results:
PMA increased MMP-9 activity from 0.10±0.08 arbitrary units (AU) to 0.94±0.18 AU (
P<
0.05). Treatment with SNAP (200 or 320 μM) attenuated the increases in MMP-9 activity induced by PMA (0.52±0.16 AU or 0.25±0.10 AU, respectively; both
P<
0.05
vs.
PMA). DetaNONOate (320 or 400 μM) exerted similar inhibitory effects on MMP-9 activity (0.57±0.13 AU and 0.45±0.06 AU, respectively; both
P<
0.05
vs.
PMA). Conversely, treatment with ODQ (10 or 32 μM) had no effects on 200 μM SNAP-induced inhibition of PMA-stimulated MMP-9 activity (0.25±020 AU and 0.14±0.11 AU, respectively; both
P<
0.05
vs.
PMA). Nevertheless, 3.2 and 5 μM BAY 11-7082 decreased PMA-stimulated MMP-9 activity (0.54±0.07 AU and 0.27±0.09 AU, respectively; both
P<
0.05
vs.
PMA).

Conclusion:
Our results suggest that NO attenuates MMP-9 production by endothelial cells in a concentration-dependent manner. While this effect is independent of soluble guanylate cyclase activation, it apparently involves inhibition of NFB activity.
Abstract 513: Control of Lysosome Trafficking and Autophagic Flux by Acid Sphingomyelinase in Coronary Arterial Myocytes: Protective Role in Atherosclerosis
Authors: Zhang Yang , Xu Ming , Li Xiang , Li Xiao-Xue , Xia Min , Xiong Jing , Boini Krishna M , Li Pin-Lan
Published: March 2022

Autophagy occurs in a variety of mammalian cell via several stages of cellular activity including induction, autophagosome (AP) formation, and autophagic flux (i.e. docking and fusion of APs with lysosomes and breakdown of autophagic vesicles). Despite extensive studies on APs formation, so far little is known about the molecular mechanisms regulating autophgic flux. Since ceramide, a hydrolysis product of sphingomyelin by lysosomal acid sphingomyelinase (Asm), is implicated in the control of lysosome function in coronary arterial myocytes (CAMs). The present study tested the hypothesis that Asm is critical for AP fusion with lysosomes in CAMs and thereby protects mice from atherosclerosis. By Western blotting and flow cytometry, proatherogenic stimulation of wild type (Asm
+/+
) CAMs with 7-ketocholesterol (7K) was found to significantly enhance LC3B expression by 4.6 folds and increase AP positive cells from 6.7% to 20%. Such increases in LC3B expression and AP positive cells were augmented in CAMs with Asm siRNA or CAMs from Asm gene deficient mice (Asm
-/-
). After overexpression of GFP-LC3B (AP marker) and RFP-Lamp1 (lysosome marker), 7K-induced fusion of APs with lysosomes, an indicator for the formation of autophagolysosomes (APLs) was abolished in Asm
-/-
CAMs compared to Asm
+/+
CAMs, which was further confirmed by flow cytometry of late autophagy using acridine orange. Consistently, an autophagy degrading protein, p62, was accumulated upon proatherogenic stimulation in Asm
-/-
CAMs, but not in Asm
+/+
CAMs. In whole animal experiments, Asm
-/-
mice but not Asm
+/+
mice on the Western diet exhibited atherosclerotic phenotype, namely, arterial wall thickening, lipid deposition, CAM dedifferentiation and extracellular matrix deposition. In coronary arterial wall of Asm
-/-
mice, there was tremendous APs accumulation and p62 aggregation and inclusion in the muscle layer. These data together indicate that the control of lysosome trafficking and fusion by Asm or its product is essential to a normal autophagic flux in CAMs and that the deficiency of Asm control of autophagic flux may be an important pathogenic mechanism of atherogenesis in coronary arteries.
Abstract 518: Vascular Smooth Muscle Na/Ca exchanger-1 (NCX1) Regulates Basal Blood Pressure and Angiotensin II-induced Hypertension
Authors: Wang Jingyu , Chen Ling , Li Meng , Zhang Jin
Published: March 2022

We have reported that vascular smooth muscle (VSM)-specific NCX1 overexpression exaggerated blood pressure (BP) elevation induced by acute Angiotensin II (Ang II) injection. Here, we aimed to study if altered VSM NCX1 expression affects chronic Ang II infusion-induced hypertension. Adult wild type (WT) and transgenic mice that express either ∼7 fold higher (TG) or ∼90% lower (KO) VSM NCX1 protein were used. Mice of each genotype were randomly divided into vehicle and Ang II groups, to which physiological saline or 400 ng/kg/min Ang II in saline were given, respectively, by a subcutaneously embedded minipump. Three weeks later, BP was measured by intra-carotid artery catheter under 1.5% isoflurane anesthesia. Of the three vehicle groups, NCX TG mice had higher (108 ± 2 mm Hg,
n
= 7,
P
< 0.05), and KO mice had lower (88 ± 2 mm Hg,
n
= 8,
P
< 0.05), baseline BP compared to WT mice (97 ± 3 mm Hg,
n
= 5). Ang II treatment caused BP elevation to a significantly greater extent in TG (∼18 ± 2%,
n
= 7,
P
< 0.05) than in WT mice (∼10 ± 2%,
n
= 4), but had negligible effect on KO mice (∼6 ± 4%,
n
= 6,
P
> 0.05). NCX1 protein expression was upregulated by Ang II treatment in TG and WT (∼2 fold), but not KO, mouse aorta. We then investigated vasoconstriction in isolated, pressurized mesenteric small arteries. Basal myogenic tone (MT, 70 mm Hg) was augmented in TG (23 ± 2% of passive diameter, PD,
n
= 5,
P
< 0.05) and attenuated in KO (15 ± 1%,
n
= 8), compared to WT arteries (17 ± 2%,
n
= 5). Myogenic reactivity (MR) and vasoconstriction induced by phenyleprine (PE) were enhanced in TG and reduced in KO arteries. Ang II treatment potentiated MR/MT but not PE-induced vasoconstriction. We conclude that: 1) Under physiological conditions, in small resistance arteries that have tone, VSM NCX1 primarily mediates Ca
2+
entry which contributes to the maintenance of vascular tone; and 2) The level of VSM NCX1 expression (function), arterial contraction, and BP are highly correlated under both physiological and certain hypertension conditions (e.g., salt-, Ang II-induced animal hypertension and human pulmonary hypertension). This indicates that NCX1 plays a critical role in the long-term control of arterial contractility and basal BP, and also might play an important role in most, if not all, forms of hypertension.
Abstract 527: Apelin-13 Inhibits Large-Conductance Ca2+-Activated K+ Channels in Cerebral Artery Smooth Muscle Cells via a PI3-Kinase-Dependent Mechanism
Authors: Modgil Amit , Yao Fanrong , O'Rourke Stephen T , Sun Chengwen
Published: March 2022

Apelin-13 has been reported to act directly on APJ receptors in vascular smooth muscle (VSM) to induce vasocontraction. However, the ionic mechanisms underlying this action at the cellular level remain unclear. Large-conductance Ca2+-activated K+ (BKca) channels in VSM cells are critical regulators of membrane potential and vascular tone. Thus, in the present study, we examined the effect of apelin-13 on BKca channel activity in VSM cells, freshly isolated from rat middle cerebral arteries. In whole-cell patch clamp mode, apelin-13 (0.001μM-1μM) caused dose-dependent inhibition of BKca in VSM cells. Apelin-13 (0.1μM) significantly decreased the BKca current density from 71.25±8.14 pA/pF to 44.52±7.10 pA/pF (n=14 cells, P<0.05). This inhibitory effect of apelin-13 was confirmed by single channel recording in cell-attached patches, in which apelin-13 (0.1μM) decreased the open-state probability (NPo) of BKca channels from 0.0239 ±.00408 to 0.0148 ± 0.00267 (n=9 cells, P<0.05) in freshly isolated VSM cells. However, in inside-out patches, application of apelin-13 (0.1μM) into the pipette solution did not alter the NPo of BKca channels (NPo of BKca: 0.72±0.06 and 0.79±0.08 before and after apelin-13 administration, n=10, p>0.05), suggesting that the inhibitory effect of apelin-13 on BKca is not mediated by its direct action on BKca. In whole cell patches, pretreatment of VSM cells with LY-294002 (10 μM), a PI3-kinase inhibitor, markedly attenuated the apelin-13-induced decrease in BKca current density by 52%. Next, we examined the effect of apelin-13 on PI3- kinase activity in rat cerebral arteries using western blot analysis. Treatment with apelin-13 (0.1μM) significantly increased the ratio of phosphorylated-Akt/total Akt by 40%, indicating that apelin-13 significantly increases PI3-kinase activity. In summary, apelin-13 inhibits BKca channel current density via a PI3-kinase-dependent signaling pathway, which may contribute to its regulatory action in the control of vascular tone.
Abstract 533: MAS Receptor Activation Contributes to the Improvement of Vascular Remodeling During Chronic Angiotensin II Type 1 Receptor Blockade in Angiotensin II Type 2 Receptor Knockout Mice.
Authors: Savoia Carmine , Arrabito Emanuele , Zezza Luigi , Sada Lidia , Nicoletti Carmine , Pucci Lorenzo , Steckelings Ulrike M , Volpe Massimo
Published: March 2022

Angiotensin (Ang)-(1-7) through MAS receptor may counteract the Ang II-induced actions. We previously demonstrated that the Ang receptor blocker olmesartan (OLM) improved vascular remodeling in SHR in part through the activation of MAS receptor. Here, we hypothesized that such an effect is independent of Ang II type 2 (AT2) receptor activation. AT2-knockout mice (AT2-K/O, 12 weeks old, n=6 per group) were treated with vehicle (control) or Ang (1-7) (15,5 pmol/kg/min) or Ang II (400 ng/kg/min) ± OLM (10 mg/kg/day) ± the MAS antagonist A-779 (11 pmol/min) for 14 days. BP was measured by tail-cuff method. In isolated mesenteric arteries pre-contracted with norepinephrine (10 μM) endothelium -dependent and -independent relaxation was assessed by dose-response curves to acetylcholine (1 nM to 100 μM) and SNP (10 nM to 1 mM) respectively. Mesenteric arteries media-to-lumen ratio (M/L) and cross sectional area (CSA) were evaluated on pressurized preparations. MAS expression in aorta was evaluated by immunoblotting. Ang II infusion increased BP in AT2-K/O (+20% vs control mice, P<0.05). OLM reduced BP in Ang II-infused mice (104.8±4.4 mmHg vs 121.3±2.8 mmHg, respectively, P<0.05). A-779 had no effect on BP in Ang II-infused mice treated with OLM. Ang (1-7) did not lower BP in AT2-K/O. Endothelium -dependent and -independent relaxations were similar in treated and control mice. M/L was similar in Ang II-infused and control mice. OLM significantly reduced M/L in Ang II-infused AT2-K/O (5.45±0.12 % vs 7.01±0.4%, respectively, -23%, P<0.05). A-779 increased M/L in Ang II-infused AT2-K/O treated with OLM (+18%, P<0.05). Ang (1-7) reduced M/L in AT2-K/O (-21%, P<0.05). CSA was similar in all the groups. MAS expression was similar in Ang II-infused and control mice. MAS expression was increased by OLM in Ang II-infused mice (+108%, P<0.05). A-779 prevented MAS increase in Ang II-infused mice treated with OLM. Ang (1-7) increased MAS expression in AT2-K/O (+81%, P<0.05). In conclusion OLM reduced M/L and improved vascular remodeling in AT2-K/O in part through the increased expression and activation of MAS, independently of BP reduction. Such an effect is indeed independent of AT2 activation, as MAS is functionally active in the vasculature of AT2-K/O.
Abstract 536: in vivo Evaluation of the Pressure-independent Aortic Stiffening in Hypertensive Rat Models
Authors: Vayssettes-Courchay Christine , Ragonnet Christophe , Isabelle Marc , Vilaine Jean-Paul
Published: March 2022

We have shown that in addition to compliance and distensibility, the analysis of the arterial distension (pulsatile diameter) via echotracking (ArtLab) provides information to discriminate the pressure-dependent and independent arterial stiffening. In the present study we evaluated these parameters in different hypertensive rat models. Mean, systolic, diastolic and pulse arterial pressure (AP) and the aortic diameter were recorded in 6 groups of pentobarbital anaesthetized rats: 20 week old spontaneously hypertensive rat (SHR), SHR treated with the NOS inhibitor L-NAME 6 mg/kg/day in drinking water (SHRLN), DOCA-salt (uni-nephrectomized, deoxycorticosterone treated rats) and VDN (VitamineD+Nicotine treated rats); all were compared to their respective controls: Wistar-Kyoto (WKY) and Wistar normotensive rats. The diameter was comparable in all groups. Except in the DOCA-salt, decreased compliance, distensibility and distension were observed, indicating an aortic stiffening. When in each rat AP was decreased acutely by clonidine i.v., reaching the control rat values, compliance and distensibility were partially restored in SHR>old SHR=SHRLN>VDN; the distension wave (area under the curve adjusted to heart rate) was restored in SHR, partially in old SHR, not in SHRLN and VDN (table). These data show, via the analysis of the aortic pulse diameter distension, that in SHRLN and VDN the increase in aortic stiffness is in part pressure-independent and due to arterial wall remodeling. The results are in line with previous structural vascular wall studies.Table : * : p<0.05 versus control normotensive rats, § : p<0.05 versus basal value before clonidine.
Abstract 54: Tissue Oxygenation Using Fractional Hypoxia Correlates Inversely With Blood Flow in Human Renovascular Disease (RVD) and Essential Hypertension (EH)
Authors: Saad Ahmed , Crane John A , Glockner James , Herrmann Sandra M , Ebrahimi Behzad , Lerman Lilach O , Textor Stephen C
Published: March 2022

Determinants of Intra-renal oxygenation in human RVD are poorly understood. Blood oxygen level-dependent (BOLD) MRI measures deoxyhemoglobin (R2*) to determine cortical and medullary R2* which have focal variations and gradients. We hypothesized that tissue fractional hypoxia (% of axial kidney area with R2*>30s
-1
) would correlate better with tissue perfusion, renal blood flow (RBF), and O2 consumption than using regions of interest (ROI).

METHODS:
Inpatient studies were performed in EH (n=68 kidneys), and high-grade RVD (n=18 kidneys, 71±5.5% occlusion) before and 3 months after stents. All were treated with ACE/ARB Rx and thiazides during 150 mEq/d Na+ intake. BOLD MR images were obtained at 3T before and after furosemide (20mg IV), and axial images analyzed to estimate R2* levels above 30s
-1
(fractional hypoxia) over the entire kidney section. Maximal R2* levels using regions of interest (ROIs) also were determined. Single-kidney cortical and medullary RBF (ml/min) perfusion (ml/min/cc) were determined by MDCT.

Results:
RBF was lower in kidneys with untreated RVD than EH (285.6±141 vs. 399±159 ml/min p<0.05), as was cortical perfusion (2.58±.63 vs. 3.50±1.05, p<.05 ). Cortical and medullary R2* did not differ, but fractional hypoxia was greater in RVD as compared to EH (18.3±12 vs. 11.5±8.5%, p<.05). Fractional hypoxia was inversely correlated with blood flow (R=-.29, p<.01) and perfusion (R=-.26, p<.01). R2* fell after furosemide (p<0.01) indicating a role for O2 consumption in determining tissue hypoxia. RBF and cortical perfusion increased after stenting (285±141 vs. 318±178 ml/min and 2.58±0.6 vs. 2.9±0.7ml/min/cc, p<0.05) respectively. Fractional hypoxia, but not cortical or medullary R2* based on ROI-methods, decreased in patients 3 months after renal revascularization (18.3 ±12 vs. 10.52%, p<0.05)

Conclusions:
These data demonstrate for the first time that fractional hypoxia in the kidney is directly related to chronically reduced blood flow and cortical perfusion in human RVD, whereas selective ROI-based measures identified only minor changes in peak R2* gradients between cortex and medulla. The reversibility of fractional hypoxia after stenting supports the use of this new method to identify critical renovascular lesions.
Abstract 605: Calcitonin Gene Related Peptide (cgrp) And Its Role In Cardiovascular And Autonomic Regulation
Authors: Mai Tu , Diedrich Andre' M , Supowit Scott , Dipette Donald J , Gangula Pandu R , Garland Emily , Robertson David
Published: March 2022

Calcitonin gene related peptide (CGRP) is the most potent microvascular vasodilator known. It exerts effects centrally as well as in the gastrointestinal, respiratory and endocrine system. However, its role in cardiovascular regulation remains controversial. We studied the hemodynamic and autonomic function in CGRP knockout (KO) mice. Blood pressure (BP), heart rate (HR) and activity level were assessed at least one week after mice were implanted with telemeters. Ten CGRP KO and ten wild type (WT) mice (3-4month old) were used in this study. After the recording period, 24-hr urine and blood were collected for assessment of catecholamine and their metabolites. To study the baroreflex sensitivity, phenylephrine and sodium nitroprusside were administered in an acute study with seven mice from each group. We found higher blood pressure at rest during daytime in the CGRP KO mice (102.4 ± 1.652mmHg, vs. 110.5 ± 3.242mmKg, p = 0.039), though not in 24-hr mean arterial pressure (MAP) (105.0 ± 1.789mmHg, vs. 112.1 ± 3.123mmHg, p = 0.05). There was no difference in HR in the two groups (KO: 566.4 ± 12.09; WT: 581.9 ± 8.544, p = 0.31). Norepinephrine was greatly elevated in both plasma and 24-hr urine in KO mice (in urine: KO: 956.7 ± 90.65 pg/ml, WT: 618.1 ± 46.15pg/ml, p = 0.004; in plasma: KO: 2505 ± 596.3pg/ml, n=6, WT: 1168 ± 98.05 pg/ml, p = 0.04). Baroreflex sensitivity was higher in the KO mice (KO: 3.217 ± 0.6852 ms/mmHg, n=7, WT: 1.398 ± 0.2583 ms/mmHg, n=7, p=0.03). The activities level was slightly lower in the KO mice (KO: 6.717 ± 1.059 counts/min, WT: 10.59 ± 1.787 counts/min p = 0.08). However, KO mice that were 6-8 month old were heavier compared to WT mice of the same age (KO: 30.65 ± 0.6009 g, n=17, WT: 27.89 ± 0.6345 g, n=17, p = 0.004). While CGRP might seem to contribute less to the physiological regulation of blood pressure in young adult mice due to their better baroreflex function, it likely contributes to blood pressure and sympathetic activation in older mice. Therefore, further study in BP, HR and metabolic syndromes in old CGRP KO mice is required.
Abstract 611: Damage-associated Molecular Pattern (DAMP) Signaling Contributes to Poor Outcomes in Diabetic Ischemic Stroke
Authors: Abdelsaid Mohammed A , Cobbs Aisha , Li Weiguo , Filosa Jessica , Webb R. Clinton , Sullivan J , Ergul Adviye
Published: March 2022

Aims:
Vascular injury including edema and hemorrhage after ischemic stroke is increased in diabetes. High-mobility group box 1 (HMGB1) is a DAMP passively released by necrotic cells that initiates pro-inflammatory signaling through binding to the TLR4 receptor. Here, we investigated the possible involvement of HMGB1 in increased vascular damage in diabetic ischemic stroke.

Methods and Results:
Focal ischemia was induced by 3 h MCAO and 21 h reperfusion in sham (n=6), (Wistar, n=5-7) or diabetic (Goto-Kakizaki, n=6) rats. Expression of HMGB1 and TLR4 was assessed by Western blot in both ischemic and non-ischemic hemispheres. Compared to sham, both control and diabetic rats showed a 2-fold increase in HMGB1 in both non-ischemic and ischemic hemispheres, respectively. While there was no change in TLR4 expression in control rats, it was increased by 2-fold in both hemispheres in diabetes. Neurovascular injury as indicated by percent (%) infarct, edema ratio (edema/infarct), incidence of hematoma and functional outcome were assessed in all groups. Infarct was greater in controls (57.7±5.8% vs. 23±1.0%), while edema ratio was greater in diabetic animals (1.8±0.2 vs. 0.2±0.06). Hematoma was present in the ischemic hemispheres of all diabetic rats and in 43% of the controls. Compared to controls, neurological deficits were greater in diabetic animals after focal ischemia (p<0.05).

Conclusions and clinical relevance:
Our findings indicate that HMGB1 and TLR4 may be involved in augmented vascular injury in diabetic ischemic stroke. As such, better understanding of the role(s) of DAMPs including HMGB1 and TLR4 will aid in the development of vasculoprotective therapeutic strategies and improve outcomes for diabetic stroke patients.
Abstract 616: Caveolin-1 (CAV1) inhibits G Protein Coupled Receptor Kinase Type 4 (GRK4) kinase activity
Authors: Van Sciver Robert E , Gildea John J , Bigler Wang Dora , Felder Robin A
Published: March 2022

In human renal proximal tubule cells, the dopamine-1 receptor (D1R) has been shown to increase cAMP production and negatively regulate sodium transport. Defects in D1R signaling have been associated with SNPs in G-protein-coupled receptor kinase type 4 (GRK4) which hyperphosphorylates and decreases D1R function, and the D1R uncoupling phenotype is fully reverted by reducing GRK4 expression with siRNA. We have also shown that CAV1 co-immunoprecipitates with GRK4, and that the normal D1R inhibition of the sodium transporter, NaKATPase alpha, can be blocked using siRNA to CAV1. We therefore hypothesized that CAV1 directly physically interacts with GRK4 and inhibits its kinase activity. Using purified human GRK4 and CAV1 proteins and an in-vitro peptide homogeneous kinase assay, we investigated the effects of caveolin-1 (CAV1) on the kinase activity of GRK4. By overexpressing and isolating a tandem affinity purified protein (TAP tagged) functional version of human CAV1 isolated from a human renal proximal tubule cell line, we were able to ensure that all of the post-translational modifications of the protein were present. Utilizing a peptide Ser/Thr kinase activity assay, we demonstrated that with an excess of CAV1, the kinase activity of GRK4 is inhibited by 37.8±1.3% (n=4, p<0.05). Using this same assay, we showed that purified human calcium calmodulin 1 (CALM1), a known inhibitor of GRK4 when using a complete cell lysate, does not significantly decrease the kinase activity of purified GRK4 using a CALM1 concentration over 100 times the published Kd value. This suggests that there is a missing co-factor or factors required for CALM1 to inhibit GRK4. In summary, we report for the first time that CAV1 inhibits the kinase activity of GRK4, and that a known inhibitor of GRK4, CALM1, does not directly inhibit GRK4 kinase activity using purified proteins in an in-vitro kinase activity assay.
Abstract 619: Dietary Calcium Intake Is Associated With Adiposity, Insulin Resistance, Hdl-cholesterol, Inflammatory State And Blood Pressure, But Not With Erythrocyte Intracellular Calcium And Endothelial Function In Healthy Premenopausal Women
Authors: Ferreira Thais , Torres Marcia , Nogueira Livia , Rodrigues Maria , Valença Debora , Soares Elaine , Carvalho Renata , Nogueira-Neto Jose , Sanjuliani Antonio F
Published: March 2022

Background:
Recent studies suggest that dietary calcium may have beneficial effects on adiposity, insulin resistance, dyslipidemia and blood pressure (BP). One potential mechanism underlying these benefits involves modifications in intracellular calcium concentration ([Ca
2+
]i).

Objectives:
To evaluate the associations of dietary calcium with adiposity, erythrocyte [Ca
2+
]i, metabolic profile, BP, inflammatory state and endothelial function in healthy premenopausal women.

Methods:
In this cross-sectional study 76 women aged 18-50 years were submitted to the evaluation of dietary intake, anthropometric parameters, body composition, erythrocyte [Ca
2+
]i, biochemical variables, endothelial function and BP. A food frequency questionnaire was used to assess the usual dietary intake. Endothelial function was evaluated by serum concentrations of adhesion molecules and by peripheral arterial tonometry (PAT) method, using Endo-PAT 2000
®
. Participants were allocated in two groups according to calcium intake: low calcium group (LCG; n=32; < 600mg/day) and high calcium group (HCG; n=44; ≥ 600mg/day).

Results:
Women in LCG compared with those in HCG exhibited, after adjustments for potential confounders (age and intake of energy, alcohol, protein, carbohydrates and lipids) higher values of body mass index (27.0 ± 1.1 vs. 25.6 ± 0.8 kg/m
2
, p = 0.02), waist circumference (87.8 ± 2.7 vs. 84.4 ± 2.1cm, p = 0.04), waist-to-height ratio (0.54 ± 0.02 vs. 0.52 ± 0.01, p = 0.02), percentage of body fat (27.0 ± 1.1 vs. 25.6 ± 0.8, p = 0.02), insulin (27.2 ± 2.9 vs. 22.6 ± 1.4 μU/ml, p = 0.02), HOMA-IR (5.7 ± 0.6 vs. 4.7 ± 0.4, p = 0.02), leptin (24.9 ± 2.5 vs. 21.5 ± 2.2ng/ml, p = 0.01) and diastolic BP (72.4 ± 2.0 vs. 68.2 ± 1.6mmHg, p = 0.04); and lower levels of HDL-cholesterol (52.9 ± 2.2 vs. 58.6 ± 1.8mg/dl, p = 0.004) and adiponectin (31.9 ± 3.5 vs. 34.6 ± 3.0μg/ml, p = 0.004). Endothelial function assessed by PAT and [Ca
2+
]i were similar in both groups. Subjects in HCG had lower odds ratio for prevalent overweight, obesity, abdominal obesity, insulin resistance, HDL-cholesterol < 60mg/dl and systolic BP > 120mmHg.

Conclusions:
The findings of the present study suggest that high calcium intake is inversely associated with some cardiovascular risk factors.