bluuebunny/crossref_metadata_2025 huggingface dataset:
Mapping of Two Quantitative Trait Loci (QTL) for Blood Pressure (BP) on Rat Chromosome (CHR.) 2 Using Congenic Strains by Telemetry
Authors: Deng Alan Y , Dutil Julie , deBlois Denis , Tremblay Johanne
Published: July 2021

P141

We previously mapped a BP QTL on rat Chr. 2 near the Na,K-ATPase (
Atp1a1)
locus in two congenic strains. In the strain, S.MNS-
Adh
/
D2Mit5,
the Dahl salt sensitive (S) Chr. region, which was replaced by that of the Milan normotensive (MNS) strain, was quite large (roughly 80 cM). Now, we made 6 substrains from the original S.MNS-
Adh
/
D2Mit5
strain, and then measured BP by telemetry for two (and will continue to measure for the rest) substrains. These two substrains do not contain overlapping chromosome regions. One, S.MNS-D2Mit6
/
D2Rat166 (S.MNS1)
,
encompasses the angiotensin receptor AT1B (
Agtr1b
) and the other substrain, S.MNS-D2Mit9/
Camk2d
(S.MNS2) includes
Atp1a
1. Both substrains showed a significant (p<0.0001) effect in reducing the mean arterial pressure (MAP) in mmHg (see graph). This proves the presence of a QTL around
Agtr1b.
The BP effects of these QTL from the two separate substrains appeared to exceed those of the same number of QTL combined in one strain. This implies that these individual QTL on the same chromosome interact epistatically with one another.
Expression of Ryanodine Receptor 3 and Trp Channels in Endothelium of Human Mesenteric Artery: A Single-Cell Rt-Pcr and Patch-Clamp Analysis
in Situ
.
Authors: Hoyer Joachim , Kuehn Meike , Degenhardt Christiane , Runkel Norbert , Paul Martin , Franklin MC Benjamin , Koehler Ralf
Published: July 2021

P146

Ca
2+
mobilization plays an important role in endothelial function by stimulating Ca
2+
-dependent synthesis of vasodilating factors. In addition to InsP
3
- mediated Ca
2+
store depletion, Ca
2+
release from ryanodine-sensitive pools and Ca
2+
-influx through cation channels of the TRP-gene family have been suggested to be involved in endothelial Ca
2+
signaling. In cultured endothelial cells (EC) the function and expression of ryanodine-receptors (RyR) and TRP channels might differ substantially from those in native endothelium of human blood vessels. We, therefore, characterized expression and function of RyR and TRP channels in EC of small human mesenteric artery (MA) by use of single-cell RT-PCR and patch-clamp techniques in situ. MA were isolated from colon specimens of patients subjected to hemicolectomy. For single-cell RT-PCR and PC experiments single human mesenteric artery EC (HMAEC) were harvested directly from the luminal vessel with the patch pipette. Expression of the RyR subtype 3, but not of the RyR1 and RyR2 subtpyes was detected in 25% of HMAEC samples. Correspondingly, in PC experiments in HMAEC, application of caffeine (0.5 mM) induced Ca
2+
-release from ryanodine-sensitive stores and subsequently activation of Ca
2+
-activated K
+
channels leading to a sustained endothelial hyperpolarization from a resting potential of 28 ± 3 mV to -46 ± 4 mV. Single HMAEC expressed the TRP subtypes, TRP1 and TRP3, but not TRP 4 and 6. The TRP1 was the predominantly expressed TRP subtype as expression was detected in 16% of HMAEC. TRP3 expression was detected in only 3% of HMAEC. In single-channel and whole-cell patch clamp experiments in HMAEC, InsP
3
-mediated Ca
2+
-store depletion in response to bradykinin (100 nM) activated TRP related nonselective cation currents leading to Ca
2+
entry. In conclusion, Ca
2+
release from ryanodine-sensitive stores mediated by RyR3 and Ca
2+
entry through TRP1 might represent important components of endothelial Ca
2+
signaling and thereby of endothelial function in intact human blood vessels.
Critical Roles of Rho Kinase and Mek/Erk Pathways for Angiotensin Ii-Induced Plasminogen Activator Inhibitor-1 Gene Expression
Authors: Takeda Kotaro , Ichiki Toshihiro , Tokunou Tomotake , Fujii Satoshi , Kitabatake Akira , Takeshita Akira
Published: July 2021

P157

Plasminogen activator inhibitor-1 (PAI-1) plays an integral role not only in the regulation of plasminogen activity and fibrinolytic system but also in the pathogenesis of atherosclerosis and hypertension. Because angiotensin II (Ang II) is also involved in these processes, we investigated its role in the intracellular signaling cascade leading to PAI-1 gene expression in vascular smooth muscle cells (VSMC). Ang II increased the PAI-1 mRNA and protein levels through Ang II type 1 receptor. Although PAI-1 mRNA stability was not increased by Ang II, PAI-1 gene promoter activity, which was measured by luciferase assay, was significantly increased by Ang II. This process did not require de novo protein synthesis. BAPTA-AM, genistein and AG1478 completely inhibited the Ang II-induced PAI-1 mRNA upregulation, suggesting that intracellular calcium, tyrosine kinase and epidermal growth factor (EGF) receptor transactivation were involved in this process. However, inhibition of protein kinase C (PKC) by calphostin C, GF109203, or prolonged exposure to PMA failed to abolish the Ang II-induced PAI-1 upregulation, suggesting PKC pathway was not involved. PD98059 suppressed Ang II-induced PAI-1 upregulation, whereas SB203580 did not, suggesting that MEK/ERK1/2 pathway rather than p38 MAP kinase pathway was crucial in this process. Furthermore, adenovirus-mediated expression of dominant negative form of Rho kinase or Rho kinase inhibitor Y27632 also completely suppressed PAI-1 induction by Ang II without affecting Ang II-induced ERK1/2 activation. These data suggest that activation of both MEK/ERK1/2 and Rho kinase pathways will be necessary for the upregulation of PAI-1 gene expression and these two pathways may act synergically to promote PAI-1 gene transcription at least at the downstream of ERK1/2 in VSMC. These findings are important biological and therapeutical implications for the evolution of arterial wall thrombus and the pathogenesis of atherosclerosis by Ang II.
20-Hydroxyeicosatetraenoic Acid Mediates Angiotensin Ii-Induced Phospholipase D Activation in Vascular Smooth Muscle Cells
Authors: Parmentier Jean-Hugues , Muthalif Mubarack M
Published: July 2021

P164

Angiotensin II (Ang II) promotes arachidonic acid (AA) release via activation of cytosolic phospholipase A2 (cPLA
2
) and D (PLD) in rabbit vascular smooth muscle cells (VSMC). Ang II also stimulates the Ras/MAP kinase pathway, and MAP kinase activates cPLA
2
and PLD in VSMC. The increase in Ras/MAP kinase activity elicited by Ang II in VSMC is mediated by 20-hydroxyeicosatetraenoic acid (HETE) and to a lesser extent by 12(S)-HETE, metabolites of AA generated by cytochrome P450 (CYP) and lipoxygenase (LO), respectively, upon activation of cPLA
2
by calcium calmodulin-dependent kinase II. The purpose of this study was to determine if Ang II-induced PLD activation stimulation in VSMC from rabbit aorta is mediated by the Ras/MAP kinase pathway via AA metabolites generated by cPLA
2
. Ang II (100 nM) increased PLD activity measured as the production of
3
H-phosphatidylethanol from phospholipids in VSMC prelabeled with
3
H-oleic. Inhibitors of PLD (C
2
-ceramide, 10 μM), phosphatidate phosphohydrolase (propranolol, 10 μM) and diacylglycerol lipase (RHC 80267, 10 μM) attenuated Ang II-induced AA release. PLD activity was not altered by propranol and RHC 80267. Therefore, Ang II-induced AA release is partly mediated through PLD activation. Ang II-induced PLD activation was decreased by MAFP (50 μM), a cPLA
2
inhibitor. Inhibitors of lipoxygenase (baicalein, 5 μM) and CYP (ODYA, 5 μM) but not cycloxygenase (indomethacin, 10 μM) attenuated Ang II-induced PLD activation. AA metabolites of CYP, 20-HETE (0.25-1 μM) and 12(S)-HETE increased PLD activity. Inhibitors of ras farnesyltransferase (FPT III, 25 μM, 18 hr) and MAP kinase kinase (UO126, 10 μM) significantly attenuated the increase in PLD activity elicited by 20-HETE. These data suggest that the CYP metabolite, 20-HETE, generated from AA following cPLA
2
activation by Ang II, stimulates Ras/MAP kinase pathway, which in turn activates PLD and release additional AA.
Enalapril (E) Attenuates Oxidative Stress (OXS) in Diabetic Rats.
Authors: de Cavanagh Elena MV , Fraga Cesar G , Toblli Jorge E , Inserra Felipe , Ferder Leon F
Published: July 2021

P183

Oxidative stress has been involved as a possible mechanism of tissue injury. The effect of E on OxS and tissue injury was studied in kidney and heart from streptozotocin (STZ)-induced diabetic rats. Male Sprague-Dawley, were divided into: STZ (65 mg/kg, single i.p. dose); STZ+E (STZ and E, 20 mg E /L drinking water), and Control (C) (saline i.p. and tap water). At 6 mo. organs were studied by light microscopy. Glomerular (GML) and tubulointerstitial lesions (TIL) were graded by a semiquantitative score (0-4). Glycemia, creatinine clearance (CrCl) and proteinuria were determined. OxS was evaluated by determining total glutathione content (GSH), protein-associated sulfhydryls (SH) and 2-thiobarbituric reactive substances (TBARS, an indicator of lipid oxidation) in kidney and heart homogenates. In STZ rats E decreased proteinuria, GML and TIL,and increased CrCl. In kidney and heart, E attenuated OxS associated to STZ. Results suggest that, in addition to previously described mechanisms, E might protect renal and cardiac tissue by attenuating OxS in STZ-induced diabetic rats.
Alterations in Blood Pressure and Heart Rate Variabililty in Transgenic Rats with Low Brain Angiotensinogen
Authors: Baltatu Ovidiu , Janssen Ben J , Plehm Ralph , Ganten Detlev , Bader Michael
Published: July 2021

P191

The brain renin-angiotensin system (RAS) system may play a functional role in the long-term and short-term control of blood pressure (BPV) and heart rate variability (HRV). To study this we recorded in transgenic rats TGR(ASrAOGEN) with low brain angiotensinogen levels the 24-h variation of BP and HR during basal and hypertensive conditions, induced by a low-dose s.c. infusion of angiotensin II (Ang II, 100 ng/kg/min) for 7 days. Cardiovascular parameters were monitored by telemetry. Short-term BPV and HRV were evaluated by spectral analysis and as a measure of baroreflex sensitivity the transfer gain between the pressure and heart rate variations was calculated. During the Ang II infusion, in SD but not TGR(ASrAOGEN) rats, the 24-h rhythm of BP was inverted (5.8 ± 2 vs. -0.4 ± 1.8 mm Hg/group of day-night differences of BP, p< 0.05, respectively). In contrast, in both the SD and TGR(ASrAOGEN) rats, the 24-h HR rhythms remained unaltered and paralleled those of locomotor activity. The increase of systolic BP was significantly reduced in TGR(ASrAOGEN) in comparison to SD rats as previously described, while the HR was not altered in TGR(ASrAOGEN) nor in SD rats. The spectral index of baroreflex sensitivity (FFT gain between 0.3-0.6 Hz) was significantly higher in TGR(ASrAOGEN) than SD rats during control (0.71 ± 0.1 vs. 0.35 ± 0.06, p<0.05), but not during Ang II infusion (0.6 ± 0.07 vs. 0.4 ± 0.1, p>0.05). These results demonstrate that the brain RAS plays an important role in mediating the effects of Ang II on the circadian variation of BP. Furthermore these data are consistent with the view that the brain RAS modulates baroreflex control of HR in rats, with AII having an inhibitory role.
Generation of a Transgenic Mouse Model Exhibiting Brain Specific Expression of Human Angiotensinogen
Authors: Morimoto Satoshi , Beltz Terry G , Johnson Kim , Davidson Robin L , Cassell Martin D , Sigmund Curt D
Published: July 2021

P194

In order to examine the importance of the brain renin-angiotensin system (RAS) in the regulation of blood pressure and the pathogenesis of hypertension, we generated a novel transgenic model expressing human angiotensinogen selectively in the brain. In the brain, angiotensinogen is abundantly expressed in astrocytes (glia) and in some neurons. The transgene (GFAP-hAGT) used consists of the coding region of the human angiotensinogen gene fused to a 2.2 Kb fragment encoding the human glial fibrillary acidic protein (GFAP) promoter, a glial-specific promoter. Three transgenic founders were identified by PCR and confirmed by Southern blot analysis. RNAse protection assay revealed that each line exhibited human angiotensinogen mRNA expression in the brain, with variable expression in tissues outside the central nervous system. In one line, transgene expression was observed primarily in the brain (10-fold greater than any other tissue) and plasma human angiotensinogen levels were only at background levels (14.8 ± 8.5 pmol/ml in GFAP-hAGT transgenic mice vs. 13.3 ± 5.7 pmol/ml in non-transgenic mice vs. 5,236.7 ± 445.3 pmol/ml in mice containing a genomic angiotensinogen construct and expressing abundant human angiotensinogen under the control of its endogenous promoter). Its physiological activity in the brain was evident from the pressor response (12 mm Hg) caused by intracerebroventricular (ICV) injection of purified human renin in conscious, unrestrained mice. The pressor response was prevented by ICV injection of angiotensin II type 1 receptor antagonist losartan. Intravenous (IV) injection of purified human renin did not elicit a pressor response. This model when combined with similar mice expressing human renin either systemically (hRen genomic construct encoded on a P1 chromosome as recently reported) or within the brain (GFAP-hREN) may provide important tools for determining the role of brain RAS in the regulation of blood pressure and in the development and maintenance of hypertension. Similar models expressing human renin and human angiotensinogen from neuron-specific promoters have been generated and are currently being analyzed.
Antisense Inhibition of Central, But not Peripheral Renin Angiotensin System Decreased Arterial Pressure in Chronic Phase of Two-Kidney, One Clip Hypertensive Rats
Authors: Kagiyama Shuntaro , Galli S M , Phillips M. Ian
Published: July 2021

P198

Introduction: The systemic renin angiotensin system (RAS) plays an important role in blood pressure (BP) regulation during the development of two-kidney, one clip hypertension (2K1C). Its contributions decrease with time after constriction of the renal artery. During the chronic phase, the peripheral RAS returns to normal, nevertheless for months the hypertension is sustained. We hypothesized that during this phase of 2K1C hypertension, the brain RAS contributes to the maintenance of high BP. Methods: Therefore, we studied in the role of brain RAS by decreasing the synthesis of angiotensinogen (AGT) and angiotensin type 1a receptors (AT1R) with intracerebroventricular (ICV) injection of antisense oligonucleotides (AS-ODN). The response of systolic BP (SBP) to AS-ODN to AGT was studied at 6 mo(Group 1) and the response to AS-ODN to AT1R at 10 mo post clipping (Group 2). Each group was divided into AS-ODN, sense or inverted ODN, and saline subgroups. All groups were implanted with ICV cannulae one week before treatment. SBP was monitored by tail cuff method. Plasma and brain angiotensin II (AngII) content was measured by radioimmunoassay in all treated 2K1C groups and in nonclipped rats. Results: The results show that in Group 1, at 6 mo post clipping, the ICV AS-ODN to AGT (200 μg/kg, n=5) significantly decreased SBP(≈−22±6 mmHg, P<0.05)compared to sense ODN and saline group (n=5). The hypothalamic AngII content in sense ODN and saline groups was significantly (P<0.05) higher than in nonclipped rats. AS-ODN to AGT reduced the elevated hypothalamic AngII level. Plasma AngII was significantly decreased in the clipped group (40±12 pg/ml) compared with nonclipped group (75±8 pg/ml). In Group 2, 10 mo post clipping, the ICV injection of AS-ODN to AT1R (250 μg/kg, n=6) significantly decreased SBP(≈−26±8 mmHg, P<0.05) for 3 days post injection, compared to inverted ODN. In contrast, intravenous AS-ODN to AT1R in dose of 250-500 μg/kg did not affect SBP. Conclusion: These results suggest that the brain RAS plays an important role in maintaining the elevated SBP in chronic hypertension phase.
Heart Protection Using a New Approach - Antisense to Angiotensin Converting Enzyme Mrna
Authors: Mohuczy Dagmara , Chen Hongjiang , Shen Leping , Mehta Jawahar L , Phillips M. Ian
Published: July 2021

P200

Introduction. Angiotensin converting enzyme (ACE), plays a major role in many cardiovascular pathophysiological conditions, such as hypertension, heart failure and myocardial ischemia. Currently used ACE inhibitors have some adverse effects and non-compliance problem. Therefore, our lab has been developing an antisense approach to inhibit the renin-angiotensin system at the gene expression level. The present study had three aims: 1. Design antisense oligodeoxynucleotide (AS-ODN) directed at ACE mRNA 2. Check tissue distribution of AS after iv injection 3. Test AS for heart protection in ischemia- reperfusion injury Methods. Epifluorescent and confocal laser-scanning microscopy, immunohistochemistry, Western blotting. Animals : mice, males, 20 g and Sprague-Dawley rats, males, 200-250 g. AS 5’ -ATTTCGTGGTGGG-3’ , inverted control (INV) 5’ -GGGTGGTGCTTTA-3’, phosphorothioates mixed with liposomes and injected iv 0.04 mg/mouse and 0.2 mg/rat. Captopril dose was 5 mg/kg. Isolated perfused heart function recordings : coronary perfusion pressure (CPP), left ventricular end-diastolic (LVEDP) and systolic pressures (LVSP). Results. Based on the rat ACE cDNA sequence, the AS was designed as 13-mer directed to the second zinc-binding domain (2998-3010 bp). Tissue distribution of phosphorothioated, fluorescein isothiocyanate (FITC)-labeled AS, 4h after iv injection, showed its presence in the lung, spleen, liver and heart, but absence in the brain. Immunostaining of the parallel heart slices with anti sarcomeric alpha-actinin antibody indicated AS localization within cardiomyocytes. ACE AS injected iv to the rats 24 h before excising the heart, which was then subjected to 25 min ischemia and 30 min reperfusion, preserved heart function. Increase in CPP and LVEDP during reperfusion was attenuated by AS as compared to INV or saline treatment (n=8 in each group, p<0.05). Moreover, ACE protein was not upregulated, as it was in the captopril-treated group. Conclusion. ACE AS is taken up by the heart tissue and provides cardioprotection by effectively preventing myocardial dysfunction caused by ischemia-reperfusion injury.
Transfer of the Human Heme Oxygenase-1 Gene Decreases Blood Pressure and Promotes Growth in Spontaneously Hypertensive Rats
Authors: Sabaawy Hatem E , Zhang Fan , Nasjletti Alberto , Laniado-Schwartzman Michal , Abraham Nader G
Published: July 2021

P203

Heme oxygenase (HO) catalyzes the conversion of heme to biliverdin, free iron and carbon monoxide (CO). Both heme and CO have been implicated in the regulation of vascular tone. We report the successful cloning of the human HO-1 cDNA (HHO-1) into a LXSN retroviral vector (LSN-HHO-1). A single intra-left ventricular delivery of 1×10
10
pfu/ml of LSN-HHO-1 to 5-day-old spontaneously hypertensive rats (SHR; n = 32) resulted in extended expression of the human HO-1 (mRNA and protein) in several tissues, including the kidney, liver, spleen, lung, heart, brain and aorta. The expression of HHO-1 was associated with a 2-3 fold increase in HO activity in these tissues. Mean blood pressure (MBP) of SHR injected with LSN-HHO-1 was significantly

lower than that of SHR injected with the control empty vector LXSN, by 4 weeks of age (144±4.6 mmHg vs 164.8±6.5 mmHg,

n=32, *
p
<0.01). SHR treated with LSN-HHO-1 demonstrated a consistent reduction in MBP of ≈20 ± 4 mmHg when compared with the control LXSN injected SHR throughout the 20 weeks of the experiment. Administration of

the HO inhibitor, stannic mesoporphyrin (Sn MP), to LSN-HHO-1-treated SHR resulted in a 15 to 18 mmHg rise of MBP, further suggesting that increased HO expression underlie, at least in part, the blood pressure lowering effect of LSN-HHO-1. Rats expressing HHO-1 showed significant reduction in the urinary excretion of the vasoconstrictor cytochrome P-450 arachidonate metabolite, 20-HETE. Moreover, gracilis muscle arterioles (≈ 55μm in diameter) isolated from HHO-1 transgenic SHR showed less contractile responses to increased intraluminal pressure than vessels isolated from LXSN-treated SHR; this effect was reversed by the addition of SnMP. Interestingly, HHO-1 transgenic rats showed significant proportionate increase in somatic growth, i.e., nose to tail length, fibula length and body weight gain. These studies demonstrate

that delivery of the human HO-1 gene by a retroviral vector results in permanent expression

of HHO-1, long-term reduction in blood pressure together with growth promoting activity in the

SHR.
Abstract 108: Title: Interaction between Systolic Hypertension and Co-Morbidities in Patients with Documented Aortic Aneurysms
Authors: Heinecke Carl , Bulibek Batyrjan , Ojha Chandra , Huang Helen , Contino Chase , Torosoff Mikhail
Published: July 2021

Background:
We investigated relation between traditional risk factors and history of systolic hypertension in patients with documented aortic aneurysms.

Material and Methods:
Study population consisted of 162 consecutive patients (37% females, 91% Caucasians) enrolled in the Aortic Aneurysm clinic at a single tertiary medical center. Medical records were reviewed retrospectively. ANOVA, chi-square, and logistic regression tests were employed. Study was approved by the institutional IRB.

Results:
Majority of patients were Caucasian males with history of tobacco abuse, hypertension, and dyslipidemia (table). Coronary artery disease, diabetes, and chronic kidney disease were infrequent in the studied cohort. Age was strongly associated with increased prevalence of systolic hypertension history (HR 1.473, 95% CI 1.123-1.932 per decade, p=0.005). On the contrary, other traditional risk factors of hypertension like male gender, diabetes, dyslipidemia, chronic kidney disease, and smoking were not associated with increased prevalence of systolic hypertension by history.

Conclusions:
In patients with documented aortic aneurysms traditional risk factors of cerebro-vascular disease, with exception for age, are not associated with hypertension history. Therefore, in a patient with hypertension history in the absence of traditional risk factors high index of suspicion for an aortic enlargement is warranted.
Abstract 115: Reduction of Trpv1 Activity in Aortic Baroreceptor Neurons in Diabetic Neuropathy: Role of Brain-derived Neurotrophic Factor
Authors: Tan Yingying , Zhang Qi
Published: July 2021

It has been well documented that diabetes mellitus is associated with cardiovascular autonomic neuropathy including dysfunction of arterial baroreflex. The mechanisms underlying diabetes-induced baroreflex dysfunction remain poorly understood. Here we investigated the function and expression of transient receptor potential vanilloid 1 (TRPV1) in aortic baroreceptor (AB) neurons isolated from streptozotocin-induced diabetic rats between 4 and 8 weeks after onset of diabetes. AB neurons in nodose ganglion were retrograde-labeled by a transported fluorescent dye, Dil. Using the whole-cell patch clamp, we found that the inward current activated by the application of capsaicin (1 μM) was significantly smaller in AB neurons from diabetic rats compared with controls. The mean peak current density of capsaicin-induced currents was 145.7 ± 24.7 pA/pF (n = 16) in diabetic neurons and 269.3 ± 31.8 pA/pF (n =15) in controls, respectively. The duration of inward current was decreased 51% in diabetic rats compared with the control group. These evoked currents were completely blocked by the capsaicin antagonist capsazepine. In addition, capsaicin-induced desensitization of TRPV1 was up-regulated, whereas TRPV1 re-sensitization was down-regulated in AB neurons from diabetic rats. Immunofluorescence staining studies demonstrated that the percentage of TRPV1-positive neurons was 50.2 ± 5.0% in control rats and 38.2 ± 1.9% in diabetic rats, respectively. This reduction in TRPV1-positive neurons in AB neurons in diabetic rats was significant (n = 11, P < 0.01). In addition, the reductions in TRPV1 currents and positive neurons s in diabetic rats were normalized by pre-treatment with anti-BDNF antibody or K252a, a TrkB tyrosine kinase inhibitor. Furthermore, incubation with BDNF caused a large reduction in TRPV1 currents in AB neurons from control rats, and the number of AB neurons with BDNF immunoreactivity was greater in diabetic than control rats. These results suggest that reduced expression and function of TRPV1 are involved in the attenuation of baroreceptor neuron excitability, and increased BDNF activity in these neurons likely contributes to the reduction in TRPV1 function through TrkB receptor stimulation in diabetic neuropathy.
Abstract 121: Cytochrome P450 1B1 Gene Disruption Prevents Cardiac Dysfunction Associated With DOCA-salt-induced Hypertension in Mice
Authors: Jennings Brett L , Estes Anne M , Yaghini Fariborz A , Fan Zheng , Malik Kafait U
Published: July 2021

Previously, we have shown that cytochrome P450 (CYP) 1B1 contributes to deoxycorticosterone acetate (DOCA) salt-induced hypertension and associated increase in vascular reactivity, endothelial dysfunction, and vascular oxidative stress in rats. This study was conducted to determine whether
Cyp1b1
gene disruption protects against cardiac dysfunction, measured by echocardiography, in this model of hypertension in mice. Eight week old male mice were uninephrectomized, subcutaneously implanted with a DOCA pellet (50mg/kg) and given 1% saline in drinking water for 4 weeks, and blood pressure (BP) was measured weekly by tail cuff method. DOCA-salt treatment increased systolic BP in

Cyp1b1
+/+

mice to a greater degree than in

Cyp1b1
-/-

mice (115 ± 3 to 188 ± 5 mmHg vs. 120 ± 5 to 152 ± 8 mmHg, respectively, P < 0.05). DOCA-salt treatment produced cardiac dysfunction, as indicated by decreased fractional shortening (FS), ejection fraction (EF), and the ratio of peak early to late diastolic mitral annulus velocities (E′/A′), and an increase in both end-diastolic volume and end-systolic volume, in

Cyp1b1
+/+

, but not

Cyp1b1
-/-

mice (Table 1). Cardiac production of reactive oxygen species, as indicated by 2-hydroxyethidium fluorescence intensity, was increased in DOCA-salt treated

Cyp1b1
+/+

mice, which was attenuated in

Cyp1b1
-/-

mice. These data suggest that
Cyp1b1
plays a pivotal role in cardiac dysfunction associated with DOCA-salt hypertension in mice by increasing oxidative stress, and that it could serve as a potential target for drugs to treat cardiovascular dysfunction associated with excess mineralocorticoid-salt hypertension.
Abstract 134: Temporal Characterization of the Development Diabetic Induced Renal Disease in Strains of Goto-kakizaki Rats
Authors: Slaughter Tiffani , Paige Adrienne , Rucker Carlos , Kyle Patrick , Kojima Naoki , Garrett Michael R , Roman Richard J , Williams Jan M
Published: July 2021

End stage renal disease is a major problem of patients suffering from diabetes and the development of novel therapeutic drugs are needed to help prevent the progression of diabetic nephropathy (DN). However, little is known about the pathogenesis of DN because the lack of an appropriate rodent model that develops progressive chronic kidney disease and renal histopathology seen in diabetic patients. The current study characterized the temporal changes in renal hemodynamics during the development of renal disease in Goto-Kakizaki (GK
Control
) diabetic rats that are resistant to the development of renal disease and a genetically modified GK substrain (GK
T2DN
) that is far more susceptible to the development of DN. Both strains have similar elevations in plasma glucose levels by 3 months of age. The GK
T2DN
strain exhibited hyperfiltration reflected by an increase in GFR (measured by the clearance of FITC-inulin) at 6 months of age compared to GK
Control
rats. GK
T2DN
rats develop progressive proteinuria that increases from 39±4 to 524±64 mg/day and a decline in GFR (from 934±68 to 370±50 μL/min/gkw) (n=21) as the rats age from 3 to 18 months of age. In contrast, proteinuria only increased to 194±33 mg/day in GK
Control
rats and GFR remained relatively unaltered over the same time period (n=19). The kidneys of GK
T2DN
rats exhibited mesangial expansion, glomerulosclerosis, and interstitial and renal fibrosis characteristic of patients with diabetes while the GK
Control
strain did not. We next utilized microarray analysis to identify differences in gene signaling pathways in the renal cortex between the two strains at the early onset of renal disease (6 months of age). We found 1266 genes that were differential expressed in signaling pathways associated with inflammation, cell proliferation, molecular transport and apoptosis between the susceptible strain (GK
T2DN
) versus the resistant strain (GK
Control
). In summary, these data indicate that the GK
T2DN
rat exhibits hyperfiltration and progressive proteinuria and chronic kidney disease and is a useful model to explore new therapies and genetic factors in the pathogenesis of DN.
Abstract 145: Clinical and Vascular Changes Related to Endothelial Dysfunction in Treated Hypertensive Patients
Authors: Trindade Michelle , Cunha Ana Rosa , Casanova Marcela , d'El-Rei Jenifer , Burlá Adriana , Burlá Marcelo , Oigman Wille , Neves Mario F
Published: July 2021

Background:
It is widely recognized that endothelial dysfunction is commonly associated to hypertension but the parameters linked to this condition among hypertensive subjects are not established yet.

Objective:
To identify clinical, metabolic and vascular changes related to endothelial dysfunction in treated hypertensive patients

Methods:
A cross-sectional study was carried out to evaluate hypertensive patients on stable drug therapy over at least 4 weeks, both genders, aged 40-70 years. Blood pressure (BP) was measured by oscilometric method, and vascular tests were performed using high-resolution ultrasonography for brachial flow-mediated dilation (FMD), Sphygmocor to obtain central hemodynamic parameters and Complior SP to determine carotid-femoral pulse wave velocity (cfPWV) which was normalized by mean BP (cfPWV-N). The 10-year risk for general cardiovascular disease from Framingham Heart Study was used to calculate vascular age. Patients (n=81) were distributed into two groups according to brachial FMD values: FMD < 10% (n=54) or FMD >= 10% (n=27).

Results:
The mean age was not different among the groups (55.1±8.3 vs 55.7±7.0 years, p>0.05). Systolic BP (142±19 vs 137±13, p>0.05) and diastolic BP (94±14 vs 92±13 mmHg, p>0.05) were similar in the two groups. The group with FMD < 10% presented greater carotid media-to-lumen ratio (0.12±0.03 vs 0.10±0.02, p<0.05), vascular age (77±14 vs 70±12, p<0.05) and higher duration of hypertension (9.9±9.19 vs 5.9±4.2, p<0.05) than the group with normal FMD. There was an inverse correlation between FMD and vascular age (r=-0.235, p<0.05). No significant differences in biochemical variables, parameters of vascular stiffness and number of criteria of metabolic syndrome were observed.

Conclusion:
The characterization of endothelial dysfunction in treated hypertensive patients was not related to age but it was more associated to duration of hypertension and vascular age. There was no evidence of vascular stiffness in these patients but the increased carotid media-to-lumen ratio may indicate the beginning of atherosclerotic process.
Abstract 147: Hypertension-related Admission and Mortality at a Referral Hospital in Tanzania - A Call to Action
Authors: Shike Naomi , Vardhana Santosh , Briant Judith , Peck Robert
Published: July 2021

Introduction
The WHO has been increasingly emphasizing and calling for research on the vast unattended burden of non-communicable diseases in the developing world. Hypertension (HTN) in particular is thought to play a growing role in morbidity and mortality in these regions, but has yet to gain significant momentum in public health initiatives.

Objective
To determine what role HTN and comorbid diseases play in admission and mortality in Bugando Medical Center (BMC), a tertiary care hospital in Tanzania serving 13 million people.

Methods
We conducted a retrospective analysis of all patients admitted to the internal medicine service at BMC over 34 months between 2008 and 2011. Data on admission diagnoses and mortality had been collected prospectively by Tanzanian doctors in hand-written logs. For patients with heart failure or stroke, the ward logs specified if this was primarily related to hypertension or other risk factors. Data were copied into an Excel database and analyzed to determine the proportion of admissions and deaths primarily related to hypertension.

Results
In 34 months 8,037 patients were admitted and 1,508 died. HTN-related disease led to 1,997 admissions (25%), while HIV-related illness led to 2,076 (26%). Similarly, HTN led to 377 deaths (25%) and HIV to 579 (38%). HTN-related disease was second only to HIV-related disease as a cause of admission and death. Among hypertensives, the most common cause of admission was congestive heart failure (446; 27%) and of death was stroke (147; 49%). In non-hypertensives, HIV-related disease was the most common cause of both admission (2029; 32%) and death (566; 46%).

Conclusions
HTN-related disease was second only to HIV as a cause of admission to our hospital and in-hospital death. Better strategies for early diagnosis and treatment of HTN are desperately need in sub Saharan Africa to prevent this morbidity and mortality. Building HTN screening and treatment on top of the extensive infrastructure for HIV disease may be a reasonable approach.
Abstract 152: John Henryism Active Coping, Blood Pressure, and Arterial Stiffness in Korean Americans
Authors: Logan Jeongok G , Barksdale Debra J , Carlson John
Published: July 2021

Introduction
It is well known that blood pressure (BP) is inversely related to socioeconomic status (SES). John Henryism Active Coping (JHAC) is defined as a strong behavioral predisposition to cope with psychosocial stressors in an active and determined manner, and has been used to explain cardiovascular disparity in African Americans. According to JHAC hypothesis, the inverse relationship between cardiovascular risk and SES is more prominent for the individuals with high JHAC. High levels of psychological stress and high prevalence of hypertension (32%) have been reported in Korean Americans (KAs).

Purpose
The study was conducted to examine the main effects and the interaction effect of SES and JHAC on cardiovascular risk (measured by BP and arterial stiffness) in KAs.

Methods
Sample
- 102 KAs (aged 21-60 years, 60% women)
Measures
-
Age, gender, BMI, SES (measured by education and income), JHAC(measured by the 12-item JHAC Scale), BP, Arterial stiffness (measured by carotid and femoral pulse wave velocity [PWV])
Data Analysis
- Two way factorial ANOVA and Duncan’s test by SAS 9.2.

Results
SBP (p=.03) and DBP (p=.01) were significantly higher in the low education group. The interaction effect of education and JHAC on SBP was significant (p=0.02), indicating that the education effect on SBP was greater in the low JHAC group. The main effect of education on PWV was not significant (p=0.64). However, the JHAC had a significant main effect on PWV (p=0.02), showing that the group with low JHAC score had higher PWV than the group with high JHAC. The interaction between education and JHAC on PWV was also significant (p=0.02), thus the effect of JHAC on PWV was greater in the low education group.

Discussion
The results demonstrated that the inverse relationship between education and cardiovascular measures was more severe in the low JHAC group, contrary to JHAC hypothesis. Furthermore, arterial stiffness, an important biomarker of cardiovascular disease, was predicted by JHAC, but not by education, showing the subjects with low JHAC had significantly higher arterial stiffness. This phenomenon was also more prominent in the subjects with low education. The findings suggest that JHAC has beneficial effect on cardiovascular health in KAs.
Abstract 166: Activation of T Cells by Dendritic Cells in Hypertension: A Potential Role of Isoketal-modified Proteins
Authors: Kirabo Annet , Wu Jing , Chen Wei , Thabet Salim R , Bikineyeva Alfiya T , Dikalov Sergey , Amarnath Venkataraman , Davies Sean S , Roberts Jackson L , Harrison David G
Published: July 2021

The adaptive immune system contributes to development of hypertension but the mechanisms or antigens involved are not known. Isoketals (IsoKs) are products of arachidonic acid oxidation that can cross-link lysine residues on proteins which in turn can be immunogenic. We sought to determine if IsoK-modified proteins serve as neoantigens presented by dendritic cells (DCs) to activate T cells in hypertension. Superoxide production by DCs was increased 5-fold by angiotensin II infusion compared to sham treated mice (334.0±49.7 versus 65.8±4.5 pmol/mg protein). This was NADPH-dependent as it did not occur in the gp91
phox-/-
mice. This increase in superoxide was associated with accumulation of IsoKs in DCs and activation of DC IL-6 production (1411.0±1330.2 versus 576.2±512.8 pg/ml). Treatment with a potent IsoK scavenger, 2-hydroxybenzylamine, markedly attenuated angiotensin II-induced hypertension (131.4 ± 9.4 mmHg versus 160.1 ± 5.1 mmHg in control mice). Finally we employed an immunization assay involving priming by adoptive transfer of DCs from either sham or angiotensin II-treated mice to naïve recipients. T cells from these recipients were then cultured for 10 days with DCs from either sham or angiotensin II treated mice. DCs from angiotensin II treated mice, but not sham mice, caused a striking activation of CD8
+
cells in this assay, as reflected by proliferation and polarization to Tc1 and Tc17 phenotypes. These studies show that angiotensin II-induced hypertension activates DCs, in large part by causing superoxide production and formation of IsoKs. We propose that IsoK-modified proteins can be presented as neoantigens by DCs, which in turn trigger T cell activation leading to hypertension.
Abstract 193: Contribution of Visceral and Epicardial Adipose Tissues to the Variation in the Cardiometabolic Risk of Apparently Healthy Young Adults
Authors: De Larochelliere Elianne , Côté Julie , Ross Marie-Kristelle , Dion-Roy Vénique , Pibarot Philippe , Pibarot Philippe , Després Jean-Pierre , Larose Eric
Published: July 2021

OBJECTIVE:
Relative contributions of abdominal versus epicardial adipose tissues to the cardiometabolic profile of young and healthy adults are unknown. We investigated the associations between subcutaneous abdominal adipose tissue (SAT), visceral abdominal adipose tissue (VAT) and epicardial adipose tissue (EAT) with markers of the cardiometabolic profile in young and apparently healthy males and females.

METHOD AND RESULTS:
Two hundred and eighty young and apparently healthy subjects (48% (134 of 280) male; age 18 to 35 years; BMI <30 kg/m2) underwent magnetic resonance imaging after voluntary recruitment in the community. We quantified SAT, VAT and EAT. Plasma cardiometabolic risk profile was assessed for each subject (blood pressure, lipid profile, fasting glucose). Associations with markers of the cardiometabolic profile were stronger for VAT and EAT than for SAT. In multivariable regression analyses including all three adipose tissues and taking into account gender and age, only VAT maintained significant associations to systolic and diastolic blood pressure, and plasma low-density lipoprotein cholesterol, total/high-density lipoprotein cholesterol ratio, triglycerides and apolipoproteins B/A1 ratio. In an effort to identify the strongest office-based predictor of visceral adiposity in this young and apparently healthy young adult sample, we determined that while BMI, waist circumference (WC) and waist-to-hip ratio were all associated to VAT in univariable analyses (p<0.05 for all), only WC remained independently associated to VAT in multivariable analysis (p<0.001). Concurrently, WC was also an independent predictor of EAT in multivariable analysis (p<0.001).

CONCLUSION
- Because visceral adipose tissues - EAT, and to a greater extent VAT - are more strongly and widely associated with the cardiometabolic profile than is SAT, we believe that assessing visceral adiposity is relevant in young and apparently healthy adults. Beyond simply measuring BMI, the addition of WC measurement in young and apparently healthy adults improves the prediction of visceral adiposity.
Abstract 222: Valsartan at a Sub-therapeutic Dose Ameliorates Myocardial Autophagy Overactivated in Renovascular Hypertension (RVH)
Authors: Zhang Xin , Li Zi-Lun , Pawar Aditya S , Crane John A , Eirin Alfonso , Lerman Amir , Lerman Lilach O
Published: July 2021

Objective:
Excessive autophagy (degradation of intracellular structures) characterizes cardiomyocyte injury in hypertensive heart disease. The effect of Angiotensin II receptor blocker Valsartan on RVH-induced myocardial autophagy remains unexplored.

Method:
Twenty-one domestic pigs (50-60 Kg) were randomized into 3 groups: RVH, RVH+Valsartan, and Control (n=7 each), and observed for 10 wks. Valsartan (320mg/d PO) was administered for 4 wks starting at 6 wks of RVH. After 10 wks, myocardial fibrosis was examined by trichrome staining, and inflammation by M1 macrophage (M1 Ø, CD163+/iNOS+) staining and TNF-α expression (immunoblotting). Autophagy was examined for Atg 12-Atg 5, Beclin-1 and its inhibitor mTOR.

Result: At 6 wks, MAP was higher in both RVH and RVH+Valsartan compared to Control (147.6 ±14.0 and 139.1±11.2 vs. 105.9±11.4 mmHg, respectively, P<0.05). At 10 wks, Valsartan did not decrease MAP compared to RVH (P=0.31), Both Beclin-1 and Atg 12-Atg 15 expressions that were elevated in RVH (P<0.05 vs. Control), indicating increased autophagy, were restored by Valsartan (Figure). Valsartan restored a compensatory increase in mTOR expression in RVH, and ameliorated myocardial fibrosis, M1 Ø, and TNF-α expression (Figure).

Conclusion:
Valsartan decreased myocardial autophagy, fibrosis and inflammation in pigs with RVH, independent of effective blood pressure control. Valsartan has novel pleiotropic effects on turnover of cellular structures in RVH-induced myocardial damage.
Abstract 239: MicroRNAs hsa-miR-584 and hsa-miR-31 Regulate Expression of Human Angiotensnogen Gene
Authors: Mopidevi Brahmaraju , Maharjan Shreekrishna , Jain Sudhir , Pandey Varunkumar G. , Kumar Ashok
Published: July 2021

Background:
Hypertension is a risk factor for stroke, myocardial infarction, and congestive heart failure. A single nucleotide polymorphism (SNP) (rs7079) in 3’UTR of human angiotensinogen (hAGT) gene is associated with elevated blood pressure. We have used TargetScan V6.0, miRanda miRNA prediction algorithms and found that two microRNAs hsa-miR-584 and hsa-miR-31 may bind differentially to rs7079 and alter the expression of hAGT gene.

METHODS:
The effect of hsa-miR-584 and hsa-miR-31 miRNAs on endogenous AGT expression levels in Hep3B cells were studied by transfection of individual miRNA mimics followed by quantitative real time PCR (Q-RT-PCR) of the hAGT gene. In addition, the 600 bp 3’UTR of hAGT gene containing either rs7079C or rs7079A was PCR amplified and cloned in to the multiple cloning site of pMIR-REPORT™ miRNA Expression Vector containing luciferase gene. These reporter constructs were then co-transfected with either miRNA mimics or inhibitors to study the effect of miRNAs on luciferase activity in Hep3B cells since these cells express hAGT gene. These experiments were also performed in HEK293 cells which do not express the hAGT gene.

Results:
Q-RT-PCR showed that hsa-miR-584 and hsa-miR-31 mimics at 50nM concentration reduced endogenous hAGT mRNA levels by 38% and 30% respectively compared to the mock (without any microRNA) or negative control (which does not have any binding site for eukaryotic 3’UTRs) in Hep3B cells. Furthermore hsa-miR-584 and hsa-miR-31 showed 40% and 25% reduced luciferase activity of the construct containing rs7079 C allele. On the other hand these miRNAs did not affect the luciferase activity in the presence of rs7079 A. When dose dependent anti miRNA inhibitors were transfected along with miRNA mimics, these mimics abolished the microRNA induced down-regulation of luciferase activity.

Conclusion:
hsa-miR-584 and hsa-miR-31 miRNAs bind strongly to the hAGT 3’UTR containing rs7079 C allele as compared to rs7079 A allele and may down regulate the expression of AGT gene containing rs7079 C allele. This may be one of the possible mechanism involved in association of rs7079 A allele with human hypertension.
Abstract 259: Sam68 Regulates Endothelin-Converting Enzyme-1b Expression and Blood Pressure Homeostasis
Authors: Zhou Junlan , Qin Gangjian
Published: July 2021

We have previously reported that E2F2, a transcription factor known for its role in cell cycle, regulates the expression of endothelin-converting enzyme-1b (ECE-1b) in endothelial cells and contributes to the maintenance of vascular contractility and blood pressure (BP). However, the molecular mechanisms underlying this novel, cell-cycle independent function of E2F2 remain largely elusive. In this study, we profiled E2F2 interactome in the nucleus of human umbilical vein endothelial cells by utilizing a proteomic approach. We identified that Sam68, a classic RNA-binding protein and Src kinase substrate, acts as an E2F2-interacting protein; co-immunoprecipitation analyses confirmed that both endogenous and ectopically-expressed Sam68 interact with E2F2. Overexpression of Sam68 repressed whereas knockdown of Sam68 increased E2F2-induced ECE-1b promoter activity and mRNA expression. Chromatin immunoprecipitation assays further confirmed that E2F2 and Sam68 co-localize on ECE-1b promoter, indicating that ECE-1b is a direct transcriptional target of E2F2 and Sam68. In vivo, Sam68 knockout (KO) mice displayed a significantly lowered BP as compared to WT littermates (tail-cuff measurements of mean BP, KO vs. WT: 104 ± 6 vs. 123 ± 7 mmHg, n=25 males of age 4-6 months, P<0.05). Together, our studies have revealed a previously unknown function of Sam68 as a transcriptional co-repressor of E2F2 and a critical regulator of BP homeostasis.