bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 230: Per Capita Proportion of Total Health Care Expenditures on Pharmaceuticals Among US Adults With Cardiovascular Disease: 2012 Medical Expenditure Panel Survey
Authors: Salami Joseph A , Valero-Elizondo Javier , Osondu Chuckwuemeka U , Ogunmoroti Oluseye , Post Janisse M , Younus Adnan , Latif Muhammad A , Ahmad Rameez , Arrieta Alejandro , Spatz Erica S , Rana Jamal S , Virani Salim S , Blanstein Ron , Blaha Michael J , Veledar Emir , Nasir Khurram
Published: March 2022

Background:
Besides inflation, the recent expansion of health insurance coverage and the limited regulation of drug pricing in the US has caused per capita pharmaceutical expenditures to increase substantially. We aim to examine the proportion of health care spending used on drugs and its association with burden of comorbidity among adults with cardiovascular disease (CVD).

Methods:
Using the 2012 Medical Expenditures Panel Survey (MEPS), adults aged 40 years with an ICD-9 diagnosis of CVD (410, 413, 414, 428, 433, 434, 435, 436, 437, 427, 440, 443, or 447) (n=1,846) from the medical condition file were studied. The burden of comorbidity was estimated using the Charlson Comorbidity Index (CCI). We used the two-part econometric model to estimate the mean healthcare and pharmaceutical expenditures and it association with CCI. The mean proportion of health care spending for pharmaceuticals was estimated and tested for trend across CCI using the variance-weighted least squares. All analyses took into account the study’s complex survey design and were weighted to the U.S. population.

Results:
Approximately 18.9 million adults aged 40 years (mean age[SD]: 69[15] years; 44% females) had a diagnosis of CVD; 37% had CCI=1, 26% had CCI=2. Overall, healthcare expenditures among adults with CVD was $248.4 billion; 26.1% ($64.8 billion) of which was spent on pharmaceuticals. Both absolute total health care expenditures and pharmaceutical expenditures significantly increased proportionally with CCI (table). However, pharmaceutical expenditures expressed as proportion of total health care spending did not increase with greater co-morbid disease burden (p-trend =0.198).

Conclusion:
Pharmaceutical expenditure is a significant contributor of healthcare spending (>1 in every 3 US dollars spent) among adults with established CVD. It is imperative to critically assess interplay of such issues as pricing and appropriate utilization of pharmaceutical agents to mitigate increasing healthcare spending among those with CVD.
Abstract 243: Risk Scoring for the Primary Prevention of Cardiovascular Disease: A Cochrane Systematic Review
Authors: Karmali Kunal N , Persell Stephen D , Perel Pablo , Lloyd-Jones Donald M , Huffman Mark D
Published: March 2022

Background:
The current paradigm for primary cardiovascular disease (CVD) prevention emphasizes absolute risk assessment to guide decision-making. Although considerable attention has been given to the derivation and validation of multivariable CVD risk scores, their effect on clinical outcomes is uncertain.

Objective:
To evaluate the effect of multivariable CVD risk scores on cardiovascular outcomes in primary CVD prevention.

Search Methods:
We searched the Cochrane Library, MEDLINE, EMBASE, and Conference Proceedings Citation Index - Science from their inception to January 2015 and imposed no language restriction. We also searched clinical trial registers and hand-searched reference lists of primary studies.

Selection Criteria:
We included RCTs and quasi-RCTs that compared the delivery of multivariable CVD risk scores to usual care among adults (≥ 18 years) free from clinical CVD.

Data Collection and Analysis:
We screened titles, selected studies for inclusion, and extracted data, including risk of bias, in duplicate. Our outcomes included: fatal and nonfatal CVD events, changes in CVD risk factor levels, adverse events, medication prescriptions in high-risk individuals, medication adherence, health-related quality of life, health-related behaviors, and costs.

Main Results:
From a total of 5,442 reports, we identified 38 studies (N=220,122 participants). Interventions ranged from simple CVD risk score presentation to multifaceted interventions incorporating multiple risk messages, clinical decision support tools, electronic reminders, patient activation material, audit and feedback, and nurse-led counseling sessions. We assessed the majority (n=32) of studies as having high or unclear risk of bias in the domains of selection, performance, detection, attrition, and reporting bias. Due to substantial heterogeneity, we were not able to perform meta-analysis for quantitative synthesis. We found no strong evidence that systematic implementation of CVD risk scores reduced fatal or nonfatal CVD events but the 2 studies reporting this endpoint were both underpowered. There were 14 studies that demonstrated improvements in CVD risk factor levels, health behaviors, or prescriptions in high-risk individuals but effect sizes were small and variable between studies. Interventions that were integrated into the electronic health record or provided “heart age” estimates tended to be most effective. There were no reported adverse effects of CVD risk scores. Effects on medication adherence, health-related quality of life, and cost were poorly reported.

Conclusions:
We found no strong evidence that CVD risk scores improve clinical outcomes. Newer models of implementing risk scores should be developed, implemented, and evaluated in adequately powered studies to improve primary CVD prevention.
Abstract 255: The Development of an Observational Registry of Treatment Patterns in US Heart Failure Patients with Reduced Ejection Fraction (HFrEF)
Authors: Fonarow Gregg C , Albert Nancy M , Butler Javed , Chan Wing W , Duffy Carol I , Hernandez Adrian F , Patterson J. Herbert , Spertus John A , Turner Stuart J , Waltman Johnson Katherine , Williams Fredonia B
Published: March 2022

Background:
Efficient heart failure management can minimize symptoms, optimize clinical outcomes and lower costs. Novel heart failure treatments can provide an important opportunity to better understand the patterns by which beneficial therapies are adopted. To date, there have been few formal studies quantifying the underlying patient and physician factors associated with the integration of novel therapies into care.

Methods:
This study is an observational, nonrandomized, multi-center US cohort study of adult patients with HFrEF. The goal is to consecutively recruit an estimated 5000 patients with HFrEF at approximately 150-200 US clinical sites. The primary objective of this study is to better understand contemporary heart failure treatment patterns, as well as the reasons and barriers for treatment changes in patients with HFrEF. Secondary outcomes include healthcare provider- and patient-reported decision-making factors and perceptions of treatment, patient-reported heart-failure-related quality of life, and health care resource utilization. The primary analysis of study outcomes - changes in chronic heart failure therapy - will occur at 12 months. Patients will be followed through a maximum duration of 24 months or until death or study withdrawal, with patient-reported data and healthcare resource utilization collected at 30 days, 3 months, 6 months, 12 months, 18 months, and 24 months.

Results:
Table 1 lists variables being collected during registry participation.

Conclusion:
This study will provide a unique opportunity to understand the initiation and titration of outpatient medications, adoption of novel therapies, and the reasons for treatment changes and outcomes in US HFrEF patients.
Abstract 257: The Direct Healthcare Expenditures of Aortic Valve Disease: Evidence From US National Survey Data
Authors: Chen Jie , Mallow Peter J , Moore Matthew , Rizzo John A
Published: March 2022

Objectives:
This study quantified the incremental direct healthcare expenditures of aortic valve disease (AVD) to insurers and individuals, stratified by asymptomatic and symptomatic disease status.

Methods:
Using 1996-2011 data from the United States (US) Medical Expenditure Panel Survey (MEPS), a nationally-representative database, this study performed multivariate analyses of the incremental annual healthcare expenditures to insurers and out-of-pocket expenditures for individuals with AVD compared to those with no valvular disease. Healthcare expenditures were reported at the individual and US aggregate levels. Individuals with AVD were identified by International Classification of Disease Codes, 9
th
revision diagnosis codes and stratified as either symptomatic or asymptomatic based on the presence of comorbid conditions. Sensitivity analyses were performed to test the robustness of the results. All expenditure data were adjusted to 2011 using the Medical Care component of the Consumer Price Index (CPI).

Results:
The MEPS database included 148 individuals with AVD. Asymptomatic individuals comprised 64% (95) of the population. Asymptomatic AVD individuals incurred $10,816 more overall annual healthcare expenditures per person when compared to those without valvular disease. Symptomatic AVD individuals incurred $12,798 more overall annual healthcare expenditures per person when compared to individuals without valvular disease. For both asymptomatic and symptomatic patients, the majority of direct healthcare costs were borne by the insurer ($10,110 and $12,022, respectively) rather than the individual ($706 and $776, respectively). When aggregated to the US population, AVD accounted for an incremental annual cost of $2 billion. Asymptomatic AVD expenditures were $0.9 billion and symptomatic, $1.1 billion. Sensitivity analysis indicated that the incremental annual expenditures ranged from $1.6 billion to $2.5 billion.

Conclusions:
These findings indicated that the incremental burden of AVD was substantial. Further research is necessary to understand the individual economic burden associated with AVD in those over 65 who comprise the majority of patients and likely a disproportionate share of the US aggregate burden.
Abstract 269: Variation in Cholesterol Documentation and Control in Different Clinics Run by the Same Provider
Authors: Gadde Sushee , Malozzi Christopher , Omar Bassam
Published: March 2022

Background:
Cholesterol level availability during a patient’s clinic visit can enhance decision-making with regards to lipid assessment and therapy, and help achieve therapeutic targets.

Methods:
Clinic encounters from a 2 clinics run by the same cardiologist; a University-based clinic and a Veterans Administration (VA) clinic were compared for availability of lipid profiles and level of lipid control in patients with documented coronary disease.

Results:
There were 93 out of 300 patient encounters (31%) with documented coronary disease in the University-based clinic and 77 out of 134 patient encounters (57%) in the VA-based clinic. University patients with CAD had 61% (57/93) lipid documentation during their encounters, while VA patients had 99% (76/77) documentation. The average total cholesterol and LDL for University patients was 179 +/- 42 mg/dL and 111 +/- 35 mg/dL respectively, compared with 154 +/- 38 mg/dL and 93+/- 35 mg/dL for the VA patients respectively (P < 0.01 for each category). Only 39% (22/57) of the University-based patients achieved < 100 mg/dL, compared with 66% (51/77) of VA patients. Blood pressures, which were consistently documented every clinic visit in both clinics, averaged 128/76 mmHg in the University clinic compared with 131/75 mmHg in the VA clinic, with no statistically significant differences.

Conclusion:
Overall, better control of cholesterol levels is seen in the VA-based clinic compare with the University-based clinic. This may partially be explained by the result availability through a unified computer system, allowing more prompt decision-making with regards to medication changes and intensified dietary counseling during the clinic encounter. This is may not be available in other settings where laboratory tests are performed by a different provider utilizing a different reporting system which may not be readily shared or available to the treating cardiologist. Other factors could include variable compliance with laboratory appointments and access to medications.
Abstract 5: Trends in Hospitalization Rates and Mortality for Acute Myocardial Infarction by Community Income Level: 1999-2013
Authors: Spatz Erica S , Beckman Adam L , Wang Yun , Desai Nihar , Krumholz Harlan M
Published: March 2022

Background:
Over the last decade, acute myocardial infarction (AMI) incidence and mortality have decreased substantially. Yet it is unknown whether these improvements were consistent across communities of different economic status and geographic regions, for which efforts to improve cardiovascular disease prevention and management may have had variable impact.

Objective:
To compare trends in risk-standardized AMI hospitalization and 1-year mortality rates from 1999 to 2013 between counties of low, average, and high median income, and to test the effects of income over time and across the 4 U.S. geographic regions.

Methods:
We performed an observational analysis comparing trends in county-level risk-standardized (age, sex, race) AMI hospitalization and 1-year mortality rates from 1999 to 2013 among Medicare beneficiaries ≥65 years, testing for effects of county-income. Counties were stratified by median income percentile using 1999 U.S. Census Bureau data, adjusted for inflation, as: low (<25th), average (25-75th) or high (>75th) income groups. Using a mixed effects model, we compared trends in AMI hospitalization and mortality rates, stratified by county income. We tested for a lagged effect among low-income counties and compared trends between the 4 U.S. regions.

Results:
Income was significantly associated with AMI hospitalization, but not 1-year mortality rates. In the 15-year period, AMI risk-standardized hospitalization and mortality rates declined across all three county income groups, though absolute hospitalization rates differed significantly by county income groups (Figure). Low-income county hospitalization rates consistently lagged behind high-income county rates by 4.3 (95%CI: 3.1-5.9) years. There was no difference in rate of decline by geographic region.

Conclusion:
In this national study of AMI trends from 1999 to 2013, Medicare beneficiaries residing in counties of all income levels achieved substantial declines in AMI hospitalization and mortality. These broad-scale declines may reflect the success of focused AMI prevention efforts or other more secular trends. Nonetheless, disparities persist with low-income counties lagging behind high-income counties - a focus for future national efforts.
L-NAME, but not Phenylephrine Enhances the Effects of Endogenous Carbon Monoxide on Vascular Tone
in Vivo
and
in Vitro
Authors: Johnson Fruzsina K , Teran Federico J , Coco Kay C , Johnson Robert A
Published: March 2022

4

Vascular endothelium and smooth muscle express heme oxygenase, which metabolizes heme to form biliverdin, free iron and carbon monoxide. Carbon monoxide promotes endothelium-independent vasodilation, but also inhibits nitric oxide formation. We previously found, that heme-derived carbon monoxide dilates isolated gracilis arterioles pretreated with a nitric oxide synthase inhibitor, N
ω
-nitro-L-arginine methyl ester (L-NAME), and lowers blood pressure in hypertensive rat models. This current study examines the hypothesis that nitric oxide modifies the effects of endogenous carbon monoxide on vascular tone both
in vitro
and
in vivo
. For this purpose
in vitro
studies were conducted on rat isolated pressurized first-order gracilis arterioles superfused with oxygenated (14%O
2
/5%CO
2
balanced with N
2
) modified Krebs’ buffer (CaCl
2
1.4mmol/L). A heme oxygenase inhibitor, chromium mesoporphyrin (CrMP, 15μmol/L), constricted gracilis arterioles (Δ-32±6μm, t
1/2max
=5min; n=6, P<.05). This effect was markedly amplified by pretreatment with 1mmol/L L-NAME (Δ-65±11μm; t
1/2max
=5min; n=7; P<.05), but not by preconstriction with 100nmol/L phenylephrine (n=6; P>.05). In addition,
in vivo
studies were performed in anesthetized rats instrumented with flow probes and arterial catheters. In these animals the heme oxygenase inhibitor CrMP (45μmol/Kg,IP) had no significant effect on hindlimb resistance (Δ 2.3±0.1 mmHg/mL/min; n=5; P>.05). But, in animals pretreated with L-NAME (100mg/Kg), CrMP increased hindlimb resistance (Δ10.7 ± 0.7 mmHg/mL/min; n=4; P<.05). In contrast, in animals infused with PE (3μg/min) to increase blood pressure and vascular tone, CrMP had no effect on hindlimb resistance (Δ1.8±0.2 mmHg/mL/min; n=5; P>.05). This study shows the vascular actions of an inhibitor of endogenous carbon monoxide formation are enhanced by an inhibitor of nitric oxide synthase, but not by an alternative vasoconstrictor. These findings suggest the endogenous heme-heme oxygenase-carbon monoxide system exerts vasodilatory influences on basal vascular tone which are uniquely enhanced in the absence of nitric oxide synthesis.
Agmatine Is an Autocrine/Paracrine Inhibitor of Pre-Synaptic Ca
++
Channels and Vasopressin Release in Vasopressinergic Neurons of the Hypothalamo-Hypophyseal Tract
Authors: Wang Gang , Gorbatyuk Oleg S , Regunathan S. , Dayanithi Govindan , Milner Teresa A , Reis Donald J
Published: March 2022

20

Agmatine (decarboxylated arginine), an organic cation, endogenous ligand at imidazoline (I-) and α
2
-adrenergic receptors, antagonist of NMDA receptors and inhibitor of nitric oxide synthase, may be a novel neurotransmitter in mammalian CNS (Reis and Regunathan, TIPS, 2000). To further establish a cellular function of agmatine we investigated whether it is expressed in, and regulates the function of, magnocellular neurons of the hypothalamic paraventricular (PVH)/supraoptic (SON) nuclei in rat. By LM, agmatine-like immunoreactivity (Ag-LI) was present in virtually all magnocellular PVH and SON neurons, where it co-localized with arginine vasopressin (AVP) and oxytocin (OXY). By EM, Ag-LI was present in large dense core vesicles in neurohypophysial nerve terminals (NNTs), known storage sites of the peptides. NNTs acutely isolated from adult rats were dispersed and voltage-activated Na
+
, K
+
, and Ca
++
currents recorded in whole terminals in amphotericin B perforated-patches. Agmatine (40-120 uM) did not affect the voltage-dependent, TTX-sensitive inward Na
+
, the 4-AP-sensitive, transient outward A-type K
+
current, or the sustained, Ca
++
-activated, maxi-conductance K
+
currents. In ∼ 60% of terminals 40 uM agmatine immediately, dose-dependently and reversibly blocked whole cell Ca
++
currents evoked by 10 mM Ca
++
with an IC
50
∼ 5 uM. Agmatine blocked the depolarization-induced release of AVP from isolated NNTs. We conclude that agmatine: (a) is co-stored with neuropeptides in hypothalamo-neurohypophysial neurons, and (b) inhibits pre-synaptic Ca
++
channels to (c) inhibit release of AVP. Agmatine is a neurotransmitter in AVP neurons of PVH and SON, wherein it functions as a paracrine/autocrine inhibitor of AVP release by blocking Ca
++
channels. Agmatine may regulate neurally integrated release of this important hormone, regulating its cardiovascular and renal functions.
Compartmentalization of no in the Juxtaglomerular Apparatus (JGA)
Authors: Ren YiLin , Garvin Jeffrey L , Carretero Oscar A , Ford Henry
Published: March 2022

23

We previously reported that NO generated by the afferent arteriolar (Af-Art) endothelium caused dilatation and that NO produced by nNOS in the macula densa (MD) blunts tubuloglomerular feedback (TGF) via a cGMP-dependent mechanism. Here we hypothesized that a) NO is compartmentalized in the JGA, with endothelial NO only acting on vascular smooth muscle cells and MD NO acting within the MD, and b) all effects of MD NO are mediated by cGMP. We microperfused the Af-Art (at 60 mm Hg) with MD attached. MDs were perfused with solutions containing either 6 mM Na and 3 mM Cl (low NaCl) or 80 mM Na and 77 mM Cl (high NaCl). We first studied the effect of the soluble guanylate cyclase inhibitor LY83583 (1 μM) on TGF when added to the MD perfusate. Increasing the NaCl concentration in the MD perfusate decreased Af-Art diameter by 3.2 ± 0.5 μm. Adding LY83583 to the MD increased TGF response to 6.3 ± 1.1 μm (p<0.031 vs control; n=6). Inhibiting MD guanylate cyclase did not alter the effect of acetylcholine on the preconstricted Af-Art. Next we studied the effect of NO produced by the Af-Art endothelium on TGF. During the control period, the TGF response was 3.0 ± 0.9 μm. After LY83583 was added to the lumen of the Af-Art, TGF was 2.3 ± 0.8 μm (n.s.; n=6). This concentration completely blocked acetylcholine-induced dilatation. Removal of the Af-Art endothelium with an antibody against factor VIII-related antigen and complement did not alter TGF response. Before removing the endothelium, Af-Art diameter decreased from 18.6 ± 1.4 to 16.1 ± 1.1 μm; after removal, Af-Art diameter decreased from 19.1 ± 1.4 to 15.9 ± 1.1 μm (n=6). To determine whether the effect of NO in the MD is entirely mediated by cGMP, TGF was studied after giving a) LY83583 or b) LY83583 plus 7NI. In the presence of LY83583 alone, the TGF response was 3.8 ± 0.8 μm. In the presence of LY83583 plus 7NI, the TGF response was 4.2 ± 1.6 μm (n.s.; n=6). We concluded that: 1) NO is compartmentalized in the JGA, and therefore MD NO acts upon the MD itself rather than diffusing to the Af-Art; and 2) all of the effects of NO in the MD are mediated by cGMP.
Small Artery Remodeling Is the First Manifestation of Target Organ Damage in Mild Essential Hypertension.
Authors: Park Jeong Bae , Schiffrin Ernesto L
Published: March 2022

35

Blood vessels participate in the development and maintenance of high blood pressure (BP), and play a role in its complications. We evaluated the prevalence of structural and functional alterations of resistance arteries and cardiac hypertrophy in mild essential hypertensive patients. Thirty-eight patients (47±1 years; 71% male; clinic BP 148±2/99±1mmHg) were compared to 10 age-matched normotensive controls (44±3 years; 40% male; BP 113±4/76±2mmHg). Arteries were studied on a pressurized myograph. Renal function was normal in all subjects. Remodeling and growth indexes of vessels were 72% and 28%, respectively. The prevalence of abnormal structure (media to lumen ratio), endothelial function (maximal acetylcholine response) and cardiac hypertrophy by ECG was 97%, 58%, and 47% (greater than mean+1SD of controls), and 63%, 34%, and 26% (greater than mean+2SD of controls), respectively. Geometry-independent vascular stiffness (the slope of incremental elastic modulus vs. media stress) was similar in both groups. By multivariate analysis, structural alterations correlated with mean BP (p<0.01). Mean BP correlated with endothelial function (p<0.01), structure (p<0.05) and cardiac hypertrophy (p<0.05). Endothelial function was impaired in the highest tertile of vascular stiffness compared to the lowest tertile (p<0.01). In conclusion, structural alterations were demonstrated in most mild hypertensive patients, followed in frequency by endothelial dysfunction and finally cardiac hypertrophy. Small artery vascular remodeling precedes other clinically relevant manifestations of target organ damage in mild essential hypertension.
Angiotensin II Increases Enos Expression in the Rat Myocardium Independent of Pressor Effect.
Authors: Moreno Heitor , Tambascia Rosa C , Sampaio Rita
Published: March 2022

P5

There is accumulating evidence that nitric oxide antagonizes angiotensin II (AII) effects at multiple regulatory levels of the cardiovascular system. In the present study, we first determined the expression of endothelial (eNOS) and neuronal (nNOS) isoforms of nitric oxide synthase (eNOS) in the ventricles of two kidney Goldblatt(2K1C, 2 months) and L-NAME treated (LN, 2 months)hypertensive rats with Western blot. Systolic arterial pressures were 130±3, 165±4, 160±5 mmHg in control (n=7) and 2K1C (n=6)and LN treated (n=6) rats, respectively. nNOS was barely detected in ventricles of normotensive and 2K1C and LN hypertensive rats. In contrast, eNOS was detected in ventricles of all groups of rats and was increased by 100±8% and by 88±12%in 1K1C and LN rats. Experiments performed with blocking peptide (100:1; blocking peptide to antibody proportion) indicated that the ∼130 kDa bands identified in membranes blotted with anti-eNOS antibody was specific for eNOS. Another group of rats was continuously infused with AII (D1= 10, n=5; D2= 40, n=4 ng/kg/min) for 6 days and arterial pressure measured daily (1 h, continuous, sampled at 100 Hz). Arterial pressure did not change significantly during infusion of D1 and increased by 22±3 mmHg during D2 infusion. Both, D1 and D2 increased eNOS expression by ∼95% while, again, no change was observed in nNOS expression. These data indicate that 1) eNOS expression is up-regulated in 2K1C and LN hypertensive rats; 2) AII infusion enhances the eNOS expression by a level similar to that observed in 1K1C and LN hypertensive rats; and 3) The influence of AII on eNOS expression is independent of its pressor effect. In conclusion,our data suggest that AII increases the expression of eNOS, independent of the its hemodynamic effect and could be responsible for the increase in eNOS expression in 1K1C hypertensive rats. In addition, this modulation is specific for eNOS since nNOS is present only in small amounts in the rat myocardium and is not modulated by AII. The mechanism responsible for the increase in eNOS expression in LN rats is not clear, but could be related to a modulation mediated by a product derived from eNOS action on its substrate.
Partial Tissue-Specific Expression of Two Human Angiotensinogen Promoter-Lacz Transgenes in Mice
Authors: Cvetkovic Branimir , Yang Gongyu , Sigmund Curt D
Published: March 2022

P13

Previous analysis of transgenic mice carrying a genomic construct encoding the human angiotensinogen (HAGT) gene indicated that the 13.7 Kb sequence harbored all
cis
elements required for the appropriate tissue and cell expression of HAGT
in vivo
. In addition, analysis of independently derived HAGT transgenic lines suggested the presence of a locus control element (LCR) in the HAGT gene since the expression levels were proportional to transgene copy number. In attempt to identify
cis
elements responsible for the regulation of HAGT expression
in vivo
we created several new transgenic mouse lines carrying either 1.5 Kb of the HAGT promoter or 1.5 kb of the promoter along with the first exon and intron of the HAGT gene fused to the LacZ reporter gene. Reporter gene expression did not correlate with transgene copy number suggesting the absence of a putative LCR element in the 1.5 Kb region proximal to the HAGT transcription start site. LacZ expression was most prominent in liver and adrenal gland, with additional expression in heart, testis, brain, and cerebellum. Most other tissues (including kidney) had little to no expression of the reporter gene. Our results indicate that the regions proximal to the HAGT transcription start site contain
cis
-acting

elements important for appropriate HAGT expression in the liver and some, but not all, extra-hepatic tissues which should express HAGT. Further dissection of the genomic HAGT sequence will be required to identify all elements involved in proper
in vivo
regulation of the HAGT gene.
Role of Enothelin B Receptors in Enhancing Endothelium-Dependent No-Mediated Vascular Relaxation During High Salt Diet
Authors: Giardina Jena B , Green Gachavis M , Rinewalt Anna N , Granger Joey P , Khalil Raouf A
Published: March 2022

P21

Basal stimulation of endothelial endothelin B (ET
B
) receptors and the ensuing release of endothelium-derived relaxing factors has been hypothesized to protect against excessive vasoconstriction and increased blood pressure particularly during high salt diet. To test this hypothesis, we investigated whether chronic infusion of ET
B
receptor antagonist is associated with impaired endothelium-dependent vascular relaxation and enhanced vascular reactivity particularly with high salt diet. Active stress was measured in aortic strips isolated from male Sprague-Dawley rats on normal sodium (NS, 1%) and high sodium diet (HS, 8%) for 7 days and untreated or infused with the ET
B
receptor antagonist A-192621 (30 mg/kg/day) for 5 days. In endothelium-intact strips, phenylephrine (Phe,10
-5
M) increased stress to 6.9±.3 in NS and to 7.2±.4x10
3
N/m
2
in HS rats. The Phe-induced stress in NS/ET
B
(8.6±.5) and HS/ET
B
rats (12.5±.4x10
3
N/m
2
) was significantly > NS and HS rats. Removal of the endothelium significantly enhanced Phe-induced stress in NS and HS, and only slightly in NS/ET
B
but not in HS/ET
B
rats. In endothelium-intact strips, acetylcholine (ACh) induced relaxation of Phe contraction was in NS > HS > NS/ET
B
> HS/ET
B
rats with an ED
50
of 0.3x10
-6
, 0.4x10
-6
, 0.7x10
-6
and 1.1x10
-6
M, respectively. Pretreatment of endothelium-intact strips with L-NAME (10
-4
M), to inhibit nitric oxide (NO) synthase, or methylene blue (10
-5
M), to inhibit cGMP production in smooth muscle, significantly inhibited ACh-induced relaxation and enhanced Phe-induced stress in NS and HS, and only slightly in NS/ET
B
but not in HS/ET
B
rats. Measurement of basal and ACh-induced nitrite/nitrate production from aortic strips showed a significant decrease in NS/ET
B
and HS/ET
B
compared to NS and HS rats. Relaxation of Phe contraction with sodium nitroprusside was not significantly different among the different groups of rats. Thus, an endothelial ET
B
receptor-mediated pathway of vascular relaxation involving release of NO is active under basal conditions and may protect against excessive vasoconstriction and increased blood pressure particularly during high salt diet.
Gaba
b
Receptor Mediated Responses in the Nucleus Tractus Solitarius (NTS) Are Altered in Acute and Chronic Hypertensive Rats.
Authors: Mifflin Steven W , Vitela Melissa
Published: March 2022

P47

In renal wrap chronic hypertensive (CHT) rats, injection of the GABA
B
agonist baclofen into the NTS evokes a greater increase in mean arterial pressure (MAP) than in normotensive (NT) controls, and in CHT rats there is an increased expression of GABA
B
receptor mRNA in the NTS (
Hypertension,
33:530). We report that the pressor response to injection of baclofen into the NTS is enhanced following acute hypertension (AHT) of only 30 minutes. Sprague-Dawley rats were anesthetized with Inactin (60mg/kg), paralyzed and artificially ventilated. Following unilateral electrolytic ablation of the NTS, microinjection of 40pmoles of baclofen into the contralateral NTS of NT (101±2mmHg) rats resulted in an increase in MAP of 21±1mmHg (n=7). During AHT (30 min of phenylephrine infusion, MAP increased to 135±3mmHg), microinjection of baclofen increased MAP by 34±2mmHg (n=13). This was significantly greater than the response before AHT (p<.01) and no different from the response in CHT rats (37±3mmHg, n=8, p>.80). Intravenous injections of norepinephrine (n=3) and angiotensin II (n=3) produced the same increases in MAP before and during AHT (p>.4) so the enhanced baclofen response is not due to a change in vascular reactivity. Baclofen has both pre- and post-synaptic effects. To eliminate the pre-synaptic component of the baclofen response sino-aortic denervations (SAD) were performed prior to the microinjections. In NT-SAD rats, baclofen injections increased MAP by 12±1mmHg (n=8), in AHT-SAD by 12±1mmHg (n=12)and in CHT-SAD rats by 20±3mmHg (n=7). To conclude, the pressor response to NTS injections of baclofen is enhanced in both CHT and AHT rats. The increase in baroreceptor afferent input to NTS during phenylephrine induced AHT provides a greater substrate for pre-synaptic inhibition by baclofen since the post-synaptic component of the baclofen response is the same in NT-SAD and AHT-SAD. The enhanced pressor response in CHT rats is associated with an increased post-synaptic component of the baclofen response, consistent with our previous finding of an increase in GABA
B
receptor mRNA in the NTS in CHT rats.
Low-Frequency Components of Renal Sympathetic Nerve Activity Are More Tightly Coupled with Blood Pressure and Renal Blood Flow in Conscious Hypertensive Rats
Authors: Sakata Katsufumi , Kumagai Hiroo , Osaka Motohisa , Matsuura Tomokazu , Oshima Naoki , Hayakawa Hirokazu , Saruta Takao
Published: March 2022

P52

We examined how various frequency components of renal sympathetic nerve activity (RSNA) regulate blood pressure (BP) and renal blood flow (RBF) in conscious Wistar-Kyoto rats (WKY, n=7) and spontaneously hypertensive rats (SHR, n=7) of 15 wk old. BP, heart rate, RSNA, and ipsilateral RBF were simultaneously recorded. Signals were sampled at 2 kHz to A/D converter for 6.50 min and the data were assessed by power spectral analysis. RSNA was augmented in SHR compared with WKY despite higher BP. Peaks of RSNA obtained by power spectral analysis were found at frequencies of 0.05, 0.80, 1-2, 6-8 Hz. RSNA power at 0.8 Hz were much larger in SHR than in WKY. Peaks of coherence of cross-spectra between BP (input) and RSNA (output), and between RSNA (input) and RBF (output) were found at frequencies of 0.05 and 0.8 Hz both in WKY and SHR. The coherence of cross-spectrum between BP and RSNA at 0-1.0 Hz was significantly higher in SHR than that in WKY, and the transfer gain of BP to RSNA was significantly larger in SHR. Similarly, the coherence between RSNA and RBF at 0-1.0 Hz in SHR was significantly higher, whereas the transfer gain of RSNA to RBF was significantly smaller in SHR. Intravenous infusion of L-arginine (50 μmol/kg/min for 60min) decreased the coherence between BP and RSNA or between RSNA and RBF to less than 0.5 in SHR. Finally, we found that low-frequency components of RSNA < 0.1 Hz were inversely related to BP and RBF, suggesting that the baroreflex system exists in the low frequency band. These results suggest that low-frequency components of RSNA are important in regulating BP and RBF. The higher coherence shown in SHR may imply that RSNA is more tightly coupled with BP and RBF. The results using L-arginine infusion suggest that endogenous NO system is important for attenuating the tight coupling (higher coherence) between the sympathetic nervous system and BP or RBF.
Cardiac Function in Endothelial Nitric Oxide Synthase Knockout Mice with Heart Failure
Authors: Liu Yun-He , Xu Jiang , Yang Xiao-Ping , Cavasin Maria A , Shesely Edward G , Carretero Oscar A
Published: March 2022

P57

Nitric oxide(NO)released by endothelial NO synthase(eNOS)is impaired during the development of heart failure (HF). We tested the hypotheses that 1)lack of eNOS (-/-) may affect cardiac function and remodeling after MI induced by coronary ligation; and 2) the cardioprotective efffect of ACEi will be lessened or absent in -/- mice. 1 month after MI, mice were treated with either vehicle or the ACEi enalapril (20 mg/kg) and this was continued for 3 months. C57BL6J (+/+) mice were used as controls. LV ejection fraction (LVEF), shortening fraction (SF), LV mass (LVm) and diastolic dimension (LVDd) were evaluated by echocardiography before and after MI. We found that LVEF (figure) and SF were decreased, and LVDd and LVm increased significantly after MI and slowly progressed with time. No difference was detected between the two strains. ACEi improved LVEF in +/+, but not in -/-; and they also decreased LVm and LVDd, and these effects were diminished in -/- mice. Our data suggest that eNOS may not play an important role in the development of HF; however, it does participate in the cardioprotective effects of ACEi.
Human Tissue Kallikrein Gene Delivery Attenuates Cardiac Glycogen Accumulation in Streptozotocin-Induced Diabetic Rats
Authors: Montanari David , Dobrzynski Eric , Chao Julie , Chao Lee
Published: March 2022

P58

The tissue kallikrein-kinin system is locally present in the heart and is reduced under diabetic conditions. In the present study, we explored the effects of kallikrein gene delivery in the heart under diabetic conditions. Intravenous injection of streptozotocin (STZ) (60mg/kg) into 8-week-old male Sprague-Dawley rats produced hyperglycemia (460 ± 10 mg/dl, n=30). Three weeks after STZ treatment, adenovirus containing the human tissue kallikrein gene (Ad.CMV-cHK) or green fluorescence protein gene (Ad.CMV-GFP) under the control of CMV promoter/enhancer were injected via the tail vein into STZ-diabetic rats. Adenovirus-mediated kallikrein gene delivery caused a significant reduction of blood pressure as compared to control rats receiving control virus (170.8 ± 3.5 vs. 187.4 ± 4.2 mmHg, n=8, P<0.01). Rats were sacrificed and tissues were collected for analysis at 16 days after gene delivery. Histological examination of heart sections stained with periodic acid-Schiff (PAS) showed marked accumulation of glycogen in cardiomyocytes of diabetic rats. Kallikrein gene delivery significantly reduced cardiac glycogen accumulation as compared to rats receiving control adenovirus. Quantitative analysis confirmed the morphological observation that kallikrein gene delivery significantly reduced cardiac glycogen levels as compared to control diabetic rats injected withAd.CMV-GFP (0.5 ± 0.07 vs. 2.15 ± 0.83 mg glycogen/mg protein, n=6, P<0.05). Cardiac cAMP levels were significantly increased after kallikrein gene delivery. This is the first study to demonstrate that kallikrein gene delivery exerts a protective effect in the reduction of glycogen accumulation in cardiomyocytes of STZ-induced diabetic rats.
Retinoid Ligands Inhibit Cell Proliferation by Modulating Cell Cycle Regulators in Human Coronary Smooth Muscle Cells
Authors: Wakino Shu , Kintscher Ulrich , Kim Sarah , Nagpal Sunil , Hsueh Willa A , Law Ronald E
Published: March 2022

P74

Retinoids inhibit vascular smooth muscle cell proliferation in vitro and intimal hyperplasia in vivo in rats. We examined the underlying mechanism of the antiproliferative effect of retinoids on human coronary smooth muscle cells (CASMC). The retinoids tested included: RAR ligands, all-trans retinoic acid (atRA) and Ethyl-p-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-l-propenyl]-benzoic acid (TTNPB); a RXR and RAR ligand 9-cis retinoic acid (9cRA); and the potent RXRα- ligand AGN4204. All retinoids inhibited DNA synthesis in CASMC stimulated with platelet derived growth factor and insulin (IC
50
: TTNPB 0.063μM, atRA 0.123μM, AGN4204 0.461μM, 9cRA 1.529μM, all p<0.01, n=6). All retinoids substantially attenuated retinoblastoma protein phosphorylation which is required for CASMC proliferation (maximal inhibition: TTNPB and atRA,
2x10
-7
M; AGN4204 and 9cRA, 2x10
-6
M all p<0.01, n=3). None of the retinoids affected the expression of CDKs 2, 4, or 6 or cyclin E. TTNPB, atRA and AGN4204 inhibited the mitogenic induction of cyclin D1 (maximal inhibition: TTNPB, 2x10
-9
M; atRA, 2x10
-7
M; AGN4204, 2x10
-6
M, all p<0.01, n=3), whereas 9cRA had no effect (n=3). All retinoids significantly attenuated mitogen-induced downregulation of CDKI p27
Kip1
, a major negative regulator of G1 cyclin/CDK activity (maximal effect: TTNPB and atRA, 2x10
-7
M; AGN4204 and 9cRA, 2x10
-6
M, all p<0.01, n=3). The present study demonstrate that retinoids have antiproliferative activity by modulating the G1→S cell cycle regulators in human CASMC, which may provide a novel therapeutic target for proliferative vascular diseases.
Ang II Infusion at 8, But not at 16 Weeks Age, Promotes Atherosclerosis in ApoE-Deficient Mice
Authors: Catanzaro Daniel F , Zhou Ying , Shi Yuexian , Chen Rong
Published: March 2022

P77

Although hypertension has long been regarded as a risk factor for atherosclerosis, the pathophysiologic mechanism is unknown. Angiotensin II can affect almost every step in the development of atherosclerosis. A number of recent studies show that angiotensin blockade reduces, and angiotensin infusion enhances atherogenesis in hyperlipidemic, ApoE-deficient mice. However, the relative importance and the time course of the processes involved remains to be elucidated. In the present study, we examined the effects of Ang II on the development of aortic root atherosclerosis in male ApoE-deficient mice. Osmotic minipumps containing saline or Ang II were implanted subcutaneously in 8 or 16 week old mice. Ang II was infused at a rate of 1000 ng/kg/min for 28 days. In all mice infused with Ang II, systolic blood pressure (SBP) measured by tail cuff increased significantly. Accordingly, plasma renin concentration (PRC) in these mice was suppressed to virtually zero. In mice in which infusion was commenced at 16 weeks age there was no increase in atherosclerotic lesion area. However, in mice in which the infusion was commenced at 8 weeks age, lesion area increased 2.3-fold (p<0.05, unpaired t-test). Total and HDL cholesterol levels were unaffected by Ang II treatment. Although, in the 16-week-old animals, there was considerable atherosclerosis, it had not reached maximum levels. Taken together, these observations suggest that Ang II may have its greatest effects on lesion initiation.
Dopamine as a Novel Antimigration Factor of Vascular Smooth Muscle Cells Through D
1a
and D
1b
Dopamine Receptors
Authors: Yasunari Kenichi , Kohno Masakazu , Minami Mieko , Maeda Kensaku , Hanehira Takao , Junichi Junichi
Published: March 2022

P82

Vascular smooth muscle cell (VSMC) migration is believed to play a key role in atherosclerosis. To elucidate the roles of rat vascular D
1A
and D
1B
dopamine receptors in atherosclerosis, the effect of antisense oligonucleotides (AS) to D
1A
(+1-+21 of rat D
1A
receptors cDNA) and D
1B
(-12-+6 of rat D
1B
receptors cDNA) receptors on dopamine-mediated suppression of platelet-derived growth factor (PDGF) BB-mediated VSMC migration evaluated by Boyden′s chamber method was studied. To avoid the degradation, phosphothioate-modified oligodeoxynucleotides were synthesized and purified by high-performance liquid chromatography. These oligonucleotides were added to serum free medium for 24 h before the start of PDGF BB stimulation with transfection using lipofectin reagent. Immunohistochemical studies (double staining) demonstrated the coexistence of D
1A
and D
1B
dopamine receptors in single VSMC derived from rat renal artery. Western blotting revealed a band of approximately 70 kD for D
1A
and D
1B
dopamine receptors. Increased VSMC migration by PDGF BB 5 ng/ml (16-fold) was suppressed significantly by coincubation with dopamine 0.025-10 μmol/l (15-59%). This suppression by dopamine 10 μmol/l was reversed by D
1A
AS 46% and D
1B
AS 51%, but by neither sense (S) nor scramble (RS) oligonucleotides to these receptors. These suppression by antisense oligonucleotides (21-51%) are dose-dependent (1-10 μmol/l) and time-dependent (0-4 hrs). Dopamine 10 μmol/l-induced cyclic AMP formation is also suppressed by D
1A
AS 50% and D
1B
AS 58%, but by neither S nor RS to these receptors. PDGF BB (5 ng/ml)-mediated activation of phospholipase D (PLD) activity (107%) measured by [
3
H]-ethanolamine and protein kinase C (PKC) activity (111%) activities measured by synthetic peptide phosphorylation were significantly suppressed by coincubation with dopamine 10 μmol/l (48%, 49%), which was reversed by D
1A
AS 45% and D
1B
AS 50%, but by neither S nor RS to these receptors. These results suggest that vascular D
1A
and D
1B
receptors inhibit migration of VSMC, possibly through cAMP formation and suppression of PLD and PKC activity.
Estradiol and Its Metabolites Differentially Induce NO Synthesis by Human Glomerular Endothelial Cells and Inhibit Glomerular Mesangial Cell Growth
Authors: Xiao Shen , Gillespie Delbert G , Baylis Christine , Jackson Edwin K
Published: March 2022

P93

Reduced NO synthesis by glomerular endothelial cells (GECs) and increased proliferation of glomerular mesangial cells (GMCs) is associated with glomerular remodeling process leading to glomerosclerosis. Moreover, estradiol (E) slows the progression of renal disease. Because E induces NO synthesis and E metabolites are as potent as E in inhibiting growth of vascular smooth muscle cells, which are phenotypically similar to GMCs, we investigated whether E and its major endogenous metabolites (2-hydroxyestradiol[2OHE] and 2-methoxyestradiol [2MeOE])inhibit growth of GMCs and induce basal NO synthesis by GECs. E and its metabolites (1nM-10μM)inhibited serum (2.5%)-induced DNA synthesis, cell number and collagen synthesis in human GMCs concentration-dependently,and with the order of potency being 2MeOE > 2OHE > E. ICI182780, an estrogen receptor antagonist (100μM) blocked the growth inhibitory effects of E, but not 2OHE and 2MeOE. Treatment with E, but not 2OHE and 2MeOE, induced NO synthesis (P<.05; assayed by the formation of
3
H-L-citrulline from
3
H-L-arginine) in human GECs, and these effects were blocked by ICI182780 and L-NMA (NO synthesis inhibitor).In conclusion, E may protect glomerosclerosis by inducing NO synthesis via an estrogen receptor-dependent mechanism and via conversion to 2OHE and 2MeOE, which inhibit GMC proliferation independently of the estrogen receptor.
The Role of Cationic Amino Acid Transporters in the Regulation of Blood Flow and Nitric Oxide in Rat Kidney
Authors: Kakoki Masao , Mattson David L
Published: March 2022

P94

Previous studies from our laboratory have indicated that nitric oxide (NO) production by isolated inner medullary collecting duct cells is dependent on cellular uptake of L-arginine via cationic amino acid (CAT) transporters (
FASEB J
13:A723,1999). In the present study we performed experiments to determine if CAT transport regulates NO production and renal hemodynamics in the renal medulla
in vivo
. The effects of renal medullary interstitial infusion of cationic amino acids (100 μg/kg/min) on renal medullary blood flow and NO levels were examined in anesthetized Sprague-Dawley rats by laser-Doppler flowmetry and a microdialysis-oxyhemoglobin trapping technique. Furthermore, we studied the effect of chronic intramedullary infusion of L-ornithine (50 μg/kg/min) on arterial pressure in uninephrectomized SD rats. Renal medullary blood flow was significantly reduced during the acute intramedullary infusion (30 min) of the cationic amino acids L-ornithine (-24±3%, N=4), L-lysine (-20±1%, N=4), and L-homoarginine (-21±2%, N=4), and during infusion of the NOS inhibitor N
ω
-nitro-L-arginine methyl ester (L-NAME; -38±4%, N=6). In contrast, infusion of L-arginine significantly increased medullary blood flow (+17±4%, N=4) and the neutral amino acids L-leucine, L-serine, and L-glycine (N=4 each) had no effect on renal medullary blood flow. NO concentration in the renal medulla was significantly decreased by the intramedullary infusion of L-ornithine (for 60 min; 336 ± 25 vs 232 ± 34 nmol/L, N=6, p<0.05). In contrast, NO concentration in renal cortex was not significantly affected by the intracortical infusion of L-ornithine (227 ± 31 vs 240 ± 60 nmol/L, p=NS). Similar changes in NO were observed with medullary and cortical infusion of L-lysine. Finally, chronic infusion of L-ornithine into the renal medulla of uninephrectomized SD rats for 7 days led to a sustained increase in mean arterial pressure (168±5 mmHg, N=5) when compared to vehicle treated rats (125±3 mmHg, N=6). These results suggest that CAT transport in the renal inner medulla may play an important role in the production of NO, the maintainence of blood flow, and the level of mean arterial pressure.