Dietary Prevention of Heart Failure in Hypertensive Metabolic Syndrome
Phase: NA
Conditions: Heart Failure
Sponsor: VA Office of Research and Development

Dietary Prevention of Heart Failure in Hypertensive Metabolic 
Syndrome 
 
 
NCT03170375 
 
 
 
 
 
2/25/2022

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Dietary Prevention of Heart Failure in Hypertensive Metabolic Syndrome 
Principal Investigator: Scott Hummel, MD 
 
BACKGROUND AND SIGNIFICANCE 
Burden of HF in Veterans: In 2014, veterans with heart failure (HF) accounted for nearly 1,200,000 VA 
outpatient visits and 40,000 primary-diagnosis hospitalizations, and decompensated HF remains the most 
common cause for hospital admission in the veteran population (Heidenreich 2016). Patients with HF suffer 
functional decline and poor quality of life, and half die within 5 years after diagnosis.(Owan et al. 2006). Within 
the VA system, mean annual costs per patient with HF were $30,719 in FY 2010 (Yoon et al. 2016). The 
overall societal burden of HF is increasing; by 2030, over 8 million Americans will have heart failure (HF), and 
will account for $70 billion/year in medical costs (Heidenreich et al. 2013). The burden and costs of incident HF 
will proportionally fall even more heavily upon the VA, since HF risk factors are more frequently present in 
veterans than in non-veterans even when matched for age (Fryar et al. 2016). These observations highlight the 
crucial role of HF prevention within the overall vision of the Veterans Health Administration to “emphasize 
prevention and population health.”  
In the more than 10,000 veterans hospitalized with new-onset HF each year, nearly 80% have systemic 
hypertension (HTN) (Heidenreich et al. 2010). HTN confers the highest population-attributable risk for incident 
HF in patients without previous myocardial infarction (Gottdiener et al. 2000), particularly when accompanied 
by the metabolic syndrome, a constellation of obesity, insulin resistance, and dyslipidemia (Nicklas et al. 2006). 
In turn, HTN and the metabolic syndrome are associated with cardiovascular abnormalities that promote HF, 
including left ventricular hypertrophy and diastolic dysfunction, increased large-arterial stiffness, and impaired 
ventricular systolic reserve (Wang et al. 2015). More than half of veterans have HTN, over 40% are obese, and 
at least 25% have metabolic syndrome; the prevalence of all of these factors is increasing over time (Fryar et 
al. 2016), and it remains unclear why only some persons in this large at-risk group develop HF. This critical 
knowledge gap impairs the development of targeted preventive strategies for HF in veterans.  
Heart failure with preserved ejection fraction, a multifactorial entity:  In older adults with HTN and 
metabolic syndrome, HF most commonly develops in the setting of preserved left ventricular ejection fraction 
(HFpEF), i.e. normal or only mildly reduced chamber contractile function (Kitzman et al. 2001). HFpEF is more 
common in women than men, but nonetheless already constitutes nearly one-third of HF in veterans (Ather et 
al. 2012). As the number of female veterans increases and the overall veteran population ages, the incidence 
of HFpEF will likely increase over time. In recent years, the mechanistic understanding of this condition 
previously termed ‘diastolic’ HF has evolved. While ventricular diastolic dysfunction does predict incident 
HFpEF and increases mortality in existing HFpEF (Persson et al. 2007, Lam et al. 2011), HFpEF is now 
appreciated as a failure of cardiovascular reserve in multiple domains (Borlaug et al. 2010), Figure 1.  
For example, increased large-arterial stiffness is frequently 
present in and can help diagnose HFpEF (Weber et al. 2013). 
Large-arterial stiffness exacerbates the effects of impaired 
systolic and diastolic functional reserve on cardiac work 
efficiency and filling pressures (Kawaguchi et al. 2003), and is a 
key determinant of exercise intolerance in HFpEF (Kitzman et 
al. 2013). Despite normal ejection fraction, global longitudinal 
ventricular strain, a more sensitive measure of systolic function 
than ejection fraction, progressively decreases across age-
matched controls, hypertensives with diastolic dysfunction, and 
HFpEF (Kraigher-Krainer et al. 2014); low longitudinal strain 
also predicts hospitalization and death in HFpEF (Shah et al. 
2015). Left atrial enlargement and dysfunction provide important 
diagnostic and prognostic information for HFpEF (Obokata et al. 
2013, Freed et al. 2016). Recent data demonstrate microvascular oxidative stress and inflammation as well as 
coronary and peripheral microvascular dysfunction in human HFpEF, which suggests a systemic syndrome 
Figure 1: Multiple contributors to HFpEF 

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induced by multiple comorbid conditions rather than a solely cardiac insult (Paulus et al. 2013, Franssen et al. 
2015, Maréchaux et al. 2016). An effective preventive strategy for HFpEF should incorporate this complexity. 
Dietary sodium restriction to prevent hypertensive heart disease: one size fits all? Dietary sodium 
restriction to prevent HTN-associated cardiovascular disease, longstanding conventional wisdom based on its 
blood pressure (BP) lowering effects, has recently become highly controversial (Oparil 2014). Despite its 
broad-based recommendation, the potential exists for harm related to neurohormonal activation during sodium 
restriction.  In prominent publications from large cohort studies, low dietary sodium intake has been associated 
with both increased and decreased long-term cardiovascular mortality (Mozaffarian et al. 2014, Mente et al. 
2016). With regard to incident HF, the findings are similarly mixed, and depend on the population studied and 
the approach used to assess dietary sodium content (Konerman et al. 2016). The methodological challenges in 
linking sodium intake to cardiovascular disease in cohort studies, including systematic measurement errors in 
sodium intake, reverse causality, and residual confounding, are well-described (Cobb et al. 2014). The Institute 
of Medicine recently concluded that the “paucity of evidence… strongly points to the need for further research 
to better define relationships between sodium intake and [cardiovascular disease] risk... (Institute of Medicine 
2013)”  
Just as importantly, this epidemiological debate obscures the possibility of individual differences in response to 
sodium intake. Controlled studies in humans have identified a salt-sensitive blood pressure (BP) phenotype, 
i.e., persons in whom BP is substantially higher during high- than low-sodium intake, and conversely those with 
salt-resistant BP, whose BP responses to changes in sodium intake are negligible. Risk factors for the salt-
sensitive phenotype include age, HTN, obesity, and the metabolic syndrome. Independent of baseline BP or 
previously diagnosed HTN, the salt-sensitive BP pattern triples the risk for cardiovascular morbidity and nearly 
doubles the long-term mortality hazard (Morimoto et al. 1997, Weinberger et al. 2001). The American Heart 
Association recently affirmed salt-sensitivity as an independent cardiovascular risk factor with important public 
health implications, and recommended further study of this phenomenon (Elijovich et al. 2016).   
 
Salt sensitivity: a novel HF risk profile? One of the most extensively studied animal models of HFpEF is the 
Dahl S (salt-sensitive) rat, an inbred model with hypertensive response to high dietary sodium intake. In the 
Dahl S rat, BP increases after 1-2 weeks of 8% sodium consumption. By 12 weeks of age, left ventricular 
hypertrophy, diastolic dysfunction, and increased arterial stiffness are present. By 16-20 weeks of age, HF 
becomes evident with increased left ventricular end-diastolic pressure and lung weight (Klotz et al. 2006). Salt-
sensitive animal models of metabolic syndrome demonstrate similar cardiovascular phenotypes (Matsui et al. 
2008, Franssen et al. 2015). The mechanisms are remarkably consistent across a broad variety of salt-
sensitive animal models: high sodium intake drives oxidative stress, microvascular inflammation, and 
cardiovascular injury; low sodium intake largely prevents such damage (Rodriguez-Iturbe et al. 2004). 
Data from human endomyocardial biopsies strongly imply that oxidative stress and microvascular inflammation 
promote the development of HFpEF in the setting of HTN and the metabolic syndrome (Franssen et al. 2015). 
In hypertensive humans, increased systemic oxidative stress is associated with reduced exercise capacity and 
HFpEF-related cardiovascular changes, including left ventricular hypertrophy, diastolic dysfunction, and arterial 
endothelial dysfunction (Giner et al. 2005, Dekleva et al. 2007). Dietary sodium restriction in salt-sensitive 
persons reduces systemic oxidative stress, but increases it in salt-resistant persons, despite causing similar 
neurohormonal activation in both groups (Laffer et al. 2006, Al-Solaiman et al. 2009). Independent of baseline 
BP, salt-sensitive hypertensives have higher left ventricular mass and larger left atrial size than salt-resistant 
hypertensives (Bihorac et al. 2000). Salt-sensitive hypertensives more commonly have peripheral 
microvascular dysfunction (de Jongh et al. 2007) and microalbuminuria, which worsens during high sodium 
intake (Bigazzi et al. 1994). Left ventricular diastolic function, large-arterial stiffness, and microvascular 
endothelial function are abnormal in salt-sensitive subjects and can improve with dietary sodium restriction 
(Musiari et al. 1999, de Jongh et al. 2007, Todd et al. 2010). These observations suggest that physiological 
differences in the response to dietary sodium could contribute to HFpEF risk.  
Improving phenotyping for salt-sensitivity: At the present time, describing the salt-sensitive phenotype in 
humans requires BP measurement under conditions of high and low-sodium balance (i.e., conditions where 
sodium intake equals excretion) following one or more weeks of constant  sodium intake (Chen et al. 2009), or 

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measuring the BP response to rapid dietary sodium loading then depletion with diuretics over a two-day 
inpatient protocol (Weinberger et al. 2001). Since the BP response to changes in sodium intake is normally 
distributed in the population, arbitrary thresholds for BP change are used to establish the salt-sensitive 
phenotype. Salt-sensitivity was originally defined as a change of ≥10% in mean BP between low- and high-
sodium conditions; other studies have used thresholds of 3-10 mmHg change in mean or systolic BP. 
Whatever threshold is used, both sets of measurements are subject to random fluctuations in BP as well as 
random error in BP assessment that can reduce reproducibility. Automated 24-hour ambulatory BP monitoring 
can reduce this variance and is recommended by experts in the field (Elijovich et al. 2016). Nonetheless, due 
to the need to institute and maintain meticulously controlled diets as an outpatient or measure the inpatient BP 
response to physiological provocation, the salt-sensitive phenotype is challenging to define in clinical practice. 
As recently stated by the American Heart Association, a biomarker for salt-sensitivity is needed and “should 
emerge from… protocols that use arbitrary BP cutoffs to define the phenotype.” Such a biomarker could then 
be “validated by [its] association with characteristics of the phenotype (Elijovich et al. 2016).”  
An ideal test for salt-sensitivity and accompanying HFpEF risk would be 
directly linked to vascular dysfunction, oxidative stress, and 
inflammation, and could identify the at-risk phenotype independent of 
BP changes and without dietary modification. The endothelial glycocalyx 
(eGC) is a gel-like proteoglycan-glycoprotein polymer which lines the 
lumen of blood vessels. The role of the eGC in diverse conditions, 
including HF with acute pulmonary edema, has recently been reviewed 
(Tarbell et al. 2016), and the direct relationship between the eGC and BP 
has previously been demonstrated in humans (Oberleithner 2012). An 
intact eGC serves several critical functions, including limiting immune cell 
interaction with vascular adhesion molecules and transducing shear 
stress into nitric oxide production (Oberleithner et al. 2007, Nijst et al. 
2015).  The eGC buffers circulating sodium cations due to its high 
negative electrical charge density (Oberleithner 2014). This buffering 
slows the entry of sodium (and accompanying water) into the interstitium 
and endothelial cells, which otherwise would directly contribute to edema 
and vascular stiffening, respectively (Oberleithner et al. 2007). Moreover, 
chronic high sodium exposure can damage the eGC (Oberleithner et al. 
2011).  Accordingly, the eGC could modulate both the mechanisms and 
consequences (vascular oxidative stress and infiltration of immune cells, 
hypertension, arterial stiffening, interstitial sodium and fluid retention) of 
salt sensitivity during acute and prolonged high sodium intake (Figure 2).  
 
Endothelial glycocalyx assessment: In vivo assessment of eGC thickness and its relationship with 
microvascular function may be performed with sublingual sidestream darkfield (SDF) microscopy (Goedhart et 
al. 2007). In this technique, a specialized camera uses 540 nm green light-emitting diodes to image red blood 
cells passing through the sublingual microvasculature. Images are collected with a blunt-tipped probe at 4-5 
locations over a 1-2 minute period, then analyzed offline. Software identifies the distribution of viable 
microvascular segments and the percentage of time they are filled with red blood cells (RBC filling %). These 
data correspond to microvascular density and perfusion, respectively. The difference between the overall 
perfused vessel diameter and the median red blood cell column width, the perfused boundary region (PBR), is 
inversely related to the thickness of the eGC in a given vessel segment. In a large population-based cohort, 
Lee et al. confirmed a strong inverse relationship between RBC filling % and PBR (linear regression r2=0.48), 
i.e. that microvascular perfusion was inversely related to eGC thickness (Lee et al. 2014).  
While SDF microscopy enables simultaneous evaluation of microvascular function and eGC thickness, this 
technology is not widely available. However, red blood cells have a very similar glyocalyx layer to the eGC. 
Following enzymatic damage to the eGC, identical changes occur in the red blood cell glycocalyx following 
repeated contact with the damaged endothelium (Oberleithner 2013). This suggests that impaired red blood 
cell glycocalyx integrity would mirror the vascular risk posed by a damaged eGC. The erythrocyte “Salt Blood 
Test” (SBT) is based on the principles: 1) red blood cell sodium buffering capacity directly relates to the 
Figure 2: Intact and damaged 
endothelial glycocalyx - schematic 

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thickness (and anionic charge) of the glycocalyx, and 2) red blood cell aggregation and sedimentation in the 
presence of dextran is affected by surface electrical charge. The SBT measures the “erythrocyte salt 
sensitivity” (ESS) from the sedimentation rate after mixing blood samples with a standardized sodium/dextran 
solution (Oberleithner et al. 2013). In other words, if the weaker negative charge of a damaged red blood cell 
glycocalyx is neutralized by adherent sodium cations, dextran-coated erythrocytes will no longer 
electrostatically repel each other and will sink faster in a capillary tube.  
Sodium is only one component of an overall eating pattern: Individual micronutrients, including sodium, 
are consumed in combination and correlate with one another. Accordingly, research and guidelines have 
shifted toward dietary patterns rather than focusing solely on single dietary components. The sodium-restricted 
Dietary Approaches to Stop Hypertension (DASH/SRD) eating plan is recommended by current guidelines to 
prevent HTN-associated heart disease (Eckel et al. 2014). Compared with the typical American diet, this 
dietary pattern contains more low-fat dairy, whole grains, fruit, vegetables, legumes, nuts, and seeds, and 
lower amounts of red meat. In addition to reducing sodium, cholesterol, simple carbohydrate, and saturated fat 
intake, the DASH/SRD should increase intake of potassium, magnesium, calcium, phytochemicals, and other 
antioxidants (Karanja et al. 1999). In salt-sensitive animals, increased intake of potassium (Kido et al. 2008), 
magnesium, calcium (Goodwin et al. 2006), and antioxidants (Bayorh et al. 2006) attenuates cardiovascular 
dysfunction and the progression to HF even if high sodium intake continues.  
In middle-aged and older adults, incident HF and survival after HF diagnosis are inversely proportional to 
DASH/SRD dietary concordance (Levitan et al. 2009, Levitan et al. 2013).  In the Nurses’ Health Study and the 
Health Professionals Follow-up Study, increased DASH/SRD adherence from baseline over a 4-year period 
was associated with lower incidence of coronary artery disease and stroke over the next 4 years. This study 
suggests that middle-aged and older adults who newly adopt the DASH/SRD can reduce the development of 
cardiovascular disease (Sotos-Prieto et al. 2015). Unfortunately, concordance with this eating pattern is 
declining over time in the U.S.(Mellen et al. 2008), and new approaches are needed. 
Implementing dietary modification to reduce cardiovascular risk: Most previous studies of the DASH/SRD 
have only provided prepared meals, lasted only several weeks, and focused primarily on BP lowering (Sacks et 
al. 2001). The PREMIER and ENCORE trials examined the effects of longer-term DASH adherence (Appel et 
al. 2003, Blumenthal et al. 2010); both reduced BP, and some cardiovascular measures improved in ENCORE. 
Neither intervention promoted sodium restriction, likely to be of increased importance in veterans with HTN and 
metabolic syndrome (Elijovich et al. 2016). Moreover, both trials were conducted in affluent, early middle-age 
participants (average age ~ 50 years) and used frequent in-person dietary counseling (13-18 visits), making 
generalizability uncertain. As the Clinical Science R&D Service approval to submit this application states, 
“given that dietary interventions are difficult and resource-intensive, [our group is] encouraged to propose an 
innovative approach that will uniquely address the research questions… the difficulty in translating the results 
of such interventions into the real world should be addressed.”  
 
All participants will receive dietary education and motivational interviewing-based counseling aimed at 
increasing dietary concordance to the DASH/SRD eating pattern. Motivational interviewing is a non-
confrontational, goal-directed, patient-centered counseling method that focuses on resolving ambivalence 
toward making behavioral changes. Open-ended questioning and reflective listening are used to help the 
participant explore their motivations and establish realistic goals specific to their personal values and individual 
life situation (Miller et al. 2012). Motivational interviewing-based counseling is frequently used within VA, e.g. in 
a nationally-disseminated and recommended training program for substance abuse clinic providers (Drapkin et 
al. 2016). These techniques are effective for dietary modification, as evidenced by a recent telephone-
delivered motivational interviewing-based intervention that increased consumption of fruits, vegetables, and 
fiber while reducing fat consumption, all key DASH diet goals. The participants in the 6-month study were 
disadvantaged rural middle-aged and older adults at risk of or with the metabolic syndrome, making the results 
particularly relevant to Michigan veterans and the current proposal (Blackford et al. 2016). 
Motivational interviewing promotes autonomy and willingness to change behavior, and hence complements 
other treatment modalities. Half of participants in the proposed study will be randomly assigned to the 
Women’s and Men’s Hypertension Experiences and Emerging Lifestyles Intervention (WHEELS-I), a program 

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designed specifically to promote DASH/SRD diet adoption via improving cognitive representations. Cognitive 
representations, enduring beliefs, attitudes, and intentions that are used to structure knowledge and guide 
behavior, are influenced by genetic, situational, and contextual factors.   Easily accessible and accurate 
cognitive representations enable knowledge integration and continuous learning and increase the ability of 
patients to cope with stressors and solve problems related to HTN management (Scisney-Matlock 1998). The 
overall intent of the WHEELS-I program is to use structured messaging to build, reframe, and integrate 
cognitive representations for the DASH/SRD dietary pattern. As originally conceived, a pen-and paper 
WHEELS-I intervention In hypertensive middle-aged and older women improved DASH/SRD cognitive 
representations (knowledge, skills and attitudes) (Scisney-Matlock 1998) and DASH/SRD dietary concordance 
per the Health Promotion Lifestyle Score at 60 and 90 days (Scisney-Matlock et al. 2006). Subsequently, an 
email-based WHEELS-I program also improved DASH/SRD adherence at 90 days, with corresponding robust 
reductions in BP (see Preliminary data below). 
 
Overall summary: This proposal would represent the first study to: 1) examine cardiovascular changes during 
short-term consumption and longer-term adoption of DASH/SRD in veterans with HTN and metabolic 
syndrome, a high-risk group for incident HFpEF; 2) evaluate investigate how cardiovascular functional and 
structural risk factors for HFpEF in this group differ by salt-sensitivity, the most consistent physiological 
phenotype in animal models of hypertension-associated HF, 3) and evaluate the performance of a novel 
biomarker for the salt-sensitive phenotype. These goals will be met while 4) testing the efficacy of cost-
sensitive strategies to promote DASH/SRD adherence in veterans with HTN and metabolic syndrome. 
C. PRELIMINARY DATA 
Cardiovascular effects of DASH/SRD: In our DASH-DHF pilot study, 13 older adults (age 72±10 years) with 
compensated HF, HTN, and metabolic syndrome consumed a provided DASH/SRD for 21 days. 24-hour BP 
monitoring, blood and urinary analyses, transthoracic echocardiography, and cfPWV were completed pre- and 
post-DASH/SRD. Echocardiographic measures of ventricular-arterial coupling (Ees:Ea ratio), which measures 
the efficiency of blood transfer from the heart to the arterial tree, were obtained. We assessed ventricular 
contractility via pressure-normalized wall stress, which strongly 
predicts mortality in HFpEF (Zhong et al. 2013). Diastolic function was 
evaluated with the Parametrized Diastolic Formalism. This kinematic 
model of ventricular filling analyzes early mitral inflow (“E-wave”) 
patterns to quantify diastolic function via relaxation (c) and stiffness-
based (k) constants. The close relationships between k and c, 
ventricular filling pressures, and cardiac mechanics have been 
invasively validated (Chung et al. 2015). 
Food diaries demonstrated excellent DASH/SRD adherence, 
corroborated by decreased urinary sodium (3352±1593 to 1478±933 
mg/24h; p<0.001) and increased urinary potassium excretion 
(2284±793 to 2925±1024 mg/24h; p=0.04). Despite treatment with 
multiple antihypertensives in nearly all subjects and overall well-
controlled baseline BP, the DASH/SRD reduced 24-hour mean BP 
from 89±10 to 84±9 mmHg; p=0.04; cfPWV declined from 12.4±3.0 to 
11.0±2.2 m/s; p=0.03. The DASH/SRD significantly improved 
ventricular stiffness and relaxation, while contractility and ventricular-
arterial coupling ratio increased (Figure 3) (Hummel et al. 2013). Despite 
doubling urinary aldosterone excretion, the DASH/SRD reduced urinary F2-isoprostanes (a measure of 
systemic oxidative stress) by 31%, a similar finding to that in salt-sensitive subjects without HF (Al-Solaiman et 
al. 2009). This decrease in F2-isoprostanes closely paralleled (r=0.76, p=0.002) achieved reductions in 24-
hour urinary sodium excretion on the study diet (Hummel et al. 2012). These data indicate that patients with 
HTN and HFpEF have a salt-sensitive phenotype, and that the DASH/SRD favorably affects several key 
predictors of functional capacity and outcomes in HFpEF. Dr. Hummel continues to collect these data (not 
presented here due to investigator blinding to treatment group) in an NIH/NIA funded study randomizing older 
patients to home-delivered DASH/SRD vs. usual care following hospital discharge from HF exacerbation.  
Whiskers: ± SD; *: p < 0.05 
Figure 3: DASH/SRD effects in HF 
*
*
*
*

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Left atrial strain and relationship to coronary microvascular function: Dr. Hummel and colleagues are 
currently investigating the relationship between coronary flow reserve (CFR, via cardiac positron-emission 
tomography) (Murthy et al. 2014), a measure of coronary microvascular function, and echocardiographic left 
atrial strain, a sensitive measure of left atrial reservoir function.  In 75 subjects (49% male, age 63±11 years, 
body mass index 31±9 mg/m2, 83% hypertensive) with normal left ventricular ejection fraction and epicardial 
coronary perfusion, left atrial strain was 28±8% and CFR was 2.0±0.7 (normal values in healthy subjects 
45±11% and >2.0, respectively). The CFR was significantly associated with left atrial strain (β: 3.3% per unit 
CFR, p=0.01) independent of age, gender, obesity, HTN, chronic kidney disease, diabetes mellitus, and 
coronary artery disease status, and CFR was significantly lower in patients with diastolic dysfunction. 
 
Measurement of endothelial glycocalyx: In an ongoing study, Dr. Hummel and colleagues are using 
sublingual SDF microscopy and automated software (Glycocheck, Maastricht, Netherlands) to assess 
microvascular function and eGC thickness in patients with HF and reduced ejection fraction. In 18 subjects (6 
advanced HF inpatients, 8 stable HF outpatients, and 4 healthy controls), we have observed an inverse 
relationship between RBC filling % and PBR (Figure 4), i.e. between microvascular perfusion and eGC 
thickness (Lee et al. 2014). 
 
We have also examined the sodium-buffering capacity of the glycocalyx. 
The mini-SBT (Care Diagnostica, Voerde, Germany) is a point-of-care 
blood test, and is performed as follows: 1) mix 50 µL of whole blood from 
an EDTA-potassium (1.8 mg EDTA/ml) tube or minivette into a vial 
containing 50 µL standardized solution of sodium chloride and dextran 
(composition proprietary due to pending patent application); 2) gently 
shake vial to mix; 3) transfer solution to a 75 mm microhematocrit capillary 
tube and place upright; 4) measure supernatant length following red blood 
cell sedimentation for 60 minutes. The mini-SBT result is calculated as: 
ESS = [supernatant length at 60 min. (mm)/24.1 mm (mean length in 
healthy controls)] * 100, and is expressed as a percentage (normal ESS = 
100%).The between-subject variance (n=90, ESS 100±34%) is larger than 
same-day within-patient variance (n=15 tests, ESS 104±6%). Increased 
variance on within-patient serial testing suggests that ESS is modulated by 
physiological state (n=109 tests over 5 months, ESS 113±16%).  
We have obtained ESS results in hypertensive HFpEF patients (21 samples from 8 patients), and 187 older 
women without HF. These data are shown with data from 98 healthy controls in Figure 5.  The ESS in HF was 
higher than in healthy controls (173±43% vs. 99±32%, p<0.001). In older women without HF, ESS values were 
intermediate to the other two groups (133±37%, p=0.03 vs. HF and p<0.001 vs. controls) The ESS trended 
higher in older women with HTN and central obesity (HTN vs. no HTN, ESS 136±38% vs. 125±37%, p=0.052, 
waist-hip ratio above vs. below median, ESS 144±40% vs. 126±36%, p=0.07).  
In addition, the ESS value in older women correlated with C-reactive 
protein, a measure of inflammation (ρ 0.25, p=0.01), the adipokines leptin 
(ρ 0.23, p=0.003) and resistin (ρ 0.18, p=0.02), and brachial-ankle pulse 
wave velocity, a measure of arterial stiffness (ρ 0.16, p=0.03). The 
relationship between ESS and brachial-ankle pulse wave velocity was 
independent of other predictors of arterial stiffness, including age, black 
race, body mass index, and the presence or absence of metabolic 
syndrome (β 8.7 cm/s per 10% increase in ESS, p=0.049).Taken together, 
these data indicate that the ESS, a measure of impaired eGC sodium-
buffering capacity, is: a) increased in HFpEF, consistent with our central 
hypothesis that salt-sensitivity contributes to HFpEF, b) increased but 
retains substantial variability in persons with expected higher prevalence of 
salt-sensitivity, and c) significantly associated with several parameters that 
predict incident HF. 
Figure 4: Microvascular perfusion 
vs. eGC thickness 
Red blood cell filling % 
         Perfused boundary region (μM) 
Figure 5: ESS values by group 
% of normal ESS
Line: median, +: mean, whiskers 5-95% 
% of normal ESS 

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Efficacy of motivational interviewing to reduce dietary sodium intake: Dr. Hummel and colleagues 
recently studied the effects of dietary sodium restriction on 24-hour ambulatory blood pressure and fluid 
volume as measured by whole-body bioimpedance in 35 patients with chronic kidney disease (age 60±12 
years, 62% male, body mass index 33±5 kg/m2, 95% hypertensive). The 4-week dietary intervention aimed to 
reduce sodium intake below 85 mmol (1955 mg)/day. The protocol involved four in-person contacts with a 
research dietitian, who used motivational interviewing techniques to lessen the subject’s resistance to change 
dietary behavior and empower them to make lower sodium food choices (Miller et al. 2012). The intervention 
significantly reduced (all p<0.001) mean 24-hour urinary sodium excretion (by 46%, 170±40 to 92±39 
mmol/24h), 24-hour systolic BP (143±22 to 125±19 mmHg) and total extracellular volume (-1.5 liters compared 
to baseline). These data show that motivational interviewing can reduce short-term sodium intake, promoting 
beneficial cardiovascular changes in obese hypertensives with chronic kidney disease. 
 
Efficacy of WHEELS-I on DASH/SRD adherence and BP reduction: An email-based WHEELS-I program 
was recently studied by Dr. Scisney-Matlock in 77 women with HTN (age 62±9 years, 77% African-American), 
randomized to usual care vs. WHEELS-I (Scisney-Matlock et al. 2015). The WHEELS-I group received email 
messages linked to web-based surveys to document momentary assessment and performance of DASH/SRD 
self-care activities. Morning messages (daily for 28 days) were tailored to cognitive representations and 
requested a brief journaling response, and evening messages (nightly for 56 days) recorded daily DASH/SRD 
adherence. The study outcomes were DASH/SRD adherence and systolic BP at 30 and 90 days. 
 
Overall participant engagement with WHEELS-I was excellent, with 78% of 
messages accessed overall and message interaction of 56% sustained at 8 
weeks. As shown in Figure 6, the adherence to DASH/SRD targets 
increased substantially in the WHEELS-I group, with the largest 
improvement from baseline and the best adherence in sodium intake.   
 
Baseline BP was similar between groups. At 30 days, systolic BP decreased 
in the WHEELS-I group, but not controls (-9±15 mmHg, p<0.001 vs. 2±15 
mmHg, p>0.05); this BP reduction persisted through 90 days (-8±17mmHg, 
p=0.004 vs. 0±13 mmHg, p>0.05). The WHEELS-I group participants with 
baseline BP >140/90 mmHg sustained a large systolic BP reduction (-13±13 
mmHg at both 30 and 90 days) (Scisney-Matlock et al. 2015). This BP effect 
magnitude is similar to that in the original DASH-Sodium trial, where all food 
was provided to participants (Sacks et al. 2001). These data suggest that the 
electronically delivered WHEELS-I program has the potential to foster long-term DASH/SRD adoption and 
accompanying favorable cardiovascular changes in a multi-ethnic, middle-aged to older adult cohort.  
 
D. APPROACH 
General Design: This 7-month dietary intervention study will recruit from the Ann Arbor Veterans Affairs 
Health System and affiliated Community-Based Outpatient Clinics (CBOCs). We hypothesize that dietary 
modification with the DASH/SRD will improve HF-associated cardiovascular dysfunction in veterans with HTN 
and metabolic syndrome, and that differential improvements will occur in those with a salt-sensitive BP 
phenotype vs. those with salt-resistant BP. The study will take place in two phases (Figure 7): Phase 1 
participants will consume their usual diet, followed by a DASH/SRD for 14 days each. Salt-sensitive vs. salt-
resistant status will be defined by 24-hour BP measurement at the end of each segment In Phase 2, all 
participants will receive motivational interviewing-based dietary counseling; half will participate in WHEELS-I to 
promote DASH/SRD adoption over 6 months. 
 
Participants will have a clinical evaluation at 4 time points (denoted by ↑ in Figure 7 and described below): at 
weeks 2 and 4 of the Phase 1 crossover intervention, and at 1 and 6 months of the Phase 2 DASH/SRD 
adoption period.  
 
Figure 6: DASH/SRD target 
adherence (% of participants) 

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The Phase 1 primary 
analysis will evaluate 
cardiovascular functional 
changes during usual 
diet and DASH/SRD diet 
periods, and test for 
interaction with salt-
sensitivity status. Phase 
2 will assess the 
performance of 
WHEELS-I in promoting 
DASH/SRD adoption, 
and evaluate cardiac 
structural changes 
between salt-sensitive 
and salt-resistant 
participants (as they were defined by BP measurement in Phase 1). After completion of Phase 2, telephone 
clinical and mailed dietary questionnaire follow-up will occur after 6 months. An optional, in-person visit may 
also be performed at this time to obtain arterial tonometry data and clinic blood pressure.  
         
Study Subjects: Veterans aged ≥45 years with HTN (here defined as screening systolic BP ≥ 130 and/or 
diastolic BP ≥ 85 mmHg, or current use of anti-hypertensive drugs) and metabolic syndrome (body mass index 
≥30 kg/m2 and/or waist circumference >94 cm). These data will be confirmed at study screening. Participants 
must also be willing to participate in the WHEELS-I program by using a smartphone application or email. 
Exclusion criteria: On-treatment, resting systolic BP of >160 mmHg (average of 3 values) at screening visit, 
previous history of HF, left ventricular ejection fraction <50%, moderate or severe valvular heart disease, 
myocardial infarction or stroke within the prior 6 months, chronic kidney disease with estimated glomerular 
filtration rate <45 ml/min/ 1.73m2, unoperated aortic aneurysm for which surgery is indicated, prior 
hyperkalemia requiring urgent treatment, hemoglobin <9 gm/dL, and investigator-determined factors: severe 
pulmonary disease (e.g. oxygen-requiring) or hepatic disease (e.g. cirrhosis), severely uncontrolled diabetes 
(hemoglobin A1c >10%), active cancer other than non-melanoma skin or low-risk prostate cancer, other 
comorbidity with expected survival <12 months, active alcohol/illicit substance abuse, and/or a history of 
persistent nonadherence to treatment.  
Recruitment strategy: Eligible veterans with HTN and metabolic syndrome documented at their most recent 
clinical encounter will be identified and allocated randomly into blocks of 10.  Beginning with the first block, 
potential subjects will be mailed a letter inviting them to participate in this study. The letter will be followed-up in 
one week by a telephone call from the study coordinator to discuss the study, pre-screen for eligibility, and 
schedule the first visit if interested in participation. In order to enroll 110 participants, we plan in-person 
screening visits for 130 pre-screened veterans who have expressed preliminary interest in the study, (i.e. we 
anticipate 20 screen-fails). Given staffing, space, and equipment availability, recruitment will take place over 
3.5 years with the final in-person clinical study visits (Figure 1) in the second quarter of year 4.   
 
 
 
 
 
 

9 
 
Study procedures: will be performed according to the study procedure timeline (Figure 8, which includes both 
phases of the study). All cardiac and vascular measures will be analyzed blinded to dietary assignment, food 
diary results, 24-hour BP, and urinary electrolytes; 1/10 of cardiovascular studies will be re-analyzed by a 
second investigator to establish reproducibility and rigor of results.  All study data will be recorded in REDCap, 
a secure research web application, under a unique study ID number assigned to participants.  
• 
History and exam:  
o Clinic BP and heart 
rate will be measured 
three times in the 
seated position and 
once in the standing 
position at each visit 
using an automated 
sphygmomanometer.  
o Standard methods 
will be used to 
measure height, 
weight (all visits), 
waist and hip 
circumference 
(screening and end of 
study). 
o Medical diagnoses, 
clinical events (e.g. hospitalizations) and medications will be updated at each visit.  
o Questionnaires:  
 Center for Epidemiologic Studies Depression Scale and the 36-item Short Form survey (SF-36) will be 
obtained, as depression and anxiety can affect quality of life and the efficacy of lifestyle change to 
reduce cardiovascular risk factors  
 The SF-36 Physical Functioning Scale and Instrumental Activities of Daily Living. These are planned 
because functional limitations affect nutrient intake; in turn, changing dietary patterns can affect 
muscle mass.   
 GOURMET Barriers questionnaire, to assess psychosocial factors related to behavior change. 
Sample attached. 
 Physical Activity Scale for the Elderly (PASE), to assess physical activity level before and after the 
intervention to ensure outcomes are not attributable to changes in physical activity. Sample attached. 
• 
24-hour BP measurement:  Participants will wear an Oscar II (Suntech Medical Inc, Raleigh, North 
Carolina) ambulatory BP monitor on the non-dominant arm for 24 hours; BP will be recorded 2 times per 
hour while awake and 1 time per hour during sleep. Acceptability of recordings for analysis will be 
assessed according to published international norms (Kikuya et al. 2007).  
• 
Blood tests: Screening: comprehensive metabolic panel, complete blood count, B-type natriuretic peptide 
(BNP), hemoglobin A1C, lipid panel. All other visits: basic metabolic panel, insulin, BNP, F2-isoprostanes, 
aldosterone, IL-6, hs-CRP, TNF-α. These measures were chosen to evaluate the effects of DASH/SRD on 
renal function, electrolytes, renin-angiotensin-aldosterone system, insulin resistance, and oxidative stress, 
as well as inflammation markers previously associated with incident HFpEF (Kalogeropoulos et al. 2010). A 
repeat lipid panel will also be obtained at the end of the study. Whole-blood samples will be used for the 
mini-SBT (see Aim 3 for description). Additional samples will be stored for post-hoc biomarker analysis, 
planned to include syndecan-1 and heparan sulfate, serological markers of endothelial glycocalyx 
degradation, and the adipokines leptin, adiponectin, and resistin.  
• 
24-hour urine: All collections: sodium, potassium, creatinine, aldosterone, F2-isoprostanes, and 
microalbumin. These measures were selected to assess DASH/SRD adherence, the relationship between 
systemic oxidative stress and renin-angiotensin-aldosterone system activation in salt-sensitive and 
resistant subjects (Al-Solaiman et al. 2009), and chronic microvascular damage. Additional samples will be 

10 
 
stored for post-hoc analysis of biomarkers. The completeness of 24-hour urinary collections will be 
assessed via creatinine excretion per the method of Knuimann et al. (Murakami et al. 2008).  
• 
Dietary assessment: Estimated nutrient intakes from the Block Food Frequency Questionnaire (collected at 
screening and at the end of study) will be analyzed (NutritionQuest, Berkeley, CA) and scored for 
DASH/SRD concordance (Mellen et al. 2008). Three-day food diaries will be performed during weeks 2 and 
4 of Phase 1 (i.e., the final week of usual diet and home-delivered DASH/SRD) and at Month 1 and Month 
6 of Phase 2; these will be initially reviewed for completeness by the study dietitian and analyzed with 
Nutrition Data System for Research (2015 version; Nutrition Coordinating Center, Minneapolis, MN) 
through the Michigan Nutrition Obesity Center (MNORC);  
• 
Arterial tonometry: Carotid-femoral pulse wave velocity (cfPWV) will be obtained with a Sphygmocor 
applanation tonometer (AtCor Medical, Itasca, IL). Pulse waveforms are obtained with gentle tonometer 
pressure above the carotid and femoral arteries, with cfPWV calculated using the distance between the two 
sites and the wave transit times, measured using the R wave from simultaneous ECG recording. Radial 
artery pulse waveforms will also be obtained to measure other parameters of potential interest, e.g. central 
pulse pressure, aortic augmentation index, and aortic characteristic impedance (Townsend et al. 2015). 
• 
Two- and three-dimensional (3D) echocardiography: B-mode 2D, M-mode, and 3D echocardiographic 
images will be obtained with the Philips EPIQ 7 ultrasound platform (Philips Electronics, Andover, MA) and 
recorded via study ID number on DVD and placed under study ID number in the Ann Arbor VA 
echocardiogram database. Standard cardiac structural, valvular, and functional parameters, including 
standard measures of diastolic function (e.g., E/A and E/e’ ratios), will be measured as per current guidelines. 
Speckle-tracking of 2- and 4-chamber apical views will be used to obtain global longitudinal strain of the left 
ventricle using the Philips QLAB program. Parametrized Diastolic Formalism analyses will be performed as 
described (Chung et al. 2015). 3D images will be used to measure left atrial volume, left ventricular mass, 
and ejection fraction (Gottdiener et al. 2004, Nagueh et al. 2009, Lang et al. 2015). 
• 
Sublingual sidestream darkfield (SDF) microscopy: Images of the sublingual microcirculation will be 
obtained and analyzed using an integrated platform that includes a SDF microscope probe and automated 
software (Glycocheck, Maastricht, Netherlands) (Lee et al. 2014).  See Aim 3 for additional description. 
• 
Safety Monitoring: Patients will perform home BP monitoring with their own personal cuff or a provided 
Omron 7 series automated cuff (Omron Healthcare, Lake Forest, IL). Monitoring will be daily during the 
run-in period and Phase 1 of the study, and at least weekly thereafter – if systolic BP is >180 or <100 
mmHg on more than one occasion, or if symptoms suggest hyper- or hypovolemia (e.g., worsening/new 
edema, dizziness), the participant will be scheduled for an in-person safety visit. Targeted history and 
physical will be performed by a clinician-investigator, with laboratory or other testing obtained based on the 
clinical presentation. Antihypertensive medications may be adjusted at this visit if needed for participant 
safety. Safety visits may also be scheduled for worsening glycemic control in diabetic participants. See 
Human Subjects Protection section for additional details.   
Study phase 1: Phenotype veterans with HTN and metabolic syndrome as salt-sensitive or salt-
resistant; determine effects of a 2-week DASH/SRD intervention vs. control diet on HFpEF functional 
cardiovascular risk factors in salt sensitive compared to salt-resistant veterans.  
Introduction: Animal models, cohort studies, and our preliminary work suggest that the DASH/SRD would 
favorably affect cardiovascular parameters that predict incident HFpEF in veterans with HTN and metabolic 
syndrome, and have a stronger effect in salt-sensitive individuals. Our working hypothesis is that the 
DASH/SRD will produce significantly greater improvement than control diet in HFpEF risk factors including 
large-arterial stiffness (carotid-femoral pulse wave velocity [cfPWV], primary outcome), left ventricular diastolic 
function, and left ventricular systolic function. We further hypothesize that the improvement in these measures 
will be greater in salt-sensitive veterans. We will test our hypotheses in a 2-sequence, 2-period, 2-treatment 
comparison of DASH/SRD vs. control diet.  
Phase 1 study design: Consented participants will have clinic BP measured at the screening visit and will 
enter a two-week usual diet period, during which antihypertensive medications, with the exception of calcium 
channel blockers, will be weaned off under Dr. Hummel’s supervision and with guidance from daily home BP 
monitoring. During this time period, the participant will consume their usual diet. Potential subjects with on-
treatment, resting, average systolic BP of > 160 mmHg at the screening visit will be excluded from further 

11 
 
participation, as will those with symptomatic hypertension or systolic BP of > 180 mmHg on more than one 
occasion during the ad-lib dietary period.   
Following the usual diet period and the week 2 visit, participants will consume the DASH/SRD diet for 14 days 
each. This time period is sufficient to demonstrate changes in BP and arterial stiffness, which peak at 2 weeks 
after dietary sodium modification (Gates et al. 2004). In addition to in-clinic BP measurement, 24-hour 
ambulatory BP will be obtained at the end of each dietary sequence. We expect that the DASH/SRD will 
reduce BP across the cohort, but differentially on an individual basis (Sacks et al. 2001).  
The salt-sensitive vs. salt-resistant phenotype will be defined as a change in 24-hour mean BP of  ≥8 mmHg 
between control and DASH/SRD dietary segments (Castiglioni et al. 2011). Based on previous studies in 
patients with HTN, we expect that 30-50% of participants will be classified as salt-sensitive (Elijovich et al. 
2016). The 24-hour ambulatory BP monitoring and 24-hour urinary collections will be used to provide an 
alternate definition of salt-sensitivity. Specifically, the quantitative Sodium Sensitivity Index will be calculated as 
the change in 24-hour mean BP divided by the change in 24-hour urinary sodium excretion between 
DASH/SRD and control diet periods (Castiglioni et al. 2011). 
The Phase 1 primary outcome is the cfPWV compared at the end of usual diet and DASH/SRD. This 
parameter was chosen because of its association with incident HF in community-dwelling elderly (Tsao et al. 
2015) and its high sensitivity and specificity for HF in older adults with unexplained exertional dyspnea (Weber 
et al. 2013). Key secondary outcomes include ventricular diastolic function (k; ventricular stiffness) and systolic 
function (global longitudinal ventricular strain) and left atrial reservoir function (left atrial strain).  
Dietary intervention: At the week 2 visit, a printed guide to the DASH/SRD produced by the U.S. Department of 
Health and Human Services will be provided, and participants will complete a standardized questionnaire that 
establishes their cognitive representation of the DASH/SRD (Scisney-Matlock et al. 2006). Baseline nutrient 
intake will be assessed with the Block Food Frequency Questionnaire and Nutrition Quest software 
(NutritionQuest, Berkeley, CA). This instrument has previously been used to assess DASH/SRD concordance 
(Mellen et al. 2008). Following this visit, participants will consume 14 days of DASH/SRD provided meals, 
designed by Dr. Hummel. These will be prepared, pre-packaged, and home-delivered twice weekly by 
PurFoods, LLC. The study DASH/SRD sodium content will be 50 mmol (1,150 mg/2,100 kcal), as used in the 
DASH-Sodium study (Sacks et al. 2001). Both diets will be calorie-adjusted to maintain stable body weight 
using the National Institutes of Health body weight planner (Hall et al. 2011).  Nutrient consumption during 
Phase 1 will be assessed by 3-day food diary during the second week of each dietary period; results will be 
corroborated by 24-hour urinary sodium and potassium. 
Sample size and power: The sample size is based on a combination of logistical considerations and power 
required to detect important differences in the primary endpoint of change in cfPWV in the context of a 
randomized crossover design.  Sample size is based on a 2-sequence, 2-period, 2-treatment crossover design 
for continuous endpoints (using paired t test for mean differences).  We conservatively assume a within-subject 
correlation of 0.70 for cfPWV measurements, based on our DASH-DHF pilot (0.81) and literature showing 
same-day correlation >0.90 (Wimmer et al. 2007) using the Sphygmocor device. We also assume approximate 
10% dropout rate during Phase 1 of our study.  Under these assumptions and using a two-sided type I error 
rate of 5%, 100 subjects, 50 allocated to the DASH/SRD-control diet sequence and 50 allocated to the control-
DASH/SRD diet sequence, provides >80% power to detect an effect size (mean treatment difference/standard 
deviation) of 0.22 and >90% power to detect an effect size of 0.26.  For comparison purposes, the effect size 
of cfPWV changes with DASH/SRD in our preliminary work in HFpEF has been 0.4-0.6; in dietary sodium 
restriction studies in healthier populations than we plan to study, the effect size is at least 0.2-0.4 (Seals et al. 
2001, Todd et al. 2010). This sample size provides >80% power to detect an effect size of 0.5 in cfPWV 
change between salt-sensitive vs. salt-resistant subjects.  This is a between-groups comparison based on a 
two-sample t test and two-sided Type I error of 5%.  
Analysis plan: We will examine the distributions of outcomes under each treatment (e.g., DASH/SRD and 
control diet), as well as by salt-sensitive phenotype.  Graphical approaches such as boxplots and scatterplots 
will be used, allowing identification of outliers, linearity, and correlation of measurements within subject and 
across time.  If the distributional assumptions of our models are not met, transformations will be employed. 

12 
 
Obvious outliers unrelated to data entry error will prompt reconfirmation of acquisition methods and quality by 
another blinded investigator.   We expect no carryover effects due to the two week duration of each dietary 
segment, a sufficient interval to match sodium intake and excretion even in subjects with chronic kidney 
disease (McMahon et al. 2013), who have more difficulty achieving this balance. 
 
An intention-to-treat approach will be used to analyze endpoints, including all randomized participants. Multiple 
imputation will be used for missing data, which assumes that outcome data are missing at random (i.e., where 
other variables from the study can account for differences in the primary outcome for observed and missing 
subjects) (Little et al. 2012).  A linear mixed effects model (Verbeke et al. 2000, Diggle et al. 2013) will be used 
to assess the effect of intervention on the change in cfPWV, with fixed-effect terms for sequence, study period, 
dietary intervention, and adjusting for baseline cfPWV and other potential confounders, e.g. age, race, body 
mass index, diabetes status, and renal function. A random-effect term will be included to account for the 
correlated measurements for each subject.  Tests to compare diet will be based on Wald and chi-squared tests 
based on maximum likelihood or restricted maximum likelihood methods. Alternative covariance structures 
(other than compound symmetry) will be explored and model fit will be examined using the Akaike Information 
Criteria (Holford 2004).  This approach will also be used for the key secondary endpoints (ventricular diastolic 
stiffness k and global longitudinal ventricular strain) and other cardiovascular parameters (e.g. E/e’ ratio). For 
binary secondary outcomes (e.g. serious adverse events) a similar mixed model approach will be employed 
(Ezzet et al. 1992). To test whether improvements in the primary and key secondary endpoints are greater in 
salt-sensitive than in salt-resistant subjects, interaction will be assessed via the linear mixed model described 
above, with a fixed-effect term for salt-sensitivity phenotype.   
 
Potential problems/alternative approaches: Despite our previous experience in collecting 24-hour BP and 
urinary data, it is possible that some attempts could be incomplete. If this occurs, the dietary intervention would 
be continued for an additional 24 hours to repeat the measurement. If despite these efforts, one or both of the 
24-hour BP or urine collection are unacceptable in an individual, we would assign the salt-sensitive phenotype 
if clinic-measured mean BP is >5 mmHg higher following the control diet vs. the DASH/SRD segment; this 
threshold has been previously used to define salt-sensitivity in metabolic syndrome (Chen et al. 2009). This 
could affect the distribution of salt-sensitivity slightly, but we expect that few patients will have incomplete data 
and there will be no bias in missing data by underlying phenotype. The detectable effect size would increase 
based on the degree of imbalance between the number of subjects defined as salt-sensitive vs salt-resistant. 
In the event that this imbalance is as high as 70%:30%, the detectable effect size with >80% power would be 
0.62 with a two-sided Type I error of 5%. If the distribution of salt-sensitivity is markedly different than 
expected, we would perform a secondary analysis using the median value or other appropriate cutpoint of the 
quantitative Sodium Sensitivity Index (Castiglioni et al. 2011). 
We do not expect substantial difficulty in obtaining cardiovascular measures; Dr. Hummel is evaluating all 
proposed echocardiographic parameters in four ongoing studies, including two NIH-funded studies using 
DASH/SRD as the intervention. A major focus of Dr. Hummel’s NIH/NHLBI K23 award has been non-invasive 
vascular assessment. In the event that one or more of the cardiovascular measures cannot be obtained, 
multiple imputation methods will be used. In the course of studies conducted by Dr. Hummel’s team, >120 
subjects have completed dietary interventions, involving provided food and/or dietary counseling, lasting three 
weeks to four months. Adherence to provided meals has been excellent overall, but it is likely that some non-
adherence will occur. The primary analysis will be intent-to-treat; if any subjects are severely nonadherent (via 
24-hour urinary sodium or food diary review) we would do secondary analyses excluding these participants.  
 
Study Phase 2: Assess the efficacy of motivational interviewing plus the WHEELS-I program vs. 
motivational interviewing alone in promoting adoption of the DASH/SRD. Determine the effects of 
DASH/SRD adoption over a 6-month period on structural cardiovascular HFpEF risk factors in salt 
sensitive vs. salt-resistant veterans.  
 
Introduction: Structural correlates of HFpEF include left ventricular hypertrophy, which strongly predicts HF 
onset in older adults, left atrial enlargement, which reflects chronically elevated ventricular filling pressures, 
and stiff central arteries. No previous studies in participants with HTN and metabolic syndrome have examined 
the relationship between the salt-sensitive phenotype and these HFpEF risk factors after long-term dietary 

13 
 
modification. Current guidelines recommend the DASH/SRD to prevent cardiovascular disease in 
hypertensives with metabolic syndrome, but adherence to this dietary pattern is declining nationwide. Our 
working hypotheses are that: 1) When compared to motivational interviewing alone, the addition of the 
Women’s and Men’s Hypertension Experiences and Emerging Lifestyles Intervention (WHEELS-I) program, an 
electronically-delivered tailored messaging intervention, will improve adoption of the DASH/SRD over a six 
month period, and 2) sustained adoption of the DASH/SRD over 6 months will produce greater improvement in 
structural cardiovascular HFpEF risk factors in salt-sensitive than salt-resistant subjects, with measures 
including left ventricular mass index (LVMI, primary outcome), left atrial volume index (LAVI), cfPWV, and BP.  
 
Phase 2 study design:  We will test our working hypotheses by providing motivational interviewing-based 
counseling to all participants and randomizing half to the WHEELS-I program, with both interventions 
described further below. The primary outcome of Aim 2 of this proposal, to be tested in Phase 2 of the protocol, 
is the intra- and inter-group mean change in DASH/SRD concordance score (Mellen et al. 2008) from the 
baseline value obtained at the beginning of the study. This score provides 1 point for fully meeting each of 9 
DASH/SRD targets and 0.5 for meeting an intermediate level of each target (e.g. fiber 14.8 g and 9.5g/1000 
kcal, respectively). The secondary dietary outcome is the group change in mean 24-hour urinary sodium 
excretion compared to baseline values. Other outcomes of interest are urinary potassium, microalbuminuria, 
and the nutrient composition from three day food diaries.  
 
We will also test if DASH/SRD adoption differentially improves HFpEF-associated structural cardiovascular 
parameters in salt-sensitive vs. salt-resistant subjects (as they were defined by BP response in Phase 1). The 
primary cardiovascular outcome of Phase 2 is the change from baseline (Week 2 visit) to 6 months in left 
ventricular mass index (LVMI, indexed to height2.7). This measure was chosen because LVMI is a strong risk 
factor for incident HF (Gottdiener et al. 2000), can decrease after 6 months of dietitian-counseled sodium 
restriction (Jula et al. 1994, Liebson et al. 1995), and is relevant to  the pathophysiology of HFpEF 
(Melenovsky et al. 2007). Key secondary outcomes include cfPWV (see Aim 1), left atrial volume, indexed for 
body surface area (LAVI) (Melenovsky et al. 2015), and 24-hour mean BP. 
 
Phase 2 data collection: History and exam, questionnaires, blood tests, 24-hour BP and urine collections will 
be performed at months 1 and 6 of Phase 2 (see Study procedures above). Dietary concordance with 
DASH/SRD, the measure of exposure in Phase 2, will be assessed via Block Food Frequency Questionnaire 
and 24-hour urine sodium excretion at month 6, and 3-day food diaries immediately prior to the Month 1 and 
Month 6 visits. Arterial tonometry, echocardiography, SDF microscopy, and the mini-SBT will be performed at 
Month 6 of Phase 2 (see also Figures 7 and 8). Following Phase 2, participants will have follow-up after 6 
months via telephone interview to assess cardiovascular events and vital status; subjects will be mailed a Food 
Frequency Questionnaire to evaluate persistence of dietary patterns after study participation; and there may be 
an optional in-person visit determined by study staff, based on need, for an additional arterial tonometry 
procedure and blood pressure check.  
 
Dietary intervention: Phase 2 of the protocol will begin within 2 weeks of the end of Phase 1, and typically will 
begin immediately (i.e., as part of the week 4 visit). During this period, participants will be placed back on their 
prior antihypertensive therapy, if any, under Dr. Hummel’s guidance. Prior to the first Phase 2 visit, subjects will 
be randomized to control (motivational interviewing only) or the addition of the WHEELS-I program. At the first 
visit of Phase 2, participants will update their cognitive representation of the DASH/SRD, and have an in-
person dietitian session during which motivational interviewing techniques will be employed to explore and 
support willingness to change dietary habits. If the participant desires, the motivational interviewing portion of 
the visit may be conducted by telephone. Motivational interviewing-based counseling will continue via 
telephone during week 2, month1 visit, and at months 2 and 4.  
The WHEELS-I program consists of three daily activities: in the morning, reading messages that reinforce the 
key DASH/SRD diet educational components (e.g., types of foods, number of servings, rationale for 
recommendations) related to a specific dietary goal, then brief journaling of emotions and thoughts related to 
this goal; in the evening, record of DASH/SRD adherence during that day. An example: goal, eat 4-5 servings 
of fruits and vegetables per day; messages, try fresh fruits in season, make a fruit salad of “light” or  “natural 
juice” canned fruits, keep dried fruits in your desk or car for snacks; journaling, “I am learning to appreciate the 

14 
 
sweetness of fruit.” The daily activities are designed to take less than five minutes. Participants assigned to the 
WHEELS-I program may choose whether to receive messages via email, in identical fashion to the study 
shown in the Preliminary Data above, or via a newly created smartphone mobile application. Both methods will 
link to the same web-based information and surveys, which will collect data and store it under subject number 
in the Amazon.gov cloud storage system, where data will be accessible only by study personnel. 
For the first 30 days of Phase 2, WHEELS-I participants will receive a structured series of messages that cycle 
through the DASH/SRD goal areas, e.g. increasing consumption of fruits and vegetables, whole grains, nuts 
and legumes, and low-fat dairy and reducing intake of sodium, red meat, and sweets. At the Month 1 dietitian 
visit, all participants will again update their DASH/SRD cognitive representation. Those assigned to WHEELS-I 
will then decide how frequently to receive morning messages for the next 30 days (evening messages will 
continue for the next 60 days). Options will include 1) continue daily morning messages, 2) morning messages 
three times a week, 3) morning messages once a week.  
These ongoing messages will be tailored toward challenges identified through motivational interviewing 
sessions and updating of the DASH/SRD cognitive representation. Message tailoring is a process that includes 
1) assessment of individual characteristics relevant to a particular behavior, 2) algorithms that use assessment 
data to generate intervention messages that address the needs of each user and 3) individual feedback that 
delivers these messages in a clear and vivid format. Messages will be tailored using the Michigan Tailoring 
System, an open-source platform developed by the Center for Health Communication Research.  
 
Throughout the first 90 days, if there are less than 3 logins weekly for evening messages noted after 2 
consecutive weeks, a message will be sent to inquire how the participant is doing and elicit any problems. At 
90, 120, and 150 days, all subjects will be provided the option to continue messages for an additional 30 days. 
Training for dietitians to facilitate WHEELS-I will be performed by Dr. Scisney-Matlock, the program’s creator. 
As recommended by the NIH Behavior Change Consortium (Bellg et al. 2004), training will be standardized, 
delivered by a single trainer, and reinforced (every 6 months). Skill acquisition will be confirmed at baseline 
and retention by facilitating dietitians observed regularly (e.g., every 10 subjects entering Phase 2) by Dr. 
Scisney-Matlock. Receipt of treatment by participants will be assessed via the proportion of messages 
accessed and responded to. At the end of 16 weeks, all app users will be sent a secure WHEELS-I feedback 
survey through REDCap, via email. All responses will be automatically uploaded and stored in the secure 
REDCap application. 
The mobile WHEELS application will be downloadable for either iOS or Android from the Apple or Google Play 
stores, respectively. The participant will receive a unique code to unlock the WHEELS-I application for 
download. The application will include an overview of DASH/SRD principles, daily notifications and reminders 
to both influence and measure adherence to the DASH/SRD as above, and positive encouragement for 
meeting goals. The system will provide a web-based management dashboard for the research team that allows 
for data collection from the mobile app or email replies, extracts and analysis by the researchers, and 
adherence to VA security requirements through the use of Amazon.gov cloud hosting. 
Sample size and power:  The sample size for Phase 2 is set by the goals of Aim 1 and influenced by drop-out 
during the 6 months of the interventions to promote DASH/SRD adoption.  We assume that an additional 10% 
of subjects will drop out during Phase 2 and that the dropout rate will be equal in salt-sensitive and salt-
resistant participants, resulting in 90 subjects with evaluable data. The primary outcome of Aim 2 of this 
proposal is the DASH/SRD concordance score from the Block Food Frequency Questionnaire in participants 
assigned to the WHEELS-I group vs. motivational interviewing alone. Based on national survey data and our 
preliminary work, we expect the mean baseline (Week 2 visit) dietary score to be 2.5±1.5. A sample size of 90 
provides >80% power to detect an intra-group change of 0.6 between baseline and the end of study, based on 
paired t-testing two-sided Type I error of 5%. This sample size provides >80% power to detect a differential 
between group change in dietary score of 0.9, based on a two-sample t test and two-sided Type I error of 5%.  
With regard to cardiovascular parameters, a sample size of 90 provides >80% power to detect an effect size of 
0.6 in the primary endpoint of change from baseline to month 6 in LVMI between salt-sensitive vs. salt-resistant 
subjects (assuming a 50:50 split of salt sensitive vs. salt-resistant participants), with a two-sided Type I error of 

15 
 
5%. This between-groups comparison is based on a two-sample t test. Conservatively assuming that the 
standard deviation of the change in LVMI between baseline and 6 months ranges between 4 to 7 g/m2.7 
(Blumenthal et al. 2010), this effect size translates to mean group differences of 2.4 to 4.2 g/m2.7. This would 
represent a 5-10% decrease in LVMI from baseline in salt-sensitive subjects (vs. no change in salt-resistant), 
similar to that in previous studies of dietary sodium restriction (Jula et al. 1994, Liebson et al. 1995). 
Analysis plan: Paired and 2-sample t-testing will be used to analyze changes in DASH/SRD concordance 
score as above, as well as changes in 24-hour urinary sodium excretion. The methods to analyze 
cardiovascular changes in Aim 2 will be identical to those used in the comparison of salt sensitivity phenotypes 
in Aim 1 (see above).  Important covariates in the models to compare changes in cardiovascular parameters by 
salt-sensitive/resistant phenotype will be the degree of DASH/SRD concordance and changes in 24-hour BP. 
 
Potential problems/alternative approaches: While motivational interviewing and the WHEELS-I program have 
been effective in our preliminary studies, we expect individual variance in DASH/SRD adoption. The primary 
analysis will be an intent-to-treat framework; secondary analyses will test interaction with the level of 
DASH/SRD adoption.(Mellen et al. 2008) We expect that participants who successfully adopt DASH/SRD will 
substantially change their eating habits. The WHEELS-I program does not target weight loss, but since sodium 
intake correlates with caloric intake (Guenther et al. 2013) it is possible that some veterans may lose weight 
during Phase 2. We do not a priori expect differential weight loss in salt-sensitive and salt–resistant subjects. 
However, since cfPWV and LVMI can both be reduced through weight loss, we will perform analyses adjusted 
for Phase 2 weight changes (Blumenthal et al. 2010). If the proportions of salt-sensitive and salt-resistant 
subjects are not equal (say 60%:40%), we would retain >80% power to detect effect size of 0.61. 
 
Specific Aim 3 (spans Study Phases 1 and 2): Determine effects of DASH/SRD feeding and longer-term 
adoption on microvascular function, and assess the relationship between endothelial glycocalyx 
integrity/sodium buffering and cardiovascular response to DASH/SRD. 
Introduction: Small-vessel oxidative stress and inflammation are critical mechanisms of cardiovascular injury 
in salt-sensitive animals, and recent data strongly implicate microvascular damage in HF pathophysiology. The 
endothelial glycocalyx (eGC) is a thin network lining the lumen of blood vessels, and if damaged may mediate 
the mechanisms and vascular consequences of salt-sensitivity. Our working hypothesis is that the DASH/SRD 
will improve microvascular function in comparison to control diet, and differentially so in salt-sensitive subjects. 
We will test this hypothesis using sublingual sidestream dark-field (SDF) microscopy. We further hypothesize 
that poor eGC integrity will identify subjects who have greater cardiovascular improvement with DASH/SRD. 
This hypothesis will be tested using SDF microscopy and a novel “mini-Salt Blood Test” (see Background and 
Significance above for description of these procedures). 
Aim 3 analysis plan: To determine the effects of the DASH/SRD intervention and adoption on microvascular 
parameters and the differential impact of salt-sensitivity, we will use the approaches described in Aims 1 and 2 
for microvascular density and RBC filling % (measures of microvascular structure and function).  To assess the 
relationship between eGC and salt-sensitivity, we will characterize the relationship between mean baseline 
PBR and mean baseline ESS (measures of eGC thickness and sodium-buffering capacity) and the salt-
sensitive phenotype (as defined in Aim 1). We will also assess the relationship of these parameters with 
cardiovascular functional and structural measures described in Aims 1 and 2.  For example, we will test 
whether mean baseline ESS differs by salt-sensitivity phenotype using a two-sample t-test or simple linear 
regression (to include adjustment for important covariates). Model performance measures will be used to 
assess the discrimination of PBR and ESS (vs. salt-sensitivity) for changes in cardiovascular function and 
structure [e.g., area under the receiver-operating-characteristic curve, net reclassification, and integrated 
discrimination improvement (Pencina et al. 2008)].  Descriptive analyses of baseline and changes in 
microvascular and cardiovascular function and structure will use graphical and summary statistics. 
Potential problems/alternative approaches: Technical factors could prevent SDF microscopy acquisition in a 
few participants. If this occurs, subjects will still have all other cardiovascular measures as well as glycocalyx 
assessment with the mini-SBT. We have successfully obtained the mini-SBT in over 200 participants. In the 
rare event of a technical failure, we would simply repeat the test. Despite supportive data as above, it is 

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possible that the mini-SBT results will not predict BP changes during DASH/SRD.  Given the mechanistic links 
between eGC function and sodium-induced vascular damage (Figure 2), it is also possible that the mini-SBT is 
more effective than BP changes in identifying subjects with beneficial cardiovascular effects during 
DASH/SRD.