High dietary sodium augments vascular tone and attenuates low-flow mediated constriction in salt-resistant adults

Abstract

Low-flow mediated constriction (L-FMC) has emerged as a valuable and complementary measure of flow-mediated dilation (FMD) for assessing endothelial function non-invasively. High dietary sodium has been shown to impair FMD independent of changes in blood pressure (BP), but its effects on L-FMC are unknown.

Purpose

To test the hypothesis that high dietary sodium would attenuate brachial artery L-FMC in salt-resistant adults.

Methods

Fifteen healthy, normotensive adults (29 ± 6 yrs) participated in a controlled feeding study. Following a run-in diet, participants completed a 7-day low sodium (LS; 20 mmol sodium/day) and 7-day high sodium (HS; 300 mmol sodium/day) diet in randomized order. On the last day of each diet, 24 hr urine was collected and assessments of 24 hr ambulatory BP and L-FMC were performed. Salt-resistance was defined as a change in 24 hr ambulatory mean arterial pressure (MAP) between the LS and HS diets of ≤5 mmHg. Resting vascular tone and L-FMC were calculated from ultrasound-derived arterial diameters.

Results

High dietary sodium increased serum sodium and urinary sodium excretion (p<0.001 for both), but 24 hr MAP was unchanged (p=0.16) by design. High dietary sodium augmented vascular tone (LS: 91 ± 23%, HS: 125 ± 56%, p=0.01) and attenuated L-FMC (LS: −0.58 ± 0.99%, HS: 0.17 ± 1.23%,p=0.008).

Conclusion

These findings in salt-resistant adults provide additional evidence that dietary sodium has adverse vascular effects independent of changes in BP.

INTRODUCTION

High dietary sodium is a clinical risk factor for which adverse effects have been typically attributed to unfavorable changes in blood pressure (BP) (Elliott et al., 1996).However, several studies in rodents and humans have demonstrated that excess sodium intake can impair arterial endothelial function independent of changes in BP (see Robinson et al., 2019 for Review).Specifically in humans, high dietary sodium has been shown to impair brachial artery flow-mediated dilation (FMD), an index of endothelial function, in healthy salt-resistant adults (DuPont et al., 2013). Conversely, sodium restriction was shown to improve both conduit and resistance vessel endothelial function, independent of improvements in BP in older adults (Jablonski et al., 2013). While the FMD test is widely used for assessing endothelial function noninvasively, it nevertheless only examines an artery’s dilatory response to augmented blood flow and shear stress (Thijssen et al., 2011).

Low-flow mediated constriction (L-FMC) has emerged as a complementary extension of FMD that examines an artery’s constrictive response to reduced blood flow and shear stress (Gori et al., 2008). L-FMC has been shown to be endothelium-dependent (Dawson et al., 2012) and offers unique information that is pertinent for assessing endothelial function. Whereas greater dilation is favorable during FMD, greater constriction is favorable in the context of LFMC. Indeed, L-FMC has been shown to be blunted in smokers, hypertensive patients, and patients with coronary artery disease compared to healthy young and middle-aged adults (Gori et al., 2008). To further characterize an artery’s functional capabilities, one can also determine its resting diameter expressed as a percentage of its vasoactive range, known as resting vascular tone (Bell, Kelley, McCoy, & Credeur, 2017; Black, Vickerson, & McCully, 2003).

To date, the effects of high dietary sodium on brachial artery L-FMC have not been examined. Therefore, the objective of this study was to investigate the effects of high dietary sodium on brachial artery vascular tone and L-FMC response in healthy, salt-resistant adults. Based on our previous observations that excess sodium attenuates FMD, we hypothesized that brachial artery L-FMC would be attenuated on a high sodium diet compared to a low sodium diet.

METHODS

Participants

Fifteen healthy, salt-resistant adults between the ages of 23 and 43 yrs (9 men, 6 women) who participated in a larger feeding study conducted in our laboratory and for which L-FMC data were available were included in this study. FMD data from this feeding study has been previously reported for salt-resistant adults (DuPont et al., 2013; Lennon-Edwards et al., 2014). All participants were classified as salt-resistant, defined as a change in 24 hr ambulatory mean arterial pressure (MAP) of ≤5 mmHg between the low and high sodium diets. Women were premenopausal and non-pregnant, as confirmed by a negative pregnancy test; however, menstrual cycle phase was not controlled for in this study. Exclusion criteria included a body mass index equal to or greater than 30 kg/m², use of tobacco products, use of vasoactive medication, and a known history of cardiovascular disease, hypertension, malignancy, diabetes mellitus, or renal impairment. The study protocol and procedures were approved by the Institutional Review Board of the University of Delaware and conform to the provisions of the Declaration of Helsinki. Written informed consent was obtained from all participants prior to enrollment in the study.

Study Design

During an initial screening visit, scheduled after a 12 hr fast, participants underwent a resting 12-lead electrocardiogram (ECG) (MAC 5500 HD Resting ECG System, GE Medical Systems, Milwaukee, WI), measurements of BP (Dash 2000, GE Medical Systems, Milwaukee, WI), and measurements of height and weight (Health O Meter 500KL, Pelstar LLC, McCook, IL). As previously described (DuPont et al., 2013), baseline sodium intake was standardized using a 7-day run-in diet (100 mmol/day), after which participants completed a 7-day low sodium diet (LS; 20 mmol/day) and a 7-day high sodium diet (HS; 300 mmol/day) in a randomized crossover manner (Figure 1). Food was prepared by a registered dietitian and diets were comprised of 50% carbohydrates, 30% fat, and 20% protein. Potassium content was similar between the LS and HS diets. On the last day of each diet, participants underwent an assessment of brachial artery L-FMC, wore an ambulatory BP monitor for 24 hr, and completed a 24 hr urine collection. Venous blood samples were collected at the screening visit and on the last day of each of the LS and HS diets.

Figure 1.

Schematic representation of protocol. Following a screening visit, participants completed a 7-day medium sodium run-in diet, followed by a 7-day low sodium diet and a 7-day high sodium diet completed in randomized order with no washout period in between. Data was collected during the last 24 hr period of the low and high sodium diets.

Low-Flow Mediated Constriction

L-FMC was assessed during the arterial occlusion phase of an FMD test, which we previously reported (DuPont et al., 2013; Lennon-Edwards et al., 2014). Briefly, a BP cuff positioned on the forearm was rapidly inflated to 200 mmHg for 5 min in order to occlude blood flow to the distal vascular bed. Using a 12 MHz linear array ultrasound transducer (Logiq e, GE Healthcare, Waukesha, WI), longitudinal images of the brachial artery and Doppler blood velocities were obtained proximal to the antecubital fossa. Resting diameter was determined as the average diameter recorded over a 30 sec resting period. The occlusion diameter was determined as the average diameter recorded over the last 30 sec of cuff inflation (i.e. arterial occlusion). Peak diameter was determined as the largest 3-sec rolling average attained following cuff deflation (i.e. reactive hyperemia). Resting, occlusion, and peak diameters were analyzed across complete heart cycles using automated edge-detection software (Cardiovascular Suite, Quipu, Pisa, Italy) and used to calculate resting vascular tone (Bell et al., 2017; Black et al., 2003) and relative L-FMC (Gori et al., 2008), as in Eqs. 1–2, respectively:

$$\text{Resting}\text{Vascular}\text{Tone}(\%)=\frac{\text{peak}\text{diameter}-\text{resting}\text{diameter}}{\text{peak}\text{diameter}-\text{occlusion}\text{diameter}}\times 100$$ [1]

$$\text{L-FMC}(\%)=\frac{\text{occlusion}\text{diameter}-\text{resting}\text{diameter}}{\text{resting}\text{diameter}}\times 100$$ [2]

Twenty-Four Hour BP and Urine

During the last 24 hr period of the LS and HS diets, an ambulatory BP monitor (Spacelabs Medical, Issaquah, WA) worn on the non-dominant arm measured BP every 20 min during waking hours and every 30 min during sleep. Laboratory BP was also measured by an automated oscillometric sphygmomanometer at the LS and HS testing visits (Dinamap Dash 2000, GE Medical Systems). During the same 24 hr period as the ambulatory BP, urine was collected and analyzed for total volume, urinary electrolytes (EasyElectrolyte Analyzer, Medica, Bedford, MA), and urine osmolality (Advanced 3D3 Osmometer, Advanced Instruments, Norwood, MA).

Venous Blood Analysis

Venous blood samples were analyzed for hemoglobin (Hb 201+ model, HemoCue,Lake Forest, CA), hematocrit (Readacrit Centrifuge, Becton Dickinson, Sparks, MD), serum electrolytes (EasyElectrolyte Analyzer, Medica, Bedford, MA), and plasma osmolality (Advanced 3D3 Osmometer, Advanced Instruments, Norwood, MA).

Statistical Analyses

All data were assessed for normality using Shapiro-Wilk tests. Descriptive characteristics were used to report the demographic and biochemical profile of our participants at baseline. Paired samples T-tests were used to compare the biochemical, renal, hemodynamic, and vascular responses between the LS and HS diets. While this study was not powered for a sexanalysis, we report the results and effect size (partial eta squared, partial η²) of a supplementary two-factor analysis of variance (diet x sex ANOVA) used to compare L-FMC across the two dietary interventions between men and women. Statistical analyses were performed using SPSS Statistics (Version 20.0, Chicago, IL) and GraphPad Prism (Version 7.0d, La Jolla, CA). Significance was set at p<0.05 and data are expressed as mean ± standard deviation (SD).

RESULTS

Participant characteristics are reported in Table 1. In comparison to the LS diet, the HS diet increased serum sodium concentration (p<0.001; Table 2) and urinary sodium excretion (LS: 33 ± 23 mmol/24 hr, HS: 264 ± 97 mmol/24 hr, p<0.001; Figure 2A). Since all participants were salt resistant by design, 24 hr MAP did not differ between the two diets (LS: 84 ± 5 mmHg, HS: 83 ± 5 mmHg, p=0.16; Figure 2B).

Table 1.

Participant Characteristics

Demographic
N (men/ women)	15 (9/6)
Age, yrs	29 ± 6
Height, cm	175 ± 8
Mass, kg	74 ± 14
BMI, kg/m2	24 ± 3
SBP, mmHg	120 ± 12
DBP, mmHg	72 ± 7
MAP, mmHg	88 ± 8
HR, bpm	70 ± 8
Biochemical
eGFR, mL/min/1.73m2	103 ± 15
Hemoglobin, g/dL	14 ± 1
Hematocrit, %	42 ± 4
Serum Sodium, mmol/L	139 ± 1
Serum Potassium, mmol/L	4.3 ± 0.3
Serum Chloride, mmol/L	104 ± 2
Serum Creatinine, mg/dL	0.9 ± 0.1
Plasma Osmolality, mOsm/kg H2O	286 ± 4
Blood Urea Nitrogen, mg/dL	12 ± 4
Fasting Glucose, mg/dL	90 ± 7
Fasting Total Cholesterol, mg/dL	177 ± 44
Fasting HDL, mg/dL	54 ± 14
Fasting LDL, mg/dL	102 ± 36
Fasting Triglycerides, mg/dL	108 ± 49

Data are mean ± SD. BMI, body mass index; DBP, diastolic blood pressure; eGFR, estimated Glomerular Filtration Rate; HDL, high-density lipoproteins; HR, heart rate; LDL, low-density lipoproteins; MAP, mean arterial pressure; SBP, systolic blood pressure.

Table 2.

Physiological Responses to Dietary Sodium

	Low Sodium	High Sodium	P
Biochemical
Hemoglobin, g/dL	13 ± 2	13 ± 2	0.39
Hematocrit, %	42 ± 2	41 ± 5	0.49
Serum Sodium, mmol/L	138 ± 2	140 ± 2	<0.001
Serum Potassium, mmol/L	3.9 ± 0.3	4.2 ± 0.5	0.07
Serum Chloride, mmol/L	102 ± 2	106 ± 3	<0.001
Renal
Urine Flow Rate, mL/min	1.3 ± 0.6	1.7 ± 0.5	0.01
Urine Osmolality, mOsm/ kg H20	322 ± 82	420 ± 139	0.01
Hemodynamic
24 hr SBP, mmHg	114 ± 9	114 ± 9	0.79
24 hr DBP, mmHg	69 ± 5	68 ± 5	0.23
Laboratory SBP, mmHg	112 ± 11	113 ± 10	0.50
Laboratory DBP, mmHg	63 ± 5	65 ± 6	0.19
Laboratory MAP, mmHg	80 ± 5	81 ± 7	0.58
Vascular
Resting Diameter, mm	3.87 ± 0.50	3.88 ± 0.49	0.92
Occlusion Diameter, mm	3.85 ± 0.51	3.89 ± 0.50	0.41
Range (peak – occlusion), mm	0.24 ± 0.16	0.21 ± 0.11	0.41

Data are mean ± SD. DBP, diastolic blood pressure; MAP, mean arterial pressure; SBP, systolic blood pressure. High dietary sodium increased serum sodium, serum chloride, urine flow rate, and urine osmolality but attenuated relative LFMC.

Figure 2.

Urine sodium excretion (A) and mean arterial pressure (B) over a 24 hr period in healthy, salt-resistant adults on a low sodium (white bar) and high sodium (black bar) diet. High dietary sodium increased urine sodium excretion (p<0.001), but by design, mean arterial pressure was unchanged (p=0.16).

There were also no differences between diets in the resting diameter (p=0.92), occlusion diameter (p=0.41), or absolute range (i.e. peak diameter − occlusion diameter, p=0.41; Table 2). Nevertheless, the HS diet augmented the resting vascular tone (LS: 91 ± 23%, HS: 125 ± 56%, p=0.01; Figure 3A) and attenuated the L-FMC response (LS: −0.58 ± 0.99%, HS: 0.17 ± 1.23%, p=0.008; Figure 3B).

Figure 3.

Brachial artery resting vascular tone (A) and relative L-FMC (B) in healthy, salt-resistant adults on a low sodium diet (white bar) and high sodium diet (black bar). Group means are presented in the left panel and individual data are presented in the right panel. High dietary sodium increased vascular tone (p=0.01) and attenuated L-FMC (p=0.008).

Brachial artery L-FMC was also variable between participants, ranging from −2.12% (vasoconstriction) to 2.75% (vasodilation). On the LS diet, using absolute 0 as a cut-off, 60% of participants vasoconstricted, 13% had no response, and 27% vasodilated. In contrast, on the HS diet, 53% of participants constricted and 47% vasodilated.

While this study was not powered for a sex comparison, supplementary analysis supported an effect of diet, but not sex, on L-FMC (two-factor ANOVA: main effect of diet, p=0.005, partial η²=0.295; no effect of sex, p=0.95, partial η²=0.003; no interactive effect of diet and sex, p=0.24, partial η²<0.001).

DISCUSSION

The novel findings from this study were that 7-days of high dietary sodium augmented resting vascular tone and attenuated L-FMC in the brachial artery of healthy salt-resistant adults. Although termed low-flow mediated constriction due to initial observations that arteries constrict during acute reductions in blood flow (Anderson & Mark, 1989; Gori et al., 2008; Levenson, Simon, & Pithois-Merli, 1987), we observed variable responses in the brachial artery. Our observation that 60% of participants constricted on the LS diet is similar to the 59% reported by Harrison et al. (2011) and the 58% reported by Aizawa et al. (2016) in studies of older adults ranging in health and cardiovascular disease risk. Interestingly, in the present study, the percentage of participants who dilated during cuff inflation (an unfavorable L-FMC response) increased from 27% on the LS diet to 47% on the HS diet. The greater prevalence of a dilatory response and the overall attenuation in L-FMC on the HS diet may, in part, have resulted from the concomitant increase in vascular tone. This is further supported by the fact that the absolute range was unchanged.

These effects of high dietary salt on brachial artery vasoreactivity closely resemble the effects of low aerobic fitness. Previously, Bell et al. (2017) demonstrated that young, healthy men with lower fitness had 24% greater vascular tone and 1.8% less brachial artery L-FMC compared to those with higher fitness. While we did not control for fitness levels in our study population, the vascular tone and L-FMC response of our participants while on the LS diet (vascular tone: 91%, L-FMC: −0.6%) were comparable to values previously reported for lower fit men (vascular tone: 95%, L-FMC: −0.7%; Bell et al., 2017). Further, whereas vascular tone is lower and L-FMC is larger with higher fitness levels (vascular tone: 71%, L-FMC: −2.5%; Bell et al. 2017), our findings suggest that high dietary sodium has the opposite effect.

One mechanism by which high dietary sodium may have increased vascular tone and attenuated L-FMC is by increasing oxidative stress (Edwards & Farquhar, 2015). The degree to which an artery is constricted under basal conditions is regulated by the release of endothelium-derived vasodilators (i.e. nitric oxide, NO; endothelium-derived hyperpolarizing factor, EDHF;prostacyclin, PGI₂) and vasoconstrictors (i.e. endothelin-1, ET-1; thromboxane A₂, TXA₂), which act to relax or contract the underlying smooth muscle, respectively (Haynes & Webb, 1994; Sandoo, Veldhuijzen van Zanten, Metsios, Carroll, & Kitas, 2010; Vallance, Collier, & Moncada, 1989). In both rodents (Lenda, Sauls, & Boegehold, 2000) and humans (Greaney et al., 2012; Ramick et al., 2019) salt loading has been shown to increase the production of reactive oxygen species, which scavenge NO thereby reducing its bioavailability (Förstermann & Münzel, 2006). Additionally, in young healthy women, 7 days of a HS diet was shown to increase plasma levels of TXA_2, a cyclooxygenase-derived vasoconstrictor (Cavka et al., 2015)_. Assuming no other compensatory dilatory responses, a decrease in NO bioavailability coupled with an increase in TXA₂ production would result in greater basal arterial constriction. However, during a low-flow state, constriction appears to be independent of NO and instead involves the coordinated inhibition of EDHF and/or PGI₂ and the increased production of ET-1 (Gori et al., 2008). Therefore, through this potential oxidative stress mechanism, the attenuations in L-FMC would be a by-product of salt-induced augmentations in vascular tone. Alternatively, high dietary sodium may be directly interfering with EDHF inhibition. High salt diets have been shown to upregulate EDHF in animal models (Goto et al., 2012; Sofola, Knill, Hainsworth, & Drinkhill, 2002); however, future research is needed to understand whether this plays a role in L-FMC in humans on a high salt diet.

Importantly, our findings in salt-resistant adults suggest that high dietary sodium is influencing vascular function independent of changes in BP. It is well established that excess sodium intake increases the risk of cardiovascular disease and mortality (Strazzullo, D’Elia, Kandala, & Cappuccio, 2009; Tuomilehto et al., 2001) and its deleterious effects have traditionally been associated with increases in BP (Kuller, 1997; Tzemos, Lim, Wong, Struthers, & Macdonald, 2008). While Americans undoubtedly consume more sodium than is recommended for the general population (3440 mg/day on average vs. 2300 mg/day max limit) (U.S. Department of Health and Human Services and U.S. Department of Agriculture., 2015),the majority of young to middle-aged normotensive adults are salt-resistant and do not demonstrate increases in BP with salt loading (Weinberger, 1996). Our findings that brachial artery tone and L-FMC were altered in salt-resistant adults adds to a body of emerging literature that suggests excess salt intake can directly impair vascular function (Boegehold, 1995; Dickinson, Clifton, & Keogh, 2011; DuPont et al., 2013; Greaney et al., 2012).

Limitations

This study did not measure markers of oxidative stress in response to high dietary sodium, thus its role in increasing vascular tone and attenuating L-FMC is speculative and necessitates further investigation. Follow-up mechanistic studies are also needed to elucidate whether attenuations in L-FMC reflect sodium-induced impairments in endothelial and/or smooth muscle function or rather a “lack of further recruitability” due to augmented vascular tone (Gori et al., 2008). It should be noted that vascular tone, calculated in accordance with Black et al. (2003) and Bell et al. (2017), exceeded 100% when the occlusion diameter was larger than the resting diameter. Nevertheless, these data reflect a dilatory response during the occlusion period, which was more prevalent on the HS diet. As this was a retrospective analysis, the Doppler sample volume was not always optimized during the cuff inflation period of an FMD test. As such, L-FMC blood flow and shear stress data were not available but would have aided in the interpretation of our findings. Lastly, this study was not powered for a sex comparison, but exploratory analysis suggested no sex-based differences.

Conclusions

Through this well-controlled, crossover feeding study, we demonstrated that salt-resistant adults have an augmented vascular tone and an attenuated brachial artery L-FMC response while on a 7-day HS diet. In light of the fact that this was a retrospective study, prospective examinations on the effects of dietary sodium on L-FMC are warranted. Nevertheless, our findings extend our previous work on the effects of salt loading on FMD and further demonstrate the need to investigate vascular responses to sodium, independent of changes in BP.

ABBREVIATIONS

BP

Blood pressure

DBP

Diastolic blood pressure

ECG

Electrocardiogram

EDHF

Endothelium-derived hyperpolarizing factor

ET-1

Endothelin-1

FMD

Flow-mediated dilation

HS

High sodium

L-FMC

Low-flow mediated constriction

LS

Low sodium

MAP

Mean arterial pressure

NO

Nitric oxide

PGI₂

Prostacyclin

SBP

Systolic blood pressure