Acute salt loading and cardiotonic steroids in resistant hypertension

CHAPTER ONE

Acute salt loading and cardiotonic steroids in resistant hypertension Igor V. Emelyanova, Alexandra O. Konradia, Edward G. Lakattab, Olga V. Fedorovab, Alexei Y. Bagrovb,c,* a

Almazov Federal Heart, Blood and Endocrinology Centre, St. Petersburg, Russia National Institute on Aging, NIH, Baltimore, MD, United States c Sechenov Institute of Evolutionary Physiology and Biochemistry, St. Petersburg, Russia *Corresponding author: e-mail address: [email protected] b

Contents 1. Introduction 2. Methods 2.1 General 2.2 Hemodynamic measurements 2.3 Salt-loading protocol 2.4 Statistical analyses 3. Results 4. Discussion Acknowledgments References Further reading

2 3 3 4 4 5 6 8 12 12 13

Abstract The study addresses the association of marinobufagenin (MBG), a natriuretic and vasoconstrictor steroid, and Na/K-ATPase (NKA) activity with pressor response to salt-loading and arterial stiffness in resistant hypertension (RH). Thirty-four patients (18 males and 16 females; 56
8 years) with RH on a combined (lisnopril/amlodipine/hydrochlorothiazide) therapy and 11 healthy age-matched normotensive subjects (7 males and 4 females; 54
2 years) were enrolled in this study. Salt-loading was performed via intravenous infusion of 1000 mL saline (0.9% NaCl) for 1 h. Arterial stiffness was measured by Sphygmocor Px device with a calculation of pulse-wave velocity (PWV). Activity of NKA was measured in erythrocytes. We demonstrated that plasma levels of MBG and magnitude of NaCl-induced MBG-dependent NKA inhibition are associated with PWV, and that this association has gender- and age-specific fashion in RH patients.

Current Topics in Membranes, Volume 83 ISSN 1063-5823 https://doi.org/10.1016/bs.ctm.2019.01.005

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2019 Elsevier Inc. All rights reserved.

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Abbreviations BP CO CS DBP MBG NKA PWV RH SBP

blood pressure cardiac output cardiotonic steroids diastolic blood pressure marinobufagenin Na/K-ATPase pulse-wave velocity resistant hypertension systolic blood pressure

1. Introduction Excessive salt ingestion plays an important role in pathogenesis of hypertension (Folkow, 1982; Weinberger, Miller, Luft, Grim, & Fineberg, 1986). One of major factors mediating compromised ability of the kidneys to excrete sodium is Na/K-ATPase (NKA) and its endogenous inhibitors, cardiotonic steroids (CS) including marinobufagenin (MBG) (Bagrov, Shapiro, & Fedorova, 2009). In the kidney, NKA and MBG represent major sodium transporting mechanism (Fedorova & Bagrov, 1997; Hamlyn & Blaustein, 1986). According to the “concept of a natriuretic hormone,” the primary role of CTS is to promote natriuresis via inhibition of sodium reabsorption in the renal tubules (Hamlyn & Blaustein, 1986). CS promote natriuresis via inhibition of the NKA in the proximal tubules and thick ascending limb (TAL) of Henle’s loop (Fedorova, Kashkin, Zakharova, Lakatta, & Bagrov, 2012). In addition to the effect of CS per se, a number of substances (natriuretic peptides, dopamine, calcitonin, etc.) exhibit natriuretic effect via modulation of NKA activity by cGMP in the TAL of Henle’s loop (Aperia et al., 1994; Bailly, 2000; Florkowski, Richards, Espiner, Yandle, & Frampton, 1994). This segment of nephron, therefore, represents a unique site for the interaction of various natriuretic compounds. Only about 15%–25% of filtered sodium is reabsorbed in TAL; the importance of this segment of nephron in sodium metabolism could be pictured by the fact that a reduction in salt reabsorption by TAL could determine pathogenesis of Bartter’s disorder (Hebert, 2003). In addition, CS exhibit growth-promoting and pro-fibrotic effects via NKAdependent signaling in cardiovascular tissues (Fedorova, Emelianov, et al., 2015; Khalaf et al., 2018; Xie & Askari, 2002). Despite significant progress in the development of antihypertensive therapies, hypertension remains a major determinant of cardiovascular mortality

Acute salt loading and cardiotonic steroids

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worldwide (Zanchetti, Grassi, & Mancia, 2009), and the achievement of blood pressure (BP) target level remains to be poor on the population level. About 10%–12% of hypertensive patients could be characterized as truly drug-resistant, i.e., when target BP cannot be achieved with a three-drug combination therapy, including a diuretic, given in adequate doses (Mancia et al., 2007). Mechanisms responsible for drug resistance are complex and are not fully understood. One of major predictors of drug resistance includes an uncontrolled salt and liquid intake (Folkow, 1982). Previously we studied CS in two human NaCl-loading studies, SALT and SARAH (Anderson et al., 2008; Fedorova, Lakatta, Bagrov, & Melander, 2015). Data from both SALT and SARAH studies suggest that MBG is an important determinant of BP, and that its reaction to salt and its relation to salt-sensitivity of BP differ by age and gender (Anderson et al., 2008; Fedorova, Lakatta, et al., 2015). While in older females (SALT study), a relative failure in MBG production may cause volume expansion and contribute to hypertension, in males aging is associated with lower baseline MBG levels, but with a greater responsiveness of MBG to HS (SARAH study) (Anderson et al., 2008; Fedorova, Lakatta, et al., 2015). The major goal of this study was to investigate whether NaCl-induced MBG response in resistant hypertension (RH) is associated with saltsensitivity of BP and with vascular stiffness. We hypothesized that (i) levels of MBG and magnitude of NaCl-induced MBG-dependent NKA inhibition would be associated with pulse-wave velocity (PWV), and (ii) that this association will exhibit gender and age specificity.

2. Methods 2.1 General We examined 21 patients with true RH (11 males and 10 females, mean age 56
8 years) from the Outpatient Resistant Hypertension Centre of the Almazov Federal Heart, Blood and Endocrinology Centre (St. Petersburg, Russia). The study was approved by local Ethic Committee and by Medstar Research Institute, and the informed consent was obtained from all participants. The inclusion criteria were: essential hypertension with a duration over 5 years, signed informed consent, stable three-drug full dose conventional antihypertensive therapy (lisnopril/amlodipine/hydrochlorothiazide) with office BP greater than 140/90 mmHg. All patients were carefully examined to exclude secondary causes of hypertension including obstructive sleep apnea; ambulatory BP monitoring was performed to exclude the

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Igor V. Emelyanov et al.

“white coat” effect. Patients with concomitant coronary artery disease, previous stroke or transient ischemic attacks, renal insufficiency, severe concomitant diseases, known poor treatment adherence, drug abuse, body mass index (BMI) over 35 kg/m2 were excluded. For 2 weeks before the test, patents were prescribed standard three-drug antihypertensive therapy in rational combinations: ACE inhibitor lisinopril 20 mg per day, calcium channel blocker amlodipine 10 mg per day and diuretic hydrochlorothiazide 25 mg per day. The diagnosis of RH was reconfirmed after 2 weeks. The control group included 11 sex- and age-matched subjects (7 males and 4 females, mean age 54
2 years).

2.2 Hemodynamic measurements Heart rate was monitored by electrocardiography and BP was measured by finger plethysmography (Finometer; TNO, Amsterdam, Netherlands) of the right index or middle finger intermittently recalibrated against oscillometry. Beat-to-beat arterial BP was noninvasively and continuously measured by a fully automated Finometer® device (Finapres Medical Systems, TNO-BMI, Netherlands), which uses a volume clamp technique and has been well-validated against intra-arterial BP measurements. Systolic BP (SBP) and diastolic BP (DBP) were measured on a beat-by-beat basis. The beat-by-beat stroke volume was derived from the blood pressure waveform using Beatscope® software (1.1a, TNO-BMI, Amsterdam, Netherlands), a program that employs the Modelflow technique computing cardiac output (CO) by simulating a nonlinear three-element Windkessel model of aortic input impedance.

2.3 Salt-loading protocol The salt-loading test was performed in the morning in a quiet room with a stable air temperature 19–22°С. Patients fasted 12
2 h before the test, and morning medications were not taken. Test was started after 30 min of rest in a supine position with an i.v. cubital catheter and with an empty urine bladder. All measurement equipment was also applied before the test. The test was divided into three parts. First, 60 min of baseline rest during which hemodynamic parameters were monitored and blood samples obtained at baseline and after 60 min of rest. Second, after 60 min of rest the infusion of 1000 mL of 0.9% NaCl was performed for 60 min (16 mL/min) followed by continuous BP and hemodynamic monitoring. Finally, the blood samples were obtained after the infusion peak, and the hemodynamic parameters

Acute salt loading and cardiotonic steroids

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were registered for an additional 60 min as a recovery period followed by the blood sampling at the end. Salt-sensitivity of BP was defined as SBP increase during the test 20 mmHg or more and/or DBP 10 mmHg or more (Luft, Weinberger, Grim, & Fineberg, 1986). Activity of NKA in erythrocytes was measured after the ex vivo incubation of the red blood cells with monoclonal anti-MBG mAb or with vehicle as described below. 2.3.1 Erythrocyte Na/K-ATPase Erythrocytes were washed three times in an isotonic medium (145 mmol/L NaCl in 20 mmol/L Tris buffer; pH 7.6 at 4°C). Erythrocytes were preincubated with Tween-20 (0.5%) in sucrose (250 mmol/L) and Tris buffer (20 mmol/L; pH 7.4, t ¼ 37°C) for 30 min, and were incubated for 30 min in the medium (mmol/L): Na 100, K 10, MgCl2 3, EDTA 0.5, Tris 50, ATP 2 (pH 7.4, t ¼ 37°C) in the final dilution 1:40. The reaction was stopped by the addition of trichloroacetic acid to final concentration 7%. Total ATPase activity was measured by the production of inorganic phosphate (Pi), and NKA activity was estimated as the difference between ATPase activity in the presence and in the absence of 5 mmol/L ouabain. Whole blood was preincubated for 1 h with a vehicle or 3E9 anti-MBG monoclonal antibody. 2.3.2 MBG immunoassay Plasma samples were extracted on Sep-Pak C-18 cartridges (Waters, Milford, MA, USA), and MBG competitive fluoroimmunoassay based on a murine monoclonal anti-MBG 4G4 antibody was performed as described recently. The cross-reactivity of 4G4 anti-MBG antibody is: MBG—100%, ouabain—0.005%, digoxin—0.03%, digitoxin <0.001%, bufalin—0.08%, cinobufagin—0.07%, cinobufatalin—40%, prednisone, spironolactone, aldosterone, proscillaridin, and progesterone <0.001%.

2.4 Statistical analyses Data analysis was performed by Statistica for Windows, version 7.0 (StatSoft). All data are presented as means
SEM. Normality of data distribution was assessed using Kolmogorov–Smirnov test. Nonparametric statistical methods were used when variables did not normally distribute. Hemodynamic data at different experimental stages were assessed by one-way or two-way analysis of variance (ANOVA) for repeated measures or by the Friedman test for nonparametric distributions, followed by the Bonferroni correction. The correlations were assessed using linear regression and Pearson correlation analysis.

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3. Results We studied 34 patients (18 males and 16 females; 56
8 years) with RH, in which BP remained over 140/80 on a combined therapy. NaCl loading was performed via intravenous infusion of 1000 mL saline for 1 h. Control group and patient characteristics are presented in Table 1. Table 2 compares hemodynamic changes in patients and in the control group during salt loading. BP and total peripheral resistance increased in both groups, although in the hypertensive group it exceeded those in controls. As presented in Fig. 1, NaCl loading resulted in the pressor response, increase in plasma levels of MBG and in a twofold inhibition of erythrocyte NKA which was blocked by anti-MBG antibody. Because anti-MBG mAb reversed NaCl-induced NKA inhibition, MBG is responsible for this NKA-inhibitory effect of NaCl loading. As presented in Table 2, plasma levels of MBG have increased in salt-loaded subjects, which was associated with inhibition of erythrocyte NKA. Table 1 Clinical and demographic characteristics. Control subjects (n 5 11)

Patients with RH (n 5 34)

54
2

56
8

28.5
0.4

32.2
5.4*

Gender, males/females

7/4

18/16

24 h systolic BP, mmHg

131.7
1.8

149.5
5.8**

24 h diastolic BP, mmHg

82.0
1.5

91.1
5.2**

Heart rate, beats/min

70.2
1.4

72.5
5.7

PWV, m/s

7.1
0.5

7.9
1.8*

Serum creatinine, μmol/L

75.5
2.2

85.8
9.1**

Glomerular filtration rate, mL/min

94.6
4.3

76.7
14.9**

Plasma sodium, mmol/L

140.7
1.4

141.6
4.9

Fasting glucose, mmol/L

5.55
0.21

6.07
0.66*

Total cholesterol, mmol/L

5.31
0.16

6.11
0.53**

Parameter

Age, years Body mass index, kg/m

2

Data are means
SEM, *P < 0.05; **P < 0.01, RH vs. controls.

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Acute salt loading and cardiotonic steroids

Table 2 Effects of acute salt loading. Parameter

Control subjects (n 5 11)

Patients with RH (n 5 34)

SBP baseline, mmHg

133
3

156
3***

DBP baseline, mmHg

82
2

90
2***

CO baseline, mL/min

5.9
0.2

5.5
0.3

Plasma MBG baseline, nmol/L

0.14
0.02

0.15
0.01

NKA baseline, μmol Pi/mL/h

3.18
0.45

2.2
0.1*

Total peripheral resistance, baseline, din  s  sm5

1087
18

1414
28*

SBP final, mmHg

145
4

180
5***

DBP final, mmHg

88
3

102
3***

CO final, mL/min

6.4
0.3

6.0
0.4

Plasma MBG final, nmol/L

0.19
0.01

0.23
0.01

NKA final μmol Pi/mL/h

2.41
0.44

1.6
0.01**

Total peripheral resistance, final, din  s  sm5

1345
24

2025
27**

Data are means
SEM. 2-way ANOVA, *P < 0.05, **P < 0.01, ***P < 0.001, RH vs. controls. baseline, before salt-loading; CO, cardiac output; DBP, diastolic blood pressure; final, after salt-loading; RH, resistant hypertension; SBP, systolic blood pressure.

Fig. 2 presents data on correlations of the magnitude of NaCl-induced NKA inhibition in the hypertensive group. While there were no association between the magnitude of NaCl-induced NKA inhibition and magnitude of BP response (A–C), delta NKA exhibited a positive linear correlation with age (D–F), and in male, but not in the female subjects, delta NKA positively correlated with PWV (G–I). Thus, in patients with RH magnitude of MBG-induced NKA inhibition associates not with the salt-sensitivity of BP per se, but with age of patients and with PVW which characterizes vascular stiffness, a major feature of cardiovascular aging. In accordance with the above finding, in male, but not in female subjects, PVW exhibited positive linear correlation with both NaCl-induced renal MBG response, and with its physiological effect, i.e., natriuretic response (Fig. 3). Importantly, a magnitude of renal MBG response did not correlate with salt-sensitivity of BP.

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Igor V. Emelyanov et al.

A

B Diastolic BP

Systolic BP 110

200

P<0.01

P<0.01 180 mmHg

mmHg

100 160 140

90

120 100

C

Bl

Plasma MBG 0.6

80

NaCl D

Bl

NaCl

Erythrocyte Na/K-ATPase

P<0.01 P<0.01

μmol Pi / ml / hour

3

nmol/L

0.4

0.2

0.0

2

1

NaCl + 3E9

Bl NaCl

0 Bl

NaCl

Fig. 1 Effect of acute NaCl loading on systolic (A) and diastolic (B) BP, on plasma MBG concentration (C) and on erythrocyte Na/K-ATPase (D) in patients with resistant hypertension. “NaCl + 3E9”—ex vivo effect of anti-MBG mAb on Na/K-ATPase in erythrocytes obtained following NaCl loading. Bl, baseline. t-test and 1-way ANOVA.

4. Discussion In the present study we demonstrate that in patients with RH on a high NaCl, MBG response do not correlate with BP response, but rather is related to PWV. Our present findings are in agreement with the growing body of evidence that, with respect to pathogenesis of hypertension, the effects of CS on growth and collagen synthesis (vascular stiffness) may be more important than the effects of CS on BP. However, it appears that CS responses on the background of established hypertension are not

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Acute salt loading and cardiotonic steroids

B

Both sexes r = 0.04 P = 0.97

20 10 0 –10

15 10 5

0.5

1.0

D

0.5

80

1.0

r = 0.45 P = 0.006

0.0

F

Males

0.0

0.5

1.0

60

40

G

1.5

0.0

PVW (m/sec)

10

5

r = 0.26 P = 0.16 0.5

1.0

1.5

Δ Na/K-ATPase (μmol Pi/ml/hr)

1.0

1.5

60

40

r = 0.48 P = 0.04

20

1.5

0.0

0.5

5

0

1.5

Females r = 0.15 P = 0.57

15

r = 0.55 P = 0.03

10

1.0

Δ Na/K-ATPase (μmol Pi/ml/hr)

I

Males 15

0.0

0.5

H

Both sexes

1.0

Females

Δ Na/K-ATPase (μmol Pi/ml/hr)

15

0

r = –0.47 P = 0.044

20

Δ Na/K-ATPase (μmol Pi/ml/hr)

0.5

80

PVW (m/sec)

20

10

Δ Na/K-ATPase (μmol Pi/ml/hr)

Age (years)

Age (years)

40

20

1.5

80

60

r = 0.05 P = 0.97

30

Δ Na/K-ATPase (μmol Pi/ml/hr)

E

Both sexes

Females 40

0 0.0

1.5

Δ Na/K-ATPase (μmol Pi/ml/hr)

Age (years)

r = –0.10 P = 0.73

0 0.0

PVW (m/sec)

C

Males 20

Δ DBP (mm Hg)

Δ DBP (mm Hg)

30

Δ DBP (mm Hg)

A

10

5

0 0.0

0.5

1.0

1.5

Δ Na/K-ATPase (μmol Pi/ml/hr)

0.0

0.5

1.0

1.5

Δ Na/K-ATPase (μmol Pi/ml/hr)

Fig. 2 Correlations of NaCl-loading-induced inhibition of erythrocyte Na/K-ATPase. Relationship between MBG-dependent inhibition of Na/K-ATPase and magnitude of NaCl-induced change of diastolic BP (A–C), age of the patients (D–F), and PWV (G–I).

anymore associated with the salt-sensitivity of BP, but rather are implicated in cardiovascular remodeling (Fig. 4). Analysis of present data suggests that involvement of CS in the response to NaCl challenge is altered with age, and that there is a gender difference in the response of CS to the saltloading. Data from the previous studies (Anderson et al., 2008; Fedorova, Lakatta, et al., 2015) suggest that MBG is an important determinant of BP and that its reaction to salt and its relation to salt-sensitivity of BP differ by age and gender. While in older females, a relative failure in MBG production may cause volume expansion and contribute to hypertension, in males, aging is associated with lower baseline MBG levels, but a greater responsiveness of

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Igor V. Emelyanov et al.

r = 0.31 P = 0.08

1.5 1.0 0.5 0.0 10 PWV (m/sec)

1.5 1.0 0.5

15

5 10 PWV (m/sec)

E

1.5 1.0 0.5

15

0

Males 150

Na excretion (mEq / 4hr)

100

15

Females

r = 0.46 P = 0.03

200

100

Na excretion (mEq / 4hr)

r = 0.40 P = 0.02

150

5 10 PWV (m/sec)

F

Both sexes

Na excretion (mEq / 4hr)

r = 0.22 P = 0.41

0.0 0

D

Females 2.0

r = 0.51 P = 0.03

0.0 5

200

C

Males 2.0

MBG excretion (nmoles per 4 hours)

MBG excretion (nmoles per 4 hours)

B

Both sexes 2.0

MBG excretion (nmoles per 4 hours)

A

50

50

r = 0.37 P = 0.16

150 100 50

0

0 5

10 PWV (m/sec)

15

0 0

5 10 PWV (m/sec)

15

0

5

10

15

PWV (m/sec)

Fig. 3 Pearson correlation of NaCl-induced MBG response and PWV (A–C), and relationship between natriuretic response to the saline loading and PWV (D–F) in patients with resistant hypertension.

MBG to administration of high salt diet (Anderson et al., 2008; Fedorova, Lakatta, et al., 2015). Subsequently, independent positive associations of SBP with urine MBG/Na ratio were found in black women, which supported idea that reduced MBG-mediated Na excretion can contribute to adverse hemodynamics (Strauss, Smith, et al., 2018). More recently it was shown that at a young age heightened endogenous MBG levels may contribute to large artery stiffness in women via pressure-independent mechanisms, increasing their risk for future cardiovascular disease (Strauss, Smith, Wei, Fedorova, & Schutte, 2018). These human observations were supported by the experimental data showing that high-salt induced aortic stiffness in normotensive animals occurred in the presence of elevated MBG (Grigorova et al., 2018). Interestingly, a decrease in a salt consumption restored aortic elasticity and diminished the risk of cardiovascular disease by reduction of production of the pro-fibrotic factor MBG (Grigorova et al., 2018). Two signaling pathways were shown to modulate pro-fibrotic effects of MBG (Fig. 4). Pro-fibrotic effects may be initiated by MBG is

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Acute salt loading and cardiotonic steroids

Ionic Pathway (NKA inhibition)

MBG

Signaling Pathway (Signal transduction via binding to NKA)

NKA Cytosolic Na+ +

Cytosolic K

c-SRC phosphorylation EGFR phosphorylation PLCγ phosphorylation

Cytosolic Ca2+ Ca2+ oscillation

TGFβ1 activation SMADs activation/ phosphorylation

PKCδ phosphorylation Fli-1 phosphorylation

Increased vascular smooth muscle cells contractility

Blood pressure increase

Phosphorylated Fli-1 releases collagen-1 DNA promoter Gene expression and proliferation Increase in collagen production Vascular fibrosis

Fig. 4 The schematic presentation of the possible molecular mechanisms of the implication of MBG in blood pressure regulation via ionic pathway (Na+/K+-mediated signaling) and stimulation of vascular fibrosis via Na+/K+-independent signaling. c-SRC, protooncogene tyrosine-protein kinase; EGFR, epidermal growth factor receptor; Fli-1, Friend leukemia integration 1 transcription factor, a negative regulator of collagen-1 production; MBG, marinobufagenin; NKA, Na/K-ATPase; PKCδ, protein kinase C delta; PLCγ, phospholipase C gamma; SMADs, mothers against DPP homologs; TGFβ, transforming growth factor beta.

SMAD-dependent TGFβ-1 signaling which underlies vascular fibrosis in salt-induced aortic stiffness in normotensive rats (Grigorova et al., 2018). Another system, inhibition of Fli1, a nuclear transcription factor and a member of ETS family is implicated in MBG-induced fibrosis (Fedorova, Emelianov, et al., 2015). Fli1 acts as a negative regulator of collagen-1 synthesis and it competes with another transcription factor, ETS-1, to maintain a balance between stimulation and repression of Col1a2 gene (Trojanowska, 2010). Preeclampsia is associated with the elevated levels of BP and MBG and fibrosis of umbilical arteries, and by a dramatic reduction in the levels of Fli1 (Fedorova et al., 2018). In a group of patients with mild hypertension in the presence of NaCl reduction levels of MBG also reduced and they were negatively related to aortic PWV ( Jablonski et al., 2013). Recently in a small group of RH patients, an addition of spironolactone to the therapy was associated with a drop in PWV and with an increase in erythrocyte NKA activity (Fedorova, Emelianov, et al., 2015). In summary, in the present study it was demonstrated that MBG plays an important role in the regulation of vascular

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stiffness, and the further clinical studies will provide additional information related to the metabolic pathways activated by MBG.

Acknowledgments Authors are grateful to Elena Frolova and Anton Bzhelyanskiy for technical support. This study was supported by Intramural Research Program, National Institute on Aging, National Institutes of Health (O.V.F., E.G.L., A.Y.B.), by a grant from Russian Federal Agency for Scientific Organizations (Mechanisms of the formation of physiological functions in phylogenesis and € Haakon, Narva, ontogenesis. Endogenous and exogenous regulation) (A.Y.B.), by OU Estonia, and by a grant from Russian Federation No. 14.740.11.0928 (I.V.E., A.O.K.).

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Grigorova, Y. N., Wei, W., Petrashevskaya, N., Zernetkina, V., Juhasz, O., Fenner, R., et al. (2018). Dietary sodium restriction reduces arterial stiffness, vascular TGF-β-dependent fibrosis and marinobufagenin in young normotensive rats. International Journal of Molecular Sciences, 19(10), pii: E3168. Hamlyn, J. M., & Blaustein, M. P. (1986). Sodium chloride, extracellular fluid volume, and blood pressure regulation. The American Journal of Physiology, 251(4 Pt 2), F563–F575. Hebert, S. C. (2003). Bartter syndrome. Current Opinion in Nephrology and Hypertension, 12, 527–532. Jablonski, K. L., Fedorova, O. V., Racine, M. L., Geolfos, C. J., Gates, P. E., Chonchol, M., et al. (2013). Dietary sodium restriction and association with urinary marinobufagenin, blood pressure, and aortic stiffness. Clinical Journal of the American Society of Nephrology, 8, 1952–1959. Khalaf, F. K., Dube, P., Mohamed, A., Tian, J., Malhotra, D., Haller, S. T., et al. (2018). Cardiotonic steroids and the sodium trade balance: New insights into trade-off mechanisms mediated by the Na+/K+-ATPase. International Journal of Molecular Sciences, 19, pii: E2576. Luft, F. C., Weinberger, M. H., Grim, C. E., & Fineberg, N. S. (1986). Effects of volume expansion and contraction on potassium homeostasis in normal and hypertensive humans. Journal of the American College of Nutrition, 5, 357–369. Mancia, G., De Backer, G., Dominiczak, A., Cifkova, R., Fagard, R., Germano, G., et al. (2007). Guidelines for the management of arterial hypertension: The task force for the management of arterial hypertension of the European Society of Hypertension (ESH) and of the European Society of Cardiology (ESC). Journal of Hypertension, 25(2007), 1105–1187. Strauss, M., Smith, W., Kruger, R., Wei, W., Fedorova, O. V., & Schutte, A. E. (2018). Marinobufagenin and left ventricular mass in young adults: The African-PREDICT study. European Journal of Preventive Cardiology, 25, 1587–1595. Strauss, M., Smith, W., Wei, W., Fedorova, O. V., & Schutte, A. E. (2018). Marinobufagenin is related to elevated central and 24-h systolic blood pressures in young black women: The African-PREDICT study. Hypertension Research, 41, 183–192. Trojanowska, M. (2010). Cellular and molecular aspects of vascular dysfunction in systemic sclerosis. Cellular and molecular aspects of vascular dysfunction in systemic sclerosis. Nature Reviews Rheumatology, 6, 453–460. Weinberger, M. H., Miller, J. Z., Luft, F. C., Grim, C. E., & Fineberg, N. S. (1986). Definitions and characteristics of sodium sensitivity and blood pressure resistance. Hypertension, 8, II127–II134. Xie, Z., & Askari, A. (2002). Na(+)/K(+)-ATPase as a signal transducer. European Journal of Biochemistry, 269, 2434–2439. Zanchetti, A., Grassi, G., & Mancia, G. (2009). When should antihypertensive drug treatment be initiated and to what level should systolic blood pressure be lowered? A critical reappraisal. Journal of Hypertension, 27, 923–934.

Further reading Blaustein, M. P., Ashida, T., Goldman, W. F., Wier, W. G., & Hamlyn, J. M. (1986). Sodium/calcium exchange in vascular smooth muscle: A link between sodium metabolism and hypertension. Annals of the New York Academy of Sciences, 488, 199–216.