Dietary Sodium Restriction Reduces Arterial Stiffness, Vascular TGF-β-Dependent Fibrosis and Marinobufagenin in Young Normotensive Rats

Abstract

High salt (HS) intake stimulates the production of marinobufagenin (MBG), an endogenous steroidal Na/K-ATPase ligand, which activates profibrotic signaling. HS is accompanied by a blood pressure (BP) increase in salt-sensitive hypertension, but not in normotensive animals. Here, we investigated whether HS stimulates MBG production and activates transforming growth factor-beta (TGF-β) profibrotic signaling in young normotensive rats, and whether these changes can be reversed by reducing salt to a normal salt (NS) level. Three-month old male Sprague–Dawley rats received NS for 4 and 8 weeks (0.5% NaCl; NS4 and NS8), or HS for 4 and 8 weeks (4% NaCl; HS4 and HS8), or HS for 4 weeks followed by NS for 4 weeks (HS4/NS4), n = 8/group. Systolic BP (SBP), pulse wave velocity (PWV), MBG excretion, aortic collagen 1α2, collagen 4α1 and TGF-β, Smad2, Smad3, Fli-1 mRNA, and total collagen abundance were measured at baseline (BL), and on weeks 4 and 8. Statistical analysis was performed using one-way ANOVA. SBP was not affected by HS (125 ± 5 and 126 ± 6 vs. 128 ± 7 mmHg, HS4 and HS8 vs. BL, p > 0.05). HS increased MBG (164 ± 19 vs. 103 ± 19 pmol/24 h/kg, HS4 vs. BL, p < 0.05) and PWV (3.7 ± 0.2 vs. 2.7 ± 0.2 m/s, HS4 vs. NS4, p < 0.05). HS8 was associated with a further increase in MBG and PWV, with an increase in aortic Col1a2 80%), Col4a1 (50%), Tgfb1 (30%), Smad2 (30%) and Smad3 (45%) mRNAs, and aortic wall collagen (180%) vs. NS8 (all p < 0.05). NS following HS downregulated HS-induced factors: in HS4/NS4, the MBG level was 91 ± 12 pmol/24 h/kg (twofold lower than HS8, p < 0.01), PWV was 3.7 ± 0.3 vs. 4.7 ± 0.2 m/s (HS4/NS4 vs. HS8, p < 0.05), aortic wall Tgfb1, Col1a2, Col4a1, Smad2, Smad3 mRNAs, and collagen abundance were reversed by salt reduction to the BL levels (p < 0.05). HS was associated with an activation of TGF-β signaling, aortic fibrosis and aortic stiffness accompanied by an MBG increase in the absence of SBP changes in young normotensive rats. The reduction of dietary salt following HS decreased MBG, PWV, aortic wall collagen and TGF-β. Thus, HS-induced aortic stiffness in normotensive animals occurred in the context of elevated MBG, which may activate SMAD-dependent TGF-β pro-fibrotic signaling. This data suggests that a decrease in salt consumption could help to restore aortic elasticity and diminish the risk of cardiovascular disease by reducing the production of the pro-fibrotic factor MBG.

The Cardiotonic Steroid Marinobufagenin Is a Predictor of Increased Left Ventricular Mass in Obesity: The African-PREDICT Study

“…Subsequent investigations into the pathophysiological role of MBG have yielded strong evidence of an adverse effect of excessive MBG on the cardiovascular system. In animal and experimental studies, MBG was shown to promote cardiac myocyte apoptosis [ 21 ], cardiovascular fibrosis, and cardiac hypertrophy [ 15 , 16 , 17 , 39 ]. While in humans, MBG was found to be associated with autonomic activity [ 40 ], microvascular dysfunction [ 41 ], arterial stiffness [ 13 , 42 ], and increased LVMi [ 19 ].…”

Microvascular function in non‐dippers: Potential involvement of the salt sensitivity biomarker, marinobufagenin—The African‐PREDICT study

“…Also, while MBG is strongly associated with salt intake in normotensive rats 46 and humans, 19 and Dahl salt-sensitive hypertension, [21][22][23] more studies are needed to establish MBG as a marker of salt sen- Bold values denote significance of P < .05.…”

“…tion44 and vascular fibrosis,45,46 respectively, ultimately impairing vasorelaxation. In support, Fedorova et al have demonstrated impaired sodium nitroprusside-induced vasorelaxation of rat aortic explants pretreated with MBG.…”

Cardiotonic Steroids—A Possible Link Between High-Salt Diet and Organ Damage

“…In normotensive rats, high dietary salt intake have been associated with the activation of TGF-β signaling within the arterial wall and increased aortic stiffness in the presence of elevated levels of the Na/K-ATPase ligand MBG despite unchanged blood pressure [43]. Moreover, the rats exposed to a reduced salt diet after the period of high salt intake exhibited a decrease in MBG levels, downregulation of the pro-fibrotic TGF-β pathway, a decrease of aortic wall collagen content and normalization of the pulse wave velocity to control levels.…”

“…Moreover, the rats exposed to a reduced salt diet after the period of high salt intake exhibited a decrease in MBG levels, downregulation of the pro-fibrotic TGF-β pathway, a decrease of aortic wall collagen content and normalization of the pulse wave velocity to control levels. The authors also demonstrated that MBG stimulates collagen production in parallel with activation of TGF-β in cultured VSMCs in vitro, in the absence of hemodynamic effects [43]. Lowering the salt intake can improve vascular elasticity and decrease the cardiovascular risk by reducing the plasma MBG concentration.…”