Silencing salusin-β attenuates cardiovascular remodeling and hypertension in spontaneously hypertensive rats

Figures

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  Figure 1. Salusin-β levels in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Plasma salusin-β levels. (B) Salusin-β contents in myocardium and mesenteric artery. (C) Salusin-β contents in hypothalamic paraventricular nucleus (PVN) and rostral ventrolateral medulla (RVLM) of the brain. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 2. Arterial blood pressure and heart rate in WKY and SHR. (A) Systolic blood pressure (SBP) and heart rate (HR) in conscious state before and after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (B) Mean arterial pressure (MAP) and HR in anesthetized state 2 weeks after the injections. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 3. Perivascular fibrosis in myocardium and cross-sectional area of cardiomyocytes in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Sections with Masson’s stain showing perivascular fibrosis in myocardium. (B) Sections with HE stain showing the size of cardiomyocytes. (C) Bar graph showing quantitative analysis of cross-sectional area of cardiomyocytes. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 4. Vascular remodeling of aorta, main renal artery and mesenteric artery in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Representative transverse section images with Masson’s stain. (B) Bar graph showing the media thickness, lumen diameter and the ratio of media thickness to lumen diameter. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 5. Norepinephrine and tyrosine hydroxylase in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Plasma norepinephrine (NE) levels. (B) NE contents in myocardium of left ventricle (LV) and mesenteric artery (MA). (C) Plasma tyrosine hydroxylase (TH) levels. (D) TH contents in LV and MA. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 6. Angiotensin II, type 1 receptors of angiotensin and angiotensin converting enzyme in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Plasma angiotensin II (Ang II) levels and Ang II contents in myocardium of left ventricle (LV) and mesenteric artery (MA). (B) Type 1 receptors of angiotensin (AT1R) expression in LV and MA. (C) Plasma angiotensin converting enzyme (ACE) levels and ACE contents in LV and MA. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 7. Superoxide anions levels and NAD(P)H oxidase activity in myocardium and mesenteric artery in WKY and SHR. The measurements were carried out 2 weeks after intravenous injection of PBS, adenoviral vectors encoding scramble shRNA (Ad-Scr-shRNA) or salusin-β shRNA (Ad-Sal-shRNA). (A) Superoxide anions levels and NAD(P)H oxidase activity in myocardium. (B) Superoxide anions levels and NAD(P)H oxidase activity in mesenteric artery. Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.

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  Figure 8. Cell proliferation and fibrosis in primary VSMCs from WKY and SHR. (A) VSMC proliferation was determined with EdU incorporation assay (×200). Red fluorescence (Edu) stands for cells with DNA synthesis, and blue fluorescence (Hoechst 33342) shows cell nuclei. (B) Fibrosis was evaluated with the collagen-I, collagen-III and fibronectin mRNA expressions. The measurements were carried out 48 h after administration of PBS, Ad-Scr-shRNA or Ad-Sal-shRNA (40 MOI). Values are mean ± S.E.M. *P < 0.05 vs. WKY. †P < 0.05 vs. PBS or Ad-Scr-shRNA. n = 6 for each group.