Short-Term Administration of Serelaxin Produces Predominantly Vascular Benefits in the Angiotensin II/L-NAME Chronic Heart Failure Model

Figures

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  Figure 1. Serelaxin Temporarily Decreases Blood Pressure in the Angiotensin II/L-NG-Nitroarginine Methyl Ester Experimental Model (A) In the Chronic:post-treatment study, telemeters were implanted on day 7. Blood pressure (BP) was recorded at baseline and throughout the duration of the experiment. In each experiment, angiotensin II (AngII) (800 ng/kg/min)/L-NG-nitroarginine methyl ester (L-NAME) (30 mg/kg/day in drinking water) treatment began at day 0. On day 7, mice were randomized to serelaxin or vehicle infusion by 3-day osmotic pump. Mice were sacrificed, and cardiac and vascular parameters were measured on day 9 in the Acute:on-treatment groups and on day 28 in the Chronic:post-treatment groups. (B) Telemetric average 24-h systolic blood pressure on day 0 and daily from day 7 to day 28. (C) Systolic blood pressure changes from baseline to day 28 during AngII/L-NAME. *p = 0.028. (B, C) Two-factor repeated-measures analysis of variance with Tukey post hoc test.

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  Figure 2. Serelaxin Does Not Reduce Left Ventricular Hypertrophy or Remodeling in the Angiotensin II/L-NG-Nitroarginine Methyl Ester Model Cardiac ultrasound was performed followed by organ harvest and histology in the Acute:on-treatment (day 9) and Chronic:post-treatment (day 28) groups in angiotensin II (AngII)/L-NG-nitroarginine methyl ester (L-NAME)–treated mice. (A) Heart and left ventricular (LV) mass normalized to tibia length. (B) Cardiac dimensions and function as measured by echocardiography. (C) Representative and summary data of cardiac myocyte cross-sectional area from hematoxylin and eosin–stained LV sections (scale bar represents 0.3 μm) and of interstitial fibrosis from picrosirius red–stained LV sections (scale bar represents 1 μm). n = 6 (day 9) or 5 (day 28) per treatment. Connected single bars over days 9 and 28 indicate analysis of main effects of time factor for vehicle and serelaxin in combination. *p < 0.01 and †p = 0.024, day 9 versus day 28; ‡p = 0.027 for main effects of treatment factor serelaxin versus vehicle. Two-factor analysis of variance with Tukey post hoc test.

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  Figure 3. Serelaxin Reduces Coronary Perivascular Fibrosis and Increases Left Ventricular Capillary Density in the Angiotensin II/L-NG-Nitroarginine Methyl Ester Model Left ventricular (LV) histology in the Acute:on-treatment group (day 9, n = 6 per group) and Chronic:post-treatment group (day 28, n = 5 per group) in angiotensin II (AngII)/L-NG-nitroarginine methyl ester (L-NAME)–treated mice. (A) Representative and summary data of perivascular fibrosis from picrosirius red–stained LV sections. Scale bars represent 1 μm. (B) Representative and summary data of capillary density from CD31 staining. Black arrows denote examples of CD31-positive capillaries. Scale bars represent 0.3 μm. Connected single bars over days 9 and 28 indicate analysis of main effects of time factor for vehicle and serelaxin in combination. *p < 0.01 treatment within day; †p < 0.001 for main effects of time factor, day 9 versus day 28; ‡p = 0.009 for main effects of treatment factor serelaxin versus vehicle. Two-factor analysis of variance with Tukey post hoc test.

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  Figure 4. Serelaxin Treatment Decreases the Mesenteric Arteriolar Contractile Response to Potassium Chloride in the Angiotensin II/L-NG-Nitroarginine Methyl Ester Heart Failure Model Mesenteric resistance arterioles were isolated on day 9 or day 28 from mice treated with AngII/L-NAME, and with either serelaxiin or vehicle. Vasoconstrictor and vasorelaxer responses were measured by wire myography. *p < 0.01 and †p < 0.05 for treatment effect within potassium chloride dose; ‡p < 0.05 for overall main effects of serelaxin versus vehicle (p = 0.004, day 9; p = 0.034, day 28). Data were compared using 2-factor repeated-measures analysis of variance with Tukey post hoc test.

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  Figure 5. Serelaxin Infusion Attenuates Vascular Fibrosis and Fibrotic Gene Expression in the Angiotensin II/L-NG-Nitroarginine Methyl Ester Heart Failure Model The abdominal aorta was isolated on day 9 (n = 6) or on day 28 (n = 5) of angiotensin II (AngII)/L-NG-nitroarginine methyl ester (L-NAME) infusion. Representative and summary data of aortic medial area from elastin-stained aortas (A) and aortic medial fibrosis from trichrome-stained aortas (B). Black arrows define the tunica media (medial area and degree of fibrosis was measured between arrows). Scale bars represent 1 μm. *p < 0.001; †p < 0.001 for main effects of treatment factor serelaxin versus vehicle; ‡p < 0.001 for main effects of time factor, day 9 versus day 28. (C) Messenger ribonucleic acid was isolated from aortas of mice treated with serelaxin or vehicle, and expression of fibrosis genes was measured by quantitative reverse transcription polymerase chain reaction. *p = 0.018 serelaxin versus vehicle day 9; †p < 0.001, serelaxin day 28 versus vehicle day 28; ‡p = 0.022 for main effects of time factor, day 9 versus day 28; §p < 0.001 for main effects of treatment factor serelaxin versus vehicle; ‖p = 0.049 and ¶p = 0.003,…