Rutaecarpine prevents hypertensive cardiac hypertrophy involving the inhibition of Nox4‐ROS‐ADAM17 pathway

Figures

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  Figure 1. Rutaecarpine (Rut) inhibited hypertensive cardiac hypertrophy in the rats subjected to abdominal artery constriction (AAC). A, HW/BW (n = 9‐10 per group). B, LVW/BW (n = 9‐10 per group). C, Cross‐sections of the hearts from the rats subjected to AAC or sham operation were stained with haematoxylin & eosin to analyse myocyte cross‐sectional area (n = 5 per group). D, Real‐time PCR analyses of hypertrophic markers (ANP and BNP, n = 5 in each group). Rut (L) represents low dosage of rutaecarpine; Rut (H) represents high dosage of rutaecarpine; HW/BW represents the ratio between heart weight and body weight; LVW/BW represents the ratio between left ventricular weight and body weight. # P < 0.05 vs Sham group, *P < 0.05 vs SHR group. One‐way ANOVA followed by Bonferroni's post hoc test was carried out for the statistical tests

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  Figure 2. Rutaecarpine inhibited cardiac fibrosis in the rats subjected to abdominal artery constriction (AAC). A, Representative microphotographs of Masson staining of heart. B, Fibrosis area (n = 5 per group). C, Col I positive area (n = 5 per group). D, Representative microphotographs of immunochemistry for col I and col III. E, Col III positive area (n = 5 per group). F, mRNA levels of col I and col III in the left ventricle (n = 4 per group). One‐way ANOVA followed by Bonferroni's post hoc test was used for the statistical tests. Rut (L) represents low dosage of rutaecarpine; Rut (H) represents high dosage of rutaecarpine; Col I represents collagen I; col III represents collagen III. # P < 0.05 vs Control group; *P < 0.05 vs AAC group

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  Figure 3. Rutaecarpine (Rut) alleviated angiotensin II (Ang II)‐induced cardiac hypertrophy in primary cardiomyocytes. Primary cardiomyocytes were treated with 100 nmol/L angiotensin II for 24 h after pretreating with rutaecarpine (10 µmol/L) for 60 min. A, Cell surface area. B and C, The mRNA levels of hypertrophic genes ANP (B) and BNP (C). # P < 0.05 vs control group; *P < 0.05 vs Ang II group. The statistical tests were undertaken by one‐way ANOVA followed by Bonferroni's post hoc test. n = 3 independent experiments

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  Figure 4. Rutaecarpine (Rut) inhibited the Nox4‐ROS‐ADAM17 pathway in cardiomyocytes treated by angiotensin (Ang II). Primary cardiomyocytes were treated with rutaecarpine (10 µmol/L) for 60 min prior to stimulation with Ang II for 24 h. A and B, The contents of H2O2 (A) and O2− level indicated by DHE fluorescence intensity (B). C, Nox4 protein level. D, Nox4 mRNA level. E, ADAM17 protein level. F, ADAM17 mRNA level. G, TNF‐α protein level in the conditioned media. H, TNF‐α mRNA level. Nox4 represents NADPH oxidase 4; ROS represents reactive oxygen species; ADAM17 represents a disintegrin and metalloproteinase‐17; TNF‐α represents tumour necrosis factor α; # P < 0.05 vs Control group; *P < 0.05 vs Ang II group. The statistical analyses were performed using one‐way ANOVA followed by Bonferroni's post hoc test. n = 3 independent experiments

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  Figure 5. Rutaecarpine (Rut) suppressed the Nox4‐ROS‐ADAM17 pathway in the left ventricle of rats subjected to abdominal artery constriction (AAC). A, The H2O2 level (n = 5 per group). B, Nox4 mRNA level (n = 5 per group). C, Nox4 protein level (n = 4 per group). D, ADAM17 protein level (n = 4 per group). E, ADAM17 mRNA level; (n = 5 per group). F, TNF‐α mRNA level (n = 4 per group). G, TNF‐α protein level (n = 5 per group). Nox4 represents NADPH oxidase 4; ROS represents reactive oxygen species; ADAM17 represents a disintegrin and metalloproteinase‐17; TNF‐α represents tumour necrosis factor α; # P < 0.05 vs Sham group, *P < 0.05 vs SHR group. The statistical tests were carried out by one‐way ANOVA followed by Bonferroni's post hoc test

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  Figure 6. Rutaecarpine (Rut) blocked the over‐activation of the extracellular signal‐regulated kinase (ERK) 1/2 pathway in hypertrophic cardiomyocytes. A, ERK1/2 activity in the left ventricle of rats with abdominal artery constriction (AAC, n = 4 per group). Rut (L) represents low dosage of rutaecarpine; Rut (H) represents high dosage of rutaecarpine; # P < 0.05 vs Sham group, *P < 0.05 vs SHR group. B, ERK1/2 activity in primary cardiomyocytes treated with 100 nmol/L angiotensin II (Ang II) for 24 h (n = 3 independent experiments). After being pretreated with Rut (10 µmol/L) for 60 min, cardiomyocytes were then stimulated with 100 nmol/L Ang II for 24 h. # P < 0.05 vs Control group; *P < 0.05 vs Ang II group. The statistical analyses were executed using one‐way ANOVA followed by Bonferroni's post hoc test