Testosterone negatively regulates right ventricular load stress responses in mice

Abstract

Right ventricular (RV) function is the major determinant of mortality in pulmonary arterial hypertension and male sex is a strong predictor of mortality in this disease. The effects of testosterone on RV structure and function in load stress are presently unknown. We tested whether testosterone levels affect RV hypertrophic responses, fibrosis, and function. Male C57BL/6 mice underwent castration or sham followed by pulmonary artery banding (PAB) or sham. After recovery, testosterone pellets were placed in a subset of the castrated mice and mice were maintained for at least two weeks, when they underwent hemodynamic measurements and tissues were harvested. Plasma levels of testosterone were reduced by castration and repleted by testosterone administration. In PAB, castration resulted in lower right ventricle/left ventricle + septum (RV/LV+S), and myocyte diameter (P < 0.05). Replacement of testosterone normalized these parameters and increased RV fibrosis (P < 0.05). Two weeks of PAB resulted in increased RV systolic pressure in all groups with decreased markers of RV systolic and diastolic function, specifically reduced ejection fraction and increased time constant, and dPdt minimum (P < 0.05), though there was minimal effect of testosterone on hemodynamic parameters. Survival was improved in mice that underwent castration with PAB compared with PAB alone (P < 0.05). Testosterone affects RV hypertrophic response to load stress through increased myocyte size and increased fibrosis in mice. Castration and testosterone replacement are not accompanied by significant alterations in RV in vivo hemodynamics, but testosterone deprivation appears to improve survival in PAB. Further study of the role of testosterone in RV dysfunction is warranted to better understand these findings in the context of human disease.

Keywords: pulmonary hypertension; right ventricle; sex hormones; testosterone.

Figure 1

(A) RV/LV+S ratio in control and PAB mice. (B) Echocardiography of diastolic RV internal diameter (RVIDd) in control and PAB mice. *P < 0.05 by two-way ANOVA for the effect of testosterone status, #P < 0.05 by two-way ANOVA for the effect of PAB n = 4-12 per group.

Figure 2

(A) Representative images of Masson-Trichrome stain in RV from Control and PAB in intact testes (CON), castration (CAS), or castration + testosterone (CAS+TEST); left column, sham operation; right column, PAB. (B) Percent fibrotic area. (C) Myocyte size. *P < 0.05 by two-way ANOVA for the effect of testosterone status, #P < 0.05 by two-way ANOVA for the effect of PAB, n = 3 per group.

Figure 3

Molecular expression levels in the RV. (A) Atrial natriuretic peptide (ANP) and (B) brain natriuretic peptide (BNP). Long bar indicates a significant difference associated with PAB by two-way ANOVA. *P < 0.05 by two-way ANOVA for interaction of PAB with castration status, n = 3 per group.

Figure 4

In vivo hemodynamic response in sham controls (Control) and with PAB with intact testes (Control), castration or castration + testosterone. Normalized data are shown for maximum power, ejection fraction, and cardiac output. Long bars indicate P < 0.05 for the effect of PAB by two-way ANOVA, n = 4-6 per group.

Figure 5

Kaplan-Meier survival analysis in long-term PAB with intact testes (CON.PAB) and castration (CAS.PAB), P < 0.05, n = 10 each group.

Figure 6

Echocardiographic measurement of cardiac output after 6 weeks of PAB in male mice with intact testes (CON.PAB) and with castration (CAS. PAB). P = non-signficant. Bars indicate the mean value.