Gestational exposure to elevated testosterone levels induces hypertension via heightened vascular angiotensin II type 1 receptor signaling in rats

Abstract

Pre-eclampsia is a life-threatening pregnancy disorder whose pathogenesis remains unclear. Plasma testosterone levels are elevated in pregnant women with pre-eclampsia and polycystic ovary syndrome, who often develop gestational hypertension. We tested the hypothesis that increased gestational testosterone levels induce hypertension via heightened angiotensin II signaling. Pregnant Sprague-Dawley rats were injected with vehicle or testosterone propionate from Gestational Day 15 to 19 to induce a 2-fold increase in plasma testosterone levels, similar to levels observed in clinical conditions like pre-eclampsia. A subset of rats in these two groups was given losartan, an angiotensin II type 1 receptor antagonist by gavage during the course of testosterone exposure. Blood pressure levels were assessed through a carotid arterial catheter and endothelium-independent vascular reactivity through wire myography. Angiotensin II levels in plasma and angiotensin II type 1 receptor expression in mesenteric arteries were also examined. Blood pressure levels were significantly higher on Gestational Day 20 in testosterone-treated dams than in controls. Treatment with losartan during the course of testosterone exposure significantly attenuated testosterone-induced hypertension. Plasma angiotensin II levels were not significantly different between control and testosterone-treated rats; however, elevated testosterone levels significantly increased angiotensin II type 1 receptor protein levels in the mesenteric arteries. In testosterone-treated rats, mesenteric artery contractile responses to angiotensin II were significantly greater, whereas contractile responses to K(+) depolarization and phenylephrine were unaffected. The results demonstrate that elevated testosterone during gestation induces hypertension in pregnant rats via heightened angiotensin II type 1 receptor-mediated signaling, providing a molecular mechanism linking elevated maternal testosterone levels with gestational hypertension.

Keywords: AGTR1; angiotensin; blood pressure; losartan; mesenteric arteries; pre-eclampsia; pregnancy; testosterone; vascular function.

FIG. 1

Changes in blood pressure in control and T-exposed pregnant rats. Mean arterial pressure was measured in conscious, free-moving control (n = 11) and T-treated rats (0.5 mg/kg/day, subcutaneously from Gestational Day 15–19; n = 7) through carotid arterial catheters. Another set of control T rats was treated with losartan (20 mg·kg⁻¹·day⁻¹; n = 6 in each group) by gavage during the course of T exposure, and then mean arterial pressure was recorded. Data points means ± SEM. *P < 0.05 versus untreated control. ^#P < 0.05 versus untreated T group.

FIG. 2

No changes in plasma angiotensin II levels were detected between control and T-exposed pregnant rats. Plasma angiotensin II levels were measured in control (n = 8) and T-treated rats (n = 8) by using enzyme immunoassay. Data points are means ± SEM.

FIG. 3

Changes in angiotensin receptor protein levels in mesenteric arteries from control and T-exposed pregnant rats. Representative Western blots for AGTR1 are shown at the top; blot density obtained from densitometric scanning of AGTR1 normalized to β-actin is shown at the bottom. Values are means ± SEM of 6 rats in each group. *P ≤ 0.05 versus control.

FIG. 4

Changes in mRNA levels of angiotensin type 1 receptor isoforms in mesenteric arteries from control and T-exposed pregnant rats. Real-time PCR was used to assess vascular Agtr1a and Agtr1b mRNA expression levels. Quantitation of vascular Agtr1a and Agtr1b mRNA expression was normalized relative to that of β-actin. Values are means ± SEM of 7 rats in each group. *P < 0.05 versus control.

FIG. 5

Contractile responses in mesenteric arterial rings of control and T-exposed pregnant rats. Endothelium-denuded mesenteric arterial rings were incubated in Krebs buffer, and then vascular contractile responses were taken to cumulative additions of KCl (A), phenylephrine (B), and Ang II (C). Contractions to Ang II were also taken in the presence of losartan (10 μM). K⁺ depolarizing contractions are presented as absolute tension (mN/mm) and as the percentage of maximal KCl contractions. Phenylephrine and angiotensin II contractions are presented as percentages of their maximal contractions, as well as percentages of 80 mM KCl contractions. Values are means ± SEM. Mesenteric arterial rings (n = 10 to 12) from 8 rats of each group were used.