Severe pulmonary hypertension in aging female apolipoprotein E-deficient mice is rescued by estrogen replacement therapy

Background: Apolipoprotein E (ApoE) is a multifunctional protein, and its deficiency leads to the development of atherosclerosis in mice. Patients with pulmonary hypertension (PH) have reduced expression of ApoE in lung tissue. ApoE is known to inhibit endothelial and smooth muscle cell proliferation and has anti-inflammatory and anti-platelet aggregation properties. Young ApoE-deficient mice have been shown to develop PH on high fat diet. The combined role of female sex and aging in the development of PH has not been investigated before. Here, we investigated the development of PH in young and middle-aged (MA) female ApoE-deficient mice and explored the role of exogenous estrogen (E2) replacement therapy for the aging females. Methods: Wild type (WT) and ApoE-deficient female mice (Young and MA) were injected with a single intraperitoneal dose of monocrotaline (MCT, 60 mg/kg). Some ApoE-deficient MA female mice that received MCT were also treated with subcutaneous E2 pellets (0.03 mg/kg/day) from day 21 to 30 after MCT injection. Direct cardiac catheterization was performed terminally to record right ventricular systolic pressure (RVSP). Right ventricular (RV), left ventricular (LV), and interventricular septum (IVS) were dissected and weighed. Lung sections were examined using trichrome and immunofluorescence staining. Western blot analyses of lung and RV lysates were performed. Results: In WT female mice, the severity of PH was similar between young and MA mice as RVSP was not significantly different (RVSP = 38.2 +/- 1.2 in young vs. 40.5 +/- 8.3 mmHg in MA, p < 0.05). In ApoE-deficient mice, MA females developed significantly severe PH (RVSP = 63 +/- 10 mmHg) compared to young females (RVSP; 36 +/- 3 mmHg, p < 0.05 vs. MA female). ApoE-deficient MA females also developed more severe RV hypertrophy compared to young females (RV hypertrophy index (RV/[LV + IVS]) = 0.53 +/- 0.06 vs. 0.33 +/- 0.01, p < 0.05). ApoE-deficient MA female mice manifested increased peripheral pulmonary artery muscularization and pulmonary fibrosis. E2 treatment of MA female ApoE-deficient mice resulted in a significant decrease in RVSP, reversal of pulmonary vascular remodeling, and RV hypertrophy. In MA female ApoE-deficient mice with PH, only the expression of ER beta in the lungs, but not in RV, was significantly downregulated, and it was restored by E2 treatment. The expression of ER alpha was not affected in either lungs or RV by PH. GPR30 was only detected in the RV, and it was not affected by PH in MA female ApoE-deficient mice. Conclusions: Our results suggest that only aging female ApoE-deficient but not WT mice develop severe PH compared to younger females. Exogenous estrogen therapy rescued PH and RV hypertrophy in aging female ApoE-deficient mice possibly through restoration of lung ERa.