C26-10 Microvascular Rarefaction Contributes to Pulmonary Hypertension in a Mouse Model of Pulmonary Fibrosis

Rationale Prognosis for patients with pulmonary fibrosis (PF) is tied closely to the severity of pulmonary hypertension (PH). However, how the fibrotic process in the lung influences vascular apoptosis and vessel rarefaction has not been defined. We hypothesized that PF increases apoptosis of smooth muscle cells (SMCs) and endothelial cells (ECs) contributing to PH. Methods We induced PF with intratracheal bleomycin (0.025 U; 3 wks prior to the study) in male and female C57Bl/6J mice (age, ∼5 mo) with saline used as a control (sham). We quantified the number of capillaries and small pulmonary arteries (≤40 µm) in lung sections labeled with lectin and Verhoeff van Gieson stain, respectively. Given the increase in ROS during PH, we measured SMC and EC death (1 µM Hoechst 33342 and 2 µM propidium iodide) and mitochondrial membrane potential (ΔΨm; TMRM) depolarization in isolated, pressurized (16 cmH2O), pulmonary arteries (diameter 100-150 µm) during exposure to H2O2 (200 µM, 50 min). Furthermore, mice were treated with the pan-caspase (apoptosis) inhibitor Q-VD (20 mg/kg; tail vein injection on days 0, 2, 4, and 7) to determine whether vascular apoptosis drives PH facilitated by PF. Results In both male and female mice, bleomycin caused a significant (P < 0.05) increase in collagen deposition and hypercellularity. Despite a similar magnitude of fibrosis, right ventricular systolic pressure (RVSP, male: 40±2 vs. 19±1; female: 31±1 vs. 18±1 mmHg) and Fulton index (male: 0.58±0.5 vs 0.31±0.12; female: 0.45±0.03 vs 0.31±0.01) were higher in males compared to females. Lung sections from PF mice exhibited fewer capillaries (male: 39±3 vs. 84±7/mm², female: 68±6 vs. 102±13 per mm²) and small arteries (male: 9±2 vs. 21±4; female: 9±1 vs. 23±4 per section). Both SMC death (male: 35±3% vs. 20±1%; female: 25±3% vs. 10±2%) and EC death (male: 20±2% vs. 8±1%; female: 11±2% vs. 4±1%) in response to H2O2 were increased in mice with PF. Consistently, ΔΨm depolarization to oxidative stress was enhanced by PF and was greater in males than in females. Chronic inhibition of apoptosis with Q-VD during PF development significantly reduced RVSP and Fulton index in males, with a trend (P = 0.10) observed in females; Q-VD did not alter the severity of fibrotic injury. Conclusion PF increases apoptosis of SMCs and ECs mediating to vessel rarefaction and the development of PH. Vessel loss is more pronounced in males contributing to enhanced PH. This abstract is funded by: NIH, AHA