Assessment of vascular leakage and leukocyte infiltration in endothelial restricted Bmpr-II–deficient mice with PAH. To assess vascular leakage (A), L1Cre(−)Bmpr2^f/f mice or L1Cre(+)Bmpr2^f/f mice were treated with SCH527123 or vehicle orally twice daily for 21 days. Evans blue dye was then administered intravenously via the tail vein for 1 hour, after which the lungs were flushed with PBS and the heart/lung block excised. Evans blue was extracted from the right lung, quantified, and expressed as micrograms per milliliter Evans blue per right lung. To assess leukocyte infiltration (B), the concentration of MPO in the right lung of L1Cre(−)Bmpr2^f/f and L1Cre(+)Bmpr2^f/f mice treated with vehicle or SCH527123 was determined by ELISA. Leukocyte infiltration was further assessed by observing leukocyte infiltration (arrows) (Ci) into the small pulmonary vessels by histologic sections stained with H&E and scoring the number and location of infiltrated leukocytes (Cii). Identification of specific leukocyte subsets was assessed in lung sections stained with specific antibodies targeting monocyte/macrophages, T cells, B cells, and neutrophils (Ciii). Data are the mean ± SEM for minimum 8 animals per group. (A) ANOVA showed a significant effect of treatment with SCH527123 and genotype (P < .001). ***P < .001, compared with L1Cre(+)Bmpr2^f/f animals treated with vehicle by Dunnett test. (B) ANOVA showed a significant effect of treatment with SCH527123 and genotype (P < .001). ***P < .001, compared with L1Cre(+)Bmpr2^f/f animals treated with vehicle by Dunnett test. Slides were mounted using VECTORSHIELD (Vector Laboratories) and viewed using a 20× objective lens. Images were captured using a DMLB microscope and digital camera. Digital image acquisition was performed using IM50 Version 4.0 imaging sofware (Leica Microsystems) and digital image slide viewing using Image Scope software (Aperio Technologies).