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Pulmonary hypertension (PH) is a leading cause of death in sickle cell disease (SCD) patients. Hemolysis and oxidative stress contribute to SCD-associated PH. We have reported that the protein thrombospondin-1 (TSP1) is elevated in the plasma of patients with SCD and, by interacting with its receptor CD47, limits vasodilation of distal pulmonary arteries ex vivo. We hypothesized that the TSP1-CD47 interaction may promote PH in SCD. We found that TSP1 and CD47 are upregulated in the lungs of Berkeley (BERK) sickling (Sickle) mice and patients with SCD-associated PH. We then generated chimeric animals by transplanting BERK bone marrow into C57BL/6J (n 24) and CD47 knockout (CD47KO, n 27) mice. Right ventricular (RV) pressure was lower in fully engrafted Sickle-to-CD47KO than Sickle-to-C57BL/6J chimeras, as shown by the reduced maximum RV pressure (P 0.013) and mean pulmonary artery pressure (P 0.020). The afterload of the sickle-to-CD47KO chimeras was also lower, as shown by the diminished pulmonary vascular resistance (P 0.024) and RV effective arterial elastance (P 0.052). On myography, aortic segments from Sickle-to-CD47KO chimeras showed improved relaxation to acetylcholine. We hypothesized that, in SCD, TSP1-CD47 signaling promotes PH, in part, by increasing reactive oxygen species (ROS) generation. In human pulmonary artery endothelial cells, treatment with TSP1 stimulated ROS generation, which was abrogated by CD47 blockade. Explanted lungs of CD47KO chimeras had less vascular congestion and a smaller oxidative footprint. Our results show that genetic absence of CD47 ameliorates SCD-associated PH, which may be due to decreased ROS levels. Modulation of TSP1-CD47 may provide a new molecular approach to the treatment of SCD-associated PH. [ABSTRACT FROM AUTHOR]

Aims: Thrombospondin-1 (TSP1) is a ligand for CD47 and TSP1-/- mice are protected from pulmonary hypertension (PH). We hypothesized the TSP1-CD47 axis is upregulated in human PH and promotes pulmonary arterial vasculopathy. Methods and results: We analyzed the molecular signature and functional response of lung tissue and distal pulmonary arteries (PAs) from individuals with (n=23) and without (n=16) PH. Compared with controls, lungs and distal PAs from PH patients showed induction of TSP1-CD47 and endothelin-1/endothelin A receptor (ET-1/ETA) protein and mRNA. In control PAs, treatment with exogenous TSP1 inhibited vasodilation and potentiated vasoconstriction to ET-1. Treatment of diseased PAs from PH patients with a CD47 blocking antibody improved sensitivity to vasodilators. Hypoxic wild type (WT) mice developed PH and displayed upregulation of pulmonary TSP1, CD47, and ET-1/ETA concurrent with down regulation of the transcription factor cell homolog of the v-myc oncogene (cMyc). In contrast, PH was attenuated in hypoxic CD47-/- mice while pulmonary TSP1 and ET-1/ETA were unchanged and cMyc was overexpressed. In CD47-/- pulmonary endothelial cells cMyc was increased and ET-1 decreased. In CD47+/+cells, forced induction of cMyc suppressed ET-1 transcript, whereas suppression of cMyc increased ET-1 signaling. Furthermore, disrupting TSP1-CD47 signaling in pulmonary smooth muscle cells abrogated ET-1-stimulated hypertrophy. Finally, a CD47 antibody given 2 weeks after monocrotaline challenge in rats upregulated pulmonary cMyc and improved aberrations in PH-associated cardiopulmonary parameters. Conclusions: In pre-clinical models of PH CD47 targets cMyc to increase ET-1 signaling. In clinical PH TSP1-CD47 is upregulated, and in both, contributes to pulmonary arterial vasculopathy and dysfunction. [ABSTRACT FROM AUTHOR]

Background: Pulmonary hypertension (PH), regardless of etiology, remains progressive and incurable despite intensive research for over half a century. Existing therapies have only modestly enhanced survival with the only available “cure” being lung transplantation. The reasons for lack of progress in this area remain unknown but suggest a deficit in understanding the overarching mechanism (s) that drive PH. Recently we reported that the thrombospondin-1 (TSP1)-CD47 ligand receptor pathway is induced in multiple pre-clinical models of PH and human disease. Mice lacking secreted TSP1 are protected from PH. Methods: Wild type and CD47 null mice were challenged with hypoxia and cardiopulmonary assessment performed. Likewise, in cell culture experiments pulmonary arterial vascular smooth muscle cells were exposed to hypoxia and molecular signal transduction assessed. Results: We know report that mice mutated to lack cell receptor CD47 are highly resistant to hypoxia-mediated PH with minimal evidence of pulmonary arterial overgrowth and no right ventricular hypertrophy compared to controls. Hypoxic CD47 null mice demonstrated increased stroke volume and cardiac output compared to a significant deterioration of these functional parameters in wild type controls. Hypoxic wild type animals also displayed concurrent loss of cMyc and upregulation of endothelin-1. In contrast, hypoxic CD47 null animals demonstrated cMyc-mediated suppression of endothelin-1 signaling. Impressively, under basal conditions endothelin-1 and the cognate receptors ETA and ETB where nearly undetectable in CD47 null lungs. Disrupting TSP1-mediated activation of CD47 in monocrotaline treated rats, and in hypoxic pulmonary arterial smooth muscle cell cultures, upregulated cMyc, inhibited endothelin-1 signaling and corrected established PH. Finally, antibody blocking CD47 in lungs from patients with end-stage PH normalized the vasodilation profile of distal pulmonary arteries to acetylcholine and sodium nitroprusside. Conclusions: The above preclinical results suggest CD47 is an important and proximate promoter of PH through cMyc-mediated inhibition of endothelin-1, while results in diseased human PH lung arterioles suggest that CD47 is a clinical target to restore end-stage pulmonary arteriole function. Disclosure: J.S.I. is Chair of the Scientific Advisory Boards and has equity interest in Vasculox, Inc. (St. Louis, MO) and Radiation Control Technologies, Inc. (Rockville, MD). [ABSTRACT FROM AUTHOR]