Genetic evidence implicates the loss of bone morphogenetic protein type II receptor (BMPR-II) signaling in the endothelium as an initiating factor in pulmonary arterial hypertension (PAH). However, selective targeting of this signaling pathway using BMP ligands has not yet been explored as a therapeutic strategy. We identified BMP9 as the preferred ligand for preventing apoptosis and enhancing monolayer integrity in both pulmonary arterial endothelial cells and blood outgrowth endothelial cells from subjects with PAH bearing mutations in BMPR-II. In vivo, we report the spontaneous generation of PAH in a mouse model bearing a heterozygous knock-in of a human BMPR-II mutation, R899X. Administration of BMP9 reversed established PAH in Bmpr2+/R899X mice, as well as in models of disease developed in response to either monocrotaline or VEGF receptor inhibition combined with chronic hypoxia. These results demonstrate the promise of direct enhancement of endothelial BMP signaling as a novel therapeutic strategy for PAH.

Objective The sphingosine-1-phosphate receptor agonist fingolimod (FTY720), that has shown efficacy in advanced multiple sclerosis clinical trials, decreases reperfusion injury in heart, liver and kidney. We therefore tested the therapeutic effects of fingolimod in several rodent models of focal cerebral ischemia. To assess the translational significance of these findings, we asked whether fingolimod improved long-term behavioral outcomes, whether delayed treatment was still effective, and whether neuroprotection can be obtained in a second species. Methods We used rodent models of middle cerebral artery occlusion and cell culture models of neurotoxicity and inflammation to examine the therapeutic potential and mechanisms of neuroprotection by fingolimod. Results In a transient mouse model, fingolimod reduced infarct size, neurological deficit, edema and the number of dying cells in the core and periinfarct area. Neuroprotection was accompanied by decreased inflammation, as fingolimod-treated mice had fewer activated neutrophils, microglia/macrophages, and ICAM-1-positive blood vessels. Fingolimod-treated mice showed a smaller infarct and performed better in behavioral tests up to 15 days after ischemia. Reduced infarct was observed in a permanent model even when mice were treated 4 hours after ischemic onset. Fingolimod also decreased infarct size in a rat model of focal ischemia. Fingolimod did not protect primary neurons against glutamate excitotoxicity or hydrogen peroxide, but decreased ICAM-1 expression in brain endothelial cells stimulated by TNFalpha. Interpretation These findings suggest that anti-inflammatory mechanisms, and possibly vasculo-protection, rather than direct effects on neurons, underlie the beneficial effects of fingolimod after stroke. S1P receptors are a highly promising target in stroke treatment.

Abstract-Hypertension and vascular dysfunction result in the increased release of endothelium-derived contracting factors (EDCFs), whose identity is poorly defined. We tested the hypothesis that endothelial cyclooxygenase (COX)-2 can generate EDCFs and identified the possible EDCF candidate. Changes in isometric tension of aortae of young and aged hamsters were recorded on myograph. Real-time changes in intracellular calcium concentrations ([Ca 2ϩ ] i ) in native aortic endothelial cells were measured by imaging. Endothelium-dependent contractions were triggered by acetylcholine (ACh) after inhibition of nitric oxide production and they were abolished by COX-2 but not COX-1 inhibitors or by thromboxane-prostanoid receptor antagonists. 2-Aminoethoxydiphenyl borate (cation channel blocker) eliminated endothelium-dependent contractions and ACh-stimulated rises in endothelial cell [Ca 2ϩ ] i . RT-PCR and Western blotting showed COX-2 expression mainly in the endothelium. Enzyme immunoassay and high-performance liquid chromatography-coupled mass spectrometry showed release of prostaglandin (PG)F 2␣ and prostacyclin (PGI 2 ) increased by ACh; only PGF 2␣ caused contraction at relevant concentrations. COX-2 expression, ACh-stimulated contractions, and vascular sensitivity to PGF 2␣ were augmented in aortae from aged hamsters. Human renal arteries also showed thromboxane-prostanoid receptor-mediated ACh-or PGF 2␣ -induced contractions and COX-2-dependent release of PGF 2␣ . The present study demonstrates that PGF 2␣ , derived from COX-2, which is localized primarily in the endothelium, is the most likely EDCF underlying endothelium-dependent, thromboxane-prostanoid receptor-mediated contractions to ACh in hamster aortae. These contractions involved increases in endothelial cell [Ca 2ϩ ] i . The results support a critical role of COX-2 in endothelium-dependent contractions in this species with an increased importance during aging and, possibly, a similar relevance in humans. Key Words: endothelium-derived contracting factors Ⅲ cyclooxygenase-2 Ⅲ thromboxane-prostanoid receptor Ⅲ aging Ⅲ aorta B esides neuronal and hormonal regulation, vascular tone is modulated locally by a delicate balance between endothelium-derived relaxing (EDRFs) and contracting (EDCFs) factors, 1,2 with the latter being less well-defined but emerging in hypertension, obesity, hyperlipidemia, diabetes, and aging. 3 A number of molecules have been proposed as possible EDCF candidates under pathophysiological conditions. These include prostaglandin (PG)H 2 , thromboxane (TX)A 2 , leukotrienes, endothelin 1, and superoxide anions. The release of these tentative EDCFs can be triggered by acetylcholine (ACh), angiotensins I/II, ADP, and ATP. 4 The contribution of additional cyclooxygenase (COX)-derived metabolites, ie, PGE 2 , PGD 2 , and PGF 2␣ , has been postulated. The precise nature of these EDCFs varies among species and vascular beds. 1,3 Two isoforms of COX have been identified in blood vessels. COX-1 is constitutively expressed and...