We have demonstrated that VEGF receptor blockade in combination with chronic hypoxia causes in rats severe angioproliferative pulmonary hypertension (SAPH) associated with arterial occlusion by proliferating endothelial cells, and we postulate that the established, lumen-occluding lesions are the result of the emergence of apoptosis-resistant proliferating cells. To study the dependence of exuberant endothelial cell proliferation on initial apoptosis, we adapted the CELLMAX artificial capillary system to analyze the effects of a VEGF receptor antagonist (SU5416) on human pulmonary microvascular endothelial cells under pulsatile shear stress. Immunohistochemical staining for caspase-3 and PCNA and flow cytometry for Annexin-V and BrdU supported our concept, since SU5416 caused initial apoptosis (35.8% at 24 h after the SU5416 addition and 4.8% in control cells) whereas the surviving cells became hyperproliferative (PCNA positive). Flow cytometry showed that apoptosis inhibition prevented the proliferation following the initial apoptosis. These lumen-filling endothelial cells were apoptosis resistant, grew without serum, and were phenotypically altered in that they express the tumor marker survivin. Hyperproliferative apoptosis-resistant cells were also generated by adding apoptosed cells instead of the VEGF receptor blocker to the CELLMAX system. In conclusion, endothelial cell death resulted in the selection of an apoptosis-resistant, proliferating phenotypically altered endothelial cell phenotype.

-Severe pulmonary hypertension (SPH) is characterized by precapillary arteriolar lumen obliteration, dramatic right ventricular hypertrophy, and pericardial effusion. Our recently published rat model of SPH recapitulates major components of the human disease. We used this model to develop new treatment strategies for SPH. SPH in rats was induced using VEGF receptor blockade in combination with chronic hypoxia. A large variety of drugs used in this study, including anticancer drugs (cyclophosphamide and paclitaxel), the angiotensin-converting enzyme inhibitor lisinopril, the antiangiogenic agent thalidomide, and the peroxisome proliferator-actived receptor-␥ agonist PGJ2, failed to decrease mean pulmonary artery pressure (PAP) or right ventricular hypertrophy. In contrast, treatment of rats with established SPH with simvastatin markedly reduced mean PAP and right ventricular hypertrophy, and this reduction was associated with caspase-3 activation and pulmonary microvascular endothelial cell apoptosis. Simvastatin partially restored caveolin-1, caveolin-2, and phospho-caveolin expression in vessel walls. In rat primary pulmonary microvascular endothelial cells, simvastatin induced caspase 3 activation and Rac 1 expression while suppressing Rho A and attenuated levels of Akt and ERK phosphorylation. We conclude that simvastatin is effective in inducing apoptosis in hyperproliferative pulmonary vascular lesions and could be considered as a potential drug for treatment of human SPH. statins HUMAN SEVERE PULMONARY HYPERTENSION (SPH), including pulmonary hypertension secondary to congenital cardiac abnormalities, collagen vascular disorders, human immunodeficiency virus and human herpes virus-8 infection (7,8,32,38,41,51), anorexigen drug intake, and so-called idiopathic pulmonary hypertension, are unfortunately diagnosed when the pulmonary vascular bed already has been extensively remodeled and is not usually responsive to vasodilators (42). Although continuous infusion of prostacyclin has prolonged the survival of many patients with SPH and improved their ability to exercise and function (15,39,44), prostacyclin does not cure the disease. Vascular lesions are frequently localized at sites distal to arteriolar bifurcations (9) and contain phenotypically altered endothelial and smooth muscle cells (25,52,53) that have lost the expression of p27, caveolin-1, and -2 protein and of prostacyclin synthase and peroxisome proliferator-activated receptor (PPAR)-␥ genes and proteins (1, 51). These endothelial cells also do not obey the "law of the monolayer" and instead form cell clusters or tumorlets (52, 53).Our rat model of SPH (48, 50) resembles the human disease because it demonstrates endothelial cell proliferation and the occlusion of small precapillary arterioles. In this model, chronic VEGF receptor blockade in combination with chronic hypoxia causes the initial apoptosis of lung endothelial cells, which is followed by the selection of phenotypically altered apoptosis-resistant endothelial cells. Endothelial cell proli...

Rationale: Severe pulmonary arterial hypertension (SPH) is a frequently lethal condition characterized by pulmonary vascular remodeling and right heart strain or failure. SPH is also often associated with autoimmune and collagen vascular disorders. Objectives: To study the effects of T cells on the development of experimental SPH. Conclusions: These studies demonstrate the importance of a complete, intact immune system in protecting against pulmonary angioproliferation in this new model of SPH as well as the importance of intact VEGF receptor signaling for lung endothelial cell homeostasis.

Tuberculosis-specific ELISPOT testing is a promising tool that should be evaluated as a potential diagnostic test for childhood tuberculosis. We caution against the use of an early decrease in response as a marker of successful antituberculous chemotherapy.

We have previously hypothesized that the development of severe angioproliferative pulmonary hypertension is associated with not only initial endothelial cell (EC) apoptosis followed by the emergence of apoptosis-resistant proliferating EC but also with proliferation of vascular smooth muscle cells (VSMC). We have demonstrated that EC death results in the selection of an apoptosis-resistant, proliferating, and phenotypically altered EC phenotype. We postulate here that the initial apoptosis of EC induces the release of mediators that cause VSMC proliferation. We cultured EC in an artificial capillary CellMax system designed to simulate the highly efficient functions of the human capillary system. We induced apoptosis of microvascular EC using shear stress and the combined VEGF receptor (VEGFR-1 and -2) inhibitor SU-5416. Flow cytometry for the proliferation marker bromodeoxyuridine showed that serum-free medium conditioned by apoptosed EC induced proliferation of VSMC, whereas serum-free medium conditioned by nonapoptosed EC did not. We also show that medium conditioned by apoptosed EC is characterized by increased concentrations of transforming growth factor (TGF)-β1 and VEGF compared with medium conditioned by nonapoptosed EC and that TGF-β1 blockade prevented the proliferation of cultured VSMC. In conclusion, EC death induced by high shear stress and VEGFR blockade leads to the production of factors, in particular TGF-β1, that activate VSMC proliferation.