Initial apoptosis is followed by increased proliferation of apoptosis‐resistant endothelial cells

Abstract

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.

Cytoplasmic provenance of STAT3 and PY-STAT3 in the endolysosomal compartments in pulmonary arterial endothelial and smooth muscle cells: implications in pulmonary arterial hypertension

“…Interestingly, the loss of cav-1 observed in plexiform lesions of idiopathic PAH was specific for these cells in that these authors did not find a decrease in cav-1 levels by Western blot assays of whole lung extracts. A similar decrease in cav-1 and cav-2 was also observed in a model of severe PAH in the rat caused by administration of the VEGF receptor antagonist SU-5416 followed by exposure of the animals to 3 wk of chronic hypoxia (1,37). Although there is evidence from several groups showing the pivotal role played by the downregulation of cav-1 in pulmonary arterial lesions of idiopathic PAH in humans and in diverse experimental models (MCT, hypoxia, and SU-5416 ϩ hypoxia), a recent report by Patel et al (34) appears to have reached a different conclusion.…”

Pathogenesis of pulmonary hypertension: a case for caveolin-1 and cell membrane integrity

“…Loss of ECs is thought to increase the proliferation of apoptosis-resistant cells and has been shown to release TGF-␤1 (a promoter of SMC proliferation) and VEGF (inhibitor of apoptosis), promoting SMC proliferation (111,112). Caveolin-1 plays an important role in TGF-␤ signaling.…”

Targeted therapy by disablingcrossroadsignaling networks: the survivin paradigm

“…Third, survivin expression is regulated by developmental signaling pathways operative in stem cells (i.e., Wnt) and it is possible that survivin antagonists may affect cancer stem cells (14), a compartment largely untouched by cytotoxics or targeted agents (15). Fourth, survivin is important in ancillary aspects of tumor formation/progression, especially angiogenesis (16,17), and survivin inhibitors have been shown to act on both the transformed population and endothelial cells in the tumor mass. Fifth, although survivin expression has been shown in cytokinestimulated hematopoietic progenitors and potentially in activated T cells, targeting this pathway did not affect normal cells or tissues in preclinical or phase I studies (see below), suggesting a favorable toxicity profile of survivinbased therapeutics.…”