Platelets, in addition to their function in hemostasis, play an important role in wound healing and tumor growth. Because platelets contain angiogenesis stimulators and inhibitors, the mechanisms by which platelets regulate angiogenesis remain unclear. As platelets adhere to activated endothelium, their action can enhance or inhibit local angiogenesis. We therefore suspected a higher organization of angiogenesis regulators in platelets. Using double immunofluorescence and immunoelectron microscopy, we show that pro-and antiangiogenic proteins are separated in distinct subpopulations of ␣-granules in platelets and megakaryocytes. Double immunofluorescence labeling of vascular endothelial growth factor (VEGF) (an angiogenesis stimulator) and endostatin (an angiogenesis inhibitor), or for thrombospondin-1 and basic fibroblast growth factor, confirms the segregation of stimulators and inhibitors into separate and distinct ␣-granules. These observations motivated the hypothesis that distinct populations of ␣-granules could undergo selective release. The treatment of human platelets with a selective PAR4 agonist (AYPGKF-NH 2 ) resulted in release of endostatin-containing granules, but not VEGF-containing granules, whereas the selective PAR1 agonist (TFLLR-NH 2 ) liberated VEGF, but not endostatin-containing granules. In conclusion, the separate packaging of angiogenesis regulators into pharmacologically and morphologically distinct populations of ␣-granules in megakaryocytes and platelets may provide a mechanism by which platelets can locally stimulate or inhibit angiogenesis. IntroductionAngiogenesis, the process of new vessel development, plays an essential role in embryogenesis, but postnatal angiogenesis is limited to sites of abnormal vascular surface. An activated vascular endothelium can be induced by tissue injury or wound healing, by hormonal cycling such as in pregnancy and ovulation, or by tumor-induced vessel growth. In all of these circumstances, platelets act as the initial responder to vascular change and provide a flexible delivery system for angiogenesis-related molecules. [1][2][3][4] The process of postnatal angiogenesis is regulated by a continuous interplay of stimulators and inhibitors of angiogenesis, and their imbalance contributes to numerous inflammatory, malignant, ischemic, and immune disorders. 5 There is a revived interest in the overlap between angiogenesis and platelets 6 because several clinical trials have now shown that anticoagulation can improve cancer survival 7,8 beyond the benefit derived from the treatment of deep vein thrombosis alone.It is known that platelets stimulate endothelial cells in culture and can promote the assembly of capillary-like structures in vitro. 9,10 Platelets may modulate angiogenesis by releasing promoters such as vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), platelet derived growth factor (PDGF), and matrix metalloproteinases (MMPs). 1,6,[11][12][13][14][15][16][17][18] The repertoire o...

SUMMARY Lung stem cells are instructed to produce lineage-specific progeny through unknown factors in their microenvironment. We used clonal three-dimensional (3D) co-cultures of endothelial cells and distal lung stem cells, bronchioalveolar stem cells (BASCs), to probe the instructive mechanisms. Single BASCs had bronchiolar and alveolar differentiation potential in lung endothelial cell co-cultures. Gain and loss of function experiments showed BMP4-Bmpr1a signaling triggers calcineurin/NFATc1-dependent expression of Thrombospondin-1 (Tsp1) in lung endothelial cells to drive alveolar lineage-specific BASC differentiation. Tsp1-null mice exhibited defective alveolar injury repair, confirming a crucial role for the BMP4-NFATc1-TSP1 axis in lung epithelial differentiation and regeneration in vivo. Discovery of this pathway points to methods to direct the derivation of specific lung epithelial lineages from multipotent cells. These findings elucidate a pathway that may be a critical target in lung diseases and provide new tools to understand the mechanisms of respiratory diseases at the single cell level.

Metastasis is responsible for the majority of prostate cancer-related deaths; however, little is known about the molecular mechanisms that underlie this process. Here we identify an oncogene-tumor suppressor cascade that promotes prostate cancer initiation and metastasis by coordinately activating Ras and NF-κB. Specifically, we show that loss of the RasGAP gene DAB2IP induces metastatic prostate cancer in a murine model. Notably, DAB2IP functions as a signaling scaffold that coordinately regulates Ras and NF-κB through distinct domains to promote tumor initiation and metastasis, respectively. DAB2IP is suppressed in human prostate cancer where expression inversely correlates with tumor grade and predicts prognosis. Moreover, we report that epigenetic silencing of DAB2IP is a key mechanism by which the polycomb-group protein EZH2 activates Ras, NF-κB, and triggers metastasis. These studies define the mechanism by which two major pathways can be simultaneously activated in metastatic prostate cancer and establish EZH2 as a driver of metastasis.