The antiangiogenic factor 16K PRL induces programmed cell death in endothelial cells by caspase activation

We asked whether the antiangiogenic action of 16K human PRL (hPRL), in addition to blocking mitogen-induced vascular endothelial cell proliferation, involved activation of programmed cell death. Treatment with recombinant 16K hPRL increased DNA fragmentation in cultured bovine brain capillary endothelial (BBE) and human umbilical vein endothelial (HUVE) cells in a time- and dose-dependent fashion, independent of the serum concentration. The activation of apoptosis by 16K hPRL was specific for endothelial cells, and the activity of the peptide could be inhibited by heat denaturation, trypsin digestion, and immunoneutralization, but not by treatment with the endotoxin blocker, polymyxin-B. 16K hPRL-induced apoptosis was correlated with the rapid activation of caspases 1 and 3 and was blocked by pharmacological inhibition of caspase activity. Caspase activation was followed by inactivation of two caspase substrates, poly(ADP-ribose) polymerase (PARP) and the inhibitor of caspase-activated deoxyribonuclease (DNase) (ICAD). Furthermore, 16K hPRL increased the conversion of Bcl-X to its proapoptotic form, suggesting that the Bcl-2 protein family may also be involved in 16K hPRL-induced apoptosis. These findings support the hypothesis that the antiangiogenic action of 16K hPRL includes the activation of programmed cell death of vascular endothelial cells

TGF-beta induces proangiogenic and antiangiogenic factors via parallel but distinct Smad pathways

Contains fulltext : 57438.pdf (publisher's version ) (Closed access)Background. Angiogenesis has a key role in numerous disease processes. One of the most important angiogenic factors is vascular endothelial growth factor (VEGF-A), whereas thrombospondin-1 (TSP-1) is a major antiangiogenic factor. Recent studies have shown that VEGF-A as well as TSP-1 is regulated by transforming growth factor-beta1 (TGF-beta1), but the mechanism remains unclear. Methods. We examined the role of TGF-beta1 and its signaling pathways in mediating expression of these two molecules. Rat proximal tubular cells (NRK52E) were stimulated with TGF-beta1 to induce VEGF-A and TSP-1 synthesis. To clarify roles of receptor-activated Smads (R-Smads), we blocked Smad signaling using overexpression of the inhibitory Smad, Smad7, and by using fibroblasts from wild-type or knockout mice. To confirm the antiantigenic role of Smads, soluble Flt-1 regulation in response to TGF-beta1 was also examined. In addition, the effect of conditioned media from NRK52E and Smad knockout cells was examined on endothelial cell proliferation. Results. Induction of VEGF-A and TSP-1 by TGF-beta1 in NRK52E cells was associated with activation of pathway-restricted R-Smads (Smad2 and 3) and blocking these Smads by overexpression of Smad7 blocked their induction. By using of Smad knockout cells, Smad3 was shown to have a key role in the stimulation of VEGF-A expression whereas Smad2 was critical for TSP-1 expression. Consistent with the hypothesis that Smad2 has an antiangiogenic function, we also demonstrated that Smad2, but not Smad3, mediated the expression of VEGF-A antagonist, soluble VEGF-A receptor sFlt-1, in response to TGF-beta1. Conditioned media from NRK52E, which was stimulated by TGF-beta1 for 24 hours, did not induce endothelial cell proliferation. However, conditioned media from Smad2 knockout induced endothelial cell proliferation, whereas endothelial cell proliferation was inhibited by Smad3 knockout-derived conditioned media. Conclusion. R-Smads have distinct roles in mediating the expression of pro- and antiangiogenic growth factors in response to TGF-beta1

Incomplete inside-out growth pattern in invasive breast carcinoma: association with lymph vessel invasion and recurrence-free survival.

Invasive micropapillary carcinoma (IMPC) is a rare subtype of epithelial tumor of the breast listed in the 2003 World Health Organization histologic classification of tumors of the breast. It is characterized by inside-out micropapillary morphology, frequent lymph vessel invasion (LVI), and lymph node metastasis; however, its etiology remains unknown. This study investigated the incomplete inside-out growth pattern (IGP) in invasive ductal carcinoma, not otherwise specified (NOS), and examined the association between incomplete IGP and clinicopathologic features, including the presence of intratumoral lymph vessels (ILV), LVI, nodal metastasis, and prognosis. Tumor tissues from 166 invasive duct carcinomas NOS and 10 IMPCs were immunostained using an anti-epithelial membrane antigen antibody to detect IGP and with D2-40 antibody to determine the presence of ILV and LVI. Incomplete IGP was detected focally in 88 (53%) of 166 invasive duct carcinomas NOS. Transition areas between IMPC and invasive duct carcinoma NOS also showed prominent incomplete IGP in 9 (90%) of 10 IMPCs. Incomplete IGP in invasive duct carcinomas NOS was associated with larger tumor size, higher frequencies of ILV, LVI, nodal metastasis, and poorer recurrence-free survival by univariate analysis. Incomplete IGP, ILV, and tumor size independently affected LVI by multivariate analysis. These findings indicate that incomplete IGP of tumor cell clusters is not uncommon and is a useful tool for predicting LVI in invasive duct carcinoma NOS of the breast