Transforming growth factor-␤ (TGF-␤) is involved in vascular formation through activin receptor-like kinase (ALK)1 and ALK5. ALK5, which is expressed ubiquitously, phosphorylates Smad2 and Smad3, whereas endothelial cell (EC)-specific ALK1 activates Smad1 and Smad5. Because ALK5 kinase activity is required for ALK1 to transduce TGF-␤ signaling via Smad1/5 in ECs, ALK5 knockout (KO) mice were not able to give us the precise mechanisms by which TGF-␤/ALK5/Smad2/3 signaling is implicated in angiogenesis. To delineate the role of Smad2/3 signaling in endothelium, the Smad2 gene in Smad3 KO mice was selectively deleted in ECs using Tie2-Cre transgenic mice, termed EC-specific Smad2/3 double KO (EC-Smad2/3KO) mice. EC-Smad2/3KO embryos revealed hemorrhage leading to embryonic lethality around E12.5. EC-Smad2/3KO embryos exhibited no abnormality of vasculogenesis and angiogenesis in both the yolk sac and the whole embryo, whereas vascular maturation was incomplete because of inadequate assembly of mural cells in the vasculature. Wide gaps between ECs and mural cells could be observed in the vasculature of EC-Smad2/3KO mice because of reduced expression of Ncadherin and sphingosine-1-phosphate receptor-1 (S1PR1) in ECs from those mice. These results indicated that Smad2/3 signaling in ECs is indispensable for maintenance of vascular integrity via the fine-tuning of N-cadherin, VEcadherin, and S1PR1 expressions in the vasculature.(Blood. 2012;119(22): 5320-5328) IntroductionAberrant vascularization leads to a number of diseases including atherosclerosis, tumorigenicity, and retinopathy, 1,2 whereas angiogenesis is essential during embryonic development as well as in adulthood. Angiogenesis is mediated by sprouting of new vessels from preexisting ones or by intussusceptive microvascular growth. In general, vascular formation is quiet in adulthood, although angiogenesis involved in wound healing, inflammation, ischemia, and the female reproductive cycle can be observed. Angiogenesis is divided into 2 phases: the activation phase and the resolution phase. The balance between physiologic stimulators (eg, vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF-2), angiopoietins, and hypoxia) and inhibitors (eg, angiostatin, endostatin, and interferon-␣) is strategic to tuning of the angiogenic switch. Proliferation of endothelial cells (ECs), increase in vascular permeability, and degradation of extracellular matrix components can be observed during the activation phase. Consequently, ECs make new capillary sprouts. In the resolution phase, the proliferation and migration of ECs ceases and is followed by reconstitution of the basement membrane and maturation of the vessels. 3 Transforming growth factor-␤ (TGF-␤) is a pivotal cytokine that contributes to the behaviors and activities of most cells from the embryonic to the adult stage. The TGF-␤ signal is initiated when the ligand binds to its own TGF-␤ type II receptor (T␤RII); thereafter, the TGF-␤ type I receptor (T␤RI or activin receptor-like kinase [ALK]...

Background Transforming growth factor (TGF)-β is a multifunctional cytokine involved in cell differentiation, cell proliferation, and tissue homeostasis. Although TGF-β signaling is essential for maintaining blood vessel functions, little is known about the role of TGF-β in lymphatic homeostasis. Methods To delineate the role of TGF-β signaling in lymphatic vessels, TβRIIfl/fl mice were crossed with Prox1-CreERT2 mice to generate TβRIIfl/fl; Prox1-CreERT2 mice. The TβRII gene in the lymphatic endothelial cells (LECs) of the conditional knockout TβRIIiΔLEC mice was selectively deleted using tamoxifen. The effects of TβRII gene deletion on embryonic lymphangiogenesis, postnatal lymphatic structure and drainage function, tumor lymphangiogenesis, and lymphatic tumor metastasis were investigated. Results Deficiency of LEC-specific TGF-β signaling in embryos, where lymphangiogenesis is active, caused dorsal edema with dilated lymphatic vessels at E13.5. Postnatal mice in which lymphatic vessels had already been formed displayed dilation and increased bifurcator of lymphatic vessels after tamoxifen administration. Similar dilation was also observed in tumor lymphatic vessels. The drainage of FITC-dextran, which was subcutaneously injected into the soles of the feet of the mice, was reduced in TβRIIiΔLEC mice. Furthermore, Lewis lung carcinoma cells constitutively expressing GFP (LLC-GFP) transplanted into the footpads of the mice showed reduced patellar lymph node metastasis. Conclusion These data suggest that TGF-β signaling in LECs maintains the structure of lymphatic vessels and lymphatic homeostasis, in addition to promoting tumor lymphatic metastasis. Therefore, suppression of TGF-β signaling in LECs might be effective in inhibiting cancer metastasis.

Transforming growth factor‐β (TGF‐β) is a potent growth inhibitor in normal epithelial cells. However, a number of malignant tumors produce excessive amounts of TGF‐β, which affects the tumor‐associated microenvironment by furthering the progression of tumorigenicity. Although it is known that the tumor‐associated microenvironment often becomes hypoxic, how hypoxia influences TGF‐β signaling in this microenvironment is unknown. We investigated whether TGF‐β signaling is influenced by long‐term exposure to hypoxia in Lewis lung carcinoma (LLC) cells. When the cells were exposed to hypoxia for more than 10 days, their morphology was remarkably changed to a spindle shape, and TGF‐β‐induced Smad2 phosphorylation was enhanced. Concomitantly, TGF‐β‐induced transcriptional activity was augmented under hypoxia, although TGF‐β did not influence the activity of a hypoxia‐responsive reporter. Consistently, hypoxia influenced the expression of several TGF‐β target genes. Interestingly, the expressions of TGF‐β type I receptor (TβRI), also termed activin receptor like kinase‐5 (ALK5), and TGF‐β1 were increased under the hypoxic condition. When we monitored the hypoxia‐inducible factor‐1 (HIF‐1) transcriptional activity by use of green fluorescent protein governed by the hypoxia‐responsive element in LLC cells transplanted into mice, TGF‐β‐induced Smad2 phosphorylation was upregulated in vivo. Our results demonstrate that long‐term exposure to hypoxia might alter responsiveness to TGF‐β signaling and affected the malignancy of LLC cells.

Dual polymerase activities of TERT as therapeutic targets RXRα promotes growth of cholangiocarcinoma through simultaneous activation of the Wnt/β-catenin and NF-κB pathways miR-1 expression predicts a poor prognosis of breast cancer wileyonlinelibrary.com/journal/cas