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Participation of the PI-3K/Akt-NF-kappa B signaling pathways in hypoxia-induced mitogenic factor-stimulated Flk-1 expression in endothelial cells.

ABSTRACT: Hypoxia-induced mitogenic factor (HIMF), a lung-specific growth factor, promotes vascular tubule formation in a matrigel plug model. We initially found that HIMF enhances vascular endothelial growth factor (VEGF) expression in lung epithelial cells. In present work, we tested whether HIMF modulates expression of fetal liver kinase-1 (Flk-1) in endothelial cells, and dissected the possible signaling pathways that link HIMF to Flk-1 upregulation.Recombinant HIMF protein was intratracheally instilled into adult mouse lungs, Flk-1 expression was examined by immunohistochemistry and Western blot. The promoter-luciferase reporter assay and real-time RT-PCR were performed to examine the effects of HIMF on Flk-1 expression in mouse endothelial cell line SVEC 4-10. The activation of NF-kappa B (NF-kappaB) and phosphorylation of Akt, IKK, and IkappaBalpha were examined by luciferase assay and Western blot, respectively.Intratracheal instillation of HIMF protein resulted in a significant increase of Flk-1 production in lung tissues. Stimulation of SVEC 4-10 cells by HIMF resulted in increased phosphorylation of IKK and IkappaBalpha, leading to activation of NF-kappaB. Blocking NF-kappaB signaling pathway by dominant-negative mutants of IKK and IkappaBalpha suppressed HIMF-induced Flk-1 upregulation. Mutation or deletion of NF-kappaB binding site within Flk-1 promoter also abolished HIMF-induced Flk-1 expression in SVEC 4-10 cells. Furthermore, HIMF strongly induced phosphorylation of Akt. A dominant-negative mutant of PI-3K, Deltap85, as well as PI-3K inhibitor LY294002, blocked HIMF-induced NF-kappaB activation and attenuated Flk-1 production.These results suggest that HIMF upregulates Flk-1 expression in endothelial cells in a PI-3K/Akt-NF-kappaB signaling pathway-dependent manner, and may play critical roles in pulmonary angiogenesis.

SUBMITTER: Tong Q

PROVIDER: S-EPMC1570355 | biostudies-literature | 2006 Jul

REPOSITORIES: biostudies-literature

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Participation of the PI-3K/Akt-NF-kappa B signaling pathways in hypoxia-induced mitogenic factor-stimulated Flk-1 expression in endothelial cells.

Tong Qiangsong Q Zheng Liduan L Lin Li L Li Bo B Wang Danming D Huang Chuanshu C Matuschak George M GM Li Dechun D

Respiratory research 20060727

<h4>Background</h4>Hypoxia-induced mitogenic factor (HIMF), a lung-specific growth factor, promotes vascular tubule formation in a matrigel plug model. We initially found that HIMF enhances vascular endothelial growth factor (VEGF) expression in lung epithelial cells. In present work, we tested whether HIMF modulates expression of fetal liver kinase-1 (Flk-1) in endothelial cells, and dissected the possible signaling pathways that link HIMF to Flk-1 upregulation.<h4>Methods</h4>Recombinant HIMF ...[more]

PMID: 16872509

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Project description:BACKGROUND: The unique responsiveness of Vgamma9Vdelta2 T-cells, the major gammadelta subset of human peripheral blood, to non-peptidic prenyl pyrophosphate antigens constitutes the basis of current gammadelta T-cell-based cancer immunotherapy strategies. However, the molecular mechanisms responsible for phosphoantigen-mediated activation of human gammadelta T-cells remain unclear. In particular, previous reports have described a very slow kinetics of activation of T-cell receptor (TCR)-associated signal transduction pathways by isopentenyl pyrophosphate and bromohydrin pyrophosphate, seemingly incompatible with direct binding of these antigens to the Vgamma9Vdelta2 TCR. Here we have studied the most potent natural phosphoantigen yet identified, (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), produced by Eubacteria and Protozoa, and examined its gammadelta T-cell activation and anti-tumor properties. METHODOLOGY/PRINCIPAL FINDINGS: We have performed a comparative study between HMB-PP and the anti-CD3epsilon monoclonal antibody OKT3, used as a reference inducer of bona fide TCR signaling, and followed multiple cellular and molecular gammadelta T-cell activation events. We show that HMB-PP activates MEK/Erk and PI-3K/Akt pathways as rapidly as OKT3, and induces an almost identical transcriptional profile in Vgamma9(+) T-cells. Moreover, MEK/Erk and PI-3K/Akt activities are indispensable for the cellular effects of HMB-PP, including gammadelta T-cell activation, proliferation and anti-tumor cytotoxicity, which are also abolished upon antibody blockade of the Vgamma9(+) TCR Surprisingly, HMB-PP treatment does not induce down-modulation of surface TCR levels, and thereby sustains gammadelta T-cell activation upon re-stimulation. This ultimately translates in potent human gammadelta T-cell anti-tumor function both in vitro and in vivo upon transplantation of human leukemia cells into lymphopenic mice, CONCLUSIONS/SIGNIFICANCE: The development of efficient cancer immunotherapy strategies critically depends on our capacity to maximize anti-tumor effector T-cell responses. By characterizing the intracellular mechanisms of HMB-PP-mediated activation of the highly cytotoxic Vgamma9(+) T-cell subset, our data strongly support the usage of this microbial antigen in novel cancer clinical trials.

Dataset Information

Participation of the PI-3K/Akt-NF-kappa B signaling pathways in hypoxia-induced mitogenic factor-stimulated Flk-1 expression in endothelial cells.

Publications

Participation of the PI-3K/Akt-NF-kappa B signaling pathways in hypoxia-induced mitogenic factor-stimulated Flk-1 expression in endothelial cells.

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