epidermal-growth-factor and icatibant

In this study, we investigated the role of the kallikrein-kinin-system in head and neck squamous cell carcinoma (HNSCC) and its implication on tumour survival, invasion, migration and response to radiotherapy.. The expression of BKB2R was studied in a series of 180 tumour samples to determine the functional significance of BKB2R in HNSCC. Additionally, four different HNSCC cell lines were treated with an irradiation dose of 8Gy following bradykinin receptor stimulation or blockage. Tumour cell survival was tested using a colony formation assay. The invasive potential of tumour cells was assessed using Matrigel invasion chambers, the cells' ability to migrate was determined with a wound-healing assay. To examine the biochemical activation of BKB2R, the epidermal growth factor receptor (EGFR) and its downstream pathways, western blot analyses were conducted.. Immunohistochemistry revealed an over-expression of BKB2R in HNSCC tumour cells in comparison to normal peritumoural tissue. Blocking the BKB2R at irradiated tumour cells led to a reduced response to radiotherapy of tumour cells and led to an activation of the EGFR and its downstream pathways, known mediators of tumour cell survival, migration and invasion. Bradykinin stimulation also resulted in a better tumour cell survival, but these effects were achieved via an EGFR-independent signalling.. Our results demonstrate that the kallikrein-kinin-system is involved in survival, invasion and migration of HNSCC cells.

Topics: Adult; Aged; Blotting, Western; Bradykinin; Bradykinin B2 Receptor Antagonists; Cell Line, Tumor; Epidermal Growth Factor; Female; Head and Neck Neoplasms; Humans; Immunohistochemistry; Kallikrein-Kinin System; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Metastasis; Signal Transduction

The intracellular effects of bradykinin are mediated through the recently cloned B2 kinin receptor which belongs to the superfamily of receptors with seven transmembrane domains. The molecular events which transduce the bradykinin signal on the post-receptor level are not understood in detail. We studied whether in human foreskin fibroblasts bradykinin treatment induces tyrosine phosphorylation of cellular proteins. Using phosphotyrosine antibodies we detected a bradykinin-dependent phosphorylation of a group of proteins of about 130 kDa and an additional signal around 70kDa after starvation of cells. The effect evoked by 10 nM bradykinin was rapid (2 min) and it was partially reduced by the B2-kinin-receptor antagonist Hoe 140 which was shown to be a weak inducer of tyrosine phosphorylation. The bradykinin-mediated tyrosine phosphorylation events were reproduced in human embryonal kidney 293 fibroblasts which were transiently transfected with the rat B2 kinin receptor, but they were not observed in untransfected 293 control cells. These data suggest that the B2 kinin-receptor subtype is involved. Upon fractionation of cells the 130kDa protein group was recovered both in the membrane and the cytosolic protein fraction. To assess the specificity of this bradykinin effect we stimulated human foreskin fibroblasts with epidermal growth factor (EGF), platelet-derived growth factor (PDGF), insulin-like growth factor (IGF-I) and insulin. While IGF-I, insulin and EGF were almost ineffective, PDGF stimulated the tyrosine phosphorylation of 130-kDa bands with a similar pattern to that produced by bradykinin. Immunoprecipitation experiments with specific antibodies against potential candidate proteins in the molecular-mass range around 130kDa revealed positive results for the focal adhesion kinase FAK and the p130 Src substrate while negative results were obtained for the GTPase-activating protein GAP, the phospholipase C-gamma1, the Janus kinase JAK-1 and vinculin. The data suggest that the tyrosine phosphorylation of FAK and the pl30 Src substrate might be involved in the B2-kinin-receptor signalling cascade.

Topics: Animals; Bradykinin; Cell Line; Cells, Cultured; Epidermal Growth Factor; Fibroblasts; Humans; Insulin; Insulin-Like Growth Factor I; Kidney; Male; Molecular Weight; Phosphoproteins; Phosphorylation; Phosphotyrosine; Platelet-Derived Growth Factor; Rats; Receptor, Bradykinin B2; Receptors, Bradykinin; Recombinant Proteins; Skin; Transfection; Tyrosine