Activation of the plasma kallikrein-kinin system on respiratory epithelium and pleural mesothelium

Julius Varano della Vergiliana

Research output: ThesisDoctoral Thesis

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[Truncated abstract] The plasma kallikrein-kinin system (KKS) is a cell-associated proteolytic mechanism of activation resulting in the release of the inflammatory peptide, bradykinin (BK). This process involves assembly of high molecular weight kininogen (HK) and plasma prekallikrein (PPK), and activation of the HK-PPK complex by prolylcarboxypeptidase (PRCP) or heat shock protein 90 (HSP90). Activation of the HK-PPK complex results in conversion of PPK to plasma kallikrein (PK) which, in turn, liberates BK from HK. This system has been comprehensively described on endothelial cells and more recently other cells have been shown to possess such a system. However, the significance of the plasma KKS on respiratory epithelium and pleural mesothelium is unclear. As such, this thesis describes the assembly and activation of the plasma KKS on these cell types. The A549 and BEAS-2B respiratory epithelial cell lines were shown to express known endothelial HK receptor-associated proteins namely, urokinase plasminogen activator receptor (uPAR), cytokeratin 1 (CK1) and gC1qR, but not Mac-1. Additionally, A549 cells bound FITC-labeled HK, which was inhibited by EDTA or 50-fold molar excess unlabeled HK. However, binding of FITC-HK was only weakly inhibited following pre-treatment with individual antibodies against uPAR, gC1qR or CK1, although about 45% inhibition was achieved when the antibodies were used in combination. In addition, sodium chlorate treatment had no effect on FITC-HK binding, indicating sulphated proteoglycans were not involved. PK activity was generated following sequential treatment of A549 and normal human bronchial epithelial (NHBE) cells with HK and PPK, resulting in the release of BK from HK. Similar results were also observed using A549 cell-free matrix and lysate. Activation of PPK on A549 and NHBE cells was inhibited by cysteine, BK, protamine sulphate and the HSP90 inhibitor, novobiocin.
Original languageEnglish
QualificationDoctor of Philosophy
Publication statusUnpublished - 2010


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