Interaction Of Tumor-Derived Tissue Factor With Endothelial Cells In The Context Of Cancer Metastasis
The metastatic cascade comprises of a series of events, which tumor cells must complete to form macrometastases at a secondary site. Due to its spread and aggressive nature, metastatic tumor is difficult to treat, and tumor metastasis remains the main cause of death in cancer patients. Most epithelial tumors over-express tissue factor (TF), the initiator of the extrinsic pathway of coagulation cascade. This over-expression of TF has been shown to tumor progression and metastasis in different studies. In this thesis, we studied two mechanisms by which tumor-derived TF can interact with endothelial cells to promote cancer metastasis. Endothelial cells act as a natural barrier between the tumor cells in circulation and secondary organs. Tumor cells must arrest on endothelial cells in order to extravasate and enter a new organ. Most previous studies have focused on the classical receptor-ligand interactions involved in leukocyte recruitment during inflammation, such as selectins and integrins. We hypothesized that TFexpressing tumor cells can mediate adhesion to the endothelium by interacting with its endogenous inhibitor, tissue factor pathway inhibitor (TFPI), which is constitutively expressed on endothelial cells. Using in vitro models, we found that high TF-expressing tumor cells bound to recombinant TFPI and endothelial cells under static conditions; however, under shear, the tumor cells adhered to recombinant TFPI, but not endothelial cells. We postulate that the TFPI expres- sion in our endothelial cell model (human umbilical vein) was too low to mediate adhesion. Unfortunately, due to the lack of availability of other primary endothelial cells that express higher TFPI (e.g. lung microvascular endothelium), we could not test this theory further. Endothelial cells, highly dynamic and responsive to its local environment, perform a range of functions in the body. Endothelial responses could be hijacked by tumor cells to promote a tumor-permissive metastatic environment. Tumor cells secrete extracellular vesicles (EV) that can interact with distant cells and re-educate them to a pro-tumor phenotype. Tumor cells secrete two types of EV - microvesicles (100-1000nm) and exosomes (30-100nm). In this part of the thesis, we evaluated the isolation of exosomes and microvesicles using common established protocols - filtration and differential centrifugation. We found that the isolation methods highly impacted the size distribution, the purity and TF-mediated procoagulant activity in the isolated fractions. We then evaluated whether tumor-derived TF-expressing EV (consist of microvesicles and exosomes) promote pro-metastatic behaviors in endothelial cells in vitro. We observed binding and uptake of EV by endothelial cells. These EV also signaled to endothelial cells and promoted a pro-adhesive (upregulation of E-selectin and intercellular cellular adhesion molecule-1), pro-inflammatory (increased secretion of monocyte chemotactic protein-1 and interleuking-8) and procoagulant (increase in TF-mediated procoagulant activity) phenotype in endothelial cells. The mechanism was dependent on TF-mediated FXa generation by EV, and activation of protease-activated receptor-1 followed by transactivation of epidermal growth factor receptor on endothelial cells.
Tissue Factor; Cancer Metastasis
Wiesner,Ulrich B.; Stokol,Tracy; Kirby,Brian
Ph. D., Biomedical Engineering
Doctor of Philosophy
dissertation or thesis