Research article
Tissue factor/FVIIa activates Bcl-2 and prevents doxorubicin-induced apoptosis in neuroblastoma cells
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* Corresponding author: Muxiang Zhou mzhou@emory.edu
- † Equal contributors
1 Division of Hematology/Oncology, Department of Pediatrics, Emory University School of Medicine, Atlanta, USA
2 Department of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
3 Department of Neurosurgery, Wuhan No.1 Hospital, Wuhan, China
BMC Cancer 2008, 8:69 doi:10.1186/1471-2407-8-69
Published: 6 March 2008Abstract
Background
Tissue factor (TF) is a transmembrane protein that acts as a receptor for activated coagulation factor VII (FVIIa), initiating the coagulation cascade. Recent studies demonstrate that expression of tumor-derived TF also mediates intracellular signaling relevant to tumor growth and apoptosis. Our present study investigates the possible mechanism by which the interaction between TF and FVIIa regulates chemotherapy resistance in neuroblastoma cell lines.
Methods
Gene and siRNA transfection was used to enforce TF expression in a TF-negative neuroblastoma cell line and to silence endogenous TF expression in a TF-overexpressing neuroblastoma line, respectively. The expression of TF, Bcl-2, STAT5, and Akt as well as the phosphorylation of STAT5 and Akt in gene transfected cells or cells treated with JAK inhibitor and LY294002 were determined by Western blot assay. Tumor cell growth was determined by a clonogenic assay. Cytotoxic and apoptotic effect of doxorubicin on neuroblastoma cell lines was analyzed by WST assay and annexin-V staining (by flow cytometry) respectively.
Results
Enforced expression of TF in a TF-negative neuroblastoma cell line in the presence of FVIIa induced upregulation of Bcl-2, leading to resistance to doxorubicin. Conversely, inhibition of endogenous TF expression in a TF-overexpressing neuroblastoma cell line using siRNA resulted in down-regulation of Bcl-2 and sensitization to doxorubicin-induced apoptosis. Additionally, neuroblastoma cells expressing high levels of either endogenous or transfected TF treated with FVIIa readily phosphorylated STAT5 and Akt. Using selective pharmacologic inhibitors, we demonstrated that JAK inhibitor I, but not the PI3K inhibitor LY294002, blocked the TF/FVIIa-induced upregulation of Bcl-2.
Conclusion
This study shows that in neuroblastoma cell lines overexpressed TF ligated with FVIIa produced upregulation of Bcl-2 expression through the JAK/STAT5 signaling pathway, resulting in resistance to apoptosis. We surmise that this TF-FVIIa pathway may contribute, at least in part, to chemotherapy resistance in neuroblastoma.