Human tissue factor pathway inhibitor-2 induces caspase-mediated apoptosis in a human fibrosarcoma cell line
| Autor: | Prakasha Kempaiah, Walter Kisiel |
|---|---|
| Rok vydání: | 2008 |
| Předmět: |
Cancer Research
Fibrosarcoma Clinical Biochemistry Down-Regulation Pharmaceutical Science Apoptosis Caspase 3 Cell Line Tumor Humans RNA Messenger Fragmentation (cell biology) education Caspase Glycoproteins bcl-2-Associated X Protein Pharmacology Inhibitor of apoptosis domain education.field_of_study biology Biochemistry (medical) Cell Biology Flow Cytometry Molecular biology Recombinant Proteins Tissue-factor-pathway inhibitor 2 Up-Regulation Cell biology Proto-Oncogene Proteins c-bcl-2 Urinary Bladder Neoplasms Cell culture Caspases Colonic Neoplasms biology.protein Signal transduction Apoptosis Regulatory Proteins |
| Zdroj: | Apoptosis. 13:702-715 |
| ISSN: | 1573-675X 1360-8185 |
| DOI: | 10.1007/s10495-008-0207-8 |
| Popis: | Human TFPI-2 is an extracellular matrix-associated Kunitz-type serine proteinase inhibitor. We previously demonstrated that a human fibrosarcoma cell line, HT-1080, does not express TFPI-2, but genetic restoration of TFPI-2 expression in these cells markedly inhibited their growth and metastasis in vivo. In the present study, either full-length recombinant TFPI-2, or its mutated first Kunitz-type domain (R24K KD1), were offered to HT-1080 cells, and the degree of apoptosis assessed by nuclear fragmentation, ethidium bromide and acridine orange staining, fluorescence activated cell sorting, immunoblotting and gene expression profiling. R24K KD1 induced apoptosis in 69% of HT-1080 cells in a 48 h period compared to 39% for TFPI-2, while a KD1 preparation lacking a reactive site arginine/lysine residue (R24Q KD1) produced only an 18% apoptosis rate, suggesting that the observed apoptosis was related to proteinase inhibition. Immunoblotting experiments indicated increased caspase 3 and 9 activation, up-regulation of pro-apoptotic Bax and suppression of anti-apoptotic Bcl-2 protein. Finally, microarray analyses of R24K KD1-treated cells indicated elevated expression of several pro-apoptotic genes and under-expression of anti-apoptotic genes. Collectively, our results demonstrate that treatment of HT-1080 cells exogenously with either TFPI-2 or R24K KD1 activates caspase-mediated, pro-apoptotic signaling pathways resulting in apoptosis. |
| Databáze: | OpenAIRE |
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