Simple and highly sensitive assay system for TNFR2-mediated soluble- and transmembrane-TNF activity
| Autor: | Shin-ichi Tsunoda, Yohei Mukai, Tetsuya Nomura, Yasuhiro Abe, Hiroyuki Miyoshi, Haruhiko Kamada, Hiroko Shibata, Hiroyuki Kayamuro, Tomoaki Yoshikawa, Shinsaku Nakagawa, Kazufumi Katayama, Yasuo Yoshioka, Yasuo Tsutsumi, Kyoko Minowa |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
Programmed cell death
Cell Survival medicine.medical_treatment Fas-Associated Death Domain Protein Recombinant Fusion Proteins Immunology Genetic Vectors Anti-Inflammatory Agents Drug Evaluation Preclinical Apoptosis Biology Transfection law.invention Mice law medicine Adipocytes Immunology and Allergy Animals Humans Receptors Tumor Necrosis Factor Type II fas Receptor Receptor Cells Cultured Tumor Necrosis Factor-alpha Cell Membrane Lentivirus Molecular biology Transmembrane protein Protein Structure Tertiary Transmembrane domain Cytokine Receptors Tumor Necrosis Factor Type I Cancer research Recombinant DNA Tumor necrosis factor alpha Biological Assay |
| Zdroj: | Journal of immunological methods. 335(1-2) |
| ISSN: | 0022-1759 |
| Popis: | Drugs that target tumor necrosis factor-alpha (TNF) are particularly important in the treatment of severe inflammatory progression in rheumatoid arthritis, Crohn's disease and psoriasis. Despite the central role of the TNF/TNF receptor (TNFR) in various disease states, there is a paucity of information concerning TNFR2 signaling. In this study, we have developed a simple and highly sensitive cell-death based assay system for analyzing TNFR2-mediated bioactivity that can be used to screen for TNFR2-selective drugs. Using a lentiviral vector, a chimeric receptor was engineered from the extracellular and transmembrane domain of human TNFR2 and the intracellular domain of mouse Fas and the recombinant protein was then expressed in TNFR1(-/-)R2(-/-) mouse preadipocytes. Our results demonstrate that this chimeric receptor is capable of inducing apoptosis by transmembrane- as well as soluble-TNF stimuli. Moreover, we found that our bioassay based on cell death phenotype had an approximately 80-fold higher sensitivity over existing bioassays. We believe our assay system will be an invaluable research tool for studying TNFR2 and for screening TNFR2-targeted drugs. |
| Databáze: | OpenAIRE |
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