Simple and highly sensitive assay system for TNFR2-mediated soluble- and transmembrane-TNF activity

被引：12

作者：

Abe, Yasuhiro ^{[1
,2
]}

Yoshikawa, Tomoaki ^{[1
,3
]}

Kamada, Haruhiko ^{[1
,4
]}

Shibata, Hiroko ^{[1
,5
]}

Nomura, Tetsuya ^{[1
,2
]}

Minowa, Kyoko ^{[1
,6
]}

Kayamuro, Hiroyuki ^{[1
,2
]}

Katayama, Kazufumi ^{[7
]}

Miyoshi, Hiroyuki ^{[8
]}

Mukai, Yohei ^{[1
,2
]}

Yoshioka, Asuo ^{[1
,3
]}

Nakagawa, Shinsaku ^{[2
]}

Tsunoda, Shin-ichi ^{[1
,4
]}

Tsutsumi, Yasuo ^{[1
,2
,4
]}

机构：

[1] Natl Inst Biomed Innovat, Lab Pharmaceut Proteom, Osaka 5670085, Japan

[2] Osaka Univ, Grad Sch Pharmaceut Sci, Suita, Osaka 5650871, Japan

[3] Osaka Univ, Ctr Adv Res & Educ Drug Discovery & Dev, Suita, Osaka 5650871, Japan

[4] Osaka Univ, Ctr Adv Med Engn & Informat, Suita, Osaka 5650871, Japan

[5] Natl Inst Hlth Sci, Div Drugs, Setagaya Ku, Tokyo 1588501, Japan

[6] Kyoto Pharmaceut Univ, Grad Sch Pharmaceut Sci, Yamashina Ku, Kyoto 6078414, Japan

[7] Tokyo Metropolitan Inst Med Sci, Allergy & Immunol Project, Bunkyo Ku, Tokyo 1138613, Japan

[8] RIKEN, Subteam Manipulat Cell Fate, BioResource Ctr, Tsukuba, Ibaraki 3050074, Japan

来源：

JOURNAL OF IMMUNOLOGICAL METHODS | 2008年 / 335卷 / 1-2期

基金：

日本学术振兴会;

关键词：

TNF; TNFR2; Fas; chimeric receptor; bioassay;

D O I：

10.1016/j.jim.2008.02.019

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

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. (C) 2008 Elsevier B.V. All rights reserved.

引用

页码：71 / 78

页数：8

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