Functional analyses for transmembrane TNF-alpha

tnf - α拮抗剂因其在活动性慢性炎症性疾病中的显著临床疗效而受到关注。在类风湿性关节炎中，英夫利昔单抗（嵌合抗tnf - α抗体）、阿达木单抗（人源化抗tnf - α抗体）和依那西普（p75 tnf - α受体融合蛋白）都是非常有效的，而在克罗恩病中，只有英夫利昔单抗和阿达木单抗可以诱导临床缓解。由于不同的临床疗效可能是由生物效应引起的，而不仅仅是可溶性tnf - α的中和，我们在这里研究了通过跨膜tnf - α的反向信号传导。利用表达mTNF的Jurkat T细胞，用cDNA阵列分析英夫利昔单抗诱导的mTNF。mTNF细胞质中2、5和27位的丝氨酸残基通过定点诱变被依次替换为丙氨酸。通过替换所有这三种细胞质丝氨酸残基，英夫利昔单抗诱导的细胞凋亡和细胞周期阻滞被完全消除。在这项研究中，我们发现英夫利昔单抗和依那西普诱导的新的生物学效应是由mTNF的反向信号介导的，这可能解释了这两种抗tnf - α的临床疗效差异。代理。我们还澄清了细胞质丝氨酸残基对mTNF的信号转导至关重要。

TNF-alpha antagonists are the center of attention for their dramatic clinical efficacy in active chronic inflammatory diseases. In rheumatoid arthritis, both infliximab (chimeric anti-TNF-alpha antibody), adalimumab (humanized anti-TNF-alpha antibody) and etanercept (p75 TNF-alpha receptor fusion protein) are highly effective, while in Crohn's disease, only infliximab and adalimumab can induce clinical remission. As the differential clinical efficacy was likely to be caused by biological effects other than mere neutralization of soluble TNF-alpha, we here investigated reverse signaling through transmembrane TNF-alpha. mTNF induced by infliximab was analysed by cDNA array in the assay system using mTNF-expressing Jurkat T cells.Cytoplasmic serine residues at positions 2, 5 and 27 of mTNF were sequentially substituted to alanine by site-directed mutagenesis. Infliximab-induced apoptosis and cell cycle arrest were completely abrogated by substitution of all of these three cytoplasmic serine residues. In this study, we revealed that novel biological effects induced by infliximab and etanercept were mediated by reverse signaling of mTNF, which might explain the differential clinical efficacy of these anti-TNF-alpha. agents. We also clarified that cytoplasmic serine residues were critical for the signal transduction through mTNF.