Hofsli E, Bakke O, Nonstad U, Espevik T
Institute of Cancer Research, University of Trondheim, Norway.
Cell Immunol. 1989 Sep;122(2):405-15. doi: 10.1016/0008-8749(89)90087-7.
The production and localization of tumor necrosis factor (TNF) in human monocytes were investigated by using monoclonal and polyclonal antibodies against recombinant human TNF together with flow cytometry and immunofluorescence microscopy. Lipopolysaccharide (LPS) induced a rapid and transient accumulation of TNF in perinuclear vesicles which was detected 20 min after the addition of LPS. The fluorescence intensity of the vesicles peaked at 40 min of LPS exposure, concomitantly with the release of TNF into the medium. Thus, our results indicate that the secretion of TNF is typical for secretory proteins as it involves passage through the secretory apparatus. Additional studies demonstrated that plasma membrane-associated TNF could not be detected in live monocytes not exposed to LPS. However, after 90 min with LPS, a small population of monocytes expressed membrane-associated TNF, and by 24 hr approximately 50% of the monocytes displayed TNF on the plasma membrane. Furthermore, our results indicate that plasma membrane-associated TNF does not represent released TNF bound back to its own receptor. Thus, our findings support the view that TNF exists as a surface trans-membrane protein in LPS-stimulated monocytes.
通过使用针对重组人肿瘤坏死因子（TNF）的单克隆抗体和多克隆抗体，结合流式细胞术和免疫荧光显微镜技术，研究了人单核细胞中TNF的产生和定位。脂多糖（LPS）诱导TNF在核周小泡中快速短暂积累，在添加LPS后20分钟即可检测到。小泡的荧光强度在LPS暴露40分钟时达到峰值，与此同时TNF释放到培养基中。因此，我们的结果表明，TNF的分泌是分泌蛋白的典型特征，因为它涉及通过分泌装置。进一步的研究表明，在未暴露于LPS的活单核细胞中未检测到与质膜相关的TNF。然而，在LPS处理90分钟后，一小部分单核细胞表达了与膜相关的TNF，到24小时时，约50%的单核细胞质膜上显示有TNF。此外，我们的结果表明，与质膜相关的TNF并不代表释放后又结合回其自身受体的TNF。因此，我们的发现支持了TNF在LPS刺激的单核细胞中以表面跨膜蛋白形式存在的观点。
