Lehmann Michael H, Masanetz Sabine, Kramer Susanne, Erfle Volker
Institute of Molecular Virology, GSF-National Research Center for Environment and Health, Ingolstädter Landstr. 1, 85764 Neuherberg, Germany.
J Cell Sci. 2006 Nov 1;119(Pt 21):4520-30. doi: 10.1242/jcs.03231. Epub 2006 Oct 17.
HIV-associated dementia (HAD) correlates with infiltration of monocytes into the brain. The accessory HIV-1 negative factor (Nef) protein, which modulates several signaling pathways, is constitutively present in persistently infected astroctyes. We demonstrated that monocytes responded with chemotaxis when subjected to cell culture supernatants of nef-expressing astrocytic U251MG cells. Using a protein array, we identified CC chemokine ligand 2/monocyte chemotactic protein-1 (CCL2/MCP-1) as a potential chemotactic factor mediating this phenomenon. CCL2/MCP-1 upregulation by Nef was further confirmed by ribonuclease protection assay, RT-PCR and ELISA. By applying neutralizing antibodies against CCL2/MCP-1 and using CCR2-deficient monocytes, we confirmed CCL2/MCP-1 as the exclusive factor secreted by nef-expressing astrocytes capable of attracting monocytes. Additionally, we showed that Nef-induced CCL2/MCP-1 expression depends on the myristoylation moiety of Nef and requires functional calmodulin. In summary, we suggest that Nef-induced CCL2/MCP-1 expression in astrocytes contributes to infiltration of monocytes into the brain, and thereby to progression of HAD.
人类免疫缺陷病毒相关痴呆（HAD）与单核细胞浸润入脑有关。辅助性人类免疫缺陷病毒1型阴性因子（Nef）蛋白可调节多种信号通路，持续感染的星形胶质细胞中持续存在该蛋白。我们证明，当单核细胞接触表达nef的星形胶质细胞U251MG细胞的细胞培养上清液时会产生趋化反应。通过蛋白质阵列，我们确定CC趋化因子配体2/单核细胞趋化蛋白-1（CCL2/MCP-1）是介导这一现象的潜在趋化因子。通过核糖核酸酶保护试验、逆转录-聚合酶链反应（RT-PCR）和酶联免疫吸附测定（ELISA）进一步证实了Nef对CCL2/MCP-1的上调作用。通过应用抗CCL2/MCP-1中和抗体并使用CCR2缺陷型单核细胞，我们证实CCL2/MCP-1是表达nef的星形胶质细胞分泌的唯一能够吸引单核细胞的因子。此外，我们表明Nef诱导的CCL2/MCP-1表达依赖于Nef的肉豆蔻酰化部分，并且需要功能性钙调蛋白。总之，我们认为星形胶质细胞中Nef诱导的CCL2/MCP-1表达促进单核细胞浸润入脑，从而促进HAD的进展。
