Differential expression of inducible NO synthase in two murine macrophage cell lines
Differential expression of inducible NO synthase in two murine macrophage cell lines
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DOI：
10.1046/j.1365-2567.1996.d01-728.x
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发表时间：
1996-10-01
期刊：
影响因子：
6.4
通讯作者：
Wietzerbin, J
中科院分区：
文献类型：
--
作者：
LePage, C;Sanceau, J;Wietzerbin, J
尽管原代巨噬细胞和大多数鼠巨噬细胞细胞例如RAW 264.7细胞通过产生大量亚硝酸盐（即由诱导型NO合酶（iNOS）产生的一氧化氮（NO）的氧化产物）来响应干扰素-γ（IFN-γ）和/或脂多糖（LPS），但其它细胞系例如P388.D1细胞不产生显著量。为了深入了解导致诱导iNOS活性的信号通路，我们比较了RAW 264.7和P388.D1细胞中的iNOS表达。我们发现IFN-γ以相似的亲和力与每种细胞系结合。此外，通过Southern印迹分析，未检测到iNOS神经元结构的差异。即使在IFN-γ和/或LPS刺激的P388.D1细胞的上清液中也没有发现显著的亚硝酸盐分泌。诱导iNOS mRNA表达。此外，IFN-γ诱导干扰素调节因子-1（IRF-1）基因，并激活该因子与iNOS基因中其靶序列的结合。这种结合最近被证明是必要的iNOS的表达。在P388.D1细胞中，我们不能检测到相应的iNOS蛋白。这些结果表明P388.D1细胞中的缺陷似乎限于控制iNOS蛋白合成的信号传导途径。这种缺陷也不影响总体IFN-γ生物学应答，而是IFN-γ和LPS共有的会聚转录后步骤。
Although primary macrophages and most murine macrophage cell lints such as RAW 264.7 cells respond to interferon-gamma (IFN-gamma) and/or lipopolysaccharide (LPS) by producing large amounts of nitrite, i.e. the oxidation product of nitric oxide (NO) produced by inducible NO synthase (iNOS), other cell lines like P388.D1 cells do not produce significant amounts. To gain insight into the signalling pathway that leads to the induction of iNOS activity, we compared iNOS expression in RAW 264.7 and P388.D1 cells. We showed that IFN-gamma binds to each cell line with a similar affinity. Furthermore, no differences in iNOS eerie structure were detectable by Southern blot analysis. Even though no significant nitrite secretion was found in the supernatant of P388.D1 cells stimulated with IFN-gamma and/or LPS. iNOS mRNA expression was induced. In addition, IFN-gamma induced the interferon regulatory factor-1 (IRF-1) gene and activated the binding of this factor to its target sequence in the iNOS gene. This binding was recently shown to be necessary for iNOS expression. However, in P388.D1 cells. we were unable to detect the corresponding iNOS protein. These results indicate a deficiency in P388.D1 cells which appears to be restricted to the signalling pathway controlling iNOS protein synthesis. This deficiency does nor affect the overall IFN-gamma biological response, but rather a convergent post-transcriptional step common to IFN-gamma and LPS.