Nitric oxide-induced apoptosis: p53-dependent and p53-independent signalling pathways
Nitric oxide-induced apoptosis: p53-dependent and p53-independent signalling pathways
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DOI：
10.1042/bj3190299
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发表时间：
1996-10-01
影响因子：
4.1
通讯作者：
Brune, B
中科院分区：
文献类型：
--
作者：
Messmer, UK;Brune, B
一氧化氮（NO）的产生在不同的实验系统中启动凋亡性细胞死亡。在RAW 264.7巨噬细胞中，典型凋亡标志物的出现与诱导型NO合酶诱导有关。从机制上讲，肿瘤抑制因子p53的积累先于凋亡DNA片段化。与使用S-硝基谷胱甘肽（GSNO），我们相关的剂量依赖性p53的上调，DNA片段后4小时和8小时，分别测量。我们的研究揭示了五种不同的NO供体在诱导凋亡和p53积累方面的效力之间的线性相关性。此外，我们探讨了NO诱导的细胞凋亡后，RAW 264.7巨噬细胞与质粒编码p53反义RNA稳定转染。响应于GSNO而下调p53水平的克隆表现出DNA片段化的显著减少。响应于脂多糖和干扰素-γ的诱导型NO合酶的表达在24小时内引起RAW 264.7巨噬细胞和新霉素载体对照中的凋亡。相反，表达p53反义RNA的克隆表现出对内源性NO的高度抗性，尽管诱导型NO合酶诱导伴随亚硝酸盐产生保持不变。对于RAW 264.7巨噬细胞，我们的研究结果建立了肿瘤抑制因子p53在NO诱导的凋亡性细胞死亡过程中的功能作用。然而，p53反义实验和p53阴性细胞系U937的使用证实了在NO介导的凋亡过程中p53独立的信号传导通路起作用。
Nitric oxide (NO) generation initiates apoptotic cell death in different experimental systems. In RAW 264.7 macrophages the appearance of typical apoptotic markers is linked to inducible NO synthase induction, Mechanistically, accumulation of tumour suppressor p53 precedes apoptotic DNA fragmentation. With the use of S-nitroglutathione (GSNO) we correlated a dose-dependent p53 up-regulation to DNA fragmentation measured after 4 h and 8 h, respectively. Our studies revealed a linear correlation between the potency of five different NO donors with respect to apoptosis induction and p53 accumulation. Furthermore, we probed for NO-induced apoptosis after stable transfection of RAW 264.7 macrophages with plasmids encoding p53 antisense RNA. Clones with down-regulated p53 levels in response to GSNO exhibited a marked reduction in DNA fragmentation, Expression of the inducible NO synthase in response to lipopolysaccharide and interferon-gamma caused apoptosis in RAW 264.7 macrophages and neomycin-vector controls within 24 h, In contrast, p53 antisense RNA-expressing clones appeared highly resistant towards endogenous NO, although inducible NO synthase induction with concomitant nitrite production remained unchanged. For RAW 264.7 macrophages our results established a functional role of the tumour suppressor p53 during NO-induced apoptotic cell death. However, p53 antisense experiments and the use of the p53-negative cell line U937 substantiated p53-independent signalling pathways operative during NO-mediated apoptosis.