低浓度MNNG诱发非DNA损伤依赖的细胞信号通路

目的：研究基因组DNA损伤与细胞信号通路激活的关系以及低浓度N-甲基-N'-硝基-N-亚硝基胍(MNNG)对细胞膜表面受体的影响。方珐：采用EUSA法来检测蛋白激酶A(PKA)的活性，不连续梯度密度离心法完成细胞脱核，以及免疫荧光法和激光共聚焦显微镜观察细胞表面受体聚集情况。结果：0．2μmol／L MNNG能在脱核细胞中引起PKA活性升高2．3倍，并可在5min内引起vero细胞表面生长因子受体和肿瘤坏死因子α受体的聚集；在脱核细胞中，表面受体聚集现象与在完整细胞中一样。结论：低浓度MNNG对PKA的激活和诱导细胞表面受体聚集均不依赖于基因组DNA的损伤，表明细胞信号转导通路的起源可能位于细胞膜或者细胞质中。

Activation of nucleus-independent signals triggered by N-methyl-N''''-nitro-N-nitrosoguanidine

OBJECTIVE: To study the effect of MNNG on inducement of non-targeted mutation and activation of several cellular signal transduction pathways, and to determine whether the activation of these signaling pathways was dependent on the DNA-damage. METHODS: Vero cells were enucleated by discontinuous density centrifugation. The PKA activities were measured by enzyme-linked immunosorbent assay. The status of cell membrane receptors was studied with immunofluorescent staining and confocal microscopy. RESULT: In enucleated cytoplasts, MNNG-treatment increased PKA activity for about 2.3-fold in accordance with the 2.7-fold up-regulation of PKA activity in whole vero cells exposed to MNNG. The clustering of cell surface receptors of epidermal growth factor and tumor necrosis factor alpha was also observed in cells exposed to MNNG; this phenomenon was also found in enucleated cells. CONCLUSION: The results indicate that the initiation of signal cascades induced by low concentration of MNNG might be associated with its interaction with cell surface receptors and/or direct activation of related signal proteins but not its DNA damage.