Synergism of nitric oxide and iron in killing the transformed murine oligodendrocyte cell line N20.1
Synergism of nitric oxide and iron in killing the transformed murine oligodendrocyte cell line N20.1
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DOI：
10.1046/j.1471-4159.1999.0721050.x
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发表时间：
1999-03-01
影响因子：
4.7
通讯作者：
Benjamins, JA
中科院分区：
文献类型：
--
作者：
Boullerne, AI;Nedelkoska, L;Benjamins, JA
在多发性硬化和实验性变态反应性脑脊髓炎中，炎性病变中产生的一氧化氮（NO）可能在少突胶质细胞的破坏中起主要作用。转化的鼠少突胶质细胞系N20.1比原代少突胶质细胞对NO产生剂S-亚硝基-N-乙酰基-DL-青霉胺（SNAP）的杀伤更具抗性。这一观察结果促使研究导致N20.1细胞中细胞死亡的机制，并将SNAP与另一种NO供体硝普钠（SNP）进行比较。我们观察到N20.1细胞对SNP的敏感性是SNAP的30倍。特异性NO清除剂2-苯基-4，4，5，5-四甲基咪唑啉-1-氧基-3-氧化物（PT 10）仅针对SNP而不针对SNAP保护。然而，二硫苏糖醇保护SNAP和SNP，表明半胱氨酸的S-亚硝基化在两个NO供体的细胞毒性中起主要作用。我们没有观察到任何过氧亚硝酸盐的形成或与SNAP或SNP的Ca 2+浓度的增加，因此排除了它们参与导致N20.1细胞死亡的机制。基于两个观察结果，（a）当与铁氰化物或亚铁氰化物而不是氰化钠共孵育时，SNP的细胞毒性作用增强，以及（B）通过去铁胺（铁氰化物螯合剂）的保护，我们得出结论，与SNAP相比，N20.1细胞对SNP的更大敏感性是由于所释放的NO与SNP的亚铁氰化物部分之间的协同作用，氰化物对细胞毒性的影响很小。最后，SNP而不是SNAP诱导一些细胞凋亡，如通过DNA梯状化和半胱天冬酶-3抑制剂的保护所示。这些结果表明，低水平的NO与增加的铁含量组合导致凋亡性细胞死亡，而不是由SNAP产生的较高水平的NO所看到的坏死性细胞死亡。
Nitric oxide (NO) produced in inflammatory lesions may play a major role in the destruction of oligodendrocytes in multiple sclerosis and experimental allergic encephalomyelitis. The transformed murine oligodendroglial line N20.1 is much more resistant than primary oligodendrocytes to killing by the NO generator S-nitroso-N-acetyl-DL-penicillamine (SNAP). This observation prompted investigation of the mechanisms leading to cell death in the N20.1 cells and comparison of SNAP with another NO donor, sodium nitroprusside (SNP). We observed that N20.1 cells were 30 times more sensitive to SNP than to SNAP. The specific NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTlO) protected against SNP only, not against SNAP. However, dithiothreitol protected against both SNAP and SNP, indicating that S-nitrosylation of cysteines plays a major role in the cytotoxicity of both NO donors. We did not observe any formation of peroxynitrite or increase of Ca2+ concentration with either SNAP or SNP, thus excluding their involvement in the mechanisms leading to N20.1 cell death. Based on two observations, (a) potentiation of the cytotoxic effect of SNP when coincubated with ferricyanide or ferrocyanide, but not sodium cyanide, and (b) protection by deferoxamine, an iron cyanide chelator, we conclude that the greater sensitivity of N20.1 cells to SNP compared with SNAP is due to synergism between NO released and the iron cyanide portion of SNP, with the cyanide accounting for very little of the cytotoxicity. Finally, SNP but not SNAP induces some apoptosis, as shown by DNA laddering and protection by a caspase-3 inhibitor. These results suggest that low levels of NO in combination with increased iron content lead to apoptotic cell death rather than the necrotic cell death seen with higher levels of NO generated by SNAP.