辛伐他汀对K562细胞Ras-MAPK信号通路的体内外研究

The investigation about Ras-MAPK signal pathway in K562 cells induced with simvastatin in vitro and in vivo

目的通过体内、外实验研究辛伐他汀对K562细胞Ras-MAPK信号通路的影响,探讨辛伐他汀诱导K562细胞凋亡的可能机制。方法体外培养CML细胞株K562细胞,做以下实验:①用流式细胞术(FCM)检测辛伐他汀处理K562细胞后其凋亡率的变化,②18只Balb/c-nu/nu裸小鼠皮下接种K562细胞,构建裸鼠的K562细胞移植瘤模型,③TUNEL法检测辛伐他汀诱导裸鼠体内K562细胞早期凋亡的变化,④RT-PCR检测裸鼠体内外K562细胞N-ras、c-Raf-1、ERK1mRNA水平的变化。结果辛伐他汀在体外能明显诱导K562细胞凋亡,Ras-MAPK通路大多数基因出现差异表达。不同剂量的辛伐他汀能够诱导裸鼠体内K562细胞凋亡,并随剂量的增加凋亡率逐渐增高(P<0.01);辛伐他汀能够引起N-ras、c-Raf-1、ERK1mRNA水平的明显差异表达(P<0.01)。结论辛伐他汀在体内外均能够引起参与K562细胞凋亡的Ras分子和Ras分子下游分子mRNA水平的变化,从一定角度说明辛伐他汀能依赖Ras-MAPK信号通路诱导K562细胞凋亡。

Aim To investigate the effect of simvastatin on Ras-MAPK signal pathway of K562 cells in vitro and in vivo, and to approach the potential mechanism of simvastatin-induced apoptosis of K562 cells. Methods K562 cells were cultured in vitro for experiments： （1） flow The apoptosis rate of K562 cells was measured by cytometry （FCM）. （2） The CML animal models were constructed by subcutaneous inoculation of K562 cells to eighteen Balb/c-nu/nu nude mice. （3）The early apoptosis changes of K562 cells in nude mice were tested by TUNEL. （4） The mRNA expression of N-ras,c-Raf-1, ERK1 in RT-PCR. Results The vitro and in vivo was tested by apoptosis of K562 cells was induced significantly by simvastatin in vitro. Most genes of Ras-MAPK signal pathway were expressed differentially. K562 cells in nude mice were induced to apoptosis by different doses of simvastatin and the ap- optotic rate increased with dose accumulation of simv- astatin（P 〈 0. 01 ）. The markedly differential expres- sion of N-ras c-Raf-1 and ERK1 mRNA involved in Ras-MAPK pathway in K562 cells was observed at dif- ferent doses of simvastatin （P 〈 0. 01 ）. Conclusion The differential expression of Ras and its downstream molecules could be induced by simvastatin in vitro and in vivo at mRNA level, which to some degree, illustrated that simvastatin could induce the apoptosis of K562 cells in a Ras-MAPK signal pathway-dependent manner.