鞘氨醇激酶-1激活ERK通路介导人结肠癌细胞株LoVo侵袭与迁移的实验  

Sphingosine Kinase-1 Regulates LoVo Cell Invasion and Migration via Activation of ERK1/2 Pathway

作　　者：钟月圆[1] 刘诗权[1] 黄杰安[1] 覃蒙斌[1] 金卉[1] 

机构地区：[1]广西医科大学第一附属医院消化内科,南宁530021

出　　处：《肿瘤防治研究》2011年第8期861-865,共5页Cancer Research on Prevention and Treatment

基　　金：国家自然科学基金资助项目(30760275);广西科学基金资助项目(0832008)

摘　　要：目的研究Sphk1对人结肠癌LoVo细胞侵袭与迁移能力的影响并探讨其作用机制。方法将人结肠癌LoVo细胞分成Sphk1激活组,Sphk1抑制组,空白对照组。以Phorbol 12-myristate 13-ace-tate(PMA)为Sphk1激活剂(终浓度为100 nM),N,N-dimethyl-D-eryt-hro-sphingosine(DMS)为Sphk1抑制剂(终浓度为50μM),NaCl(终浓度为0.9%)为空白试剂处理LoVo细胞24 h后,用Transwell Boyden小室模型测定LoVo细胞的相对侵袭率与迁移率;用Western blot方法测定细胞Sphk1、ERK1/2与p-ERK1/2蛋白水平的变化;用ELISA方法检测细胞培养上清中MMP-2、MMP-9及uPA的蛋白含量;用半定量RT-PCR检测细胞中MMP-2、MMP-9和uPA的mRNA水平。结果 Sphk1激活剂可促进LoVo细胞侵袭与迁移,同时明显增强LoVo细胞中Sphk1、ERK1/2及p-ERK1/2的蛋白表达,并促进MMP-2、MMP-9及uPA的mRNA表达与蛋白分泌。Sphk1抑制剂则抑制LoVo细胞侵袭与迁移,同时抑制Sphk1、ERK1/2与p-ERK1/2的蛋白表达,并抑制MMP-2、MMP-9及uPA的mRNA表达与蛋白分泌。结论 Sphk1可促进人结肠癌细胞株LoVo细胞的侵袭与迁移,其机制可能与激活ERK1/2信号通路从而促进MMP-2、MMP-9及uPA mRNA表达与蛋白分泌有关。Objective To investigate the effect of Sphk1 on colon cancer cell invasion and migration. Methods Human colon cancer LoVo cells were divided into three group： LoVo cells were treated using 100 nM Phorbol 12-myristate 13-acetate（PMA） as the Sphk1 activation group,50 μM N,N-dimethyl-D-erythro-sphingosine（DMS） as suppression group,and 0.9%NaCl as control group.Cell invasiveness and migration were detected by Transwell boyden chamber model.Sphk1,ERK1/2,p-ERK1/2 protein expressions were detected by Western blot,MMP-2,MMP-9 and uPA protein levels in the culture medium were detected by enzyme-linked immunosorbent assay（ELISA）,and MMP-2,MMP-9 and uPA mRNA expressions in LoVo cells were detected by semi-quantitative reverse transcription-polymerase chain reaction.Results The Sphk1 activator induced the expression of Sphk1 and obviously enhanced LoVo cell invasion and migration capacity,accompanied with the up-regulating of ERK1/2,p-ERK1/2 protein expressions： moreover the protein in culture medium and the mRNA in cells levels of MMP-2,MMP-9 and uPA were elevated.On the contrary,the inhibitor obviously suppressed the protein expression of Sphk1 and cell invaseness and migration,associated with the suppressing of ERK1/2,p-ERK1/2 protein expressions;furthermore,the protein and the mRNA levels of MMP-2,MMP-9 and uPA were down-regulated.Conclusion Sphk1 was able to promote the invasion and migration of LoVo cells,through the activation of ERK1/2 signaling pathway,MMP-2,MMP-9 and uPA mRNA expression were up-regulated.

关 键 词：鞘氨醇激酶-1 人结肠癌细胞 侵袭 迁移 

分 类 号：R730.2[医药卫生—肿瘤] R735.35[医药卫生—临床医学]