摘要
目的观察异硫氰酸苯乙酯(PEITC)对结肠癌细胞SW480增殖侵袭的分子机制。方法体外培养结肠癌细胞系SW480,分别用10,30和50μmol/L PEITC作用24 h,MTT法检测细胞的增殖;伤口愈合实验和侵袭实验分别检测PEITC对SW480细胞迁移与侵袭的影响。荧光共振能量转移法检测基质金属蛋白酶9(MMP-9)的活性,RTPCR检测MMP-9 mRNA表达情况;Western blot检测PI3K和PTEN的表达和Akt、m TOR磷酸化,以及NF-κB核转位情况。荧光素酶报告基因分析NF-κB的活性。最后,建立裸鼠异种移植瘤动物模型,观察PEITC对异种移植瘤生长的抑制作用。结果 PEITC能在不影响细胞活性的条件下显著抑制SW480细胞侵袭和迁移(P<0.05),也能抑制MMP-9的酶活性以及mRNA表达(P<0.05)。同时PEITC能抑制PI3K的表达以及抑制Akt和m TOR磷酸化(P<0.05),上调PTEN表达。PEITC也能抑制NF-κB核转位,并能降低其活性(P<0.05)。此外,PEITC也能抑制裸鼠异种移植瘤的生长(P<0.05)。结论 PEITC是一种潜在的抗结肠癌细胞转移药物,它可能通过影响PI3K/Akt和NF-κB通路发挥作用。
Objective To investigate the anti-invasive mechanisms of phenethyl isothiocyanate (PEITC) in human colon cancer SW480 cells.Methods Human colon cancer SW480 cell line was cultured,and treated with 10,30,50 μmol/L PEITC for 24 h.Cell viability,cell migration and invasion were analyzed by MTT assay,scratch healing assay and transwell membrane assay.Activity of matrix metalloproteinase-9 (MMP-9) was analyzed by a fluorescence resonance energy transfer kits,and MMP-9 mRNA expression was detected by RT-PCR.Activation of phosphoinisitide-3-kinase (PI3K) /Akt/mammalian target of rapamycin (mTOR) and phosphatase and tensin homologue (PTEN),and nuclear translocation of nuclear factor kappa B (NF-κB) were examined by Western blot.NF-κB activity was detected by luciferase reporter assay.Furthermore,a mice model with xenograft tumors was constructed,and the effect of PEITC on tumor growth was analyzed.Results PEITC significantly reduced SW480 cells invasion and migration without affecting the viability of cells (P 〈 0.05).PEITC also markedly inhibited MMP-9 activity and expression at both protein and mRNA levels (P 〈 0.05).PEITC attenuated PI3 K and phosphorylation of AKT and mTOR,whereas PTEN increased.In the meantime,PEITC inhibited NF-κB transcriptional activity (P 〈 0.05).In addition,PEITC diminished NF-κB nuclear translocation.Furthermore,PEITC could significantly inhibit xenograft tumors growth in nude mice (P 〈 0.05).Conclusions PEITC is a potential anti-invasive agent by inhibiting MMP-9 to be involved in PI3K/AKT and NF-κB pathways.
出处
《基础医学与临床》
CSCD
北大核心
2014年第7期927-932,共6页
Basic and Clinical Medicine
基金
湖南省科技计划项目(2013SK3120)
湖南省高校科技创新团队资助项目[湘教通(2010)212号]