肿瘤研究与临床
腫瘤研究與臨床
종류연구여림상
CANCER RESEARCH AND CLINIC
2013年
10期
649-653,658
,共6页
郭红霞%吴素慧%贾睿%尚海霞
郭紅霞%吳素慧%賈睿%尚海霞
곽홍하%오소혜%가예%상해하
巨噬细胞移动抑制因子%白细胞介素8%基质金属蛋白酶9%转染%子宫颈肿瘤
巨噬細胞移動抑製因子%白細胞介素8%基質金屬蛋白酶9%轉染%子宮頸腫瘤
거서세포이동억제인자%백세포개소8%기질금속단백매9%전염%자궁경종류
Macrophage migration inhibitory factor%Inrerleukin-8%Martix metalloproteinase-9%Transfection%Uterine cervical neoplasms
目的 研究过表达巨噬细胞移动抑制因子(MIF)对子宫颈癌SiHa细胞中白细胞介素8(IL-8)、基质金属蛋白酶9(MMP-9)表达及细胞侵袭迁移能力的影响.方法 化学合成MIF cDNA,设计含Xhol和BamHI酶切位点的引物序列,利用聚合酶链反应(PCR)方法 扩增MIF基因片段,构建人pEGFP-N1/MIF真核表达载体,采用脂质体转染法将载体转染到SiHa细胞中；酶联免疫吸附法(ELISA)检测各组上清液中MIF的表达,采用实时荧光定量PCR、免疫细胞化学法分别检测各组细胞中MIF、IL-8、MMP-9 mRNA和蛋白的表达,Boyden小室穿膜迁移实验检测过表达MIF对细胞迁移能力的影响.结果 转染pEGFP-N1/MIF组细胞上清液中MIF蛋白表达明显升高(F组别=8267.564,P＜ 0.01)；转染pEGFP-N1/MIF组细胞中MIF、IL-8、MMP-9 mRNA及蛋白也均过表达(F值分别为7019.619、2148.094、3303.540、1565.114、2807.300、523.466,均P＜0.01)；Pearson相关分析表明,MIF与IL-8、MMP-9之间mRNA和蛋白的表达均存在正相关(r值分别为0.865、0.895、0.934、0.908,均P＜0.01),转染pEGFP-Nl/MIF组细胞的迁移能力明显增强(F=3430.898,P＜ 0.01).结论 子宫颈癌SiHa细胞中过表达MIF能提高子宫颈癌细胞的侵袭、转移能力,其机制可能与IL-8、MMP-9表达上调相关.
目的 研究過錶達巨噬細胞移動抑製因子(MIF)對子宮頸癌SiHa細胞中白細胞介素8(IL-8)、基質金屬蛋白酶9(MMP-9)錶達及細胞侵襲遷移能力的影響.方法 化學閤成MIF cDNA,設計含Xhol和BamHI酶切位點的引物序列,利用聚閤酶鏈反應(PCR)方法 擴增MIF基因片段,構建人pEGFP-N1/MIF真覈錶達載體,採用脂質體轉染法將載體轉染到SiHa細胞中；酶聯免疫吸附法(ELISA)檢測各組上清液中MIF的錶達,採用實時熒光定量PCR、免疫細胞化學法分彆檢測各組細胞中MIF、IL-8、MMP-9 mRNA和蛋白的錶達,Boyden小室穿膜遷移實驗檢測過錶達MIF對細胞遷移能力的影響.結果 轉染pEGFP-N1/MIF組細胞上清液中MIF蛋白錶達明顯升高(F組彆=8267.564,P＜ 0.01)；轉染pEGFP-N1/MIF組細胞中MIF、IL-8、MMP-9 mRNA及蛋白也均過錶達(F值分彆為7019.619、2148.094、3303.540、1565.114、2807.300、523.466,均P＜0.01)；Pearson相關分析錶明,MIF與IL-8、MMP-9之間mRNA和蛋白的錶達均存在正相關(r值分彆為0.865、0.895、0.934、0.908,均P＜0.01),轉染pEGFP-Nl/MIF組細胞的遷移能力明顯增彊(F=3430.898,P＜ 0.01).結論 子宮頸癌SiHa細胞中過錶達MIF能提高子宮頸癌細胞的侵襲、轉移能力,其機製可能與IL-8、MMP-9錶達上調相關.
목적 연구과표체거서세포이동억제인자(MIF)대자궁경암SiHa세포중백세포개소8(IL-8)、기질금속단백매9(MMP-9)표체급세포침습천이능력적영향.방법 화학합성MIF cDNA,설계함Xhol화BamHI매절위점적인물서렬,이용취합매련반응(PCR)방법 확증MIF기인편단,구건인pEGFP-N1/MIF진핵표체재체,채용지질체전염법장재체전염도SiHa세포중；매련면역흡부법(ELISA)검측각조상청액중MIF적표체,채용실시형광정량PCR、면역세포화학법분별검측각조세포중MIF、IL-8、MMP-9 mRNA화단백적표체,Boyden소실천막천이실험검측과표체MIF대세포천이능력적영향.결과 전염pEGFP-N1/MIF조세포상청액중MIF단백표체명현승고(F조별=8267.564,P＜ 0.01)；전염pEGFP-N1/MIF조세포중MIF、IL-8、MMP-9 mRNA급단백야균과표체(F치분별위7019.619、2148.094、3303.540、1565.114、2807.300、523.466,균P＜0.01)；Pearson상관분석표명,MIF여IL-8、MMP-9지간mRNA화단백적표체균존재정상관(r치분별위0.865、0.895、0.934、0.908,균P＜0.01),전염pEGFP-Nl/MIF조세포적천이능력명현증강(F=3430.898,P＜ 0.01).결론 자궁경암SiHa세포중과표체MIF능제고자궁경암세포적침습、전이능력,기궤제가능여IL-8、MMP-9표체상조상관.
Objective To investigate the effects of macrophage migration inhibitory factor (MIF) overexpression on the expression of interleukin-8 (IL-8),martix metalloproteinase-9 (MMP-9) and invasion of human cervical cancer SiHa cells.Methods Chemical synthesis MIF eDNA gene,designed primer sequence including XhoI and BamHI enzyme sites,MIF gene was amplified by polymerase chain reaction (PCR),constructed eukaryotic expression vector pEGFP-N1/MIF and transfected into SiHa cells using Lipofectamine and won over-expression of MIF.The expression of MIF in supernatant fluid was detected by ELISA,the expression of MIF,IL-8,MMP-9 in both mRNA and protein levels were detected by real-time fluorescence quantitative-PCR and immunocytochemistry respectively.The effect of over-expressed MIF on migration was detected by Boyden small chamber.Results The expression of protein in supernatant fluid transfected with pEGFP-N1/MIF was significantly increased (Fgroup =8267.564,P ＜ 0.01),the expression of MIF,IL-8,MMP-9 in both mRNA and protein in SiHa cells transfected with pEGFP-N1/MIF were significantly increased (F values were 7019.619,2148.094,3303.540,1565.114,2807.300,523.466,P ＜ 0.01),and there was a positive correlation among MIF,IL-8,MMP-9 expression in both mRNA and protein (r values were 0.865,0.895,0.934,0.908,P ＜ 0.01).Invasion ability in SiHa cells transfected with pEGFP-N1/MIF was obviously increased (F=3430.898,P＜ 0.01).Conclusion The over-expression MIF gene in SiHa cells can promote cervical cancer cell invasion and metastasis of ability,which could be associated with the upregulation of IL-8 and MMP-9 expression.
