摘要
目的:构建裸鼠人肝癌种植瘤模型,观察沉默特异性蛋白3(specificity protein 3,Sp3)对裸鼠人肝癌种植瘤侵袭能力的影响,并探讨其机制.方法:将Sp3 shRNA转染人肝癌HepG2细胞,并设无关序列阴性组及空白组为对照,建立3组裸鼠人肝癌种植瘤模型.观察裸鼠肿瘤的成瘤率,瘤体进行HE染色观察其病理学改变,采用Real-time PCR及免疫组织化学检测瘤组织中Sp3、β-连接素(β-catenin)、E-钙黏附蛋白(E-cadherin)及基质金属蛋白酶9(matrix metalloproteinase 9,MMP-9)mRNA和蛋白的表达情况.结果:实验组的成瘤率明显低于其他两组(60%vs 100%,100%);HE观察实验组肿瘤细胞侵袭能力明显降低.荧光定量PCR检测结果显示,种植瘤中实验组的Sp3、β-catenin及MMP-9 mRNA相对表达量明显低于空白组及阴性组,差异有统计学意义(F=29.692,P=0.001;F=21.894,P=0.002;F=109.414,P<0.001);实验组瘤体中E-cadherin mRNA的相对表达量高于其他两组(F=66.983,P<0.001).免疫组织化学结果显示,实验组Sp3蛋白表达为30±5.69、β-catenin蛋白表达为28±5.13、MMP-9蛋白表达为97±10.41均低于空白组及阴性组;E-cadherin蛋白表达为132±4.36明显高于其他两组,差异有统计学意义(P=0.000).结论:Sp3可通过上调β-catenin及其下游基因MMP-9表达,同时下调E-cadherin的表达,增强人肝癌细胞的侵袭能力.
AIM： To observe the influence of specificity protein 3 （Sp3） silencing on invasion of HepG2 cells in a xenogeneic graft mouse model, and to explore the possible mechanisms involved. METHODS： A shRNA-Sp3 or non-relevantshRNA was transfected into HepG2 cells using a lentiviral vector. Non-transfected HepG2 cells were used as controls. The three groups of cells were injected into nude mice. Tumor formation rate was determined, and the pathological mor- phology of cells in three groups was observed. The mRNA and protein expression of Sp3, β-catenin, E-cadherin and matrix metallopepti- dase 9 （MMP-9） was detected by real-time PCR and immunohistochemistry. RESULTS： The tumor formation rate was low- er in the shRNA-Sp3 group than in the non-rel- evant group and control group （60% vs 100%, 100%）. The ability of invasion was weaker in the shRNA-Sp3 group. The expression of Sp3, β-catenin and MMP-9 mRNAs in the shRNA- Sp3 group was significantly lower than that in the non-relevant group and control group （F = 29.692, 21.894, 109.414; P = 0.001, 0.002, 〈 0.001）. The expression of E-cadherin mRNA was significantly higher in the shRNA-Sp3 group than in the other two groups （F = 66.983, P 〈 0.001）. The expression levels of Sp3 （30 ± 5.69）, β-catenin （28 ± 5.13） and MMP-9 proteins （97 ± 10.41） were significantly lower than those in the non-relevant group and control group （P = 0.000 for all）. The expression of E-cadherin pro- tein （132 ± 4.36） was significantly higher in the shRNA-Sp3 group than in the other two groups （P = 0.000）. CONCLUSION： Sp3 may influence the inva- sion of HepG2 cells in vivo by up-regulating the expression of β-catenin and MMP-9 and down- regulating the expression of E-cadherin.
出处
《世界华人消化杂志》
CAS
北大核心
2014年第6期813-818,共6页
World Chinese Journal of Digestology
基金
广西科学研究与技术开发计划基金资助项目
No.桂科攻1298003-2-5
广西新世纪十百千人才工程专项基金资助项目
No.2007214
广西壮族自治区卫生厅自筹经费科研课题基金资助项目
No.Z2012052~~
