摘要
目的研究同源异型盒基因B7(HOXB7)介导Wnt/β-Catenin信号通路在结直肠癌侵袭转移中的作用与机制。方法构建慢病毒转染重组体质粒HOXB7/psin,包装质粒PIK和阴性对照质粒psin,通过转染技术获得稳定过表达HOXB7的结直肠癌细胞株(SW480/HOXB7,293FT/HOXB7)及对照细胞株(SW480/vector,293FT/Vector)。采用实时荧光定量PCR(qRT-PCR)和Western blot法检测HOXB7 mRNA和蛋白的表达,采用免疫荧光染色技术评估HOXB7对结直肠癌细胞株β-Catenin的调控作用,采用qRT-PCR和Western blot法检测Wnt信号通路下游靶基因及蛋白的表达,采用小室试验评价各组细胞转移侵袭能力。结果 SW480/HOXB7,293FT/HOXB7细胞株的HOXB7 mRNA和蛋白表达水平较SW480/vector,293FT/Vector细胞株显著提高(P均<0.05)。HOXB7过表达后,细胞核中β-Catenin表达水平显著升高,并可见明显的入核现象,而在细胞浆中β-Catenin表达水平无明显变化。随着HOXB7的过表达,293FT、SW480细胞株中DKK1、cyclin D1、LEF1等Wnt信号通路下游基因及蛋白的表达均出现不同程度的上调现象。SW480/HOXB7细胞株穿过滤膜的细胞数显著高于SW480/vector细胞株(P<0.05)。结论 HOXB7介导Wnt/β-Catenin信号通路的激活能够通过调控下游基因,提高结直肠癌细胞侵袭能力,增加肿瘤转移的风险。
Objective To study the effect and mechanism of homeobox B7 （HOXB7） in invasion and metastasis of color-ectal cancer by mediating Wnt/β-Catenin pathway. Methods Constructing lentivirus-recombinant plasmid HOXB7/psin, packaging plasmid PIK and negative control plasmid psin, the colorectal cancer cell strains 293FT and SW480 of stably overexpressing HOXB7 （ SW480/HOXB7,293FT/HOXB7 ） and control cell strains （ SW480/vector, 293FT/Vector） were gotten by transfection technique. Real time fluorescence quantitative polymerase chain reaction （qRT-PCR） and Weatern blot method were used to respectively detect the expression levels of HOXB7 mRNA and protein. Immunofluoreseence stai-ning technique was used to evaluate the regulation effect of HOXB7 to β-Catenin of colorectal cancer cell line. qRT-PCR and Weatern blot method were used to detect the expression levels of downstream target gens and proteins of Wnt signaling pathway. The chamber test was used to evaluate the ability of cellular metastasis and invasion. Results The expression levels of HOXB7 mRNA and proteins in SW480/HOXB7 and 293FT/HOXB7 cell strains were significantly higher than those in SW480/vector, 293FT/Vector cell strains （all P 〈 0.05 ）. After overexpression of HOXB7, the expression level of β-Catenin in cell nucleus obviously increased with obvious nuclear import phenomenon, but its expression level, in the cytoplasm was unchanged. With the over expression of HOXB7, the expression levels of downstream gens and proteins of DKK1, cy-clinD1 and LEF1 of Wnt signaling pathway in 293FT and SW480 cell strains presented up-expressions of deferent degrees. The cells number passed through the filter membrane in SW480/HOXB7 cell strain was significantly higher than that in SW480/vector cell strain（P 〈 0.05 ）. Conclusion The activation of HOXB7-mediated Wnt/β-catenin signaling pathway could increase the invasive ability of colorectal cancer cells and the risk of tumor metastasis.
出处
《中国临床研究》
CAS
2016年第11期1453-1457,共5页
Chinese Journal of Clinical Research
基金
国家自然科学基金资助项目(81303135)
陕西省科技厅自然科学基金青年人才项目(2015JQ8288)
