骨桥蛋白在人胆囊腺癌中的表达

Expression of osteopontin in human gallbladder adenocarcinoma

目的应用抑制差减杂交联合cDNA芯片技术筛选人胆囊腺癌组织中差异表达基因,进一步探讨筛选出的骨桥蛋白在胆囊腺癌中的表达及作用。方法胆囊腺癌组织和非肿瘤组织作为对比材料进行抑制差减杂交,构建正、反向差减杂交文库,应用差减片断的PCR产物制备cDNA芯片,筛选胆囊腺癌组织中差异表达基因。采用实时荧光定量技术和免疫组化方法验证和分析筛选出的胆囊腺癌中高表达基因骨桥蛋白在胆囊腺癌和正常胆囊组织中的转录水平和蛋白水平的表达。结果成功构建了胆囊腺癌差减文库。cDNA芯片杂交结果显示骨桥蛋白在胆囊腺癌中高表达,经实时荧光定量技术证实。免疫组化结果表明骨桥蛋白在胆囊腺癌和癌旁胆囊组织中高表达,在正常胆囊组织中低表达(P<0.05),其表达部位主要在细胞核,在低分化胆囊腺癌中骨桥蛋白的表达有增高趋势。结论抑制差减杂交方法构建的胆囊腺癌差减杂交文库富含胆囊腺癌差异表达基因。胆囊腺癌中高表达基因骨桥蛋白可能与胆囊细胞的生长、转移和侵袭能力有关。

Objective To examine the differential gene expressions between paracancerous gallbladder tissue and the human gallbladder adenocarcinoma（HGA） by suppression subtractive hybridization combined cDNA microarry and identification of the expression of osteopontin in HGA.Methods The differential gene expression subtracted cDNA library of paracancerous gallbladder tissue and HGA was constructed by suppression subtractive hybridization technique.The cDNA fragments of differentially expressed genes were prepared substrate of cDNA microarray. Based on the data of cDNA microarrays,which indicated an over-expression of osteopontin in human gallbladder adenocarcinoma,Real-time quantitative PCR and immunohistochemistry was used to identify and analyze the Transcription and protein expression levels of osteopontin in human gallbladder adenocarcinoma and normal gallbladder tissues.Results The subtracted cDNA library of gallbladder adenocarcinoma was successfully constructed.Osteopontin significantly elevated expression in HGA.The result of real-time PCR and immunohistochemistry showed that osteopontin was highly expressed in the tumor tissues but not expressed in the normal gallbladder tissues.Osteopontin was mainly seen in the nuclease of tumor cells and increscent tendency of expression in poorly differentiated adenocarcinoma.Conclusion The subtracted cDNA library contains differentially-expressed genes in the tissues of gallbladder adenocarcinoma.Osteopontin is involved in the gallbladder adenocarcinoma malignant transformation and relates with the transferability and invasion ability of the gallbladder adenocarcinoma cells.