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蛋白质组学筛查人胰腺癌候选相关免疫原性膜抗原的有效验证

Validation of candidate immunogenic membrane antigens of human pancreatic cancer screened by proteomics

摘要:

目的 对前期蛋白质组学筛查、鉴定的人胰腺癌相关免疫原性候选膜抗原进行有效验证.方法 前期实验中,将胰腺癌患者血清提取的IgG与人胰腺癌细胞株SW1990的膜蛋白进行免疫印迹杂交,共获得了9个阳性蛋白位点,现应用基质辅助激光解吸离子化飞行时间质谱分析与肽质指纹库对5号和6号位点进行鉴定.应用RT-PCR、real-time PCR验证二者在不同的人胰腺癌细胞株中的基因表达水平.分别提取人正常胰腺组织和胰腺癌组织的mRNA,应用real-time PCR检测prohibitin 2在人正常胰腺组织中和胰腺癌组织中的基因表达差异.结果 5号和6号阳性蛋白位点经质谱分析鉴定分别是prohibitin 2和prohibitin.RT-PCR和real-time PCR证明,候选膜抗原prohibitin 2和prohibitin在人胰腺癌细胞株SW1990、AsPc、P3的细胞中均有明确的基因表达,尤其在P3细胞中mRNA表达水平最高(t=7.442,P<0.01).并且prohibitin 2在人胰腺癌组织中的基因表达水平明显高于正常胰腺组织(t=0.893,P<0.01).结论 人胰腺癌细胞膜蛋白prohibitin 2和prohibitin在人胰腺癌细胞株SW1990、AsPc、P3中均有明确的基因表达,并且prohibitin 2在人胰腺癌组织中的基因表达水平明显高于人正常胰腺组织,二者可能是具有免疫原性的胰腺癌相关膜抗原.

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abstracts:

Objective To validate those obtained immunogenic membrane antigens candidate of human pancreatic cancer in the performed research.Methods In the pre-studies,serum IgG purified from clinically collected sera of pancreatic cancer patients underwent immunoblot with human pancreatic caner cell line SW1990 membrane protein,totally obtained 9 positive protein spots.Number 5 and 6 positive dots of immunoblot were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and peptide mass fingerprinting matching.The candidate membrane antigens were further validated in cell lines by RT-PCR and real-time PCR.RNA of human normal pancreatic tissue and pancreatic cancer tissue was extracted respectively,different gene expression level of prohibitin 2 was studied by real-time PCR.Results Number 5 and 6 positive dots were identified as prohibitin 2 and prohibitin.RT-PCR and real-time PCR all showed that gene of prohibitin 2 and prohibitin were expressed in the human pancreatic cancer cell line SW1990,AsPc and P3 respectively,especially in P3 cell with highest expression( t =7.442,P < 0.01 ).In addition,gene expression level of prohibitin 2 was significant higher in human pancreatic cancer than that of normal pancreatic tissue ( t =0.893,P < 0.01 ). Conclusions Prohibitin 2 and prohibitin are both differently expressed in the pancreatic cancer cell line SW1990,AsPc and P3.Prohibitin 2 is obvious highly expressed in human pancreatic cancer tissue. Prohibitin 2 and prohibitin might be the candidate immunogenic membrane antigens of human pancreatic cancer.

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作者: 王维斌 [1] 赵玉沛 [1] 廖泉 [1] 张太平 [1] 徐徕 [1] 吴元德 [2]
第一作者: 王维斌
作者单位: 100730,中国医学科学院北京协和医学脘北京协和医院基本外科 [1] 中国医学科学院基础研究所 [2]
期刊: 《中华外科杂志》2012年50卷3期 260-263页 MEDLINEISTICPKUCSCD
分类号: R735.9
栏目名称: 论著
DOI: 10.3760/cma.j.issn.0529-5815.2012.03.018
发布时间: 2012-04-20
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