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结直肠癌奥沙利铂耐药相关蛋白的筛选与鉴定
         
Screening and identifying oxaliplatin-resistance-associated proteins in colorectal cancer cell lines

摘    要
目的:采用蛋白质组学技术筛选结直肠癌(colorectal cancer,CRC)中对化疗药物奥沙利铂(oxaliplatin,L-OHP)耐药的相关蛋白,以期为CRC的个体化治疗提供生物标志物。方法:建立L-OHP耐药细胞株HT-29/L-OHP,提取细胞总蛋白,采用二维凝胶电泳(two-dimensional gel electrophoresis,2-DE)和基质辅助激光解吸电离-飞行时间质谱(matrix assisted laser desorption-ionization time-of-flight tandem mass spectrometry,MALDITOF-MS)筛选并鉴定与亲本HT-29细胞差异表达的蛋白,并应用蛋白质印迹法对鉴定获得的部分蛋白进行验证。结果:建立CRC亲本细胞株HT-29和L-OHP耐药细胞株HT-29/L-OHP蛋白的2-DE图谱,共获得表达差异2倍以上的蛋白质点38个,经MALDI-TOF-MS分析,有37个得到鉴定,与亲本细胞相比,表达上调的有17个,表达下调的20个;蛋白质印迹法检测结果显示,多聚胞嘧啶结合蛋白1[poly (C)-binding protein-1,PCBP1]和TUBB2A(tubulin beta-2A)蛋白在HT-29/L-OHP细胞中表达上调,膜联蛋白A3(annexin A3,ANXA3)和磷酸化应激诱导蛋白(stress-induced-phosphoprotein 1,STIP1)表达下调,与MALDI-TOF-MS的实验结果相一致。结论:筛选获得37个与CRC对L-OHP耐药的相关蛋白,为进一步研究CRC对L-OHP耐药机制提供了有力的实验依据。
标    签 结直肠肿瘤   抗药性   肿瘤   电泳   凝胶   双向   质谱分析法   奥沙利铂   HT-29细胞   Colorectal neoplasms   Drug resistance   neoplasm   Electrophoresis   gel   two-dimensional   Mass spectrometry   Oxaliplatin   HT-29 cells  
 
Abstract
Objective:To screen and identify oxaliplatin-resistance-associated proteins in CRC (colorectalcancer) cell lines using proteomics technologies in order to find new biomarkers for individual therapyof CRC. Methods:Oxaliplatin-resistant human CRC cell line HT-29/L-OHP (oxaliplatin) was established.The total proteins in HT-29 and HT-29/L-OHP cells were extracted. The differentially expressed proteinsbetween HT-29 and HT-29/L-OHP cells were screened and identified using 2-DE (two-dimensional gelelectrophoresis) and MALDI-TOF-MS (matrix assisted laser desorption-ionization time-of-flight tandemmass spectrometry). Some proteins obtained were validated by Western blotting. Results:The 2-DE mapsof total proteins in HT-29 and HT-29/L-OHP cells were established. Of the 38 protein spots identified asdifferentially expressed proteins (over two-fold,P>0.05) between HT-29 and HT-29/L-OHP cells, 37protein spots were positively identified by MALDI-TOF-MS (17 proteins were up-regulated and 20 proteinswere down-regulated as compared with the parental HT-29 cells). The result of Western blotting showedthat the PCBP1[poly (C)-binding protein-1] and TUBB2A (tubulin beta 2A) proteins were up-regulatedwhile ANXA3 (annexin A3) and STIP1 (stress-induced-phosphoprotein 1) proteins were down-regulated inHT-29/L-OHP cells. The result of Western blotting was consistent with that of proteomics. Conclusion:Therewere 37 oxaliplatin-resistance-associated proteins in CRC identified in this study which may provide usefulevidence in further research on mechanism of oxaliplatin-resistance in CRC.

中图分类号 R735.34   DOI 10.3781/j.issn.1000-7431.2013.03.003

 
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所属栏目 基础研究

基金项目 江苏省自然科学基金资助项目(编号:Bk2008192);江苏省卫生厅面上项目(编号:H200652)

收稿日期 2012/12/11

修改稿日期 2013/1/30

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引用该论文: ZONG Ming-zhu,FENG Wan-ting,DU Nan,YE Li-lin,TAO Shan-dong,FU Xian-hua,HE Jing-dong,ZHOU Jian-wei. Screening and identifying oxaliplatin-resistance-associated proteins in colorectal cancer cell lines[J]. Tumor, 2013, 33(3): 223~228
宗明珠,冯婉婷,杜楠,叶丽霖,陶善东,付宪华,何敬东,周建伟. 结直肠癌奥沙利铂耐药相关蛋白的筛选与鉴定[J]. 肿瘤, 2013, 33(3): 223~228


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参考文献
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【3】WAGGONER S A, JOHANNES G J, LIEBHABER S A.Depletion of the poly(C)-binding proteinsalphaCP1 and alphaCP2 from K562 cells leads top53-independent induction of cyclin-dependentkinase inhibitor (CDKN1A) and G1 arrest[J]. J BiolChem, 2009, 284(14):9039-9049.
 
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