青年和老年人结肠上皮的比较蛋白质组学研究  

Comparative Proteomics Analysis of Normal Colonic Epithelium in Young and Old People

作　　者：郑杰[1] 朱果[1] 陈主初[1] 李明[1] 程爱兰[1] 阮林[1] 刘迎福[1] 袁伟键[2] 张鹏飞[1] 肖志强[1] 

机构地区：[1]中南大学湘雅医院卫生部肿瘤蛋白质组学重点实验室,长沙410078 [2]中南大学湘雅医院消化科,长沙410008

出　　处：《生物化学与生物物理进展》2007年第1期63-72,共10页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金资助项目(30240056;30370642).~~

摘　　要：衰老的大肠上皮不仅多种生理功能下降,而且对大肠癌在内的多种衰老相关的肠道疾病易感性显著增加,但结肠上皮衰老及衰老的结肠上皮对癌症易感的分子机制仍然不清楚.为此,应用双向凝胶电泳(2-DE)技术分离青年人及老年人的正常结肠上皮的总蛋白质,图像分析识别差异表达的蛋白质点,基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)对差异表达的蛋白质点进行鉴定,免疫组化和实时定量(real-timequantitative)PCR检测部分差异蛋白,在青年人和老年人结肠上皮中的表达水平.得到了分辨率较高、重复性较好的青年人和老年人结肠上皮的2-DE图谱,质谱分析鉴定了17个结肠上皮衰老相关的蛋白质,免疫组化和real-timequantitativeRT-PCR证实了部分差异蛋白质的表达水平.研究结果提示,线粒体功能受伤、抗氧化能力下降是结肠上皮衰老的重要原因,4个差异蛋白质即guaninenucleotide-bindingproteinbetasubunit-likeprotein(Rack1)、stress-70protein、40SribosomalproteinSA和chlorideintracellularchannelprotein1可能与衰老的结肠上皮对癌症易感有关.The aging process of human colonic epithelium involves a slow decline in physiological vigor and an increasing susceptibility to age-related diseases, especially, colon cancer, but the molecular mechanisms of the aging and susceptibility of aged colonic epithelium to carcinogenesis is still unclear. Identification of aging related proteins in colonic epithelium will help to reveal the molecular mechanisms of colonic epithelial aging and ageelated colonic diseases. Therefore, the total proteins of human normal colonic epithelial tissues from 10 young and 10 old men were separated by two-dimensional gel electrophoresis（2-DE）, respectively. PDQuest software was applied to analyze 2-DE images, the differentially expressed protein spots of colonic epithelium between young and old groups were identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF-MS）, and the expression levels of partial identified proteins were determined by real-time quantitative PCR and immunohistochemistry. Well-resolved, reproducible 2-DE maps of human colonic epithelial tissues from young and old men were established, 17 aging related proteins were identified by MALDI-TOF-MS, and the differential expression levels of partial identified proteins were confirmed by real-time quantitative PCR and immunohistochemistry. The results indicate that injury of mitochondrial function and decline of antioxidant capability are important reasons for the aging of human colonic epithelium, and four differential proteins （guanine nucleotide-binding protein beta subunit-like protein, stress-70 protein, 40 S ribosomal protein SA and chloride intracellular channel protein 1） may be involved in susceptibility of aged colonic epithelium to carcinogenesis.

关 键 词：结肠上皮 衰老 蛋白质组 肿瘤易感性 双向凝胶电泳 质谱 免疫印迹 实时定量PCR 

分 类 号：R735.35[医药卫生—肿瘤]