体外培养对人皮肤角质形成细胞蛋白质组的影响  

Effects of cultivation in vitro on the proteome of human skin keratinocytes

作　　者：吴晓萍[1] 曾耀英[1] 聂燕芳[1] 刘钧澄[2] 

机构地区：[1]暨南大学组织移植与免疫实验中心,广州市510632 [2]中山大学附属第一医院小儿外科,广州市510080

出　　处：《实用医学杂志》2010年第10期1704-1706,共3页The Journal of Practical Medicine

基　　金：国家高技术研究发展计划项目863计划项目(编号:2007AA02Z110);国家自然科学基金项目(编号:30973671);广东省自然科学基金项目(编号:9151064001000031);温州市科技计划项目(编号:Y20090244)生物化学与分子生物学广东省重点学科资助项目

摘　　要：目的:探讨体外长期培养对人皮肤角质形成细胞蛋白质组的影响。方法:制备原代和第7代体外培养的人皮肤角质形成细胞的总蛋白样品进行双向凝胶电泳。采用PDQuest图像分析软件对获得的双向凝胶电泳图谱进行分析。串联飞行时间质谱鉴定选取的差异表达蛋白质点。半定量RT-PCR对差异表达蛋白质的mRNA水平进行确证。结果:获得了蛋白质分离效果较好的双向凝胶电泳图谱,串联飞行时间质谱鉴定选取的2个在第7代角质形成细胞中表达显著下调的蛋白质分别为PA-FABP(psoriasis-associated fatty acidbinding protein5)和Galectin-7,半定量RT-PCR确证第7代角质形成细胞中PA-FABP和Galectin-7的基因转录水平较原代角质形成细胞相应的基因转录水平显著降低。结论:体外长期培养人皮肤角质形成细胞可能影响一些与细胞的脂肪酸代谢及细胞凋亡相关蛋白的表达。Objective To investigate the effects of long-term cultivation in vitro on the proteome of keratinocytes. Methods The whole-cell proteins were prepared from the primary and the seventh-passage human skin keratinocytes and applied to two dimension gel electrophoresis （2-DE）. The 2-DE patterns were analyzed by PDQuest software. The selected differentially expressed protein spots were analyzed by tandem time-of-flight mass spectrometry. The mRNA levels of two differentially expressed proteins were further detected by semi-quantitative RT-PCR. Results The 2-DE patterns with high resolution were obtained. Two differentially expressed proteins down-regulated in the seventh-passage keratinocytes were identified as psoriasis associated fatty acid binding protein （PA-FABP） and galectin-7. The results of RT-PCR suggested that the transcription levels of both PA-FABP and galectin-7 were much lower in the seventh-passage keratinocytes than those in the primary keratinocytes. Conclusion Long-term cultivation of human skin keratinocytes in vitro might affect the expression of some proteins related to fatty acid metabolism and apoptosis.

关 键 词：电脉 凝胶 双向 人皮肤角质形成细胞 质谱 RT-PCR 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]