Liu Lijie, Guo Jing, Yuan Li, Cheng Mei, Cao Lihua, Shi Hui, Tong Hui, Wang Ning, De Wei
Department of Biochemistry, Nanjing Medical University, Nanjing, Jiangsu Province, China.
Dev Growth Differ. 2007 Oct;49(8):669-81. doi: 10.1111/j.1440-169X.2007.00961.x.
To identify proteins involved in pancreatic development, we used a differential proteomics approach by comparing pancreatic extracts from four biologically significant stages of development: embryonic day (E) 15.5, E18.5, postnatal (P) days 0 and adult. By two-dimensional gel electrophoresis (2D-E) and MALDI-TOF MS (Matrix Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry) following database searching and protein annotation, 15 proteins were identified as being differently expressed in the pancreas between the four phases. The expression pattern and the localization of alpha-fetoprotein (AFP), one of significant changed proteins observed, were further determined. Four isoforms of AFP (72 kDa, 60 kDa, 48 kDa and 37 kDa) were found by Western blotting in the pancreas tested, most of them showed a stronger signal in E18.5 followed by a steady decrease and only a 60-kDa isoform was detected in the adult pancreas. Immunolocalization for AFP revealed that a positive reactivity was detectable at E15.5 pancreas, became stronger in the cytoplasm of mesenchyme cells at E18.5, and declined after birth to a nearly undetectable level in adults. The dynamic expression of AFP in rat pancreas from different stages indicates that AFP might be involved in some aspects of pancreatic development.
为了鉴定参与胰腺发育的蛋白质,我们采用了差异蛋白质组学方法,比较了发育过程中四个具有生物学意义阶段的胰腺提取物:胚胎第(E)15.5天、E18.5天、出生后(P)第0天和成年期。通过二维凝胶电泳(2D-E)和基质辅助激光解吸/电离飞行时间质谱(MALDI-TOF MS),并经过数据库搜索和蛋白质注释,鉴定出15种蛋白质在这四个阶段的胰腺中表达存在差异。对观察到的显著变化的蛋白质之一甲胎蛋白(AFP)的表达模式和定位进行了进一步测定。通过蛋白质免疫印迹法在测试的胰腺中发现了AFP的四种同工型(72 kDa、60 kDa、48 kDa和37 kDa),其中大多数在E18.5时信号较强,随后稳定下降,在成年胰腺中仅检测到一种60 kDa的同工型。AFP的免疫定位显示,在E15.5胰腺中可检测到阳性反应,在E18.5时间充质细胞的细胞质中反应更强,出生后下降,在成年时降至几乎无法检测的水平。AFP在不同阶段大鼠胰腺中的动态表达表明,AFP可能参与了胰腺发育的某些方面。