Li Zeng-gang, Ji Hong, Fu Wei-neng, Zhao Yan-yan, Jin Chun-lian, Ji Shi-jun, Sun Kai-lai
Department of Medical Genetics, China Medical University, Shenyang, Liaoning, 110001 PR China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2007 Feb;24(1):52-8.
To explore the etiology of idiopathic talipes equinovarus (ITEV) in all-trans retinoic acid (ATRA) induced clubfoot-like deformity in rat fetuses with two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS).
Clubfoot-like deformity model in rat fetuses was induced with ATRA (135 mg/kg) in gestation day (GD10) pregnant Wistar rats. 2-DE was applied to separate the total proteins of ankle joint tissue, ankle joint bone and spinal cord of the animal models. The Coomassie Brilliant Blue staining gels were analyzed by 2-DE software PDQuest 7.1.0. Selected differential protein spots were identified with peptide mass fingerprinting based on matrix-assisted laser adsorption/ionization time-of-flight mass spectrometry and database searching. xiap, tnnt1 and col2 alpha 1, three genes of the differential proteins, were identified furthermore. Apoptosis study was made in terminal deoxynucleotidyl transferase nick end labeling.
There were many differential expressed proteins in the clubfoot-like deformity model. Out of the differentially expressed proteins,16 protein spots were identified to be differentially expressed in the clubfoot-like deformity model with MS. Three of the 16 protein spots, xiap, tnnt1 and col2 alpha 1 were confirmed to be significantly down-regulated by the RT-PCR, and Xiap was further confirmed to be significantly down-regulated with immunohistochemistry. Another randomly selected gene, ngfr, did not express differently in ATRA-induced clubfoot-like deformity in rat fetuses. The rates of the apoptosis in the spinal, bone of the clubfoot-like deformity fetuses was 5.4 and 10 times of those of the normal fetuses respectively.
The results suggest that there are certain differently expressed proteins in ankle joint tissue, ankle joint bone and spinal cord of the ATRA-induced clubfoot-like deformity in rat fetuses, and Xiap, sTnT, and Col2 alpha 1 show a significant correlation with ITEV. Ngfr is not correlation with ITEV. Apoptosis plays a key role in the development of ITEV and related to the decreased expression of the Xiap.
采用双向凝胶电泳（2-DE）和质谱（MS）技术，探讨全反式维甲酸（ATRA）诱导胎鼠马蹄内翻足样畸形（ITEV）的病因。
在妊娠第10天（GD10）的怀孕Wistar大鼠中，用ATRA（135mg/kg）诱导胎鼠马蹄内翻足样畸形模型。应用2-DE分离动物模型踝关节组织、踝关节骨和脊髓的总蛋白。考马斯亮蓝染色凝胶用2-DE软件PDQuest 7.1.0进行分析。基于基质辅助激光吸附/电离飞行时间质谱和数据库搜索，用肽质量指纹图谱鉴定选定的差异蛋白点。进一步鉴定差异蛋白的三个基因，即xiap、tnnt1和col2α1。采用末端脱氧核苷酸转移酶缺口末端标记法进行凋亡研究。
马蹄内翻足样畸形模型中有许多差异表达蛋白。在差异表达蛋白中，16个蛋白点经MS鉴定在马蹄内翻足样畸形模型中差异表达。16个蛋白点中的3个，即xiap、tnnt1和col2α1，经RT-PCR证实显著下调，Xiap经免疫组化进一步证实显著下调。另一个随机选择的基因ngfr在ATRA诱导的胎鼠马蹄内翻足样畸形中无差异表达。马蹄内翻足样畸形胎儿脊髓、骨骼的凋亡率分别是正常胎儿的5.4倍和10倍。
结果表明，ATRA诱导的胎鼠马蹄内翻足样畸形的踝关节组织、踝关节骨和脊髓中存在一定的差异表达蛋白，Xiap、sTnT和Col2α1与ITEV显著相关。Ngfr与ITEV无关。凋亡在ITEV的发生发展中起关键作用，且与Xiap表达降低有关。
