基于iTRAQ技术的伴放线聚集杆菌细胞致死性扩张毒素诱导Jurkat细胞的凋亡研究

摘要

目的：探索伴放线聚集杆菌（Aggregatibacter actinomycetemcomitans， A．a）的一种毒力因子（cytolethal distending toxin， CDT）诱导人T淋巴细胞发生凋亡过程中表达改变的信号分子。方法培养Jurkat细胞，并进行分组：不做处理的正常细胞对照组、野生型A．a CDT刺激Jurkat细胞的野生组、突变型A．a CDT刺激Jurkat细胞的突变组，应用iTRAQ技术检测A．a CDT作用Jurkat细胞24 h过程中表达差异蛋白。结果通过筛选，我们得到20个凋亡相关的蛋白（变化倍数＞1．3），其中表达上调的有8个（ CASP8／CD3E／YY1／SH3BGRL3／P53／CASP3／ UBE2I／ TNFRSF10B ），表达下调的有12个（ F5／RPL18A／DAD1／MTCH2／HIST1H1E／CNBP／RBM25／SLC16A1／KDM2A／CD99／RAC2／TMX1）。结论本研究检测出了伴放线聚集杆菌细胞致死性膨胀毒素诱导人 T 淋巴细胞凋亡过程中表达差异的蛋白，并筛选了一条可能的信号通路 UBE2I／P53／TNFRSF10B／CASP8／CASP3，为下一步研究及临床治疗局限型侵袭性牙周炎提供思路。%Objective To explore the key signaling molecules when cytolethal distending toxin ( CDT) , a kind of Aggregatibacter ac⁃tinomycetemcomitans (A.a) induced apoptosis of T lymphocytes in vitro. Methods Jurkat cells were cultured and then divided into groups:the control group, the wild group (Jurkat cells were stimulated by wild A.a CDT), and the mutant group (Jurkat cells were stimulated by mutant A.a CDT). After human Jurkat cells were treated by A.a CDT for 24 hours, iTRAQ (Isobaric tags for relative and absolute quantitation) labeling technique was used to test differentially expressed apoptosis⁃relating proteins. Results 20 apoptosis⁃re⁃lated differentially expressed proteins ( fold change ratio>1.3) were identified, including 8 proteins ( CASP8/CD3E/YY1/SH3BGRL3/P53/CASP3/UBE2I/ TNFRSF10B ) which were upregulated and 12 proteins ( F5/RPL18A/DAD1/MTCH2/HIST1H1E/CNBP/RBM25/SLC16A1/KDM2A/CD99/RAC2/TMX1) which were downregulated. Conclusion This study elucidated the signaling mole⁃cules when A.a CDT induced apoptosis of human Jurkat cells at the molecular level, and one possible signaling pathway was selected ( UBE2I/P53/TNFRSF10B/CASP8/CASP3) , providing thoughts for the future studies and the treatment of localized aggressive peri⁃odontitis.