利用同位素标记相对和绝对定量技术筛查神经管畸形胚胎孕鼠血清差异蛋白质谱
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摘要
目的筛查神经管畸形相关血清标记物,为阐明神经管畸形的发病机制提供理论依据。方法应用维甲酸在大鼠神经管闭合的关键期(孕10 d,E10)给药,建立脊柱裂大鼠模型,选取致畸早期孕11、13 d (E11、E13)显性脊柱裂胚胎孕鼠和正常胚胎孕鼠血清各5例,应用同位素标记相对和绝对定量技术(iTRAQ)和液相色谱质谱鉴定技术分析2组蛋白质,定量信息以蛋白的丰度比差异倍数1.5倍以上(P <0.05)为差异蛋白质。结果质谱共得到谱图157 422张,鉴定出390种蛋白质,发现E11维甲酸致畸组与正常对照组差异蛋白40种,E13维甲酸致畸组与正常对照组差异蛋白26种。结论筛选出致畸早期大量神经管畸形和正常孕鼠血清差异蛋白,差异蛋白通过直接或间接相互作用,促进或抑制神经管畸形的发生与发展。
Objective To screen for a potential biomarker of neural tube defects and understand the pathogenesis of this defect. Methods The rat model of spina bifida aperta(SBA) was induced with all-trans retinoic acid at E10. Serum samples were collected from normal pregnant rats and those harboring SBA fetuses at E11 and E13(n = 5 each). The isobaric tag for relative and absolute quantitation(iTRAQ) proteomic approach was used to identify and quantify the differentially expressed proteins in the two groups. The quantitative protein ratios with P < 0.05 and fold-changes > 1.5 were considered significant in Mascot. Functional annotations of proteins were conducted. Results A total of 390 proteins corresponding to 157 422 tandem mass spectrometry spectra were identified. A total of 40 proteins at E11 and 26 proteins at E13 had significant differential expression between pregnant rats carrying SBA fetuses and normal controls.Conclusion Differential serum proteome profiles associated with neural tube defects at the early stage of malformation can be identified using the iTRAQ proteomic approach,and various proteins might be involved in abnormal neurological development.
Objective To screen for a potential biomarker of neural tube defects and understand the pathogenesis of this defect. Methods The rat model of spina bifida aperta(SBA) was induced with all-trans retinoic acid at E10. Serum samples were collected from normal pregnant rats and those harboring SBA fetuses at E11 and E13(n = 5 each). The isobaric tag for relative and absolute quantitation(iTRAQ) proteomic approach was used to identify and quantify the differentially expressed proteins in the two groups. The quantitative protein ratios with P < 0.05 and fold-changes > 1.5 were considered significant in Mascot. Functional annotations of proteins were conducted. Results A total of 390 proteins corresponding to 157 422 tandem mass spectrometry spectra were identified. A total of 40 proteins at E11 and 26 proteins at E13 had significant differential expression between pregnant rats carrying SBA fetuses and normal controls.Conclusion Differential serum proteome profiles associated with neural tube defects at the early stage of malformation can be identified using the iTRAQ proteomic approach,and various proteins might be involved in abnormal neurological development.
引文
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[4]CHOOLANI M,NARASIMHAN K,KOLLA V,et al.Proteomic technologies for prenatal diagnostics:advances and challenges ahead[J].Expert Rev Proteomics,2009,6(1):87-101.DOI:10.1586/14789450.6.1.87.
[5]HERR F,BAAL N,ZYGMUNT M.Studies of placental vasculogenesis:a way to understand pregnancy pathology?[J].Z Geburtshilfe Neonatol,2009,213(3):96-100.DOI:10.1055/s-0029-1224141.
[6]ASLAM B,BASIT M,NISAR MA,et al.Proteomics:technologies and their applications[J].J Chromatogr Sci,2017,55(2):182-196.DOI:10.1093/chromsci/bmw167.
[7]SHAN L,FAN Y,LI H,et al.Proteomic analysis of amniotic fluid of pregnant rats with spinabifida aperta[J].J Proteomics,2012,75(4):1181-1189.DOI:10.1016/j.jprot.2011.10.033.
[8]FAN Y,WANG L,ZHOU F,et al.Comparative proteomics of spinal cords of rat fetuses with spina bifida aperta[J].J Proteomics,2011,75(2):668-676.DOI:10.1016/j.jprot.2011.09.006.
[9]WONG YK,ZHANG J,HUA ZC,et al.Recent advances in quantitative and chemical proteomics for autophagy studies[J].Autophagy,2017,13(9):1472-1486.DOI:10.1080/15548627.2017.1313944.
[10]NAGALLA SR,CANICK JA,JACOB T,et al.Proteomic analysis of maternal serum in down syndrome:identification of novel protein biomarkers[J].J Proteome Res,2007,6(4):1245-1257.
[11]GRAVETT MG,NOVY MJ,ROSENFELD RG,et al.Diagnosis of intra-amniotic infection by proteomic profiling and identification of novel biomarkers[J].JAMA,2004,292(4):462-469.
[12]SONG HJ,ZHANG P,GUO XJ,et al.The proteomic analysis of human neonatal umbilical cord serum by mass spectrometry[J].Acta Pharmacol Sin,2009,30(11):1550-1558.DOI:10.1038/aps.2009.140.
[13]ABDI F,QUINN JF,JANKOVIC J,et al.Detection of biomarkers with a multiplex quantitative proteomic platform in cerebrospinal fluid of patients with neurodegenerative disorders[J].J Alzheimers Dis,2006,9(3):293-348.
[14]TOYO-OKA K,SHIONOYA A,GAMBELLO MJ,et al.14-3-3epsilon is important forneuronal migration by binding to NUDEL:a molecular explanation for Miller-Dieker syndrome[J].Nat Genet,2003,34(3):274-285.
[15]PERRI E,PARAKH S,ATKIN J.Protein disulphide isomerases:emerging roles of PDI and ERp57 in the nervous system and as therapeutic targets for ALS[J].Expert Opin Ther Targets,2017,21(1):37-49.
[16]SOARES MORETTI AI,MARTINS LAURINDO FR.Protein disulfide isomerases:redox connections in and out of the endoplasmic reticulum[J].Arch Biochem Biophys,2017,617:106-119.DOI:10.1016/j.abb.2016.11.007.