应用质谱技术检测牙龈炎治疗前后唾液及龈沟液多肽组的变化

Pre-and post-treatment changes of gingvitis-associated peptidomic profiles in the saliva and gingival crevicular fluid by mass spectrometry

目的通过基质辅助激光解吸/电离飞行时间质谱(matrix-assisted laser desorption-ionization time-of-flight mass spectrometry,MALDI-TOF MS)探索菌斑性龈炎患者治疗前后唾液和龈沟液多肽组的特征性变化,为多肽标志物应用于疾病监测奠定基础。方法从北京大学口腔医学院·口腔医院预防科2017年1至9月就诊患者中选择符合标准的17例菌斑性龈炎患者(男性6例,女性11例,年龄24~62岁),分别于基线、洁治后1周记录受试者的探诊出血、出血指数,收集唾液、龈沟液样本,处理后利用MALDI-TOF MS检测样本中多肽表达水平差异。纳米液相色谱-电喷雾-串联质谱技术(nano-liquid chromatography-electrospray ionization-tandem mass spectrometry,nano-LC/ESI-MS/MS)用于识别差异多肽的蛋白质来源。结果17例菌斑性龈炎受试者治疗前后的唾液样本经比较筛选出4个具有统计学意义的差异多肽峰(P<0.05),这4个差异多肽峰在治疗后均呈下降趋势,质荷比分别为1030.6、1043.4、1053.4和1064.6;治疗前后的龈沟液样本经比较筛选出5个具有统计学意义的差异多肽峰(P<0.05),其中多肽峰1055.5和1168.3在治疗后强度显著下降(P<0.05),而3363.7、3480.9和3489.5则呈显著上升趋势(P<0.05)。唾液、龈沟液的个别差异多肽峰之间、差异多肽峰强度与探诊出血、出血指数之间呈正相关关系,利用这些差异多肽峰进行的主成分分析显示治疗前与治疗后个体的分布集中趋势不同,龈沟液差异多肽峰代表的个体分离趋势略明显。经nano-LC/ESI-MS/MS鉴定,龈沟液中多肽峰1055.5来源于血清白蛋白,唾液中1064.6来源于补体C3。结论唾液和龈沟液中存在与牙龈炎治疗相关的差异多肽峰,可成为潜在的生物学标志物。

Objective To explore peptidomic changes of peptides in saliva and gingival crevicular fluid(GCF)before and after treatment of gingivitis.Methods From January 2017 to September 2017,seventeen participants at the age of 24-62(6 males and 11 females)at Department of Preventive Dentistry,Peking University School and Hospital of Stomatology with gingivitis were recruited in the present study.Their clinical parameters were measured and recorded.Saliva and GCF samples were collected from each of the participants at the baseline and 7 days after ultrasonic supragingival scaling,respectively.Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry(MALDI-TOF MS)was employed to detect the changes of peptidomic profiles,while ano-liquid chromatography-electrospray ionization-tandem mass spectrometry(nano-LC/ESI-MS/MS)was performed to identify the possible proteins from which the peptides might derive.Results Initially,four peptide peaks[mass-to-charge ratio(m/z)values:1030.6,1043.4,1053.4 and 1064.6]were screened out exhibiting a decreasing trend after treatment(P<0.05).Besides,five peptide peaks from gingival crevicular fluid(P<0.05)exhibited differential expression,among which 1055.5 and 1168.3 demonstrating a decrease after treatment,while 3363.7,3480.9 and 3489.5 increased overtime.Certain positive correlations were detected between some peptides and clinical parameters.Principle component analysis using the above mentioned peptide peaks showed a distinct distribution before and after treatment and peptides from GCF showed a slightly better capacity to discriminate patients before and after treatment.The peptides with m/z values of 1055.5 in GCF and 1064.6 in saliva were identified to be segments of serum albumin and complement C3,respectively.Conclusions Several differentially expressed peptides were detected in saliva and GCF by MALDI-TOF MS,exhibiting the potentiality to act as biomarkers in gingivitis patients.