背景:2型糖尿病(T2DM)使冠心病(CHD)的风险增加2-4倍,并与内皮功能障碍有关,血脂异常,胰岛素抵抗,和慢性高血糖症.这项调查的目的是评估,通过多标记质谱方法,循环蛋白作为糖尿病相关心血管损害进展的生物标志物的预测作用。
方法:该研究考虑了34名T2DM和CHD患者,31例T2DM和无CHD患者,和30名诊断为冠心病的无糖尿病患者。通过基于多重靶向液相色谱质谱(LC-MS)的测定法分析受试者的血浆样品,即多反应监测(MRM),允许同时检测来自感兴趣的蛋白质的肽。基因本体论(GO)分析用于识别生物过程中富集的GO术语,分子功能,或细胞组件类别。使用非参数多变量方法对患者样品进行分类,并评估分析的蛋白质组的相关性。
结果:在人血浆样品中成功定量了总共81种蛋白质。基因本体论分析评估了与血液微粒相关的术语,细胞外泌体和含胶原的细胞外基质。使用方差分析(ANOVA)对患者组之间的蛋白质组谱差异的初步评估鉴定出81种蛋白质中的13种具有显著差异。多变量分析,包括聚类分析和主成分分析,确定了13种蛋白质的相关分组。第一个主要簇包括载脂蛋白C-III,载脂蛋白C-II,载脂蛋白A-IV,视黄醇结合蛋白4、溶菌酶C和胱抑素C;第二种包括,尽管有分组,α2巨球蛋白,afamin,激肽原1,玻连蛋白,维生素K依赖性蛋白S,补体因子B和甘露聚糖结合凝集素丝氨酸蛋白酶2。使用标称逻辑回归获得的13种选定蛋白质的受试者工作特征(ROC)曲线表明三组受试者之间存在显着总体差异(p<0.001)。ROC曲线下面积(AUC)范围为0.91-0.97,敏感性和特异性为85%至100%。
结论:靶向质谱方法表明13种多种循环蛋白可能是2型糖尿病相关心血管损害进展的生物标志物,在灵敏度和特异性方面具有出色的分类结果。
Type 2 diabetes mellitus (T2DM) increases the risk of coronary heart disease (CHD) by 2-4 fold, and is associated with endothelial dysfunction, dyslipidaemia, insulin resistance, and chronic hyperglycaemia. The aim of this investigation was to assess, by a multimarker mass spectrometry approach, the predictive role of circulating proteins as biomarkers of cardiovascular damage progression associated with diabetes mellitus.
The study considered 34 patients with both T2DM and CHD, 31 patients with T2DM and without CHD, and 30 patients without diabetes with a diagnosis of CHD. Plasma samples of subjects were analysed through a multiplexed targeted liquid chromatography mass spectrometry (LC-MS)-based assay, namely Multiple Reaction Monitoring (MRM), allowing the simultaneous detection of peptides derived from a protein of interest. Gene Ontology (GO) Analysis was employed to identify enriched GO terms in the biological process, molecular function, or cellular component categories. Non-parametric multivariate methods were used to classify samples from patients and evaluate the relevance of the analysed proteins\' panel.
A total of 81 proteins were successfully quantified in the human plasma samples. Gene Ontology analysis assessed terms related to blood microparticles, extracellular exosomes and collagen-containing extracellular matrix. Preliminary evaluation using analysis of variance (ANOVA) of the differences in the proteomic profile among patient groups identified 13 out of the 81 proteins as significantly different. Multivariate analysis, including cluster analysis and principal component analysis, identified relevant grouping of the 13 proteins. The first main cluster comprises apolipoprotein C-III, apolipoprotein C-II, apolipoprotein A-IV, retinol-binding protein 4, lysozyme C and cystatin-C; the second one includes, albeit with sub-grouping, alpha 2 macroglobulin, afamin, kininogen 1, vitronectin, vitamin K-dependent protein S, complement factor B and mannan-binding lectin serine protease 2. Receiver operating characteristic (ROC) curves obtained with the 13 selected proteins using a nominal logistic regression indicated a significant overall distinction (p < 0.001) among the three groups of subjects, with area under the ROC curve (AUC) ranging 0.91-0.97, and sensitivity and specificity ranging from 85 to 100%.
Targeted mass spectrometry approach indicated 13 multiple circulating proteins as possible biomarkers of cardiovascular damage progression associated with T2DM, with excellent classification results in terms of sensitivity and specificity.