Antibody-Free Mass Spectrometry Identification of Vascular Integrity Markers in Major Trauma.
Antibody-Free Mass Spectrometry Identification of Vascular Integrity Markers in Major Trauma.
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DOI：
10.1089/neur.2021.0007
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发表时间：
2021
影响因子：
2.4
通讯作者：
Rodland K
中科院分区：
文献类型：
--
作者：
Hinson HE;Jacobs J;McWeeney S;Wachana A;Shi T;Martin K;Rodland K
抗体介导的蛋白质生物标志物检测策略很常见，但可能限制发现。我们假设使用无抗体蛋白质组学来检测遭受重大创伤的患者血浆中的蛋白质生物标志物是可行的。对前瞻性观察试验中遭受重大创伤的受试者子集进行了分析。根据患者提出的简明损伤严重程度评分 (AIS)，将患者分配到四组中的一组。使用灵敏的无抗体选择性反应监测 (SRM) 质谱 (MS)，掺入同位素标记的合成肽，对一组 10 个预期靶点进行靶向蛋白质定量。基于血浆免疫亲和力耗竭和 PRISM 分级分离，采用总体分层灵敏度分析方法进行肽检测和定量。分析包括 44 名患者，其中 82% 为男性，平均年龄为 50 (±19) 岁。一半患者患有孤立性头部损伤 (n = 22)，其余患者经历多处损伤或多发伤 (n = 14)、孤立性身体损伤 (n = 2) 或轻伤 (n = 6)。在未去除处理的血浆中检测并定量了来自 3 种蛋白质（血管粘附分子 1 [VCAM1]、细胞间粘附分子 1 [ICAM1] 和基质金属蛋白酶 9 [MMP9]）的肽。在耗尽的血浆中检测到了来自 2 种蛋白质（血管生成素 2 [Ang2] 和纤溶酶原激活剂抑制剂 - 1 [PAI1]）的肽并进行了定量，而 10 个预期靶标中的其余 5 个未检测到。与轻度损伤组 (4) 相比，VCAM1 (p = 0.02) 和 MMP9 (p = 0.03) 在重度损伤组 (1-3) 中显着上调，而其他组则没有。创伤性脑损伤患者（TBI；第 1 组和第 2 组）与无 TBI 患者（第 3 组和第 4 组）之间的蛋白质表达没有差异。我们在严重受伤的患者中检测到反映血脑屏障破坏的蛋白质非特异性上调，表明无标记 MS 技术是可行的并且可能提供信息。
Antibody mediated strategies for protein biomarker detection are common, but may limit discovery. We hypothesized that the use of antibody-free proteomics is feasible for detecting protein biomarkers in plasma of patients sustaining major trauma. A subset of subjects with major trauma from a prospective observational trial were analyzed. Patients were assigned to one of four groups based on their presenting Abbreviated Injury Severity Score (AIS). Sensitive, antibody-free selective reaction monitoring (SRM) mass spectrometry (MS), with spiked-in isotopically labeled synthetic peptides, was used for targeted protein quantification of a panel of 10 prospective targets. An overall tiered sensitivity analytical approach was used for peptide detection and quantification based upon plasma immunoaffinity depletion and PRISM fractionation. Forty-four patients were included in the analysis, of which 82% were men with a mean age of 50 (±19) years. Half had isolated head injury (n = 22), with the remaining patients experiencing multiple injuries or polytrauma (n = 14), isolated body injury (n = 2), or minor injury (n = 6). Peptides from 3 proteins (vascular adhesion molecule 1 [VCAM1], intercellular adhesion molecule 1 [ICAM1], and matrix metalloproteinase 9 [MMP9]) were detected and quantified in non-depleted processed plasma. Peptides from 2 proteins (angiopoietin 2 [Ang2] and plasminogen activator inhibitor-1 [PAI1]) were detected and quantification in depleted plasma, whereas the remaining 5 of the 10 prospective targets were undetected. VCAM1 (p = 0.02) and MMP9 (p = 0.03) were significantly upregulated in in the major trauma groups (1–3) versus mild injury group (4), whereas the others were not. There were no differences in protein expression between patients with traumatic brain injury (TBI; groups 1 and 2) versus those without TBI (groups 3 and 4). We detected non-specific upregulation of proteins reflecting blood–brain barrier breakdown in severely injured patients, indicating label-free MS techniques are feasible and may be informative.