慢性乙型肝炎肝脏炎症分期与血清多肽分子的相关性分析

Correlation of Serum Peptide Markers with Liver Inflammation Stages in Patients with Chronic Hepatitis B

背景:准确诊断肝组织炎症程度对慢性乙型肝炎(CHB)的预后评估具有重要临床意义。肝活检具有较多缺陷,因此迫切需寻求非创伤性、准确可靠的血清诊断标记物。目的:应用血清多肽组学法筛选CHB患者肝脏炎症分期的多肽诊断标记物。方法:建立146例前瞻性队列,包括126例CHB患者和20名对照者。采用Scheuer病理学标准进行肝脏炎症分期;液相色谱法串联质谱法(LC-MS/MS)分离和筛选血清差异多肽峰;多反应监测(MRM)法分析血清差异多肽分子m/z 520.3离子对与炎症程度、病毒载量的关系。计算受试者工作特征曲线下面积(AUROC),评估差异多肽诊断炎症分期的敏感性和特异性。结果:LC-MS/MS法筛选出9个差异表达多肽峰。差异多肽分子m/z 520.3为二羟丙酮激酶(DAK)片段,其4个离子对(520.3/176.1、520.3/353.7、520.3/459.8和520.3/503.3)的SPAR值随着肝脏炎症程度加重而逐渐减少,但与病毒载量(HBeAg阳性和HBV-DNA)无关。多肽离子对520.3/176.1、520.3/503.3区分G2～G4期与G0～G1期的AUROC值分别为0.840、0.856;区分G3～G4期与G0～G2期的AUROC值分别为0.886、0.888。结论:血清DAK的多肽分子m/z 520.3能有效诊断CHB的肝脏炎症程度。

Background： Diagnosis for the stage of liver inflammation is crucial in the assessment of prognosis in patients with chronic hepatitis B （CHB）. Considering the limitation of liver biopsy, exploration of reliable non-invasive serum biomarkers for stage of liver inflammation is needed. Aims： To screen peptide markers for the stage of liver inflammation in CHB patients by serum peptidomics. Methods： A total of 146 subjects including 126 CHB patients and 20 healthy controls were enrolled in this prospective clinical queue. Liver inflammation grade was determined by Scheuer＇ s classification. Liquid chromatography combined with mass spectrometry （LC-MS/MS） were used to separate and identify differentially expressed peptide peaks. Multiple reaction monitoring （MRM） was used to quantify ion pairs derived from peptide m/z 520.3, and its correlation with different stages of liver inflammation and virus load was analyzed. Area under receiver operating characteristic curve （AUROC） was calculated to assess the sensitivity and specificity of peptide m/z 520.3 in the diagnosis of inflammation stage. Results： Nine differentially expressed peaks were found through LC-MS/MS analysis. SPAR of 4 ion pairs （520.3/176. 1, 520.3/353.7, 520.3/459.8 and 520.3/503.3） from peak with m/z of 520.3, which was identified to be a fragment of dihydroxyaeetone kinase （ DAK）, significantly decreased from non-inflammation （GO） to severe inflammation stage （G4）, but was not correlated with virus load （HBeAg positivity and HBV-DNA）. AUROC of model 520.3/176.1 and 520.3/503.3 were 0. 840, 0.856 （ G2-G4 versus GO-GI ） and 0.886, 0.888 （ G3-G4 versus GO-G2 ）, respectively. Conclusions： A peptide fragment of DAK （ m/z of 520.3 ） can be served as a promising new candidate biomarker, which can be used to discriminate liver inflammation stages in CHB patients.