我们先前的研究提示4.7 kDa和4.4 kDa的肽段对于诊断肾移植急性排斥有意义。该研究的目的在于明确这些多肽的特征并评估其作为生物标记的意义。这些多肽现通过串联质谱法和蛋白芯片免疫测定明确为人β-防卫素-1(4.7 kDa)和α-1-抗胰凝乳蛋白酶(4.4 kDa)。我们采用表面增强激光解吸离子化时间飞行质谱（SELDI-TOF MS）发现尿中上述两种蛋白大量存在，并发现发生排斥反应的患者相对于临床平稳的患者尿中β-防卫素-1下降而α-1-抗胰凝乳蛋白酶上升。两种肽结合比例用于诊断排斥反应的受试者工作特征曲线(ROC)的曲线下面积(AUC)为0.912。纵向分析也证实在急性排斥反应发生时β-防卫素-1上升而相反的α-1-抗胰凝乳蛋白酶下降。通过放免法测定发现临床平稳的患者和发生排斥的患者尿中β-防卫素-1水平不同，前者为176.8 +/- 122.3 pg/mL而后者为83.2 +/- 52.2 pg/mL，ROC AUC 0.749（p < 0.01）。免疫组化证实急性排斥反应的患者肾实质β-防卫素-1表达下降。结论：尿中排泄的β-防卫素-1和α-1-抗胰凝乳蛋白酶比值是全新的可能作为急性排斥的生物标记。

We previously demonstrated that 4.7 kDa and 4.4 kDa peptides are useful in diagnosing acute rejection in renal transplant recipients. The aim of this study was to characterize these polypeptides and assess their potential as biomarkers. The polypeptides were identified as human beta-Defensin-1 (4.7 kDa) and alpha-1-antichymotrypsin (4.4 kDa), by tandem mass spectrometry and ProteinChip immunoassay. The urinary abundance of both polypeptides, assessed using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS), revealed a reduction in beta-Defensin-1 while alpha-1-antichymotrypsin increased in patients with rejection (p < 0.05) compared with clinically stable transplants. The area under the curve (AUC) for the receiver operator characteristic (ROC) curve for the diagnosis of rejection for the ratio of both peptides combined was 0.912. Longitudinal analysis confirmed a reduction in beta-Defensin-1 with a reciprocal increase in alpha-1-antichymotrypsin as rejection developed. The difference in urinary beta-Defensin-1 levels quantified by radioimmunoassay was 176.8 +/- 122.3 pg/mL in stable patients compared with 83.2 +/- 52.2 pg/mL in patients with acute rejection, with an ROC AUC of 0.749 (p < 0.01). Immunohistochemistry (IHC) confirmed reduced beta-Defensin-1 expression in the renal parenchyma of patients experiencing acute rejection. In conclusion, the ratio of beta-Defensin-1 and alpha-1-antichymotrypsin excretion in the urine is a novel, potentially useful candidate biomarkers of acute rejection.

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