Exosomal Fetuin-A identified by proteomics: A novel urinary biomarker for detecting acute kidney injury
Exosomal Fetuin-A identified by proteomics: A novel urinary biomarker for detecting acute kidney injury
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DOI：
10.1038/sj.ki.5001874
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发表时间：
2006-11-01
影响因子：
19.6
通讯作者：
Star, R. A.
中科院分区：
文献类型：
--
作者：
Zhou, H.;Pisitkun, T.;Star, R. A.
含有顶膜和细胞内液的尿液外泌体通常从所有肾单位节段分泌到尿液中，并且可能携带肾功能障碍和结构损伤的蛋白质标志物。我们的目的是发现尿液外泌体中的生物标志物，以检测死亡率和发病率较高的急性肾损伤（AKI）。给动物注射顺铂。通过差速离心分离尿液外泌体。通过凝胶电泳中的二维差异评估蛋白质变化，并通过质谱法鉴定变化的蛋白质。通过蛋白质印迹法对注射顺铂的大鼠的个体尿液样本中鉴定出的候选生物标志物进行了验证；双侧缺血和再灌注（I/R）；容量耗尽；以及患有或不患有 AKI 的重症监护病房 (ICU) 患者。我们鉴定出 18 种蛋白质在顺铂注射后 8 小时增加，9 种蛋白质减少。大多数候选者无法通过蛋白质印迹法进行验证。然而，注射顺铂后，外泌体胎球蛋白-A 在第 2 天（血清肌酐增加和肾小管损伤前 1 天）增加了 52.5 倍，并在第 5 天（肾损伤峰值）时保持升高 51.5 倍。通过免疫电子显微镜和洗脱研究，胎球蛋白-A 位于尿外泌体内部。尿胎球蛋白-A 在 I/R 早期（2-8 小时）增加 31.6 倍，但在肾前性氮质血症中则没有增加。与无 AKI 的患者相比，三名患有 AKI 的 ICU 患者的尿液外泌体胎球蛋白-A 也有所增加。我们的结论是：（1）尿液外泌体的蛋白质组学分析可以为诊断 AKI 提供候选生物标志物；（2）尿液胎球蛋白-A 可能是结构性肾损伤的预测生物标志物。
Urinary exosomes containing apical membrane and intracellular fluid are normally secreted into the urine from all nephron segments, and may carry protein markers of renal dysfunction and structural injury. We aimed to discover biomarkers in urinary exosomes to detect acute kidney injury (AKI), which has a high mortality and morbidity. Animals were injected with cisplatin. Urinary exosomes were isolated by differential centrifugation. Protein changes were evaluated by two-dimensional difference in gel electrophoresis and changed proteins were identified by mass spectrometry. The identified candidate biomarkers were validated by Western blotting in individual urine samples from rats subjected to cisplatin injection; bilateral ischemia and reperfusion (I/R); volume depletion; and intensive care unit (ICU) patients with and without AKI. We identified 18 proteins that were increased and nine proteins that were decreased 8 h after cisplatin injection. Most of the candidates could not be validated by Western blotting. However, exosomal Fetuin-A increased 52.5-fold at day 2 (1 day before serum creatinine increase and tubule damage) and remained elevated 51.5-fold at day 5 (peak renal injury) after cisplatin injection. By immunoelectron microscopy and elution studies, Fetuin-A was located inside urinary exosomes. Urinary Fetuin-A was increased 31.6-fold in the early phase (2-8 h) of I/R, but not in prerenal azotemia. Urinary exosomal Fetuin-A also increased in three ICU patients with AKI compared to the patients without AKI. We conclude that (1) proteomic analysis of urinary exosomes can provide biomarker candidates for the diagnosis of AKI and (2) urinary Fetuin-A might be a predictive biomarker of structural renal injury.