ANALYSIS OF FINE-NEEDLE ASPIRATION BIOPSIES BY FLOW CYTOMETRY IN LIVING-RELATED PEDIATRIC LIVER TRANSPLANT PATIENTS

肝移植的预后有所改善。然而，急性排斥仍然是一个主要问题。细针穿刺活检（FNAB）是诊断急性排斥反应的一种非常安全和有用的方法，在经验丰富的中心达到与核心肝活检相似的诊断结果。本研究的目的是通过流式细胞术分析FNAB样本与外周血淋巴细胞（PBL）的比较，定量评估同种异体移植物的免疫状态。方法：对10例儿童活体同种异体肝移植进行FNAB监测，每隔2 ~ 3天监测一次，每当怀疑有排斥反应时监测一次。用流式细胞术分析同种异体移植物PBL和抽吸活检的淋巴样细胞亚群。结果：两种样品在轻中度排斥反应时，CD4CD25、CD8CD25和CD4DR的表达均略有升高（3-6%）。然而，排斥前1 ~ 3天，与PBL相比，FNAB样品中CD8DR阳性细胞显著增加（p< 0.05）。在严重排斥事件中，不仅CD4DR、CD8DR，而且CD4CD25、CD8CD25阳性细胞也增加（20-30%），特别是在FNAB样本中。这些发现与生化实验室参数密切相关，在移植物中活化淋巴细胞（CD4CD25、CD4DR、CD8CD25、CD8DR阳性细胞）的增加可以比血清值升高提示肝功能障碍早2-5天观察到。此外，在非免疫性移植物损伤（如动脉血栓形成或胆漏）期间，没有明显的系列变化。结论：流式细胞术研究FNAB样本可作为预测同种异体肝移植（即急性排斥反应）免疫激活的可靠方法。

Introduction: The outcome of liver transplants has improved. However, acute rejection is nevertheless a major problem. Fine-needle aspiration biopsy (FNAB) is a very safe and helpful procedure for the diagnosis of acute rejection, achieving a similar diagnostic yield as core liver biopsy in experienced centers. The purpose of this study is to evaluate an immunological state of allografts quantitatively using FNAB samples in comparison with peripheral blood lymphocytes (PBL) by flow cytometric analysis.Methods: Ten living-related liver allografts for children were monitored with FNAB at 2-3 day intervals and whenever rejection crisis was suspected. Lymphoid cell subpopulations from PBL and aspiration biopsies of allografts by flow cytometry were analyzed.Results: The expression of CD4CD25, CD8CD25 and CD4DR increased slightly (3-6%) in case of mild to moderate rejection in both samples. However, 1 to 3 days prior to rejection, CD8DR positive cells increased significantly in FNAB samples compared to that in PBL (p<0,05). In severe rejection episodes, not only CD4DR, CD8DR but CD4CD25, CD8CD25 positive cells also increased (20-30%) especially in FNAB samples. These findings correlated well with biochemical laboratory parameters, and the increase of activated lymphocytes (CD4CD25, CD4DR, CD8CD25, CD8DR positive cells) in the grafts could be observed already 2-5 days earlier than elevated serum values indicated liver dysfunction. Furthermore, there was no significant serial change during non-immunological graft damages (I.e. arterial thrombosis or bile leakage).Conclusion: Flow cytometry study of FNAB samples could be a reliable method to prospect immunological activation in liver allografts (I.e. acute rejection episodes).