Mixed chimera status in patients with Wiskott-Aldrich syndrome (WAS) after hematopoietic stem cell transplantation ; evaluation by flow cytometric analysis of intracellular WAS protein expression.

Wiskott-Aldrich综合征（WAS）是由X染色体上的WAS蛋白（WASP）基因缺陷引起的。以前我们报道过流式细胞术分析细胞内WASP表达（FCM-WASP）对WAS患者和携带者的诊断是有用的。在这项研究中，我们应用FCM-WASP评估混合嵌合体（MC）状态的12例WAS患者接受造血干细胞移植（HST）。在HST之后，通过该方法可以容易地区分供体和供体来源的外周血单核细胞（PBMC），因为观察到供体细胞是WASP+<bright>，而缺陷受体细胞是WASP+<dim>。此外，通过双色FCM-WASP，MC状态可以通过细胞谱系来表征。12例WAS患者中有6例HST后为MC状态，而其他患者为完全嵌合体状态。在所检查的每个细胞系中观察到MC。然而，在PBMC中，受体细胞最常见于单核细胞群中。最后，为了研究这些患者中供体和受体T细胞的幼稚/记忆状态，使用抗CD 45 RA或CD 45 RO进行三色FCM-WASP。结果表明，与WASP^<bright>T细胞相反，大多数WASP^<dim>T细胞在HST后一年多仍保持幼稚状态（CD 45 RA ^+/RO^~）。在HST前WAS患者中未观察到幼稚/记忆T细胞比例失衡。我们的结论是，FCM-WASP是一个潜在的有用的方法进行HST的WAS患者的临床随访。这项研究也可能对WASP在造血发育过程中的作用有重要意义。

The Wiskott-Aldrich syndrome (WAS) is caused by defects in the WAS protein (WASP) gene on the X-chromosome. Previously we reported that flow cytometric analysis of intracellular WASP expression (FCM-WASP) was useful for the diagnosis of WAS patients and carriers. In this study, we applied FCM-WASP to evaluate the mixed chimera (MC) status of 12 WAS patients who underwent hematopoietic stem cell transplantation (HST). After HST, donor and recipient-derived peripheral blood mononuclear cells (PBMC) could be easily distinguished by this method, as the donor cells are observed to be WASP^<bright>, while the defective recipient cells are WASP^<dim>. Furthermore, by two-color FCM-WASP, the MC status could be characterized by cell lineage. Six of 12 WAS patients were revealed to have the MC status after HST, while others had the complete chimera status. The MC was observed in every cell lineage examined. However, among PBMC, recipient cells were most commonly observed in the monocyte population. Finally, to investigate the naive/memory status of donor and recipient T cells in these patients, three-color FCM-WASP using anti-CD45RA or CD45RO was performed. It was demonstrated that, in contrast to WASP^<bright> T cells, most WASP^<dim> T cells remained naive (CD45RA^+/RO^~) more than one year after HST. No imbalance in the ratio of naive/memory T cells was observed in WAS patients before HST. We conclude that FCM-WASP is a potentially useful method for clinical follow-up of WAS patients who underwent HST. This study may also have significant implications regarding the role of WASP during hematopoietic development.