Assessment Of Lymphocytes In Patients With Autoimmune Ly

一个广泛的流式细胞术评估继续与自身免疫性淋巴组织增生综合征（ALPS）患者及其大家庭成员，扩展的双阴性T细胞和B细胞群体的特征的基础上。双阴性T细胞已被证明是α β TcR、CD 57+、HLA-DR+和CD 45 RA+。这项研究已经扩展到更完整地表征双阴性T细胞，包括所有ALPS患者中的B220表达和γ-δ TcR T细胞。此外，我们扩大了B细胞的表征，针对记忆B细胞，在这些患者中使用CD 27和B220评估，并记录了与免疫球蛋白重链表达改变相关的CD 27 + B细胞减少。目前正在收集这些数据以便公布。免疫调节性T细胞的功能研究未能提供一致的体外抑制指示系统。但是我们已经开发了Fox P3的定量RTPCR测定，并从ALPS Ia型患者、家庭成员和对照组获得的T细胞中收集mRNA。目前正在对这些样本进行Fox P3 mRNA的评估。此外，我们还验证了细胞内Fox P3染色的流式细胞术测定。该测定法证明了大部分表达Fox P3的细胞是CD 4/CD 25 T细胞。对对照受试者和ALPS患者中Fox P3表达细胞的评估表明，尽管CD 4/CD 25 T细胞受到抑制，Fox P3表达在ALPS中是正常的。最后，我们还将Fox P3 PCR和基于流式细胞术的检测扩展到ATL细胞，这是基于这种疾病代表免疫调节性T细胞的白血病扩增的可能性。ATL研究显示，在通过流式细胞术测量的FoxP 3表达水平方面，各种ATL患者之间存在显著差异，这已经通过RTPCR测试证实。我们目前正在评估这些差异的可能解释，包括体外IL-2独立性

An extensive flow cytometric evaluation continues of patients with autoimmune lymphoproliferative syndrome (ALPS) and their extended family members, on the basis of characterization of the expanded double-negative T-cell and B-cell populations. Double-negative T-cells have been demonstrated to be alpha beta TcR, CD57+, HLA-DR+, and CD45RA+. This study has been extended to characterize the double-negative T-cells more completely including B220 expression and gamma-delta TcR T-cells in all ALPS patients. In addition, we have expanded characterization of the B cells, directed at memory B cells using CD27 and B220 assessment in these patients and have documented a decrease in CD27+ B cells associated with altered repertoire of immunoglobuine heavy chain expression. These data are currently being assembled for publication. Functional studies of immunoregulatory T cells have failed in providing a consistent ex vivo indicator system for inhibition. But we have developed a quantitiative RTPCR assay for Fox P3 and have collected mRNA from T cells obtained from ALPS type Ia patients, family members and controls. These samples are now in the process of being evaluated for Fox P3 mRNA. In addition we have validated a flow cytometric assay for intracellular Fox P3 staining. This assay demonstrated that the bulk of Fox P3 expressing cells are CD4/CD25 T cells. Evaluation of the Fox P3 expressing cells in control subjects and ALPS patients suggests that Fox P3 expression is normal in ALPS despite the depression CD4/CD25 T cells. Finally, we have also extended the Fox P3 PCR and flow based assays to ATL cells based on the possibility that this disease represents a leukemic expansion of immunoregulatory T cells. The ATL studies show substantial differences between various ATL patients in terms of the level of FoxP3 expressed as measured by flow cytometry which has been corroborated by RTPCR testing. We are currently evaluating possible explanations for these differences including in vitro IL-2 independence