STUDIES OF PRIMARY IMMUNODEFICIENCY DISEASES

我们实验室的研究集中在定义B的性质上 常见变异型免疫缺陷（CVI）， 原发性获得性人体免疫缺陷状态，特征为 低丙种球蛋白血症（血清中免疫球蛋白水平低），以及 功能性抗体应答受损。 我们以前的研究 来自CVI患者的纯化B细胞显示，尽管细胞具有 正常的增殖能力，它们表现出分化缺陷 在多个层面上。 例如，与正常B细胞相比， 循环的CVI B细胞含有减少数量的表面携带IgG 伊加细胞表面IgM B相应增加 细胞，表明在同种型转换的体内缺陷。 此外，本发明还提供了一种方法， 这些细胞不能分化成免疫球蛋白， 产生细胞。 在此期间，我们开始了CVI B细胞的研究， 就其已知的转录因子和辅因子的表达而言， 是B细胞转换和终末分化所必需的。 在 特别地，我们集中于B细胞特异性转录因子， 10月2日，由于基因靶向而缺乏这种蛋白质的小鼠， 与CVI患者的表型相似。 在最初的研究中，我们从患者的纯化B细胞中制备RNA， 用SAC加IL-2刺激后， 使用Oct-2基因特异性引物通过RT-PCR检测RNA。 CVI患者 和对照RNA产生相同的信号，表明没有明显的 Oct-2 RNA表达异常。 然后我们评估了10月2日 蛋白质功能的电泳迁移率变动分析 使用来自患者的刺激的新鲜B细胞的核提取物， 对照，以及来自患者的EBV转化的B细胞系， 对照 所用的靶寡核苷酸是核心寡核苷酸。 Oct-2启动子位点。 我们发现从病人和 对照产生适当大小的EMSA信号， 强度。 这些结果表明，CVI B细胞并不表现出一种 Oct-2基因缺失，也没有表现出异常 Oct-2与核心Oct-2启动子位点的结合。

Studies in our laboratory have focused on defining the nature of the B cell defects in the disease, Common Variable Immunodeficiency (CVI), a primary acquired human immunodeficiency state characterized by hypogammaglobulinemia (low levels of immunoglobulin in the serum), and impaired functional antibody responses. Our previous studies of purified B cells from CVI patients show that although the cells have a normal capacity to proliferate, they manifest differentiation defects at multiple levels. For example, when compared to normal B cells, circulating CVI B cells contain reduced numbers of surface bearing IgG and IgA cells with a commensurate increase in surface bearing IgM B cells, suggesting an in vivo defect in isotype switch. In addition, these cells fail to undergo differentiation into immunoglobulin producing cells. During this period we have initiated the study of CVI B cells with respect to their expression of transcription factors and cofactors known to be necessary for B cell switching and terminal differentiation. In particular, we have focused on the B cell-specific transcription factor, Oct-2, sine mice lacking this protein as a result of gene targeting have a phenotype similar to that of patients with CVI. In initial studies, we prepared RNA from purified B cells of patients and controls following stimulation with SAC plus IL-2 and amplified the RNA by RT-PCR using primers specific for the Oct-2 gene. CVI patient and control RNA gave rise to identical signals indicating no gross abnormality in the expression of Oct-2 RNA. We then evaluated Oct-2 protein function by performing electrophoretic mobility shift assays using the nuclear extracts of stimulated fresh B cells from patients and controls, as well as EBV-transformed B cell lines from patients and controls. The target oligonucleotide utilized was that of the core Oct-2 promoter site. We found that extracts from both patients and controls yielded an EMSA signal of appropriate size and identical intensity. These results indicate that CVI B cells do not manifest a gross deletion of the Oct-2 gene, nor do they manifest an abnormality of Oct-2 binding to the core Oct-2 promoter site.