Bray R A, Samberg N L, Gewurz H, Potempa L A, Landay A L
Department of Immunology/Microbiology, Rush Medical College, Chicago, IL 60612.
J Immunol. 1988 Jun 15;140(12):4271-8.
Previously, we have shown that antibodies specific for C-reactive protein determinants, not present on the native molecule, termed neo-CRP, also react with a significant percentage of PBL. In the present study, cells were evaluated by flow cytometry using alpha-neo-CRP antisera and mAb specific for lymphocyte subsets. With use of either monocyte-depleted PBL or Percoll-enriched large granular lymphocytes, we observed an overlap between cells reactive with alpha-neo-CRP and cells bearing the surface markers CD16, CD11b, Leu-7, and/or Leu-19, which are expressed on NK cells. In addition, we showed co-expression of the neo-CRP antigen with CD19, CD20, and HLA-DR, cell surface markers which are expressed on B lymphocytes. The major proportion of CD3+ cells failed to exhibit co-expression of neo-CRP. Single parameter flow cytometric analyses demonstrated that cells reactive with alpha-neo-CRP exhibited a bimodal staining pattern based on fluorescence intensity: high intensity neo-CRPbright and low intensity neo-CRPdim. Two-color analysis revealed that neo-CRPbright cells co-expressed CD19, CD20, and HLA-DR, whereas neo-CRPdim cells co-expressed CD16, CD11b, Leu-7, and Leu-19. Anti-neo-CRP also reacted with PBL obtained from patients with CD16+ lymphoproliferative disorders and from patients with chronic lymphocytic leukemia of B cell origin, but not with cells from patients with T cell or myeloid leukemias. The alpha-neo-CRP cells from patients with NK cell expansions showed dim fluorescence, whereas patients with B cell expansions showed bright fluorescence, consistent with the staining patterns observed with normal PBL. In addition, cell lines of T cell, B cell, NK cell, myeloid, and erythroid origin were evaluated for reactivity with alpha-neo-CRP. The cloned NK cell line NK 3.3 reacted as neo-CRPdim, but the B cell lines BL41, BL41/95, T1, T2, and CESS all reacted as neo-CRPbright. The cell lines K562, Molt-4, Hut-78, HL-60, U-937, and THP-1-0, which lack characteristic NK and B cell markers, did not react with alpha-neo-CRP. Additional study of the two-color histograms revealed a distinct diagonal staining pattern that was observed only when cells were co-stained with alpha-neo-CRP and either alpha-CD16 (alpha-Fc gamma RIII) or antibody IV3 (alpha-CDw32; alpha-Fc gamma RII). This finding suggests a 1:1 relationship between Fc gamma R on both NK and B cells and determinants recognized by alpha-neo-CRP.(ABSTRACT TRUNCATED AT 400 WORDS)
此前，我们已经表明，针对C反应蛋白决定簇（天然分子上不存在，称为新C反应蛋白）的特异性抗体，也能与相当比例的外周血淋巴细胞（PBL）发生反应。在本研究中，使用α-新C反应蛋白抗血清和针对淋巴细胞亚群的单克隆抗体，通过流式细胞术对细胞进行评估。使用单核细胞耗竭的PBL或经Percoll富集的大颗粒淋巴细胞，我们观察到与α-新C反应蛋白反应的细胞和带有NK细胞表面标志物CD16、CD11b、Leu-7和/或Leu-19的细胞之间存在重叠。此外，我们还显示新C反应蛋白抗原与B淋巴细胞表面表达的细胞表面标志物CD19、CD20和HLA-DR共表达。大部分CD3+细胞未表现出新C反应蛋白的共表达。单参数流式细胞术分析表明，与α-新C反应蛋白反应的细胞基于荧光强度呈现双峰染色模式：高强度新C反应蛋白明亮型和低强度新C反应蛋白暗淡型。双色分析显示，新C反应蛋白明亮型细胞共表达CD19、CD20和HLA-DR，而新C反应蛋白暗淡型细胞共表达CD16、CD11b、Leu-7和Leu-19。抗新C反应蛋白抗体也能与来自CD16+淋巴细胞增殖性疾病患者和B细胞来源的慢性淋巴细胞白血病患者的PBL发生反应，但不能与T细胞或髓系白血病患者的细胞发生反应。NK细胞扩增患者的α-新C反应蛋白细胞显示暗淡荧光，而B细胞扩增患者的细胞显示明亮荧光，这与正常PBL观察到的染色模式一致。此外，还评估了T细胞、B细胞、NK细胞、髓系和红系来源的细胞系与α-新C反应蛋白的反应性。克隆的NK细胞系NK 3.3表现为新C反应蛋白暗淡型，但B细胞系BL41、BL41/95、T1、T2和CESS均表现为新C反应蛋白明亮型。缺乏特征性NK和B细胞标志物的细胞系K562、Molt-4、Hut-78、HL-60、U-937和THP-1-0不与α-新C反应蛋白发生反应。对双色直方图的进一步研究揭示了一种独特的对角线染色模式，仅当细胞与α-新C反应蛋白和α-CD16（α-FcγRIII）或抗体IV3（α-CDw32；α-FcγRII）共染色时才会观察到。这一发现表明NK细胞和B细胞上的FcγR与α-新C反应蛋白识别的决定簇之间存在1:1的关系。（摘要截短至400字）
