Cytokine production pattern of CD4+ T cells and its relation to anti-DNA antibody production in patients with systemic lupus erythematosus

人们对系统性红斑狼疮 (SLE) 个体患者新鲜分离的 CD4 T 细胞中一系列细胞因子的细胞内积累进行了研究，以确定这些细胞在体内可能产生哪些因子。使用细胞因子特异性抗体和流式显微荧光测定法的间接免疫荧光技术在单细胞水平上鉴定产生细胞因子的细胞。来自 SLE 患者（而非正常受试者）的 CD4 T 细胞被证明能够自发产生白细胞介素 (IL)-2、IL-4、IL-6、干扰素-γ 和肿瘤坏死因子-α。通过进行双色免疫荧光研究，我们观察到细胞不产生、同时产生一种或多种细胞因子的多样化生产模式。结果表明，SLE 患者体内的 CD4 T 细胞可能在体内多克隆地重新激活，因此这些多克隆激活的 CD4 T 细胞的扩大群体会自发地产生多种细胞因子。导致 B 细胞等免疫效应细胞的持续激活。这可能在人类SLE的发生和维持中发挥重要作用。在我们对细胞因子产生模式的研究过程中，我们发现SLE患者的外周血淋巴细胞（PBL）上细胞粘附分子（例如LFA-1和ICAM-L）过度表达。此外，SLE患者的血液中还存在一种新的CD4 T淋巴细胞亚群，该亚群同时表达CD45RA和ICAM-1。将抗LFA-1或抗ICAM-1单克隆抗体（mabsi）引入新鲜分离的SLE PBL的体外培养物中，导致自发多克隆IgG产生和自发IgG抗DNA抗体产生均受到抑制。因此，SLE淋巴细胞的LFA-1和ICAM-1分子的过度表达促进T-B细胞通讯，从而导致它们产生过量抗体。这一发现可能为mabs治疗性免疫干预提供可能的策略。狼疮性肾炎的小鼠模型表明，阳离子抗DNA自身抗体具有致肾炎的潜力，我们还研究了活动性狼疮性肾炎患者体内是否也产生了IgG类阳离子抗DNA抗体，并分析了此类抗体产生的机制，其中高阳离子性抗DNA抗体在狼疮性肾炎患者血清抗体的等电聚焦免疫印迹中表现突出。在用preunisolone成功治疗后，与循环中阳离子抗DNA抗体的消失有关，而且，通过基因中的许多点突变产生了阳离子抗DNA抗体，包括用种系序列的中性氨基酸替换阳离子氨基酸，结果表明这些抗体可能是通过抗原驱动的方式产生的。

The intracellular accumulation of a series of cytokines in freshly- isolated CD4+ T cells in individual patients with systemic lupus erythematosus (SLE) has been studied to determine what factors may be produced by these cells in vivo. The cytokine-producing cells were identified at a single cell level using by cytokine-specific antibodies and an indirect immunofluorescence technique with flow microfluorometry. CD4+ T cells from patients with SLE, but not normal subjects, were shown to spontaneously produce interleukin (IL)-2, IL-4, IL-6, interferon-gamma, and tumor necrosis factor-alpha. By performing two-color immunofluorescence studies, we observed a variegated production pattern with cells making no, one or several cytokines simultaneously. The results indicate that CD4+ T cells in patients with SLE may reactivated polyclonally in vivo, and thus that the expanded population of these polyclonally activated CD4+ T cells spontaneously produces several cytokines which may. cause sustai … More ned activation of immunological effector cells such as B cells. This may have important roles in both the development and maintenance of human SLE.In the course of our studies of the cytokine production pattern, we found overexpression of cell adhesion molecules such as LFA-1 and ICAM-+L on peripheral blood lymphocytes (PBL) of SLE patients. Moreover, there existed a novel CD4+ T lymphocyte subset, which expressed CD45RA and ICAM-1 simultaneously in the blood of patients with SLE. Introduction of anti-LFA-1 or anti-ICAM-1 monoclonal antibodies (mabsi into the in vitro cultures of freshly isolated SLE PBL resulted in inhibition of both the spontaneous polyclonal IgG production and spontaneous IgG anti-DNA antibody production. Thus, overexpression of LFA-1 and ICAM-1 molecules of SLE lymphocytes facilitates T-B cell communication, thereby causing them to produce excessive antibodies. This finding may be offer a possible strategy for therapeutic immune intervention by mabs.Extensive studies in the murine model of lupus nephritis have shown that cationic anti-DNA autoantibodies have nephritogenic potential. We have also investigated whether cationic anti-DNA antibodies of IgG class are also produced in vivo in patients with active lupus nephritis, and analyzed the mechanism by which such antibodies are produced. The highly cationic anti-DNA antibodies of IgG class were prominent in the isoelectric focusing-immunoblots of serum antibodies from the patients with lupus nephritis. Decreased proteinuria after successful treatment with preunisolone was associated with disappearance of the cationic anti-DNA antibodies in the circulation. Moreover, the cationic anti-DNA antibodies were produced through numerous point mutations in the genes, including substitution of neutral amino acids of germline sequences for cationic amino acids. The results indicate that these antibodies may be produced by antigen-driven manner. Less