Nontuberculous mycobacterial infection with concurrent IgG4‐related lymphadenopathy

Disseminated nontuberculous mycobacteria (NTM) infection with concurrent IgG4‐related lymphadenopathy has not been reported. We described a patient with neutralizing autoantibodies to interferon‐gamma (IFN‐γ) and elevated levels of serum IgG4 presenting with generalized lymphadenopathy and reactive dermatosis. Histologically, lymph nodes (LNs) showed effaced nodal architecture with polymorphic infiltrates, mimicking angioimmunoblastic T‐cell lymphoma. Both the absolute number and the ratio of IgG4+ plasma cells to IgG+ plasma cells were increased. Mycobacterium abscessus was isolated from cultures of LNs, and demonstrated by polymerase chain reaction‐restriction fragment length polymorphism. The skin biopsy showed neutrophilic dermatosis, consistent with Sweet syndrome. The patient met the criteria of both adult‐onset immunodeficiency syndrome and IgG4‐related lymphadenopathy. This case provides evidence of disseminated NTM infection with concurrent type III IgG4‐related lymphadenopathy in the patient with anti‐IFN‐γ autoantibodies.     

自身抗体在原发性胆汁性肝硬化中的应用分析

目的评价自身抗体对原发性胆汁性肝硬化患者诊断的应用价值,探讨该病与自身抗体的关系。方法用欧蒙印迹法、间接免疫荧光法、免疫金标法检测血清AMA-M2,ANA,ENA和ds-DNA,对结果进行统计、分析,与17例PBC、32例非PBC和35例健康对照进行比较。结果PBC组AMA-M2阳性率为94.1%,特异性为100.0%,非PBC组及健康对照组检出率为零。ANA,ENA和Ads-DNA检测结果在PBC组、非PBC肝硬化组、健康对照组分别为58.8%,56.2%和5.7%,前两组与对照组差异有显著性(P<0.01)。ENA检出率,PBC组23.5%、非PBC肝硬化组28.1%、对照组检出率为0%。结论AMA-M2是PBC的重要血清学标志,同时也是该病的重要诊断手段之一。PBC患者的AMA-M2阳性与临床病情无关,ANA,ENA和Ads-DNA对PBC的诊断无特异性帮助。     

Development of B cells producing natural autoantibodies to thymocytes and senescent erythrocytes

Natural antibodies produced by CD5⁺ B-1 B cells include those with specificity for senescent erythrocytes (anti-BrMRBC, anti-PtC) and for thymocytes (anti-thymocyte autoantibody, ATA). Here we describe work from our laboratories studying two prototypic examples, V_H11V9-encoded anti-BrMRBC and V_H3609V21c-encoded ATA. Using V_H11- transgenic mice, we discovered that certain natural autoantibodies utilize V_H genes that are selected against in bone marrow B cell development, but not fetal liver, effectively restricting their generation to fetal/neonatal life. Studies with ATA- transgenic mice demonstrated a critical requirement for self antigen in the accumulation of B cells with this specificity and for the production of high levels of serum ATA. Finally, analysis of B cell development in ATA- transgenic mice revealed two distinct responses by B cells to expression of this B cell receptor (BCR): most developing B cells in spleen of adult mice were blocked at an immature stage and only escaped apoptosis by editing their BCR to eliminate the ATA specificity; nevertheless, high levels of serum ATA were observed, indicating that some B cells differentiated to antibody-forming cells without altering their specificity. Thus, our studies reveal mechanisms for restricting the generation of B cells producing natural autoantibodies, demonstrate a key positive selection step in their development, and show that most developing B cells in adult mice bearing such specificities fail to reach a mature stage.     

Selective killing of B-cell hybridomas targeting proteinase 3, Wegener's autoantigen

Wegener's granulomatosis (WG) is a rare disease characterized by granulomatous lesions, small vessel vasculitis and the presence of anti-neutrophil cytoplasmic autoantibodies (C-ANCAs) in the sera of affected patients. Their main target antigen is proteinase 3 (PR3), a neutrophil and monocyte-derived neutral serine protease. Since the standard treatment of this severe autoimmune disease, with cyclophosphamide and corticosteroids, is associated with potential side-effects, the development of a more specific immunotherapeutic agent is warranted. The key role of ANCA in the pathogenesis of vasculitis and the effectiveness of anti-CD20 antibodies in patients with refractory WG points towards the importance of B cells in WG. We thus evaluated a new approach to selectively eliminate PR3-specific autoreactive B cells by targeting the B-cell receptor. For this purpose we used a bifunctional recombinant fusion protein consisting of the antigen PR3 and a toxin. The cytotoxic component of this novel fusion protein was the ribonuclease angiogenin, a human toxin with low immunogenicity. The toxin was stabilized by exchanging the catalytically relevant histidine in position 44 with glutamine to eliminate the autoproteolytic activity. PR3H44Q was fused either to the N terminus or to the C terminus of angiogenin. The recombinant proteins were expressed in 293T cells. Binding assays demonstrated the appropriate size and recognition by anti-PR3 antibodies. Using TUNEL technology, we demonstrated that these autoantigen toxins kill proteinase 3-specific B-cell hybridomas selectively by inducing apoptosis. The data indicate that autoantigen-toxins are promising tools in the treatment or co-treatment of autoimmune diseases in which the antigen is known.     

Tolerance established in autoimmune disease by mating or bone marrow transplantation that target autoantigen to thymus

Autoimmune diseases are a significant cause of death and morbidity, affecting up to 5% of the population. At present, there is no cure. Autologous bone marrow transplantation has been promoted as a treatment for achieving disease reversal and long-term remission. However, clinical trials in progress in Europe and North America report a significant risk of relapse. Here, we have addressed whether we can establish tolerance in an active autoimmune disease model by thymic expression of autoantigen. We show that tolerance and disease resistance can indeed be established in transgenic mice that spontaneously develop granulocyte macrophage colony stimulating factor-induced autoimmune gastritis, by mating them with disease-resistant transgenic mice that target autoantigen to the thymus. T cells from these double-transgenic mice are non-responsive to gastric antigen in vitro and fail to initiate disease following transfer to naive recipients. Further, we show that transplantation with bone marrow from disease-resistant transgenic mice renders recipient mice with gastritis tolerant to autoantigen as shown by a dramatic fall in autoantibody levels and T cell non-responsiveness to antigen in vitro. We suggest that genetically modified bone marrow targeting autoantigen to the thymus may be used to establish tolerance and prevent relapse of autoimmune disease following autologous bone marrow transplantation.     

Autoantibody Responses in Chinese Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is one of the most common cancers in the world and is particularly prevalent in Henan, China. The objective of this study was to analyze the frequency and specificity of autoantibodies associated with HCC in Henan. In the present study, 137 sera from HCC patients, 77 sera from other liver diseases, and 30 sera from normal human individuals were examined for autoantibodies using immunohistochemistry, Western blotting, and immunoprecipitation assays. Autoantibodies were detected in 80 of 137 (58.4%) HCC sera. Antinucleolar antibodies were seen more frequently in HCC compared to other liver diseases (9.5% vs. 1.3%, P < 0.05). Two nucleolar proteins—fibrillarin and NOR-90/hUBF—were identified as autoantigens. The frequency of autoantibodies in HCC sera with known hepatitis C virus (HCV) infection was significantly higher than that in sera without HCV infection (84.2% vs. 57.7%, P < 0.01). Another interesting finding was that autoantibodies to a 90-kDa cytoplasmic antigen were found in 21% of HCC patients. This is the first report on the frequency and specificity of autoantibodies in sera from Chinese patients with HCC. The data support that autoimmune responses to certain cellular proteins may be a by-product in the transformation to HCC, and further studies of novel targeted autoantigens in Chinese HCC may provide insights into how these proteins might be involved in malignancy.     

Serologic analysis of ovarian tumor antigens reveals a bias toward antigens encoded on 17q

We utilized SEREX immunoscreening to identify a set of novel tumor antigens that are associated with human serous ovarian cancer and may prove useful for the early detection and treatment of this disease. Extensive screening with a panel of sera from 25 late-stage ovarian cancer patients against 3 independent cDNA libraries identified a set of 9 antigens that were immunogenic in more than 1 patient and not in a panel of 20-45 normal female serum donors. These antigens include p53, NY-ESO-1, UBQLN1, HOXB6, TOP2A, putative helicase-RUVBL (RUVBL), HMBA-inducible (HEXIM1), DDX5 and HDCMA. Ten of 25 ovarian cancer patients (40%) expressed serum IgG to at least 1 of these antigens, while 14% (4/25) had antibodies to 2 or more antigens. Unexpectedly, 4 antigens identified in this screen, DDX5, HEXIM1, TOP2A and HOXB6, are encoded within a region of 17q that also includes the genes for HER2/neu, Homeobox-B7 and BRCA1. Real-time RT-PCR analysis showed that mRNA for HER2/neu and 3 SEREX-defined antigens, TOP2A, HOXB6 and DDX5, was more abundant in ovarian tumors than most normal tissues, including normal and benign ovarian tissues, suggesting that elevated expression of genes encoded within this region of chromosome 17 is a common event in ovarian tumors. Thus, these abnormal expression patterns combined with the endogenous immune response suggests that these antigens represent potential targets for immunotherapy.     

Identification of the amino acids on a neuronal glutamate receptor recognized by an autoantibody from a patient with paraneoplastic syndrome

Autoantibodies from a patient with paraneoplastic disease were identified previously to bind to the glutamate receptor (GluR) subunit GluR5 and to function as potential allosteric modulators of receptor activity (Gahring et al. [1995] Mol Med 1:245-253). In the present study we have used deletion mapping and mutagenesis to define the residues in GluR5 bound by this autoreactivity. The autoantibody contact residues include residues K497, N508, K510, E512, and to a lesser extent Q507. Residues 507-512 confer autoantibody specificity of the autoreactivity to GluR5. These residues have been shown in crystallographic studies (Armstrong et al. [1998] Nature 395:913-917) to participate in a loop structure, whereas residue K497 is located on a beta-strand. Notably, this binding spans tyrosine 504, a residue important in forming the agonist-binding site. We propose that autoantibody binding of essential residues in this GluR5 autoantigenic region defines a subunit-specific allosteric regulatory site on neuronal glutamate receptors and suggests how receptor dysfunction and region-specific neuronal death in the brain can progress in certain autoimmune neurological diseases.     

系统性红斑狼疮患者高表达的CD154促进自身抗体分泌

【目的】探讨系统性红斑狼疮（SLE）患者外周血单个核细胞（PBMC）CD154表达水平、及CD40/CD154对SLE患者分泌自身抗体的影响和地塞米松（Dex）对此的作用。【方法】选择SLE活动期患者（10例）和正常对照者（11例）,分离PBMC,检测其CD154的表达水平;培养PBMC,应用CD40 mAb和Dex进行干预,使用ELISA法检测培养液上清抗dsDNA水平。【结果】①活动期组PBMC CD154表达水平明显高于对照组6.4%±2.1%vs 1.5%±1.2%,（P〈0.05）;②活动期组培养液上清抗dsDNA抗体水平明显高于对照组[（11.8±6.8）U/mL vs（5.6±2.3）U/mL（RPMI 1640）,（11.4±7.0）U/mL vs（6.3±2.1）U/mL（IgG）,P均〈0.05];③CD40 mAb使两组培养液上清抗dsDNA抗体水平明显增高[活动期组：（18.6±7.7）U/mL vs（11.8±6.9）U/mL,对照组：（8.3±4.4）U/mL sv（5.6±2.3）U/mL,P均〈0.05]。④在活动期组,Dex明显抑制CD40 mAb引起的PBMC培养液上清抗dsDNA抗体水平的增高[（8.8±5.2）U/mL vs（18.6±7.7）U/mL,P〈0.05],使其低于非特异性处理时水平[（8.8±5.2）U/mL vs（11.8±6.8）U/mL,P〈0.05];Dex不影响对照组。【结论】活动期SLE患者PBMC表达的CD154水平升高,此能够促进抗dsDNA抗体的分泌,而Dex能够抑制其作用。