T lymphocyte interaction with immunoglobulin G antibody in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease with multiple immune disturbances whose mechanisms remain unclear. We examined the interaction of antilymphocyte antibodies with cultured normal T lymphocytes. T cells were prepared by E-rosetting after petri-dish removal of adherent cells and cultured for 2-7 d in the presence of SLE sera or normal human sera. Cultured T cells were washed and sonicated, and the amount of cell-associated IgG was quantitated by radioimmunoassay or enzyme-linked immunoassay (ELISA) methods. T cells cultured with 27 of 39 SLE sera showed marked increments of associated immunoglobulin G (IgG) although this was not observed with sera from mixed connective tissue disease patients containing high titers of ribonucleoprotein antibody or normal donors. The effective factors for IgG association in SLE sera were absorbed with normal peripheral blood lymphocytes or T cells. Anti-T cell IgG cytotoxic activity strongly correlated with T cell IgG association (P less than 0.01). T cell-associated IgG was not removed by stripping of cell membrane IgG from living cells by acid buffer treatment; indirect immunofluorescence of cells fixed after 2-4 d of culture revealed cytoplasmic IgG staining. IgG anti-T cell antibodies appeared to associate inside the cell membrane or to penetrate into the cytoplasm of cells. T cell Fc receptor blocking by heat-aggregated IgG or anti-beta 2-microglobulin antibody did not alter IgG cell association. Since pepsin-digested SLE sera showed no T cell association activity, whole IgG antibody molecules appeared to be necessary for interaction with cultured T cells. In addition, reduction and alkylation of active SLE sera completely nullified T cell reactivity. When normal T cells were cultured with SLE sera showing marked IgG T cell association, viability of cultured T cells decreased rapidly after 4 d, which suggests that IgG anti-T cell antibodies were associated with cell destruction. IgG cell-associating antilymphocyte antibodies present in SLE sera may cause T cell disturbances in vivo and may be related to the lymphocytopenia present in SLE patients.     

Characterization of autoantibody activities in sera anti-DNA antibody and circulating immune complexes from 12 systemic lupus erythematosus patients

To examine autoantibodies present in patients with active systemic lupus erythematosus (SLE), sera, circulating immune complexes (CIC), and antibodies purified on DNA-immunoadsorbent were tested by enzyme immunoassay. A panel of self-antigens, including DNA, histones (HIS), glomerular basal membrane (GBM), thymus cell extract (TCE), actin (ACT), myosin (MS), and tubulin (TUB), was used to define their specificities. IgM antibodies against all antigens of the panel were detected in sera, CIC, and in antibodies eluted from the DNA-immunoadsorbent and demonstrated a large polyreactivity. IgG antibodies showed restricted activities against DNA, HIS, GBM, and TCE in sera and a large polyreactivity in CIC. Inhibition experiments were performed to assess their mono- or polyreactivities. Among the IgG autoantibody population recognizing DNA, two populations of IgG antibodies were detected in the sera and in the affinity purified anti-DNA: one recognizes DNA, HIS, and GBM, and the other binds to DNA and to cytoskeletal proteins. These autoantibody populations were found in CIC, which also often contained high amounts of IgG antibodies recognizing ACT and MS. A third population of IgG antibody that recognizes only TCE and could not be inhibited by DNA or other antigens was found in serum and CIC. Our data demonstrate the existence of several populations of autoantibody in serum and CIC of SLE patients: (1) IgM polyreactive autoantibodies, (2) IgG polyreactive autoantibodies recognizing DNA and cytoskeletal proteins, (3) IgG specific to DNA, which cross react with HIS and GBM, and (4) IgG specific to TCE antigens. © 1996 Wiley-Liss, Inc.     

The Raji cell radioimmune assay for detecting immune complexes in human sera.

A sensitivie and simple procedure for the detection and quantitation of soluble complement (C)- fixing immune complexes in sera of patients with various disease states has been developed by utilizing C receptors on Raji cells. These cells lack membrane-bound immunoglobulin but have receptors for IgG Fc, C3b, C3d, and possibly with other C proteins. Uptake experiments showed that both aggregated human gamma globulin (AHG) and 7S IgG bound to receptors for IgG Fc; however, AHG reacted with C bound to cells only via receptors for C and this binding was much more efficient than via IgG Fc receptors. AHG was used as an in vitro model of human immune complexes and its uptake by Raji cells was quantitated by 125I-radiolabeled antihuman IgG. The limit of sensitivity of this test was 6 mug AHG/ml serum. The ability of Raji cells to detect AHG in serum depended on the amount of radioactive antibody used and the size of aggregates. The presence of an excess of C somewhat inhibited binding of AHG containing C to Raji cells. The efficient binding of AHG by receptors for C on Raji cells was used for the detection and quantitation of immune complexes in human sera. Raji cells were incubated with sera to be tested and then reacted with excess radiolabeled antihuman IgG; the amount of radioactivity bound to the washed cells was determined and referred to a standard curve of radioactive antibody uptake by cells previously incubated with increasing amounts of AHG in serum. Thereby immune complexes were detected and quantitated in serum hepatitis, systemic lupus erythematosus, vasculitis, subacute sclerosing panencephalitis, dengue hemorrhagic fever, and malignancies.     

Neutrophil-binding immunoglobulin G in systemic lupus erythematosus.

The objectives of these studies were to quantify the amounts of immunoglobulin (Ig)G bound to peripheral blood neutrophils from patients with systemic lupus erythematosus (SLE) and to determine the contributions of soluble immune complexes or anticell antibodies to the levels of IgG neutrophil-binding activity in SLE sera. Neutrophil-bound IgG, determined by a sensitive antiglobulin inhibition assay, was elevated in 7 out of 14 SLE patients compared with values obtained in 23 normal controls. The levels of IgG neutrophil-binding activity in sera were elevated in 22 of 38 patients with SLE over the values seen with 36 normal sera. No correlation was found between the peripheral blood neutrophil counts in the SLE patients and the values for IgG adherent to the cells or serum cell-binding activity. The sera from 18 patients with SLE were fractionated by gel filtration. Elevated levels of IgG neutrophil-binding activity were found in 11 of the 18 G-200 excluded pools and in 13 of the G-200 IgG pools. In nine sera elevated levels were observed in both pools. F(ab')2 fragments of IgG from SLE sera bound to normal polymorphonuclear leukocytes in greater amounts than F(ab')2 fragments of IgG from normal sera. A significant correlation existed between the values of IgG neutrophil-binding activity found in SLE sera and those obtained with both the G-200 excluded and IgG pools. Sucrose density gradient fractionation of four sera from SLE patients confirmed the presence of both large (greater than 19S) and intermediate-sized (7S-19S) cell-binding immune complexes as well as of monomeric IgG antibodies to neutrophils. The levels of IgG neutrophil-binding activity in the SLE sera correlated well with the results obtained with the Raji cell assay for immune complexes as well as with the titer of antibodies to nuclear antigens. These data indicate that circulating neutrophils from patients with SLE commonly have increased amounts of cell-bound IGG. The elevated levels of IgG neutrophil-binding activity in the sera of these patients are caused by both soluble immune complexes and antibodies reactive with neutrophils.     

Antiglycolipid autoantibody detected in the sera from systemic lupus erythematosus patients.

A high incidence of autoantibody against the neutral glycolipid "asialo GM1" was observed in sera from patients with systemic lupus erythematosus (SLE) with neurological disorders, using an immunoflocculation test. The sera from 14 out of 17 cases of SLE with neurological disorders showed antibody activity against asialo GM1 but not against the following glycolipids: asialo GM2 GM1, and galactocerebroside. In another 87 cases of SLE without any history of seizures, as well as 61 cases of other autoimmune diseases (rheumatoid arthritis, progressive systemic sclerosis, mixed connective tissue disease, etc.) and 20 cases of various neurological diseases (epilepsy, multiple sclerosis, etc.), no antibody could be detected. In general, the antibody titer was high several months, even years, before and/or after the seizure, though the titer was low at the time that patients showed definite neurological symptoms. Immunochemical characterization with Sephadex G-200 chromatogrphy and protein A-Sepharose CL-4B affinity column indicated that the antiasialo GM1 was probably an autoantibody belonging to the immunoglobulin G class. The above results suggest that this newly found autoantibody plays a role in the pathogenesis of neurological disorders accompanying SLE.     

Natural antibodies in sera of patients with systemic lupus erythematosus

Sera obtained from fifty-five patients with active systemic lupus erythematosus (SLE) and from four patients with mixed connective tissue disease (MCTD) previously shown by immunofluorescence and by double immunodiffusion to possess antinuclear antibodies, were tested for the presence of natural antibodies of IgG, IgA, and IgM isotypes. Antibody activity to actin, myosin, DNA, TNP, albumin, and tubulin was examined, using an enzyme-linked immunosorbent assay (ELISA). It was found that, in comparison with the antibody titers in normal sera, most of the SLE and MCTD sera possessed statistically greater amounts of IgG, IgA, and IgM antibodies directed against all the antigens tested. Furthermore, the IgG, IgA, and IgM antibody activity to DNA and TNP, compared to that found with all the other antigens, was significantly higher. Antibodies reacting with a saline extract of calf thymus (ECT) were studied by ELISA and by immunodiffusion. No correlation was observed between the natural antibody titers and the serum antibody levels to ECT detected either by ELISA or by immunodiffusion.     

Specific concentration of polynucleotide immune complexes in the cryoprecipitates of patients with systemic lupus erythematosus.

Although the association of cryoglobulinemia with hypocomplementemia and tissue injury in systemic lupus erythematosus is well recognized, composition of cryoprecipitates in terms of circulating antigens and antibodies in this disease is less clear. To clarify this question, cryoprecipitates from patients with SLE were examined with sensitive assay techniques for certain antipolynucleotide antibodies and DNA antigen. DNA antibodies were highly enriched relative to serum levels in the majority of cryoprecipitates. DNA antigen was also demonstrable. Antibody to ribonucleoprotein, although less frequently present, was similarly enriched in certain cryoprecipitates. In contrast, anti-double strand RNA, which was commonly detectable in relatively high titer in serum, was only minimally concentrated in a minority of cryoprecipitates. Absorption experiments using red blood cells heavily coated with polynucleotide antigen indicated that a major proportion of the IgG in certain cryoprecipitates was specific antibody. The data strongly suggest that the cryoprecipitates in systemic lupus erythematosus represent circulating immune complexes that are soluble at 37 degrees C and come out of solution in the cold. The marked concentration of immune complexes in the cryoglobulin offers a simple and direct method for determination of the nature of the complexes. The accumulated evidence obtained in the present study indicates that these complexes closely reflect, in their composition, the circulating immune complexes which are most significant pathogenetically in renal tissue injury.     

An in vitro assay for detection of glomerular binding IgG autoantibodies in patients with systemic lupus erythematosus.

The deposition of anti-dsDNA antibodies in the glomerulus is believed to play a critical role in the pathogenesis of nephritis in SLE. However, an absolute correlation between serum levels of anti-dsDNA antibodies and renal disease has not been found. Recently a glomerular binding assay (GBA) has been developed to detect IgG binding to isolated rat glomeruli. We have used the GBA to study sera from four groups of SLE patients: (A) + anti-dsDNA antibodies, active nephritis; (B) - anti-dsDNA antibodies, active nephritis; (C) + anti-dsDNA antibodies, no nephritis; and (D) - anti-dsDNA antibodies, no nephritis. The serum anti-dsDNA antibodies in group A and group C patients could not be distinguished on the basis of isotype, charge, or cross-reactivity with histones. Nevertheless, the mean intensity of glomerular immunofluorescence was significantly higher in group A than in the three other patient groups and distinguished between patients with serum anti-dsDNA antibodies who had nephritis and those without clinically apparent nephritis. GBA reactivity was unaffected by DNase treatment of sera, but was partially inhibited by preincubation with dsDNA. These findings are consistent with the hypothesis that some anti-dsDNA antibodies cross-react with glomerular components and that the presence of this cross-reactivity is associated with, and may be responsible for, the development of nephritis. In addition, we have identified a group of SLE patients with renal disease and typical renal histopathology and immune deposits who do not have serum anti-dsDNA antibodies or antibodies that directly bind to glomeruli in the GBA. The mechanism of renal immune deposition in these patients remains to be determined.     

Evidence for a therapeutic effect of plasmapheresis in patients with systemic lupus erythematosus.

Fourteen patients with active systemic lupus erythematosus (SLE) have been treated with plasmapheresis at a rate of two litres daily on three to four days per week, over a period of two to three weeks. Plasma was replaced isovolemically with either fresh frozen plasma or with human plasma protein fractions. Ten patients were receiving treatment with prednisone at the time of plasmapheresis, and four had received no prior treatment. Eight patients showed evidence of either clinical improvement or clinical and immunochemical improvement, at the time of plasmapheresis. In the three patients who showed high levels of circulating complexes before treatment, there was a sudden fall in the level of circulating immune complexes, which was quantitatively greater than could be explained by the amount removed. This suggests that in some patients with SLE, clearance of complexes by the mononuclear phagocytic system is initially blocked by high levels of circulating complexes and that one effect of plasmapheresis may be to relieve this blockade. Five patients showed a clinical response to plasmapheresis despite the fact that tests for immune complexes were negative. Three patients showed no response to plasmapheresis, and three were regarded as unevaluable. In a limited number of patients, who show a high level of circulating immune complexes, and whose condition is deteriorating despite treatment with corticosteroids, there may be an important therapeutic role for plasmapheresis.     

Lamin B autoantibodies in sera of certain patients with systemic lupus erythematosus

Sera from four patients with systemic lupus erythematosus containing antibodies that yield nuclear rim staining of HEp-2 cells by indirect immunofluorescence were identified and characterized. Each serum contained autoantibodies reacting strongly with lamin B on western blots. One of the four sera displayed weaker reactivity with lamins A and C, while the other three displayed only minimal reactivity with lamins A and C. Titers of antilamin antibodies ranged from 1:1,250 to 1:36,250. Two of the sera also reacted at a dilution of 1:20 with cytoplasmic filaments of PTK-2 cells, suggesting that a small fraction of the autoantibodies in these sera may bind to alpha-helical domains of the lamins that are homologous to those of intermediate filaments. The majority of the antilamin antibodies in these patients' sera are specific for portions of the lamin B molecule that are not homologous to lamins A and C, however. The findings suggest that autoantibodies to the nuclear lamina may, in some instances, be responsible for a rim pattern in the fluorescent antinuclear antibody assay. In addition, autoantibodies to the nuclear lamina in sera of certain patients with systemic lupus erythematosus may be useful for defining the molecular structure and biological functions of lamin B, as well as for studying mechanisms of autoimmunity.     

系统性红斑狼疮患者的血清蛋白电泳谱及免疫球蛋白测定分析

目的探讨系统性红斑狼疮(systemic lupus erythematosus,SLE)患者的血清蛋白电泳谱、免疫球蛋白和补体的变化情况.方法 对64例系统性红斑狼疮患者与53例健康者血清进行琼脂糖凝胶电泳及免疫球蛋白和补体测定,并对其各测定结果进行分析.结果 清蛋白、α1-球蛋白、α2-球蛋白、γ-球蛋白、免疫球蛋白IgG、IgA、IgM高于对照组,而C3、C4比对照组低,β-球蛋白无差异.结论 血清蛋白电泳、免疫球蛋白和补体的联合检测对系统性红斑狼疮的诊断和疗效观查有重要意义.     

A serum inhibitor of immune regulation in patients with systemic lupus erythematosus.

Normal mononuclear leukocytes were incubated with serum from patients with active systemic lupus erythematosus (SLE) and healthy subjects and then studied on lymphoproliferative tests. Serum from SLE patients that contained an autoantibody to a subpopulation of thymus-derived (T) lymphocytes inhibited suppressor T-cell activity induced with concanavalin A. These sera did not inhibit lymphoproliferative responses or suppression by monocytoid cells. Mitogen-activated suppressor cells were not inhibited with serum from SLE patients or healthy subjects lacking T-cell autoantibody. This abnormality may contribute to the altered immune response that occurs with SLE.     

Decreased circulating thymus-derived cells with receptors for the Fc portion of immunoglobulin G in systemic lupus erythematosus.

Thymus-derived cells with receptors for the Fc portion of immunoglobulin G (Fcgamma+ T cells) have recently been found to have a suppressor function, a function that is decreased in systemic lupus erythematosus (SLE). Fcgamma+ T cells were found significantly diminished in 21 untreated SLE patients, particularly in the 7 patients who had active disease. Most Fcgamma+ T cells were separated with a subpopulation of T cells with low affinity for sheep erythrocytes. Decrease of this subpopulation was dependent on the decrease in Fcgamma+ T cells. Non-T cells with Fcgamma receptors were also diminished in SLE patients, but their decrease did not correlate with disease activity. The decrease in suppressor-cell function in SLE may be a result of loss, rather than of dysfunction, of the suppressor Fcgamma+ T cells.     

SLE患者血清氧化LDL水平检测分析

目的检测、分析SLE患者血清氧化低密度脂蛋白(ox-LDL)水平。方法采用ELISA检测47例SLE患者和42例健康对照者血清ox-LDL,同时对受检者常规血脂水平进行测定。结果SLE患者ox-LDL水平显著高于对照组[(143.40±65.36)mg/L vs(65.77±26.19)mg/L,P<0.01],总胆固醇、三酰甘油、LDL-胆固醇(LDL-C)和载脂蛋白B(apo B)等常规血脂水平也明显升高。以对照组ox-LDL水平的x-+s值为cut off值,SLE组37例高于此值(敏感性78.7%),健康对照组6例异常(特异性85.7%)。多因素分析显示,患者组ox-LDL水平与LDL-C显著正相关。结论SLE患者血清ox-LDL水平升高,检测ox-LDL水平有助于预测SLE患者As的发生风险。     

抗凝血酶原抗体与系统性红斑狼疮的临床相关性

目的检测抗凝血酶原抗体（aPT）在系统性红斑狼疮（SLE）患者血清中的含量，以进一步辅助SLE的诊断及治疗。方法应用间接ELISA法检测系统性红斑狼疮患者血中的aPT（IgG及IgM型），应用商品抗磷脂抗体筛查试剂盒检测患者的总抗磷脂抗体（IgG及IgM型），同时计算其阳性率和优势比（OR，95％置信区间）。结果30例SLE患者的IgG、IgM、IgG＋IgM-aPL阳性率分别为36．7％、26．7％和46．7％，对照组分别为2．5％、5．0％和7．5％，P〈O．05，两者差异有统计学意义。IgG、IgM、IgG＋IgM-aPT的阳性率分别为20．0％、16．7％和26．7％；健康对照组分别2．5％、2．5％和5．O％，P值均〈O．05，差异有统计学意义，不论抗磷脂抗体（aPL）或aPT在SLE患者中均高于正常人。结论aPL和aPT在SLE患者血清中有较高的水平，检测这些患者血中的aPT及aPL，将有助于进一步完善抗磷脂综合征的临床研究及这些疾病的预防、诊断及治疗。     

The detection of circulating immune complexes containing immunoglobulin G

An immune complex assay using radiolabelled immunospecific antibodies against human IgG and polyethylene glycol precipitation (IgG-PEG assay) is described. The material reactive in this assay was evaluated using aggregated immunoglobulins, immune complexes prepared in vitro, sera of patients with a variety of disorders and normal human serum. Sucrose density gradient ultracentrifugation showed that only large-sized immune complexes (greater than 25 S) were detected. Comparison of the results of the IgG-PEG assay with those of the C1q binding assay showed a highly significant positive correlation (p less than 0.001). It was found that rheumatoid factors do not influence the results of the IgG-PEG assay. The method described in this study detects specifically immune complexes containing IgG and might be extended to the detection of other constituents of circulating immune complexes.     

系统性红斑狼疮患者外周血淋巴细胞G蛋白—cAMP信号机制的研究

目的 研究系统性红斑狼疮（SLE）患者外周血淋巴细胞（PBL）增殖反应与cAMP的关系以及G蛋白介导机制,方法 淋巴细胞增殖法,cAMP放兔测定及G蛋白功能分析。结果 SLE患者PBL增殖反应增强与其cAMP水平的降低有关,百日咳毒素（PT）可部分升高cAMP水平。结论 PT敏感的Gi蛋白-cAMP信号机制参与了SLE患者PBL的异常增殖反应。     

Listerial infections in patients with systemic lupus erythematosus.

Infection with Listeria monocytogenes is known to occur more frequently in immunosuppressed patients, including those receiving high-dose prednisone or cytotoxic therapy for collagen vascular disease. We reviewed three cases of listeriosis and systemic lupus erythematosus (SLE) seen at our institution, in addition to five cases reported in the English literature. Seven of the eight patients had non-CNS listerial infections. All patients but one had associated risk factors of either renal failure or pregnancy. From our review, we found that listeriosis is uncommon in SLE, and patients without renal failure or pregnancy do not seem to be at increased risk for listeriosis. Although most patients were treated with high-dose prednisone, with or without cytotoxic drugs, the role of immunosuppression by these drugs as a risk factor for listeriosis remained unclear.     

Interaction between anti-DNA and anti-DNA-binding protein autoantibodies in cryoglobulins from sera of patients with systemic lupus erythematosus

We have previously shown that sera from some patients with SLE and related disorders contain autoantibodies to a DNA-binding protein complex designated p70/p80. The present study shows that anti-p70/p80 autoantibodies are frequently accompanied by anti-DNA antibodies and cryoglobulins. When the cryoglobulins were isolated, they were found to be specifically enriched in both anti-p70/p80 and anti-DNA activities. The anti-p70/p80 and anti-DNA antibodies were found to be distinct populations of autoantibodies rather than a single crossreactive species, since they could be separated from one another by chromatography on DNA-cellulose. Certain human anti-DNA mAbs could inhibit the binding of autoimmune polyclonal anti-p70/p80 antibodies to p70/p80, suggesting that anti-DNA antibodies might also associate with the variable regions of some anti-p70/p80 antibodies in the cryoglobulins. Binding of one murine anti-p70/p80 mAb (111-12) also was inhibited by certain human anti-DNA mAbs, but the binding of another murine mAb (162-11) to a different epitope of p70/p80 was not. These studies suggest that certain anti-DNA antibodies may interact with the variable regions of a population of anti-p70/p80 antibodies. The cryoglobulins found in the sera containing both anti-p70/p80 and anti-DNA antibodies may represent immune complexes consisting, in part, of idiotype and antiidiotype.