Idiopathic thrombocytopenic purpura (ITP) is characterized by the development of a specific anti-platelet autoantibody immune response mediating the development of thrombocytopenia. Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of a wide variety of autoantibodies. In 15-20% of SLE cases, patients develop thrombocytopenia which appears to be autoimmune in nature (SLE-TP). To better understand the pathogenesis of the thrombocytopenia associated with SLE, we investigated the overlapping platelet and cellular immune features between SLE and ITP. Thirty-one patients with SLE, eight with SLE-TP, and 17 with ITP, were studied and compared to 60 healthy controls. We evaluated platelet-associated IgG, platelet microparticles, reticulated platelets, platelet HLA-DR expression, in vivo cytokine levels, lymphocyte proliferation, and the T lymphocyte anti-platelet immune response in these patients. Patients with SLE-TP and those with ITP had increased platelet-associated IgG, an increased percentage of platelet microparticles, a higher percentage of reticulated platelets and larger platelets, suggesting antibody-mediated platelet destruction and increased platelet production. More than 50% of patients with ITP had increased HLA-DR on their platelet surface whereas subjects with SLE-TP did not. Analysis of serum cytokines demonstrated increased levels of IL-10, IL-15 and TNF-alpha in patients with SLE, but in those with ITP, only increased levels of IL-15 were seen, no increases in any of these cytokines were observed in patients with in SLE-TP. The ability of lymphocytes to proliferate in response to phorbol myristate acetate (PMA) stimulation was increased in SLE-TP, but was normal in both SLE and ITP. Lymphocytes from subjects with ITP displayed an increased ability to proliferate on exposure to platelets, in contrast, those with SLE-TP did not. While the number of subjects evaluated with SLE-TP was small, these data reveal a number of differences in the immunopathogenesis between SLE-TP and ITP.

Adult↗

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Characterization of red blood cells strongly coated in vitro by C3 via the alternative pathway.

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Agglutination Tests↗

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Differences in specificities of anti-C3d sera raised to C3d antigens prepared in different ways.

Anti-C3d sera were raised to different C3d antigens or to the same C3d antigen by different methods. Although identical by immunoprecipitation studies, the various anti-C3d sera showed differences in specificities against bound C3d antigen. Such differences were observed wih red blood cells coated with C3d in vivo and in vitro. Antisera made to the C3d-KAF antigen detected fewer molecules/cell on C3d-tryp cells than did antisera made to the C3d-tryp antigen. The converse was true for C3d-KAF cells. "Saturation" experiments indicated that different anti-C3d detected different "subpopulations" of bound C3d. C3d bound to red blood cells in vivo was, in at least one case, detectable by some anti-C3d sera but not by others. Such differences in anti-C3d specificity may be important in determining the optimal characteristics of anti-C3d antiglobulin serum for routine laboratory use.

Animals↗

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Red blood cell-bound C3d in normal subjects and in random hospital patients.

Red blood cells (RBC) drawn into EDTA from 103 normal adults, 26 newborn infants, and 203 randomly selected hospital patients, and into citrate-phosphate-dextrose (CPD) from 87 Red Cross donors, were evaluated for RBC-bound C3d by agglutination with anti-C3d and by a radiolabeled anti-antiglobulin technique. Positive agglutination reactions were observed only with RBC from 26 hospital patients; however, by the radiolabeled antiglobulin method, C3d was demonstrable on all RBC, but its amount varied widely among different subjects. A normal range for RBC-bound C3d was established. RBC-bound C3d levels were not related to the age (18-88 years) or sex of the subjects and remained unchanged over 9 months. Red blood cells taken into CPD had more RBC-bound C3d than cells taken into EDTA; cord RBC had less than adult RBC. In 39 percent of the hospital patients (excluding those with autoimmune hemolytic anemia) RBC-bound C3d was above the normal range and a good correlation was found between values obtained by the radiolabeled anti-antiglobulin method and by agglutination titration scores. There was a threshold level of RBC-bound C3d below which agglutination reactions were negative. Evaluation of RBC-bound C3d in health and disease is important for determining optimal characteristics of anticomplement antiglobulin reagents.

Adolescent↗

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Red blood cell-bound C3d in selected hospital patients.

A radioactive anti-antiglobulin technique was used to measure C3d bound to the red blood cells of 227 hospitalized patients in 129 patients, with a wide variety of diseases, normal levels of RBC-bound C3d were found. Seventy-two patients had moderately elevated RBC-bound C3d; they generally did not have autoimmune hemolytic anemia but had diseases in which complement is thought to be activated. Patients with markedly elevated RBC-bound C3d (26 patients) usually had autoimmune hemolytic anemia with a positive antiglobulin test. Some patients with only moderately elevated levels of RBC-bound C3d had autoimmune hemolytic anemia and a negative antiglobulin test in individual patients the level of RBC-bound C3d correlated with both the severity of disease and the response to treatment. RBC-bound C3b was detected in two patients with a very high level of RBC-bound C3d. This study provides background data for assessing the significance of complement activation and fixation to RBC in health and disease.

Anemia, Hemolytic, Autoimmune↗

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Membrane-bound immunoglobulins and complement components on young and old red cells.

In order to characterize changes in membrane-bound immunoglobulins and complement components, red cells (RBCs) were separated into young and old populations by simple centrifugation. Old RBCs had reduced mean corpuscular hemoglobin volume, increased mean corpuscular hemoglobin concentration, and reduced sialic acid. Using radioactive anti-antiglobulin techniques, old RBCs were shown to have more IgG, IgM, IgA, and C3d on their surfaces than did young RBCs; there was no increase on old RBCs of C3b, factor B, C4b, or C5. Similar results were observed with RBCs strongly coated with C3d in vivo from a patient with cold agglutinin disease. RBCs taken into ethylenediamine tetraacetate, washed thoroughly in saline, and then stored for prolonged periods in Alsever's solution or kept in autologous ethylenediamine tetraacetate plasma, at 4 degrees C, showed no increase in RBC-bound C3d with increased storage time. If, however, blood was taken into citrate-phosphate-dextrose and maintained at 4 degrees C in autologous plasma, a significant increase in RBC-bound C3d was observed in the mixed-cell population with prolonged storage time. Order donor blood units, taken into citrate-phosphate-dextrose and stored at 4 degrees C as packed red cells, showed higher levels of RBC-bound C3d in the mixed-cell population than did units stored for a shorter time. In no case did donor unit RBCs give a positive direct antiglobulin test on serologic testing with anti-C3d. The findings complement data already collected on membrane and cytoplasmic changes in aging RBCs and may contribute to an understanding of RBC senescence.

Anticoagulants↗

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Comparison of a modified manual hexadimethrine bromide (Polybrene) and a low-ionic-strength solution antibody detection technique.

Manual hexadimethrine bromide (Polybrene) tests (Polybrene in low-ionic medium) were used in parallel with manual low-ionic-strength solution (LISS) procedures for the routine testing of patient samples referred to a general hospital blood bank. Of 5646 consecutive sera tested, 5167 (91.5%) did not react with either technique; 320 sera (5.7%) reacted in both methods. The Polybrene technique detected 63 antibodies which did not react in the LISS methods. One hundred sera did not react in the Polybrene test, but did react in the LISS methods. Sera showing discrepant results between the two methods were further tested in a reference laboratory. Polybrene tests appeared to be better in avoiding reactions due to clinically nonsignificant antibodies. The LISS methods, however, appeared to be more sensitive in detecting antibodies of potential clinical significance.

Blood Banks↗

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Autoimmune hemolytic anemia with concurrence of warm and cold red cell autoantibodies and a warm hemolysin.

This report describes the laboratory findings and clinical course of a patient with thrombophlebitis, venous gangrene, and autoimmune hemolytic anemia. Three concomitant red cell autoantibody activities were detected: a low-titer, high-thermal-amplitude, IgM anti-I cold agglutinin; an IgG warm 'incomplete' panagglutinating autoantibody; and an IgM warm hemolysin.

Anemia, Hemolytic, Autoimmune↗

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Hemolytic warm IgM autoagglutinins in autoimmune hemolytic anemia.

The authors report a patient with fulminant autoimmune hemolytic anemia due to a rare warm IgM autoagglutinin more reactive at 37 degrees C than at lower temperatures and secondary to systemic lupus erythematosis. The patient's clinical course and the serologic and immunochemical characteristics of the antibody are described, including the possibility that transfusions of small amounts of incompatible red cells may have contributed to the hemolysis. The consequences of using the initial serologic test results as the basis for therapy are discussed.

Agglutinins↗