Search

In the Bernard-Soulier syndrome (BSS), the giant platelets are said to have increased phosphatidylserine (PS) surface exposure in the resting state and shortened survival in the circulation. When normal platelets are activated, they undergo many biochemical and morphological changes, some of which are apoptotic. Herein, we investigated apoptotic-like events in BSS platelets upon activation, specifically, PS exposure, microparticle (MP) formation, cell shrinkage, and loss of mitochondrial inner membrane potential (DeltaPsi(m)). Platelets from two unrelated BSS patients were examined in whole blood; agonists used were collagen, thrombin, PAR1- or PAR4-activating peptides (APs), or combinations of collagen with thrombin, and the PAR-APs. Flow cytometry was used to measure PS exposure (annexin A5 binding), platelet-derived MPs (forward scatter; events

Indirect platelet radioactive antiglobulin tests using anti-IgG and anti-C3 were performed in 483 individuals. The subjects' diagnoses and clinical states were correlated with sensitizing protein (IgG and/or C3). Sensitivity of the technique was increased by increasing the serum to cell ratio and there was good correlation between direct and indirect antiglobulin tests in 56 patients. Seven percent of normal healthy non-thrombocytopenic untransfused males and 88% of patients in whom autoimmune thrombocytopenia was clinically suspected showed a positive test result. In the indirect antiglobulin test, mean +/- SD fg anti-C3d/platelet was 1.75 +/- 0.28 (N = 58) with serum from normal subjects, 3.91 +/- 1.86 (N = 123) with serum from patients with idiopathic thrombocytopenic purpura and systemic lupus erythematosus, and 2.78 +/- 0.91 (N = 52) with serum from patients with lymphoproliferative disease. The differences were significant with P less than 0.001. Levels of bound anti-IgG were in general accord with those of other reports; there was no significant relationship with serum immunoglobulin levels. When the indirect antiglobulin tests were positive for both IgG and C3, there was a significant correlation between IgG and C3 bound to normal platelets (r = 0.671; P less than 0.01). Although sera from 39% of patients whose thrombocytopenia was not initially suspected to be on an autoimmune basis gave a positive test result, indirect platelet radioactive antiglobulin tests using anti-IgG and anti-C3 may be convenient tools in the evaluation and follow-up of immune thrombocytopenias.

With platelet activation, there is modulation of platelet surface molecule expression. In flow cytometric analyses of in vivo platelet activation, results are often confounded by activation induced in vitro by the preparative procedures. It is particularly important therefore to prevent or retard platelet activation as soon as possible after withdrawal of the blood sample. Taking blood into paraformaldehyde, or fixing the cells with paraformaldehyde as soon as possible after withdrawal, has been employed to prevent platelet activation in vitro, but paraformaldehyde-fixed platelets cannot be further used in functional studies. We investigated the efficacy of Diatube-H, a commercially available combination of platelet antagonists (theophylline, adenosine, and dipyridamole), in preventing or retarding platelet activation in vitro, along with its effects on modulation of platelet membrane glycoproteins (GP) and adhesion molecules. In contrast to blood taken into EDTA, blood taken into Diatube-H vacutainer tubes could be stored at room temperature for up to 4 hr prior to paraformaldehyde fixation without significant in vitro platelet activation, as measured by CD62P, CD63 and modulation of GPIb and GPIIbIIIa surface expression. Hence, paraformaldehyde fixation could be deferred for several hours, permitting transport of samples from distant sites. Studies of thrombin-induced platelet activation indicated that platelets taken into Diatube-H remained functional i.e. were able to be activated. Expression of the CD29, CD49b and CD31 adhesion molecules on the platelet surface was unaffected by storage in Diatube-H. The results suggest that Diatube-H may be a useful reagent for flow cytometric studies of platelets when the samples cannot be processed immediately.

Intravenous anti-D is often used in the treatment of autoimmune thrombocytopenic purpura (AITP), but little is known about its mechanisms of action. To investigate anti-D's potential in vivo mechanism(s) of action, a small group (N = 7) of children with chronic AITP was studied. The children initially received either 25 or 50 microg/kg of WinRho-SD in a four-cycle cross-over trial, and peripheral blood samples from the first and third cycles were assessed for cytokine levels at pre-treatment, 3 hr, 1 day, and 8 days post-treatment. Results showed that platelet counts significantly increased in all the children by day 8 post-treatment. Analysis of serum by ELISA showed that there was a significant but transient rise in both pro- and anti-inflammatory cytokine/chemokine levels (e.g., IL1RA, IL6, GM-CSF, MCP-1 alpha, TNF-alpha and MCP-1) by 3 hr post-treatment in both cycles which returned to baseline levels by 8 days post-treatment. These results suggest that anti-D administration may initially activate the RES in the form of cytokine/chemokine secretion, which is subsequently followed by an increase in platelet counts. It is possible that the induced cytokine/chemokine storm may have an effect on several physiological processes such as those mediating either adverse effects or potentially RES phagocytic activity.

Natural killer (NK) cell activity was studied in 17 patients with primary chronic idiopathic autoimmune thrombocytopenic purpura (ATP). Fifteen of 17 patients had a significantly reduced NK cytotoxicity against 51chromium labeled K562 target cells (mean LU20% = 18 +/- 20 in patients versus 65 +/- 25 in controls, P less than 0.001). NK activity was also significantly reduced in all of six patients with secondary ATP as compared with normal controls (LU20% 28 +/- 15, respectively, P less than 0.005). The NK activity in both patient groups correlated with the duration of therapy being received (r = 0.60, P less than 0.001). Immunophenotypic analysis of peripheral blood mononuclear cells from patients with ATP revealed that CD8- cells bearing CD57 (HNK-1, Leu 7) and CD3- cells bearing CD56 (Leu 19) were quantitatively within the normal range. These findings indicate that patients with ATP have a functional defect in NK cytolytic activity.

This report describes a patient with immunoblastic sarcoma and autoimmune hemolytic anemia with reticulocytopenia and red cell hypoplasia. Both IgG and IgM red cell autoantibodies were present in the patient's serum, and both antibodies had the uncommon specificity of anti-IT. This report appears to be the first to describe IgG auto anti-IT and autoimmune hemolytic anemia in a patient who did not have Hodgkin's disease; the laboratory findings and clinical course are described. The reticulocytopenia and red cell hypoplasia may have been immunologically mediated. The patient responded to plasma exchange, corticosteroids, and chemotherapy.

Fifty-nine HIV-1 antibody positive and 58 antibody negative hemophiliacs were evaluated over a 2 year study period to gain insight into the natural history and prognosis of HIV-1 disease in members of this risk group. Mean CD4 (Leu 3+) cell counts calculated at 6 month intervals decreased gradually in seropositive patients (from 403 to 311/microliters) whereas CD8 (Leu 2+) counts remained stable but above the normal range. CD4 cell counts correlated closely with advancing CDC clinical stage; CD8 numbers showed no such association, but were markedly lower in the six patients with overt AIDS. Serum P24 antigenemia was associated with low CD4 cell counts and with advanced clinical stage (58% of antigenemic and 14% of non-antigenemic seropositive patients were in stage IV). In addition to CD4 cell counts, significant reductions in Leu 11+ natural killer cell (NK) subsets and in Leu 3 + 8 - cells occurred in seropositive patients over the study period; Leu 2 + DR + cells increased significantly. When expressed as a percentage of lymphocytes, the reduction in Leu 19 + NK cells was also significant, as were the increases in Leu 4 + DR + cells and Leu 12 + 8 + B cells. In summary, declining CD4 cell numbers and percentages are valuable markers of progressive HIV-1 disease in hemophiliacs, but may not always accurately reflect the degree of disease activity. Progressive changes in additional variables such as serum P24 antigen, and numbers and percentages of NK cell subsets and (as AIDS supervenes) CD8 cell numbers, may allow more precise monitoring of HIV-1 disease. This will, in turn, facilitate the design of optimal individualized strategies for therapeutic intervention.

This report describes a patient with recurrent thrombotic thrombocytopenic purpura (TTP) in whom complement activation was observed during the acute episodes. Serum C3, C4, and CH50 were reduced, and there was deposition of C3 on red cells and platelets. These findings returned to normal during clinical remission.