Determination of antiplatelet antibodies on the surface of platelets by direct radioimmune method in patients with different types of immune thrombocytopenia

Direct radioimmune assay (RIA) have been developed for detection of antibodies associated wild platelet membrane. Platelets from 12 patients with idiopathic thrombocytopenic purpura (ITP) and 27 patients with chronic lymphocytic leukemia (CLL) (platelet count (100,000 in 1 microliters) have been tested. Antibodies on platelets surface have been detected in all 12 patients with ITP and in 21 patients with CLL. In 6 CLL patients the number of immunoglobulins associated with platelets surface does not increase control level. It is possible, that in some CLL patients development of thrombocytopenia is mediated not only by platelet associated antibodies but by other mechanisms, one of which can be linked with the depression of megakaryocytes growth in bone marrow. Direct RIA for measurement of antibodies on platelet surface detect antiplatelet antibodies with higher frequency than indirect enzyme-linked-immunosorbent-assay (ELISA), developed earlier for assessment of antiplatelet antibodies in serum. Increase of platelet count in CLL patients after steroid and cytostatic treatment correlated with the decrease of platelet surface associated antibodies.     

Autoantibodies in chronic ITP

Chronic ITP is a syndrome of destructive thrombocytopenia due in most cases to antiplatelet autoantibodies. In the present studies we have studied 74 patients with chronic ITP using a new immunobead assay. Of these, 59 (79.7%) had demonstrable platelet-associated autoantibodies: 48 against platelet glycoprotein IIb/IIIa and 11 against glycoprotein Ib/IX. Plasma autoantibodies were studied in all patients and 32 (43.2%) had positive results; in each case the patient also had platelet-associated autoantibodies directed to the same antigen. We conclude that the majority of patients with chronic ITP have autoantibodies against platelet membrane glycoproteins and that the immunobead assay is a sensitive and reproducible method for their detection which is applicable to the routine hospital laboratory.     

Effect of six weekly transfusions on canine marrow grafts: tests for sensitization and abrogation of sensitization by procarbazine and antithymocyte serum.

We have previously shown that blood transfusions can immunize a dog and lead to rejection of a subsequent marrow graft despite lethal total body irradiation (TBI). Sensitization to histocompatibility antigens induced by two prior transfusions of whole blood could be overcome by a regimen of procarbazine and anti-thymocyte serum (ATS) preceding TBI. The current study investigated a) whether this regimen could abrogate sensitization induced by six weekly transfusions given from days --50 to --15 preceding a marrow graft, and b) whether platelet survival studies and two in vitro tests of immunity could predict marrow graft rejection. All donor-recipient pairs were histoincompatible, unrelated, and of different breed. Twenty-two recipients received platelet concentrate transfusions and eight received whole blood transfusions. Recipients were given 1200 R TBI and a graft of marrow and peripheral blood leukocytes from the transfusion donor on day 0. Three of 15 recipients (20%) given procarbazine, 12.5 mg/kg i.v. on days --8, --6, and --4, and ATS, 0.6 ml/kg subcutaneously on days --7, --5 and --3, Rejected their grafts, whereas 11 of 15 dogs (73%) not given procarbazine and ATS rejected their grafts (p less than 0.01). Serum lymphocytotoxic antibodies, peripheral leukocyte migration inhibition, and in vivo donor platelet recovery and survival were studied in those recipients receiving six weekly transfusions and in 18 other recipients receiving a single donor transfusion 3 months before marrow grafting. No significant correlation was found among these in vitro and in vivo tests of sensitization. Sensitization to marrow grafts was not reliably detected by the presence of cytotoxic antibodies or leukocyte migration inhibition. Platelet survival, however, was positively correlated with the results of marrow grafting in 12 of 15 (80%) evaluable recipients (p approximately 0.15).     

Characteristics of Helicobacter pylori-induced gastritis and the effect of H. pylori eradication in patients with chronic idiopathic thrombocytopenic purpura

BACKGROUND: The association between Helicobacter pylori infection and idiopathic thrombocytopenic purpura (ITP) has been reported widely. We investigated the prevalence of H. pylori infection, its virulence profile and the effectiveness of its eradication in patients with ITP. MATERIALS AND METHODS: Twenty patients with ITP, 20 with peptic ulcer (10 gastric ulcer (GU), 10 duodenal ulcer (DU)) and 20 with NUD were studied. The virulence profile of the strains was assessed by genotyping for cagA, vacA, iceA, and hpyIIIR/hrgA and by assaying for IL-8 and DNA fragmentation after incubation with AGS cells. Infected patients and two uninfected ITP patients received triple therapy and platelets were counted before and 1 month, 6 months, 1 year, and 2 years after eradication therapy. RESULTS: H. pylori infection was found in 17 ITP (85%), 20 ulcer (100%) and 13 NUD (65%) patients. Biopsies and strains were collected from five ITP, 20 ulcer and 13 NUD patients. The ITP patients had a pangastritis or corpus-predominant gastritis pattern. All H. pylori isolates, from ITP, ulcer and NUD patients, were cagA(+) and vacA s1/m1, and did not differ in levels of IL-8 induction or DNA fragmentation. Fifteen ITP (88%) and 17 ulcer (85%) patients had successful eradication of H. pylori. Ten of these 15 (67%) H. pylori-eradicated ITP patients had platelet recovery. There was no significant change in platelet count in the two ITP patients in whom eradication failed or in the two originally H. pylori-uninfected ITP patients, or in the treated ulcer patients. Age at onset of ITP was the main determinant of platelet recovery: 100% of patients diagnosed after the age of 60 recovered compared with only 22% of those diagnosed before 50. CONCLUSIONS: H. pylori-infected ITP patients have a corpus-predominant pattern of gastritis but the virulence profile of their strains does not differ from that of ulcer or NUD patients. Eradication of H. pylori infection is a good therapeutic option for some patients with chronic ITP, especially for those who develop ITP in older age.     

Severe thrombocytopenia in Epstein-Barr virus-induced mononucleosis.

Severe thrombocytopenia is a rare complication of Epstein-Barr virus-induced infectious mononucleosis. We evaluated the clinical and laboratory data from seven patients seen between 1976 and 1985 whose lowest platelet counts varied from 3 to 25 x 10(9) per liter. Five of the seven patients were initially thought to have either acute leukemia or idiopathic thrombocytopenic purpura; eventually, however, primary Epstein-Barr virus infections were confirmed in all patients. Two of six patients tested had antiplatelet antibodies during the acute phase of their illnesses. Eight additional patients with acute disease who had only mild thrombocytopenia (94 to 144 x 10(9) per liter) were also tested for platelet antibodies with negative results. Steroid therapy was administered to three patients and platelet transfusions to one. All seven patients recovered with no serious hemorrhagic sequelae.     

Efficacy of Amifostine in Treating Patients with Idiopathic Thrombocytopenia Purpura

Idiopathic Thrombocytopenic Purpura (ITP) is an autoimmune disease characterized by the production of antibodies against platelet surface antigens, resulting in platelet destruction. ITP is generally treated using glucocorticoids, splenectomy, immunosuppressants, platelet transfusions, and also rituxan and rituximab. However, as these treatments are not effective in some refractory ITP patients, especially the elderly, who are also at greater risk of cerebral hemorrhage, we have undertaken this study to find a safe and effective way of treating these patients. In a clinical protocol, we have examined the efficacy of the cytoprotective adjuvant, amifostine, on 24 ITP patients, consisting of 21 Chinese (age: 13–92 years), and 3 Caucasians (age: 46–73 years). In order to prevent the side effects associated with amifostine treatment, an alternative dosing and anti-emetic regimen was developed as part of this protocol, which significantly improved patient acceptance. The protocol consisted of daily intravenous infusions of amifostine 5 × 400 mg per week, for a total of 4–5 weeks. All the patients experienced a long-lasting and continuing remission, defined as platelet counts greater than 100,000. Two patients relapsed: one after an upper respiratory tract infection, and another due to Helicobacter pylori. However, both these patients had complete remission, after they were treated again with amifostine. In this clinical study, we report for the first time, the successful use of amifostine for ITP treatment in refractory patients. In conclusion, amifostine may have good therapeutic effect on ITP patients, especially in refractory and/or elderly. The long-term clinical outcome and the mechanism of action of this drug still need further investigation.     

ITP in pregnancy: use of the bleeding time as an indicator for treatment

ITP is a relatively common disorder seen in pregnancy. Current recommendations for management of patient with ITP recommend maintaining the platelet count above 50 x 10(9)/L and the bleeding time less than 20 min. It has been well documented that the bleeding time in ITP is disproportionately shortened in many patients relative to the platelet count. We present a prospective study of 24 ITP patients in whom the bleeding time was used as an indicator for therapeutic intervention in pregnancy. Indications for therapy with prednisone and/or intravenous gammaglobulin were the following: significant clinical hemorrhage due to thrombocytopenia; bleeding time of greater than 20 min at the baseline platelet count; for normalization of hemostasis prior to delivery or surgical procedure. Caesarean section was performed only in cases in which there were obstetrical indications for this mode of delivery or when the fetal platelet count (obtained by fetal scalp vein sample) was less than 50 x 10(9)/L. Of 24 patients with ITP, eight had significant thrombocytopenia (platelet count less than 50 x 10(9)/L) throughout pregnancy. Only two patients required prolonged prednisone therapy. Both suffered side effects of chronic prednisone administration. Four patients were treated with prednisone for a short course (10-14 days) at term to improve hemostasis for delivery. One patient was treated with intravenous gammaglobulin at term in an effort to prevent severe neonatal thrombocytopenia. Seven patients required caesarean section; the remaining 17 patients underwent vaginal delivery. Only one minor bleeding complication was seen - a small wound hematoma post caesarean section. In summary, using the bleeding time as an indicator for therapeutic intervention, treatment of ITP in pregnancy can be minimized. Thus, therapy related toxicity can be avoided.     

Gammaglobulin treatment and anti-IgA antibodies in IgA-deficient patients.

Antibodies to IgA may cause severe anaphylactic reactions during blood transfusions. Tests for anti-IgA antibodies were carried out on six patients with IgA deficiency (five of whom also had hypogammaglobulinaemia) who had received continuous gammaglobullin treatment for chronic or recurrent infections for three to eight years. Three patients had minute amounts of IgA, and three had none (less than 0.01 microgram/ml). Only one patient had anti-IgA. Her antibody titre did not change during treatment. No patient had any untoward effects of treatment, which relieved the symptoms of infection in every case. IgA determinations should be performed by more accurate methods than radial immunodiffusion when evaluating the risks of giving gammaglobulin to patients with hypogammaglobulinaemia and IgA deficiency. Probably the stimulus provided by intramuscular gammaglobulin in such patients is insufficient for the formation of anti-IgA antibody.     

不同剂量西洛他唑强化抗血小板对冠脉支架术后血小板高反应性的影响

目的：观察临床应用不同方案强化抗血小板治疗改善冠脉支架术后血小板高反应性的可行性、安全性及有效性。方法：选择2009年3月至2011年2月在沈阳军区总医院、中国医科大学第一附属医院、解放军第463医共入选560例冠脉支架术后血小板高反应性（HPR Highon-treatment Platelet Reactivity）患者,在给予阿司匹林300mg／天,氯吡格雷150mg／天,3天后HPR仍未缓解者,随机分为两组,一组在强化抗血小板治疗即阿司匹林300mg,氯吡格雷150mg的基础上加用小剂量西洛他唑（50mg,2／日）,另一组在标准两联方案即阿司匹林300mg,氯吡格雷75mg的基础上加用西洛他唑100mg,2／日,3天后测定HPR的缓解情况。结果：大剂量氯吡格雷治疗3天后HRP的缓解卒为54-3％（304／560）,接受不同西洛他唑剂量治疗3天后又有58．6％的患者HPR缓解,但是西洛他唑50mg组和100mg组HRP缓解率无差别（59．4％VS57．8％,P=0．80）。两组患者30天随访均无死亡及卒中事件,无主要及次要出血事件。结论：强化抗血小板治疗可改善冠脉支架术后的血小板高反应性且未增加出血风险,但其临床获益还需更长时间的随访结果进一步明确,两种强化抗血小板治疗方案对改善冠脉支架术后HPR的作用相似。     

CD34(+)-selected stem cell boost for delayed or insufficient engraftment after allogeneic stem cell transplantation

BACKGROUND: Poor graft function without rejection may occur after stem cell transplantation (SCT). CD34(+) stem cell boost (SCB) can restore marrow function but may induce or exacerbate GvHD. We therefore investigated the feasibility and efficacy of CD34(+)-selected SCB in some patients with poor graft function. We present the results for eight patients (median age 46 years) transplanted initially for myelofibrosis, acute leukemia, myeloma and NHL. Six patients had received HLA-matched and two mismatched grafts (PB, BM; n=5, 3). After a median of 128 days post-transplant, the median leukocyte and platelet counts were, respectively, 2.05/nL and 18/nL. None had achieved platelet counts >50/nL even though donor chimerism was >95% in seven recipients. METHODS: Positive selection of CD34(+) stem cells was performed on a CliniMACS device, observing GMP and achieving a median of 98.5% purity. The patients received a median of 1.7 x 10(6)/kg CD34(+) cells and 2.5 x 10(3)/kg CD3(+) T lymphocytes. RESULTS: Hemograms at days +30, +60 and +90, respectively, showed steadily increasing median leukocyte (2.55, 3.15 and 4.20/nL) and platelet (29, 39 and 95/nL) counts. After a median follow-up of 144 days, five patients remained alive. No patient had developed acute or chronic GvHD. One patient died of leukemic relapse and two others of systemic mycosis. DISCUSSION: These preliminary results point to the possibility of safely improving graft function using CD34(+) positively selected stem cells without necessarily increasing the incidence of GvHD in patients with poor graft function post-SCT. Experience with more patients and longer follow-up should clarify the optimal role for this procedure.     

Prediction of the effect of immunoglobulin therapy in ITP

The correlation between the response to high-dose immunoglobulin therapy (IVIg) and the sequestration pattern of Indium-labeled platelets (In-PLT) in the body was studied in 9 patients with chronic idiopathic thrombocytopenic purpura (ITP). Patients that has prominent platelet sequestration in the spleen responded to IVIg. In these patients, splenic sequestration decreased by 20-30% after IVIg without significant changes in hepatic sequestration. This finding suggests that the blocking of splenic Fc receptors with immunoglobulin minimized the destruction of sensitized platelets. However, patients who had almost equal platelet sequestration in the liver and spleen did not respond to IVIg. In these patients, hepatic sequestration decreased after IVIg, whereas splenic sequestration increased. Thus, it appears that estimating the platelet sequestration pattern using In-PLT is useful for predicting the effects of IVIg.     

Advances in ITP--therapy and quality of life--a patient survey

Background

Current guidelines recommend glucocorticoids and splenectomy as standard 1^st and 2^nd line treatments for chronic immune thrombocytopenia (ITP). We sought to find out how German ITP-patients are treated with respect to these guidelines.

Methods

Members of a patient support association ≥18 years with a self-reported history of chronic ITP>12 months were surveyed with a web-based questionnaire.

Results

122 questionnaires were evaluated. 70% of patients had chronic ITP for more than 5 years and 20% an average platelet count of ≤30·10⁹/L. 41% of the patients reported haematomas or petechiae more than once or twice and up to 12 times or more per year and 17% oropharyngeal and nasal bleeds. 11% had been admitted to hospital during the last 12 months. 88% had received or currently receive glucocorticoids, 27% were splenectomised. IVIG had been given to 55%, rituximab to 22%, anti-D to 12%, ciclosporin to 7%, while complementary and alternative medical treatments had been used by 36%. 50 women responded to questions concerning pregnancy. 14 (28%) had been advised not to become pregnant. 23 reported pregnancies and 10 (44%) required treatment for their ITP during pregnancy.

Conclusion

Glucocorticoids are the most common therapy for chronic ITP but complementary and alternative treatments already come second and less than ⅓ of patients are splenectomised. This and the frequent use of complementary medicines suggests patients'' dissatisfaction with conventional approaches. Many patients receive off-label therapies. There is a major need for adequate counselling and care for pregnant ITP-patients.     

Lupus anticoagulant and antiplatelet properties of human hybridoma autoantibodies

The clinical association of lupus anticoagulant antibodies with thrombocytopenia and thrombosis was the rationale for investigating the in vitro reactivity of these human hybridoma lupus anticoagulant antibodies with platelets. Fifty human hybridoma antibodies from 13 patients with systemic lupus erythematosus, 2 women with multiple spontaneous abortions, and 4 normal individuals were analyzed for lupus anticoagulant, antiplatelet, anti-DNA, and antiphospholipid reactivities. Of the hybridoma antibodies studied, 25 had lupus anticoagulant activity, 21 had antiplatelet reactivity, and 7 of these antibodies had both lupus anticoagulant and antiplatelet properties. No correlation was found between lupus anticoagulant antibody activity and antiplatelet, anti-denatured DNA, anticardiolipin, anti-egg phosphatidylethanolamine, antiphosphatidylserine, antiphosphatidylinositol, and antiphosphatidylcholine reactions. In contrast, antiplatelet activity was strongly correlated with antiphosphatidylethanolamine (rho = 0.761, p less than 0.001), anticardiolipin (rho = 0.748, p less than 0.001), and anti-dDNA (rho = 0.745, p less than 0.001) reactivities. Pretreatment of platelets with deoxyribonuclease, ribonuclease, trypsin, or phospholipases A2 and C resulted in different effects on the binding of individual hybridoma antibodies to platelets, suggesting that antiplatelet antibodies may recognize different epitopes on the platelet membrane. Our data demonstrate that most hybridoma lupus anticoagulant antibodies did not bind directly to platelets in vitro. This suggests that additional serum factors may be required in vivo to explain the association of these antibodies with thrombocytopenia and thrombosis.     

The disulfide-rich region of platelet glycoprotein (GP) IIIa contains hydrophilic peptide sequences that bind anti-GPIIIa autoantibodies from patients with immune thrombocytopenic purpura (ITP)

Immune thrombocytopenic purpura (ITP) is an autoimmune blood disease caused by autoantibody-mediated destruction of blood platelets. Platelet glycoprotein (GP) IIb/IIIa is a common target for antiplatelet autoantibodies. The present studies were undertaken (1). to confirm whether the disulfide rich repeat region of GPIIIa contains target epitopes for antiplatelet antibodies in patients with ITP; (2). to determine whether these antigens were defined by peptide sequences in the absence of post-translational modification; and (3). to correlate observed immunologic reactivity with the recently solved X-ray crystallographic structure of an analogous integrin complex, the vitronectin receptor, alpha(V)beta(3). Recombinant fusion proteins of four GPIIIa extracellular sequences were prepared and purified. Immunoblotting results with purified recombinant peptides showed potent reactivity of 16 of 24 ITP patient serum anti-GPIIb/IIIa antibodies with the fusion protein containing the GPIIIa sequence of residues from 468 to 691. These results are consistent with a report by Kekomaki et al. that a 50 kDa chymotryptic digestion product of GPIIIa isolated from blood platelets contains target epitopes for serum antiplatelet antibodies in 16 of 33 ITP patients. Smaller peptides including residues 446-501 and residues 593-691 each reacted with only 5 of the 24 patient sera; furthermore all but 3 of these interactions were very weak. Visualization of the conformation of the extracellular portion of alpha(V)beta(3) reveals the location of the 222-residue antigenic GPIIIa (beta(3)) peptide 'B' at the immediately extracellular region of the protein that includes a beta-tail domain and several integrin-EGF domains. In summary, predictions of hydrophilicity, surface accessibility and antigenicity and the three dimensional structure of the beta(3) integrin correlate with autoantibody binding to a recombinant GPIIIa peptide 'B' containing residues 468-691.     

Platelet autoantigens: identification and characterization using immunoblotting

Antiplatelet autoantibodies are important in the etiology of idiopathic (or immune) thrombocytopenic purpura (ITP). Studies using immunoblotting techniques have been helpful in identifying the antigenic target proteins for the antibodies. Antibodies against the glycoprotein (GP) IIIa portion of the GPIIb/IIIa complex were the first to be demonstrated by this approach. Similar GPIIIa autoantigens have also been found to be the most frequent targets of ITP antibodies. Not all anti-GPIIIa antibodies are directed against the same epitope on GPIIIa. A subset of anti-GPIIIa antibodies found in patients with an acquired qualitative platelet dysfunction actually interfere with fibrinogen binding to normal platelets. Antibodies directed against targets on GPV have been found in patients with acute ITP of childhood. In patients with ITP associated with lupus erythematosus, antibodies which bind to intracellular proteins of apparent molecular weights of 66 and 108 kDa have been detected. Thus, ITP antibodies can have a variety of target antigens. Study of larger series of patients will determine whether identification of platelet autoantigens correlates with clinical course of ITP.     

Editor's preface

OBJECTIVE: To describe the process used to notify pediatric patients who received transfusions of blood or blood products at our institution before donor blood was routinely screened for antibodies to HIV (1985) and hepatitis C virus (1990), and to evaluate the effectiveness of the notification program. DESIGN: Patients who had received transfusions were identified through the hospital''s medical records and the records from the Transfusion Medicine Laboratory. Patients were contacted by registered mail to provide notification of transfusion. A questionnaire was included with the notification to obtain information about the patient''s awareness of the transfusion and whether he or she had undergone or planned to undergo testing for HIV and hepatitis C virus. SETTING: Tertiary care university-affiliated teaching hospital in Hamilton, Ont. PATIENTS: Patients 16 years of age or younger who had received blood products between February 1978 and November 1985. Patients who had received only albumin or immune serum globulin were not included as these products were not associated with viral transmission in Canada. RESULTS: Notification letters were sent to 1546 patients. Of these letters 522 (33.8%) were returned undelivered. Of the 1024 patients contacted 493 (48.1%) responded to the questionnaire, of whom 157 (31.8%) were not aware of their transfusion. A total of 130 (26.4%) of the respondents had already undergone testing for HIV, and 342 (69.4%) indicated that they would undergo such testing as a result of the notification. In contrast, only 30 (6.3%) of 474 respondents had undergone testing for hepatitis C virus, but 425 (89.7%) indicated that they would undergo such testing. Overall, the patients'' response to the notification was neutral or positive; however, a number of patients expressed dissatisfaction and anxiety. CONCLUSIONS: The high proportion of patients who were unaware that they had undergone transfusion and who decided to undergo testing for HIV and hepatitis C virus as a result of notification supports the use of notification programs such as this one.     

Human monoclonal autoantibodies to characterize platelet antigens in immune-mediated thrombocytopenia

Idiopathic thrombocytopenic purpura is characterized by antiplatelet antibodies which mediate the rapid destruction of these cells by the reticuloendothelial cell system. Low serum titers of autoantibodies and the polyclonal nature of human serum make it difficult to identify platelet target antigens with plasma antibodies. To circumvent these problems, we have utilized the techniques of EBV transformation and somatic cell hybridization in order to isolate human monoclonal antibodies from patients with ITP. In this paper we describe the use of human monoclonal autoantibodies to characterize an activation specific antigen on GPIIIa and an autoantigen on the GPIb complex. Ultimately, we hope to determine whether these autoantibodies emerge from a pool of naturally occurring antibodies to activation or senescence antigens, or are triggered by environmental agents such as bacteria or virus, which are comprised of antigens similar to those found on the platelet membrane.     

Blood transfusion and renal allograft survival.

Thirty-two first renal transplantations with cadaveric allografts were reviewed to see how many of the recipients had received blood transfusions preoperatively. There was a significant difference in transplant survival between patients who had and patients who had not received blood transfusion before transplantation; this difference was entirely due to acute rejection within three months after transplantation in patients who had not received transfusion. Other factors studied had no effect on survival.     

HLA compatibility and survival of 51 chromium-labeled allogeneic thrombocytes

In 37 patients with thrombocytopenia (mostly with ITP) the survival time of 51Cr-labeled allogeneic platelets was investigated. The HLA antigens were typed in donors and recipients and the presence of HLA cytotoxins and specific thrombocyte antibodies in sera of patients were examined. In 7 cases the identity of 2 HLA antigens, in 15 cases that of 1 HLA antigen and in 15 cases the HLA incompatibility between donor and patient were found. The survival of platelets did not depend on the degree of HLA compatibility, unless the HLA cytotoxins in sera of patients appeared. The HLA, as well the specific platelet antibodies brought about the shortened platelet survival to 1 day and less. The importance of these observations for platelet kinetics is discussed.     

Ability of plasma from patients with immune thrombocytopenic purpura (ITP) to promote the growth of megakaryocyte progenitors from normal bone marrow.

The ability of plasma from ITP patients (before and after splenectomy) to support the growth of megakaryocyte progenitors was compared with that from healthy subjects. Plasma Factor Index-Megakaryocyte PFI-Mk (ITP) which expressed resultant colony growth was significantly lower before splenectomy, but it normalized after splenectomy. (PFI-Mk) (ITP) did not relate neither to megakaryocyte nor to platelet counts. A positive correlation has been observed between megakaryocyte and platelet numbers in healthy subjects and in ITP patients after splenectomy, but not before splenectomy. The proportion of immature megakaryocytes was markedly higher in ITP marrow before splenectomy. This study indicates, that in ITP apart from antibodies directed to platelets and megakarocytes a low plasma stimulatory activity affected megakaryocytopoiesis.