Abnormalities of cytoplasmic Ca2+ in platelets from patients with uremia

Uremic patients have a hemorrhagic tendency, often associated with prolonged bleeding times and decreased platelet function in vitro. Whether these defects result from abnormalities in plasma factors affecting platelet activity, platelet surface receptors, intracellular platelet mediators, or other aspects of platelet behavior is unknown. To examine the possibility that the abnormality in platelet function may result from aberrations in Ca2+ homeostasis, blood was obtained from 29 patients with severe uremia. The platelets were washed, loaded with the Ca2+ -sensitive probes indo-1 and aequorin, gel-filtered, and resuspended in either plasma or buffer. Of the 29 patients, seven had template bleeding times prolonged to 11 minutes or more, but platelet aggregation in plasma was not consistently impaired in these patients. However, in aequorin-loaded platelets from the patients with long bleeding times, the highest elevation of cytoplasmic calcium [( Ca2+]i) in response to the Ca2+ ionophore A23187, arachidonate, adenosine diphosphate (ADP), or epinephrine was lower than that seen in platelets from both uremic patients with less prolonged bleeding times and normal volunteers. The reduced [Ca2+]i response was associated with decreased aggregation of gel-filtered platelets suspended in buffer. Suspending washed aequorin-loaded uremic platelets in normal plasma for 20 minutes did not reverse the decreased agonist-induced rise in [Ca2+]i; platelets from a normal donor resuspended in uremic plasma aggregated and produced a normal increase in [Ca2+]i in response to agonists. We conclude that the platelet defect seen in some patients with uremia is associated with a decreased rise in platelet [Ca2+]i after stimulation and that this is a manifestation of an intrinsic platelet defect.     

Prognostic implications of cytogenetic studies in an intensively treated group of children with acute lymphoblastic leukemia

We assessed the prognostic significance of leukemia cell cytogenetics by analyzing bone marrow aspirates obtained at time of diagnosis in 165 children on a single protocol for acute lymphoblastic leukemia (ALL). These children were assigned to six mutually exclusive cytogenetic categories as follows: (1) hyperdiploid, with 50 or more chromosomes (n = 35); (2) hyperdiploid, with 47 to 49 chromosomes (n = 11); (3) diploid (n = 42); (4) pseudodiploid (n = 34); (5) hypodiploid (n = 9); and (6) insufficient data (n = 34). At a median follow-up of 5 years, there were no statistically significant differences between any of these cytogenetic categories in either event-free or overall survival. Those children with chromosomal translocations (n = 26) appeared to fare the same as those lacking translocations (n = 105). The absence of karyotypic prognostic significance was observed not only within the overall group, but also when the results were stratified by standard- risk and high-risk status. Of the specific structural chromosome changes that we studied, only the Philadelphia chromosome (Ph) appeared to confer a poor prognosis, although there were too few such cases to achieve statistical significance. Although we did not detect the event- free survival differences that have been described previously in hyperdiploid, hypodiploid, and pseudodiploid childhood ALL, our findings must be viewed as preliminary given the small number of children in some of the cytogenetic categories. We think that the prognostic implications of these cytogenetic features might have been nullified by improvements in therapy.     

Immunologic characterization of a helper T-cell lymphoma

The lymphocytes of a patient with a T-cell non-Hodgkin's lymphoma with peripheral blood involvement and polyclonal hypergammaglobulinemia were characterized in terms of surface markers and immunologic functions. Using the fluorescence-activated cell sorter and employing various monoclonal antibodies against T-cell surface antigens, it was shown that almost all of the patient's peripheral blood lymphocytes were positive for OKT4 and 9.3, antibodies that recognize helper T-cell subset. The circulating lymphoma cells had typical characteristics for T cells; they formed spontaneous rosettes with sheep erythrocytes and stained with the pan-T-cell antibodies 9.6 and 10.2, but did not react with other anti-T-cell monoclonal reagents such as OKT3, UCHT-1, and 3A1. The cells appeared to be mature by the fact that they did not stain with OKT6, and terminal deoxynucleotidyl transferase was undetectable. Functionally, they were able to provide "help" for antibody production, and they could be stimulated to produce moderate amounts of interleukin-2, while unable to proliferate in response to mitogens. Morphologically, some of the lymphocytes showed a deeply cleaved nucleus.     

Mb1, a plasma membrane antigen selectively expressed by U-937 cells

To examine the plasma membrane characteristics of an immature monocytic cell capable of proliferation, we have developed a murine monoclonal antibody that identifies an antigen, Mb1, found on the surface of U- 937. In immunofluorescence analyses, Mb1 is not expressed by peripheral blood monocytes (freshly isolated, lymphokine-activated, or cultured for seven days), neutrophils, or any other circulating element. It is also absent on human bone marrow mononuclear cells, including the CFU- GM. Among a series of malignant cells from 50 patients with acute myeloid leukemia (including 22 with monocytic or myelomonocytic leukemia), no Mb1 expression was detected. Continuous human cell lines of B or T cell origin were also negative, as were the myeloid lines HL- 60 and K562. Apart from U-937, which uniformly expresses Mb1 in high antigen density, only KG-1 (a myeloblastic line) exhibits Mb1 in low antigen density. Exposure of U-937 to phorbol diester (TPA) under conditions that induce features of macrophage differentiation (including the expression of Mo1) results in a significant reduction in Mb1 expression. Mb1 expression is also reduced as a result of culture of U-937 in medium containing anti-Mb1 antibody (antigenic modulation). On sodium dodecyl sulfate-polyacrylamide gel electrophoresis of radiolabeled immunoprecipitates, Mb1 appears to be a dimeric protein with an estimated molecular weight of 80 kd (43 kd under reducing conditions). Antigenic activity on U-937 is destroyed by treatment with trypsin or papain but is regenerated after 24 hours' culture in enzyme- free medium. Mb1 is a constituent plasma membrane protein of U-937, and its degree of expression relates to the state of cellular differentiation.     

Evaluation of E-rosetting human lymphocytes with OKT11 and other monoclonal antibodies

Monoclonal antibody OKT11 was found to compete with sheep red blood cells for binding sites on human lymphocytes. Preincubation of lymphocytes with OKT11 eliminated E-rosette formation. In a study of 142 peripheral blood samples ranging from 1% to over 90% E-rosette- positive cells, comparison to the percent OKT11-positive cells yielded a correlation coefficient of 0.93. In normal donors, subsets of OKT11+ cells were identified using two-color immunofluorescent staining methods with OKT3, OKT4, and OKT8. On the average, approximately 13% of OKT11+ lymphocytes were OKT3- and 13% of OKT11+ lymphocytes were OKT4- and OKT8-. Based on our double antibody fluorescence intensity data, low antigen density OKT11+ lymphocytes were OKT3-. OKT4+ and OKT8+ lymphocytes in normal peripheral lymphocytes have similar OKT11 antigen density.     

Anti-inhibitor Coagulant Complex (Autoplex) for treatment of factor VIII inhibitors in hemophilia

Fourteen individuals with severe hemophilia complicated by factor VIII inhibitors (1 to 132 Bethesda Units) were treated for 33 bleeding episodes with a new activated prothrombin complex concentrate, Anti- Inhibitor Coagulant Complex (Autoplex, Hyland, Glendale, Calif.). Excellent or good results were observed in 21 of 25 minor bleeding episodes treated, which included joint, soft tissue, and mucous membrane hemorrhages. Eight major bleeding problems (an epidural bleed, a puncture wound, 2 serious soft tissue hemorrhages, 2 lacerations, and 2 major surgical procedures) were treated with excellent (6) or good (2) results. No serious complications were encountered, but two children developed transient hypofibrinogenemia following Autoplex infusion. Although some shortening of the prothrombin time and activated partial thromboplastin time was noted after infusion of Autoplex, there is no useful laboratory test for monitoring therapy. Despite the unknown mechanism of action for bypassing factor VIII, Autoplex appears to be a useful and needed interim product and is safe and effective. In view of the possible potentiation of thrombosis concurrent use of fibrinolytic inhibitors should be avoided.     

Frequent somatic deletion of the 13q12.3 locus encompassing BRCA2 in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) has consistent 13q chromosomal abnormalities detected by conventional cytogenetics. Using interphase cytogenetics we show deletion of a 1-megabase 13q12.3 locus, encompassing the BRCA2 gene, in 80% of 35 CLL cases studied. Homozygous deletion of BRCA2, located within the minimal deletion consensus, was detected in a significant population of cells in 60% of the cases. Deletion of the previously described 13q14 locus (analyzed with RB1 and D13S25 probes) was seen in 63% of the cases. Homozygous deletion of RB1 was seen in one case. Seven of the cases (32%) with D13S25 deletion had a population of cells with homozygous deletion. Deletions at the 13q12 and 13q14 loci result from distinct events because they were not contiguous. These data provide evidence for the existence of a new tumor suppressor locus in B-cell CLL located at 13q12.3. BRCA2, located within the minimal deletion consensus, is a candidate for the gene whose somatic inactivation could play a role in the initiation and or progression of B-cell CLL.     

Fatal graft-versus-host disease in a patient with lymphoblastic leukemia following normal granulocyte transfusion

A woman with lymphoblastic lymphoma was treated with combination chemotherapy. She subsequently became febrile while granulocytopenic and was given unirradiated granulocyte transfusions from normal, unrelated donors. She recovered, but 12 days later noted the onset of progressive skin rash, hepatic dysfunction, diarrhea and pancytopenia and, 22 days after her last granulocyte transfusion, died of gram negative septicemia. Histologic examination of multiple tissues including the skin, liver, and intestinal tract showed changes characteristic of acute graft-versus-hose disease (GVHD). Y-chromatin analysis of the patient's peripheral blood just before death indicated the presence of male cells. HLA typing of lymphocytes and skin fibroblasts from the patient and lymphocytes from the family and granulocyte donors was also consistent with engraftment of cells from one of the male granulocyte donors. This donor most likely was homozygous for one of the patient's halotypes, perhaps facilitating engraftment of his cells and subsequent development of transfusion- induced acute GVHD. Until more precise guidelines can be established, we recommend that all cellular blood products given to patients receiving intensive chemotherapy be irradiated with 1500 rad.     

Position paper for the organization of ECMO programs for cardiac failure in adults

Extracorporeal membrane oxygenation (ECMO) has been used increasingly for both respiratory and cardiac failure in adult patients. Indications for ECMO use in cardiac failure include severe refractory cardiogenic shock, refractory ventricular arrhythmia, active cardiopulmonary resuscitation for cardiac arrest, and acute or decompensated right heart failure. Evidence is emerging to guide the use of this therapy for some of these indications, but there remains a need for additional evidence to guide best practices. As a result, the use of ECMO may vary widely across centers. The purpose of this document is to highlight key aspects of care delivery, with the goal of codifying the current use of this rapidly growing technology. A major challenge in this field is the need to emergently deploy ECMO for cardiac failure, often with limited time to assess the appropriateness of patients for the intervention. For this reason, we advocate for a multidisciplinary team of experts to guide institutional use of this therapy and the care of patients receiving it. Rigorous patient selection and careful attention to potential complications are key factors in optimizing patient outcomes. Seamless patient transport and clearly defined pathways for transition of care to centers capable of providing heart replacement therapies (e.g., durable ventricular assist device or heart transplantation) are essential to providing the highest level of care for those patients stabilized by ECMO but unable to be weaned from the device. Ultimately, concentration of the most complex care at high-volume centers with advanced cardiac capabilities may be a way to significantly improve the care of this patient population.