The platelet alloantigen Zwa or PlA1 is expressed by cultured endothelial cells

Recently, the synthesis by cultured human endothelial cells of a membrane protein complex immunologically related to platelet glycoprotein (GP) IIb/IIIa complex was demonstrated. Since platelet GP IIIa is known to carry the platelet alloantigen Zwa or PlA1, studies were performed to establish whether this antigen is also expressed on endothelial cells. The present report describes the results of these studies, which provide evidence for the presence of the Zwa or PlA1 antigen on the surface of cultured human endothelial cells. This evidence is based on the following observations: (1) cultured endothelial cells react with anti-Zwa (PlA1) antibodies as shown by indirect immunofluorescence; (2) two proteins are precipitated by anti-Zwa (PlA1) antibodies from lysates of 125I-labelled endothelial cells with an electrophoretic mobility corresponding with that of GP IIb and IIIa; (3) anti-Zwa (PlA1) reacts specifically, as shown by immunoblotting of sodium-dodecylsulphate polyacrylamide gels of solubilized endothelial cells, with a protein with a mobility similar to that of platelet GP IIIa.     

Normal synthesis and expression of endothelial IIb/IIIa in Glanzmann's thrombasthenia

Glanzmann's thrombasthenia is a bleeding disorder, inherited in an autosomal recessive way and characterized by an absence or deficiency of the platelet glycoprotein (GP) IIb/IIIa complex. Recently, we and others demonstrated that cultured human umbilical vein endothelial cells synthesized a membrane protein complex similar to the platelet GP IIb/IIIa complex. In this article, we demonstrate that endothelial cells isolated from the umbilical vein of a newborn with Glanzmann's thrombasthenia, as compared with normal endothelial cells, show no difference in their ability to synthesize and express this GP IIb/IIIa complex. Our results indicate that Glanzmann's thrombasthenia is not accompanied by an "endotheliopathy."     

The 5q- chromosome abnormality in haematological disorders:|a collaborative study of 34 cases from the Netheralands

Summary . Clinical, haematological and cytogenetic data of all patients with an acquired 5q - abnormality, diagnosed in four cytogenetic laboratories, were studied. The 5q- arose de novo in 24 patients, while 10 patients exhibited it after radiation and/or chemotherapy. Sixteen cases with a dysmyelopoietic syndrome (DMPS) and one case with acute nonlymphocytic leukaemia (ANLL) had no chromosome abnormalities other than 5q- . All nine patients with refractory anaemia (RA) belonging to the de novo group had a normal cell line. So far none of them has transformed to acute leukaemia, while three patients with refractory anaemia with excess of blasts (RAEB), pure red cell aplasia (PRCA) and acquired idiopathic sideroblastic anaemia (AISA), having the 5q- in 100% of the cells showed such a transformation. The second group consisted of nine patients with a 5q- and additional chromosome abnormalities: three of them died of progression of the disease, while four died of unrelated causes. Eight out of nine patients with a 5q- and acute leukaemia, either de novo (five) or secondary (four) to radiation and/or chemotherapy, had additional chromosome abnormalities. Among the changes (partial) monosomy of chromosome 7 was most frequently found. The median survival time of the group of patients with an isolated 5q- in the presence of a normal cell line was significantly longer than that of patients with additional chromosomal changes with or without acute leukaemia. All patients with an isolated 5q- abnormality showed the previously reported megakaryocytic abnormalities (small mono- or bilobulated). In most of the other patients these megakaryocytic abnormalities were also found, whereas others were definitely normal in this respect. A striking and unexplained preponderance of females (1 2 out of 16) was found in the group with an isolated 5q- anomaly. All deletions studied with banding techniques appeared to be interstitial but the breakpoints were variable. The shortest region of overlap is near band 5_q22.     

Fibrinopeptide A and the phosphate content of fibrinogen in venous thromboembolism and disseminated intravascular coagulation

Concentrations of plasma fibrinopeptide A (FPA) were measured by radioimmunoassay in 50 patients with venous thromboembolism or disseminated intravascular coagulation or both. A consistent discrepancy was observed in values obtained with two anti-FPA antisera. Analysis of extracts from plasma of these patients by high-performance liquid chromatography (HPLC) revealed the presence of a phosphorylated and an unphosphorylated form of the A peptide. Differences in concentrations of FPA measured with the two antisera could be accounted for by their different reactivity with phosphorylated FPA (FPA-P). The differences were abolished by treatment with alkaline phosphatase. A good correlation was observed between the FPA-P content of free A- peptide material and of fibrinogen in plasma as determined by HPLC (r = .88, P less than .001, n = 11). In patients with elevated FPA levels, the mean FPA-P content of fibrinogen was significantly higher (P less than .002, n = 13) than in patients with normal FPA levels (n = 8) and in healthy controls (n = 14). Phosphorus in fibrinogen did not correlate with fibrinogen degradation products or fibrinogen levels and became normal on adequate anticoagulation. Therefore, blood-clotting activation may lead to a high phosphate content of fibrinogen and of free FPA in plasma.     

Longitudinal analysis of point mutations of the N-ras proto-oncogene in patients with myelodysplasia using archived blood smears.

We performed a longitudinal analysis of point mutations of the N-ras proto-oncogene in patients with myelodysplasia and a follow-up of at least 2.5 years after diagnosis. Point mutations at codons 12, 13, and 61 of the N-ras oncogene were analyzed after in vitro amplification of N-ras specific sequences followed by dot-blot hybridization. Lysed cells scraped from archived blood and bone marrow smears were used as template for a polymerase chain reaction. In 3 of 90 patients tested (3.3%), a mutation in codon 12 could be detected in the most recent blood smears. All available blood and bone marrow samples of these patients were subsequently analyzed for the occurrence of that particular mutation. In all three cases the mutation was not detectable at diagnosis, but was acquired later during the course of the disease. In two of these patients this event was associated with rapid deterioration and transformation to acute leukemia. However, the third patient showed a protracted course during a period of 5 years after acquisition of the mutation. These results indicate that activation of the N-ras protooncogene in these three patients represents a secondary phenomenon associated with disease progression in some cases, but compatible with stable disease in others.     

Chronic myeloid leukemia in myasthenia gravis after long-term treatment with 6-mercaptopurine

A woman with myasthenia gravis and a thymoma did not respond sufficiently to thymectomy. She was treated with 6-mercaptopurine. Withdrawal of this treatment was several times followed by an aggravation of myasthenic symptoms. After more than 12 1/2 years treatment she developed Ph1-positive chronic myeloid leukemia (CML). No other case of CML following immunosuppressive treatment has been described. Because the therapeutic agent is potentially leukemogenic, the possibility cannot be definitely excluded that the development of CML is not a mere coincidence.     

A BCR-ABL oncoprotein p210b2a2 fusion region sequence is recognized by HLA-DR2a restricted cytotoxic T lymphocytes and presented by HLA-DR matched cells transfected with an Ii(b2a2) construct.

Peptides corresponding to the fusion site in 210 kD BCR-ABL protein b3a2 (p210b3a2) were previously shown to bind to several HLA class I and II alleles. We have found that b3a2 peptide-specific CD4-positive T-helper cells were able to recognize p210b3a2-positive chronic myelogenous leukemia (CML) blasts in a DR4 restricted manner. Until now, there were no reports of b2a2 breakpoint-specific human T-cell responses. Here we show that repetitive stimulation of T lymphocytes with a 17mer peptide covering the fusion region in p210b2a2 also leads to specific T-cell responses. CD4 and CD4/CD8 double-positive clones obtained from a b2a2 peptide-specific cell line were cytotoxic and proliferative in an HLA-DR2a (DRB5*0101) restricted fashion. Autologous Epstein-Barr virus (EBV) transformed cells, expressing BCR-ABL(b2a2) on transfection, and allogeneic HLA-DR matched p210b2a2-positive cells from CML patients were, however, not lysed. BCR-ABL peptide-specific T-cell clones did respond to autologous EBV cells transfected with invariant chain (li) cDNA in which the HLA class II-associated invariant chain peptide (CLIP) was replaced by a BCR-ABL b2a2 fusion oligonucleotide sequence, illustrating the potential of these T cells to recognize an endogenous BCR-ABL(b2a2) ligand.