Diminished presentation of complement regulatory protein CD55 on red blood cells from patients with hereditary haemolytic anaemias.

INTRODUCTION: Hereditary haemolytic anaemias (HHA) encompass a heterogeneous group of anaemias characterized by decreased red blood cell survival. The aim of this study was to evaluate the status of red blood cell (RBC) surface molecules known or previously proposed to participate in preventing premature RBC clearance, analysing erythrocytes from patients with two types of HHA: hereditary spherocytosis (HS) and microcytosis. MATERIAL/METHODS: Relative binding of five monoclonal antibodies (mAbs), anti-CD55, anti-CD59, anti-CD44, anti-CD47 and anti-CD58, was evaluated in erythrocytes of patients with HS and hereditary microcytosis, using flow cytometry. The amount of CD55 protein was assessed by semi-quantitative Western blots densitometry analysis. RESULTS: The majority of both HS and microcytic patients demonstrated significant reduction of anti-CD55 binding by erythrocytes (average 23% and 19%, respectively, P < .001), with no concomitant anti-CD59-binding deficiency. Anti-CD44, anti-CD47 and anti-CD58 binding was within the healthy control range or was slightly decreased. CONCLUSIONS: This study provides evidence supporting the presence of erythrocytes deficient in CD55 presentation in HS and hereditary microcytosis. Moreover, deficiency of CD55 antigen presentation on RBC does not correlate with the amount of CD55 in RBC membrane. Further studies using molecular techniques will clarify the exact participation of CD55 deficiency in premature RBC clearance in HHA.

Association of HLA haplotypes with paroxysmal nocturnal hemoglobinuria.

The pathologies of paroxysmal nocturnal hemoglobinuria (PNH) are primarily caused by somatic mutation in the PIG-A gene in hematopoietic stem cells resulting in glycosyl phosphatidylinositol deficiency and accumulation of phosphatidylinositol (PI) in plasma membranes. The mechanism of pathologic clone domination over normal hematopoietic clones in PNH patients is not yet understood. Forty-four PNH patients, including 9 with aplastic anemia traits (AA/PNH), 31 without full aplasia in bone marrow (de novo PNH, or dn/PNH), and 4 with unclassified PNH, and 200 ethnically matched controls were tested for the HLA A, B, C, DRB1, and DQB1 alleles and haplotype associations. The top block association analysis showed the primary association of PNH with 3 haplotype fragments: the class I fragment A*2501-Cw*1203-B*1801 (risk ratio [RR], 6.64; P=.00012), and 2 class II fragments: DRB1*1501-DQB1*0602 (RR, 7.09; P=.0000015) and DRB1*0401-DQB1*0301 (RR, 6.76, P=.0093). The stratified analysis revealed that the A*2501-Cw*1203-B*1801 haplotype associated preferentially with AA/PNH, and its component HLA molecule showed immunodominant antiapoptotic peptides derived from PI-activated phospholipase D; whereas the DRB1*1501-DQB1*0602 haplotype was associated strongly with dn/PNH and presented immunodominant class II-derived autopeptides. We concluded that certain HLA haplotypes were associated with PNH much more strongly than their allelic components. At least 3 HLA haplotype blocks (A*2501-Cw*1203-B*1801, DRB1*1501-DQB1*0602, and DRB1*0401-DQB1*0301) were primarily associated with PNH. Our results supported the hypothesis of the roles in AA/PNH of antiapoptotic and in dn/PNH of autoimmune mechanisms.

B cells CD19+ in patients with B-cell chronic lymphocytic leukaemia and autoimmune haemolytic anaemia.

The study presents results of B and T lymphocytes population analysis in patients with chronic lymphocytic leukaemia B cells and autoimmune haemolytic anaemia (CLL-B + AIHA). We evaluated the following groups of patients: (1) with newly recognized CLL-B and co-existent AIHA (untreated), (2) after short-term treatment with corticosteroids, (3) after treatment with chemotherapy and corticosteroids. The control groups were made of patients with CLL-B without AIHA. The populations of lymphocytes and determination of cells immunophenotype were performed by means of flow cytometry. The analysed data were obtained from 25 patients. The untreated patients with CLL-B + AIHA presented significantly more numerous population of neoplastic cells CD19+ CD5+ in comparison with patients without AIHA. The patients with AIHA showed a reduced percentage of B CD19+ CD22+ cells in comparison with those without AIHA. Untreated patients with AIHA or after a short-term corticosteroid treatment showed a higher ratio of the number of CD19+ CD5+ cells to the number of T CD4+ and T CD8+ lymphocytes than CLL-B patients without AIHA. It can be presumed that the differences found may be related to the pathogenesis of the autoimmune haemolysis syndrome in patients with CLL-B.

Hemoglobin F in primary myelofibrosis and in myelodysplasia.

Our study investigated two groups of adult patients with established diagnoses of primary myelofibrosis (21 patients) and myelodysplastic syndromes (MDS) (21 patients). The objective was to assess fetal hemoglobin (HbF) concentration and to investigate correlations with organomegaly and extramedullary hematopoiesis and with the level of anemia and blood transfusion requirement. In all patients, the diagnosis was confirmed by histopathological examination. Patients with myelofibrosis were investigated by ferrokinetics using 59Fe. The percentage of marrow sideroblasts was assessed in patients with refractory anemia with ringed sideroblasts. Increased values of HbF were found to occur both in patients with myelofibrosis and with MDS, although a higher incidence and higher concentrations were evident in patients with myelofibrosis. Statistically significant increases in HbF concentration were found when there was accompanying organomegaly, as compared to patients without this feature. The average HbF concentration in both groups of patients under study was twice as high in cases with as in those without marrow fibrosis. The difference was statistically significant. Increased HbF levels appear to correlate with extramedullary hematopoiesis. HbF concentration did not correlate with the level of anemia or with requirement for blood transfusion.

Degradation of glycophorin A of human erythrocytes in patients with myelo- or lymphoproliferative disorders: possible role of neutrophil proteases.

We have previously reported that glycophorin A (GPA) of human erythrocytes (carrying blood group M and N determinants) was totally digested by incubation of erythrocytes with human neutrophil elastase (HNE) and cathepsin G (CathG). The membrane-bound GPA fragments fractionated by SDS-PAGE gave characteristic patterns of bands detected by immunoblotting with the monoclonal antibody PEP80. Erythrocytes were incubated with HNE and CathG at low enzyme concentrations, similar to those found in vivo. Characteristic electrophoretic patterns of bands derived from a partial GPA digestion were observed and these patterns were different for both enzymes and different from those obtained after total GPA digestion. GPA was also partially digested by incubation of erythrocytes with granulocytes in the presence of Ca2+ and calcium ionophore and electrophoretic pattern of digestion products was similar to that obtained with low doses of HNE. No GPA digestion products were detected after treatment of erythrocytes with plasmin and kallikrein. Untreated erythrocytes of 21 patients with various myelo- or lymphoproliferative disorders were tested by SDS-PAGE of RBC membranes and immunoblotting with the anti-GPA PEP80 antibody. GPA degradation products, resembling those formed by a mild CathG treatment of control RBC, were detected in nine patients. GPA fragmentation was in some cases accompanied by a reduced expression of blood group MN determinants. No distinct relation was observed between the occurrence of GPA degradation in erythrocytes and increases in plasma concentrations of HNE-alpha1-proteinase inhibitor (alpha1-PI) complex considered to be an indication of a release of neutrophil proteinases in vivo. However, the results suggested that a partial GPA degradation in haematological proliferative disorders may occur due to limited proteolysis by neutrophil proteinases, most likely by CathG.

[Therapeutic plasmapheresis in patients with multiple myeloma]. / Plazmafereza lecznicza u chorych na szpiczaka plazmocytowego.

The effectiveness of therapeutic plasmaphereses with antineoplastic chemotherapy was evaluated in 25 cases of plasmocytic myeloma in stage III of progression of the proliferative process. The indication to plasmapheresis was hypergelification of serum with clinical symptoms of central nervous system disturbances, renal failure in various stages of progression, intensification of coronary symptoms, bleeding tendency. Good effects with reduced level of total protein and monoclonal protein in serum by 30-80% with regression of clinical symptoms caused by serum hypergelification were obtained in 11 cases. In the remaining patients clinical improvement of varying degree was noted when the level of total and monoclonal protein in the serum fell by 10-29%.

[Chemotherapy and immunomodulating treatment of patients with multiple myeloma]. / Chemioterapia i leczenie immunomodulacyjne u chorych na szpiczaka plazmocytowego.

Prospective studies were carried out on the effectiveness of various treatment methods in 208 patients with plasmocytic myeloma. In 102 patients induction therapy was based exclusively on melphalan, in 106 cases polychemotherapy was used including vincristine, melphalan, carmustine, cyclophosphamide and prednisone. The differences in the per cent of patients with good response to treatment and in the survival time after treatment beginning were statistically not significant between these groups which suggests that polychemotherapy begun from the diagnosis of the disease is justified in patients with large mass of the neoplasm and poor prognostic factors. In 45 patients chemotherapy was supported by administration of immunomodulatory agents, including calf thymus extract in 25 cases, levamisole in 18 and interferon in 2. It was observed that maintenance of remission with chemotherapy and with immunomodulatory agents calf thymus extract or levamisole prolonged the survival of the patients. In cases of leucopenia the use of calf thymus extract facilitated chemotherapy by stimulation of myelopoiesis.