RARS with fibrosis and del(20q) transformed into ALL.

Transformation of myelodysplastic syndrome (MDS) into acute myelogenous leukemia occurs in approximately 30 % of cases, while progression into acute lymphoblastic leukemia (ALL) is rare. We report on a 67-year-old man with the diagnosis of MDS, subtype refractory anemia with ring sideroblasts (RARS), karyotype 20q- , JAK-2 negative and grade III fibrosis on the bone marrow biopsy, who evolved into ALL 33 months after the diagnosis of MDS. RARS is one of the subtypes of MDS with most indolent course. Deletion of the long arm of chromosome 20 (20q-) is considered as good prognosis by the International Prognostic Scoring System, an important scoring system for predicting survival and evolution of MDS. Primary MDS with bone marrow fibrosis may represent a distinct clinicopathological and is supposed to have an unfavorable prognosis. The combined analysis of these features makes this rare report still more challenging and illustrates that biology of MDS is yet to be discovered.

Clinical significance of Th1/Th2 ratio in patients with myelodysplastic syndrome.

In this study, we attempted to evaluate the clinical significance of T helper 1 (Th1)/T helper 2 (Th2) ratio in patients with myelodysplastic syndrome (MDS), five refractory anaemia (RA), four refractory anaemia with ringed sideroblasts (RARS), 31 refractory cytopenia with multilineage dysplasia (RCMD), nine refractory anaemia with excess blast-1 (RAEB-1) and seven refractory anaemia with excess blast-2 (RAEB-2). Intracellular interleukin-4 (Th2 cytokine) and interferon-gamma (Th1 cytokine) production was assessed in CD4+ T lymphocytes activated by phorbol 12-myristate 13-acetate and ionomycin using flow cytometry. Mean Th1/Th2 ratios in each MDS group were as follows: RA/RARS, 8.8 (95% CI, 5.8-11.8), RCMD, 14.7 (95% CI, 9.5-19.9), RAEB-1, 10.6 (95% CI, 4.6-16.6), RAEB-2, 12.8 (95% CI, 3.0-22.7) and control 12.8 (95% CI, 9.6-16.1). There were no significant differences in Th1/Th2 ratio in the RA/RARS, RCMD, RAEB-1 and RAEB-2 subgroups when compared to controls. Because Th1/Th2 ratio in the RCMD group was widely distributed, we divided RCMD patients according to Th1/Th2 ratio into three groups (low, normal and high Th1/Th2 groups). There were no differences in severity of cytopenia among the three above groups. However, the percentage of CD8 cells in the low Th1/Th2 group was significantly lower than those in the high group (P < 0.01). These data suggest that Th1/Th2 imbalance induces CD4/CD8 imbalance, and serves as a marker of the biological interplay in immune regulation.

Immature and mature monocyte-derived dendritic cells in myelodysplastic syndromes of subtypes refractory anemia or refractory anemia with ringed sideroblasts display an altered cytokine profile.

Dendritic cells (DC) are pivotal for T cell-mediated immunity. We investigated the early and terminal maturation of monocyte-derived DC (MoDC) in myelodysplastic syndromes (FAB subtypes refractory anemia (MDS-RA) or refractory anemia with ringed sideroblasts (MDS-RARS)). Immature MoDC were obtained by culture of monocytes with GM-CSF and IL-4 for 8 days. To obtain mature MoDC, TNF-alpha was added during the final three culture days. T cell proliferation and T cell cytokine secretion in mixed lymphocyte reactions (MLR) unveiled a strong reduction of allostimulatory capacity of mature but also of immature MoDC from MDS patients. Immature MoDC from MDS patients exhibited an almost normal immunophenotype, but secreted substantially less IL-12 and more IL-10 in response to LPS/IFN-gamma than normal controls. Terminal addition of TNF-alpha to GM-CSF/IL-4 treated monocytes failed to extinguish cytokine production by MDS MoDC and failed to induce the expected membrane upregulation of costimulatory and other ligands as in normal controls. While our data provide further support for previous studies that have indicated an impaired differentiation of immature towards mature MoDC, they also clearly demonstrate a qualitatively and quantitatively altered cytokine secretion at the level of immature MoDC, which may in part explain the reduced allostimulatory capacity of these cells. These alterations may contribute to immune modulation of the clinical phenotype of marrow failure in MDS, and may have to be considered when designing DC-based immunotherapeutic strategies for MDS.

[Auto- and alloreactivity of T lymphocytes in myelodysplastic syndrome]. / Auto- a aloreaktivita T lymfocytu u myelodysplastického syndromu.

BACKGROUND: Successful therapy with ATG and cyclosporine A in some myelodysplastic syndrome (MDS) patients led us to study the existence of T cells attacking autologous hemopoietic cells. In our study, we attempted to give the direct prove of autoreactive T cells in MDS (autoreactivity analysis). Simultaneously, we analysed the capacity of MDS patients to respond to allogeneic cells from unrelated individuals (alloreactivity analysis). METHODS AND RESULTS: Autoreactive lymphocytes directed against own bone marrow mononuclear cells were analysed using the modification of cell mediated cytotoxic reaction. With one exception we did not confirm the presence of autoreactive T cells among 10 patients examined. Analysis of alloreactivity was performed by means of standard cell mediated cytotoxic reaction and mixed lymphocyte reaction. Surprisingly, the cytotoxic response to allogeneic cells was negative in 11 MDS patients from 16 analysed. When comparing refractory anaemia (RA) and refractory anaemia with ring sideroblasts (RARS) patients, the proportion of negative results was higher in RA (78 %) than in RARS (40 %). In mixed lymphocyte reaction, the response of MDS cells to allogeneic cells of unrelated individual was positive in all tested patients. The preliminary testing of TNF and IFNgamma secretion examined in supernatants of effector cells showed impaired levels of both cytokines in RA and normal levels in RARS in accordance with the findings achieved in alloreactivity analysis. CONCLUSIONS: Autoreactive T cells were not found in MDS patients using our experimental arrangement. Analysis of alloreactivity showed the defect in effector--cytotoxic--phase of cell mediated cytotoxic reaction in the majority of MDS patients. The initial phase of this reaction represented in vitro by mixed lymphocyte reaction gave normal results. The possible reasons of disturbed alloreactivity and its relevance to immunity in MDS are commented in discussion.

Apoptosis in refractory anaemia with ringed sideroblasts is initiated at the stem cell level and associated with increased activation of caspases.

Treatment with granulocyte colony-stimulating factor plus erythropoietin may improve haemoglobin levels in patients with ringsideroblastic anaemia (RARS) and reduce bone marrow apoptosis. We studied bone marrow from 10 RARS patients, two of whom were also investigated after successful treatment. Mononuclear, erythroid and CD34+ cells were analysed with regard to proliferation, apoptosis, clonogenic capacity and oncoprotein expression, in the presence or absence of Fas-agonist, Fas-blocking antibody 2 and caspase-3 inhibitor. During culture, RARS bone marrow cells showed higher spontaneous apoptosis (P < 0.05) and caspase activity (P < 0.05)) than bone marrow cells from healthy donors. Eight out of nine patients had reduced growth of erythroid colony-forming units (CFU-E) (< 10% of control) and granulocyte-macrophage CFU (CFU-GM) (< 50% of control) from CD34+ cells. Fas ligation increased apoptosis and decreased colony growth equally in RARS and controls, but caused significantly more caspase activation in RARS (P < 0.01). Fas-blocking antibody showed no significant inhibitory effect on spontaneous apoptosis or ineffective haematopoiesis, as measured using phosphatidylserine exposure, the terminal deoxynucleotide transferase-mediated dUTP-biotin nick-end labelling technique, caspase activity or clonogenic growth. Caspase inhibition reduced apoptosis, increased proliferation and enhanced erythroid colony growth from CD34+ cells in RARS, but showed no effect on normal cells. CFU-E improved > 1000% after successful treatment. Thus, erythroid apoptosis in RARS is initiated at the CD34+ level and growth factor treatment may improve stem cell function. Enhanced caspase activation at the stem cell level, albeit not mediated through endogenous activation of the Fas receptor, contributes to the erythroid apoptosis in RARS.

Coincident myelodysplastic syndrome and T-cell large granular lymphocytic disease: clinical and pathophysiological features.

Myelodysplastic syndrome (MDS) and T-cell large granular lymphocytic disease (T-LGL) are bone marrow failure disorders. Successful use of immunosuppressive agents to treat cytopenia in MDS and LGL suggests a common pathophysiology for the two conditions. Of 100 patients with initial diagnoses of either MDS or T-LGL referred to the National Institutes of Health for immunosuppressive treatment of cytopenia, nine had characteristics of both T-LGL and MDS (T-LGL/MDS). Fifteen patients with T-LGL received cyclosporin (CSA) (10 responses). Eight out of nine patients with T-LGL/MDS received CSA (two responses) and one patient received ATG (one response). Of 76 patients with MDS, eight received CSA (one response) and 68 received ATG (21 responses). The response to immunosuppression was significantly lower in patients with T-LGL/MDS and MDS than in patients with T-LGL disease alone (28% vs. 66%, P = 0.01). The proportion of T-helper cells and T-suppressor cells with an activated phenotype (HLA-DR(+)) was increased in patients with T-LGL, T-LGL/MDS and MDS, but the increase in activated T-suppressor cells in patients with T-LGL/MDS was not statistically significant. Autoreactive T cells may suppress haematopoiesis and contribute to the cytopenia in T-LGL and some patients with MDS, leading to T-LGL/MDS. The lower response rate of MDS or T-LGL/MDS to immunosuppression, compared with T-LGL alone, may reflect the older age and intrinsic stem cell abnormalities in MDS and T-LGL/MDS patients.

Allogeneic peripheral stem cell transplantation in a case of hereditary sideroblastic anaemia.

We report on a case of pyridoxine refractory hereditary sideroblastic anaemia (HSA) in a 19-year-old man who underwent peripheral blood stem cell transplantation (PBSCT) from his HLA-identical brother. By using short tandem repeat polymorphism, 100% donor cells were observed in peripheral blood on day +21; bone marrow showed mixed chimaerism from day +21 to day +221, when 100% cells of donor origin were observed. The patient developed extensive chronic graft-versus-host disease with favourable response to treatment. When the haemoglobin range was normal, a programme of phlebotomies reduced serum ferritin levels. Three years after transplantation, the patient has an ECOG rating of 0, with completely normal haemoglobin values (15 g/dl). To our knowledge, this is the first PBSCT reported in a case of hereditary sideroblastic anaemia.

[Relation between circulating immune complexes and serum ferritin in hemosiderosis of different etiologies]. / Vzaimosviaz' mezhdu tsirkuliruiushchimi immunnymi kompleksami i ferritinom syvorotki pri gipersiderozakh razlichnoi étiologii.

Parameters of iron metabolism and humoral immunity were studied in patients with chronic diffuse diseases of the liver (cirrhosis, chronic hepatitis), beta-thalassemia major, dyserythropoiesis, hereditary hemochromatosis. High ferritin content has been recorded in the plasma of these patients, that leads to the formation of antibodies to this protein followed by the production of circulating immune complexes inducing metabolic disorders that aggravate the pathologic process. Plasmapheresis and deferoxamine therapy result in a decrease of ferritin and circulating immune complex content in the plasma, that produces a favourable effect on the patients' condition.

Analysis of leucocyte differentiation antigens in blood and bone marrow in patients with refractory anaemia (RA) and RA with sideroblasts. Prognostic implications of sequential and follow-up data.

34 patients with primary myelodysplastic syndrome (MDS), initially diagnosed as subtypes refractory anaemia (RA) and RA with ringed sideroblasts (RA-S), were followed to investigate the distribution of lymphoid and myeloid differentiation antigens in the blood and bone marrow in search of potential prognostic significance with regard to progression to RA with an excess of blasts (RAEB) or acute myeloid leukaemia, and for relations to clinical, morphological and cytogenetic findings. Patients who later progressed to RAEB had significantly decreased percentages of anti-T8 positive T-suppressor cells in the blood at diagnosis compared to those who did not (p = 0.05). Sequential analysis showed a decrease with time also in the percentages of anti-T8-positive cells (p = 0.05). In the bone marrow, progressing patients initially showed significantly increased percentages of anti-My9-positive immature myeloid cells (p less than 0.001), and the percentages of anti-My9-positive cells in the bone marrow increased with time (p less than 0.005). Analysis of the pooled data revealed a statistically significant relation between increasing percentages of anti-My9-positive cells and the frequencies of clonally abnormal (p less than 0.001) and abnormal (p = 0.004) metaphases.

Immunologic indices in myelodysplastic syndromes.

Immunocompetence was evaluated in 36 untreated and noninfected patients affected with myelodysplastic syndromes (MDS). T-cell number and activity were evaluated by counts of total T-cells and T-lymphocyte subsets, and by measure of DNA synthesis in response to phytohemagglutinin and Concanavalin A. B-cells were evaluated as surface immunoglobulin- (SIg+) bearing cells and by serum immunoglobulin levels. Granulocyte activities were evaluated by responses to chemotaxis and to nitroblue tetrazolium test. Complement activity was measured by classic hemolytic complement assay. In addition, circulating immune complexes were detected in serum. MDS were associated with a significant decrease in the absolute numbers of total T (E-rosetting and T3+) cells, T4+, and T8+ cells and a dramatic decrease of the responses to Concanavalin A. An impairment of either chemotaxis or of nitroblue tetrazolium (NBT) test was frequently encountered. An increase in the levels of IgG or IgA was also a frequent feature. The findings reveal that all patients with a high degree of T-cell impairment have refractory anemia associated with an excess of medullary blast cells. All in all, the data suggest that the counts of the absolute number of cells bearing the T3 and T8 phenotypes could be of prognostic value: the higher the number, the better the patient's survival.

Is the HLA-linked haemochromatosis allele implicated in idiopathic refractory sideroblastic anaemia?

In order to assess the previously reported association of HLA-linked idiopathic haemochromatosis with idiopathic refractory sideroblastic anaemia (IRSA), the prevalence of HLA-A3 antigen in a group of 22 patients with IRSA was compared to that observed in healthy controls and in patients with homozygous idiopathic haemochromatosis and to that calculated for a population heterozygous for idiopathic haemochromatosis. The prevalence of A3 in patients with IRSA (0.23) was quite similar to that observed in controls (0.29) and significantly different from that observed in homozygous (0.73; P less than 10(-5] and heterozygous (0.57; P less than 10(-3] haemochromatosis. Serum iron, transferrin saturation, serum ferritin and liver iron concentration showed no difference in IRSA patients with or without A3. It is concluded that there is neither systematic association between the haemochromatosis allele and IRSA nor systematic implication of such an allele in the development of iron overload observed in IRSA.

Granulocyte and monocyte surface membrane markers in the myelodysplastic syndromes.

The expression of lineage specific surface antigens on granulocytes and monocytes was quantitated using monoclonal antibodies in 16 healthy adults and 21 patients with myelodysplastic syndromes. In nine of 19 patients the granulocytes showed a decrease in myeloid or an increase in monocyte antigen expression or both. In 11 of 19 patients the monocytes showed a decreased expression of monocyte antigens or an increase in myeloid antigens or both. The data suggest that in the myelodysplastic syndromes the granulocyte-macrophage progenitors do not develop along two divergent lines but differentiate with the emergence of dual characteristics.

Functional defects in phagocytic cells from patients with iron overload.

Phagocytic functions were studied in patients with iron overload. Phagocytosis of radiolabelled opsonised Staphylococcus aureus by mononuclear (MN) leucocytes and polymorphonuclear (PMN) leucocytes was measured in 15 and 16 patients, respectively. The intracellular killing capacity of MN and PMN leucocytes of seven and nine patients, respectively, and chemotaxis of PMN leucocytes of eight patients, were assessed also. These cellular functions were compared with phagocytic functions of controls tested on the same day, and with the normal ranges of phagocytic cell functions obtained with MN and PMN leucocytes from 48 and 59 healthy donors, respectively. One or more phagocytic functions were impaired in 62.5 per cent of the patients. Comparison of the various phagocytic functions in patients and simultaneously tested controls showed a significant decrease of the mean phagocytic capacity of the patients' MN and PMN leucocytes (P less than 0.015 and P less than 0.03, respectively), as well as the mean bactericidal activity of the MN leucocytes (P less than 0.05) and the mean chemotactic responsiveness of the PMN leucocytes (P less than 0.025). Patients with excess iron must be regarded as compromised hosts, not only because of the increased availability of iron for bacterial growth, but also because of the associated functional impairment of monocytes and granulocytes.

Function of peripheral blood and bone-marrow monocytes in preleukemic patients: normal phagocytosis and intracellular killing of Candida albicans.

Studies were performed to evaluate the function of peripheral blood and bone marrow monocytes from 15 patients with preleukemia and 16 healthy controls. The patients were grouped according to the criteria of the FAB collaborative group. No abnormality in phagocytosis and killing of Candida albicans by peripheral blood and bone marrow monocytes was found in patients compared to normal controls. Normal opsonization by autologous serum was found. No differences were found in this respect between the three groups of patients.

A study of the routine use of the Imugard IG500 Leukocyte Removal Filter illustrating a possible cause of transfusion reactions and a method of prevention.

The results of the use of the Imugard IG500 Leukocyte Removal Filter in the transfusion of 100 patients requiring leucocyte poor blood are presented. The patients were suffering from a variety of disorders. Many of them had previously received leucocyte poor blood prepared by other methods. Five patients developed non-haemolytic transfusion reactions. Subsequent investigations suggested that these were reactions to the plasma proteins present in the leucocyte poor blood prepared by filtration. A simple method of removal of these plasma proteins combined with filtration is described. The resultant product has been transfused, without incident, into the patients who previously reacted.

Sézary syndrome associated with sideroblastic anaemia.

A 66-year-old man with acquired sideroblastic anaemia and Sézary syndrome is described. This is believed to be the first such case reported in the literature. We speculate on the possibility that a common stem cell defect could account for the simultaneous development of the two abnormalities. Other features of this case of Sézary syndrome are discussed in relation to previous reports on patients with this disease.

Antibody-mediated acquired sideroblastic anemia: response to cytotoxic therapy.

A 5 1/2-yr-old child developed severe anemia with erythroid hypoplasia and 50% ringed sideroblasts in his bone marrow. A serum inhibitor of erythropoiesis was demonstrated, utilizing syngeneic and autologous bone marrow in a plasma-clot culture system. The IgG fraction of the patient's serum effected similar suppression of erythroid colony formation. Prednisone therapy was ineffective, but following treatment with cyclophosphamide, normal erythropoiesis was established, ringed sideroblasts disappeared, and his serum no longer inhibited erythropoiesis in vitro. Cyclophosphamide was discontinued, and the patient has remained hematologically normal. This patient is an example of antibody-mediated sideroblastic anemia successfully treated with a cytotoxic drug.