Ketamine modulates hippocampal neurochemistry and functional connectivity: a combined magnetic resonance spectroscopy and resting-state fMRI study in healthy volunteers.

A growing body of evidence suggests glutamate excess in schizophrenia and that N-methyl-d-aspartate receptor (NMDAR) hypofunction on γ-aminobutyric acid (GABA) interneurons disinhibiting pyramidal cells may be relevant to this hyperglutamatergic state. To better understand how NMDAR hypofunction affects the brain, we used magnetic resonance spectroscopy and resting-state functional magnetic resonance imaging (MRI) to study the effects of ketamine on hippocampal neurometabolite levels and functional connectivity in 15 healthy human subjects. We observed a ketamine-induced increase in hippocampal Glx (glutamate+glutamine; F=3.76; P=0.04), a decrease in fronto-temporal (t=4.92, PFDR<0.05, kE=2198, x=-30, y=52, z=14) and temporo-parietal functional connectivity (t=5.07, PFDR<0.05, kE=6094, x=-28, y=-36, z=-2), and a possible link between connectivity changes and elevated Glx. Our data empirically support that hippocampal glutamatergic elevation and resting-state network alterations may arise from NMDAR hypofunction and establish a proof of principle whereby experimental modelling of a disorder can help mechanistically integrate distinct neuroimaging abnormalities in schizophrenia.

Immunophenotypic profile and clinical characteristics in patients with advanced stage mantle cell lymphoma.

The objective of this study was to evaluate immunophenotypic profile along with clinical follow-up in patients with advanced stage mantle cell lymphoma (MCL), and their possible influence on overall survival (OS). Bone marrow (BM) cell and/or peripheral blood mononuclear cell flow cytometric analyses of the following antigens were performed: HLA-DR, CD19, CD20, CD22, CD23, CD25, CD10, SmIg, kappa, lambda, CD79b, CD38, FMC7, CD3, CD2, and CD5. There were 14 patients in IV CS, and 26 patients in CS V. All patients were treated with CHOP. Immunological markers showed a typical phenotype (CD5+ CD23-, Cyclin D1) in all cases. Pathohistological type of BM infiltration was predominantly diffuse (72.5%), and in remainder of patients, nodular. Comparison of patients with leukemic phase of MCL with CSIV (BM), has shown significantly higher expression of CD19, CD20, and CD23, followed by permanently negative expression of CD23. Patients with blastic variant of MCL had higher expression of CD23, compared to typical MCL (P < 0.001). Median OS was 20 months, and there were no significant OS-differences between CS IV and leukemic phase patients. Survival analyses showed that negative prognostic influence had high IPI (P < 0.01), presence of extranodal localization (P < 0.01), and diffuse type of BM involvement (P < 0.01). Using Cox regression according to OS, IPI had independent prognostic value (P < 0.001). Our results demonstrated that in the advanced MCL patients the most powerful prognostic factor was IPI, while extranodal localization and type of BM infiltration were of a limited value.

TNF-alpha induced changes in cell membrane antigen expression on K-562 cells associated with increased lactate dehydrogenase (LDH) release.

TNF-alpha is a pleiotropic cytokine, which induces death of sensitive cells, whose effect depend on cell membrane receptor expression, cell cycle phases, as well as on intracellular ratio of pro- apoptotic and anti-apoptotic molecule expression. Since determination of LDH release from cultured cells in vitro, reflects early membrane alterations, we estimated and compared LDH release from cultured cells with changes in cell membrane antigen expression on K-562 cells after TNF-alpha treatment by flow cytometry. The significant increase in LDH release activity and cytotoxicity values was associated with decrease in membrane molecule expression for CD45 and CD30 as well as for low expressed CD45RA and CD38 after TNF-alpha treatment. However, percentage of decrease of all examined molecules is not uniform, and appears to depend on the respective level of pre treatment values expression and molecule type. These results indicated the complexity of events on cell membrane, including association between increasing LDH release and decrease of antigen expression of membrane molecules following TNF-alpha mediated processes.

Acute panmyelosis with myelofibrosis: clinical, immunophenotypic and cytogenetic study of twelve cases.

The clinical, cytogenetic, and immunophenotypic features in 12 adult patients with acute panmyelosis with myelofibrosis (APMF; ICD-0-3: 9931/3; C42.1) are reported (median age: 57 years; f/m = 1.4). The white cell count (WBC) was normal in 3 patients; 9 had leucopenia. The median hemoglobin value was 64.5 g/l, and median platelet count 12 x 10(9)/l. Bone marrow biopsy showed a hypercellular marrow in 10/12 patients with a significant infiltration of pathological blasts (range: 30 - 60%). All the cases had marked reticulin fibrosis. Immunophenotyping of bone marrow blast cells showed the expression of early (CD34) and lineage-unspecified antigens (HLA-DR) in 6/7, and 7/7 patients, respectively. "Early" myeloid antigens (CD13, CD33) were seen in 6/7 and 4/6 patients respectively. Monocyte antigen (CD14) was expressed in 3/7 patients. Megakaryocyte antigen (CD61) and erythroid cell antigen (GpA) were each expressed in only 1 patient. Two patients had expression of CD34, HLA-DR and "early" myeloid antigens by their bone marrow blast cells and 1 of these also had a co-expression of the antigens from a differentiated monocytic cell proliferation (lysozyme+, CD68+). Nonspecific chromosomal aberrations were recorded in 8/10 patients. The median survival was 2 months. These findings suggest an immature myeloid phenotype of blast cells in APMF. In 6/9 patients a leukemic cell differentiation into monocytic, megakaryocytic or erythroid lineage was also demonstrated.

Long-term survival in acute lymphoblastic leukaemia in adults treated according to the LALA 87 protocol.

BACKGROUND: Between January 1989 and July 1995, a prospective study of the therapeutic efficacy of the LALA 87 protocol in adult acute lymphoblastic leukaemia (ALL) has been conducted. METHODS: A total of hundred and twelve patients with ALL have been analysed. The median age of the patients was 40 years (range: 15-65), the gender ratio (M/F) was 66/46, and the morphologic FAB (French-American-British) profile was L1 in 30 (26.9%) patients, L2 in 71 (63.3%) and L3 morphology in 11 (9.8%) of the patients. The LALA 87 protocol includes five phases: induction, consolidation, reinforcement, maintenance and central nervous system (CNS) prophylaxis with intrathecal methotrexate and irradiation. The induction phase comprised daunorubicin 50 mg/m(2) (days 1-3), cyclophosphamide 600 mg/m(2) (days 1 and 8), vincristine 1.5 mg/m(2) (on days 1, 8, 15 and 22) and daily oral prednisone on days 1-21. Maintenance therapy was given for 2 years and consisted of different drugs as reinforcement, daily 6-mercaptopurine and weekly methotrexate. RESULTS: Complete remission (CR) was achieved in 81 (72.3%) of the patients. The causes of induction failure were partial response in 10 (8.9%), and hypoplastic death in 12 patients (10.7%), and 9 were non-responders (8.0%). Of the 81 patients who achieved CR, 62 relapsed (76%). Among the relapsed patients, 9 developed CNS disease in spite of CNS prophylaxis during induction chemotherapy. Median follow-up for the living patients was 110 months. Median disease-free survival (DFS) was 16 months; 19 patients are still in remission with an estimated 10-year DFS (24%). Adverse prognostic factors were >50 years of age, immunologic subtype and cytogenetic profile. CONCLUSION: The results support the strategy of applying more effort and other treatment modalities in the therapy of ALL.

Clinical, Immunophenotypic and Cytogenetic Features of Megakaryocytic Blast Crisis of Chronic Myeloid Leukemia: A Single Institution Study.

We present 15 patients with megakaryocytic (Mk) blast crisis (BC) of a Philadelphia (Ph) chromosome positive CML confirmed by immunophenotype analysis between 1989-2000. The primary aim of this study is to define clinical, immunological, cytogenetic and laboratory characteristics of Mk BC in Ph positive CML. We have done retrospective analysis regarding basic clinical findings, immunologic phenotype, cytogenetic studies and platelet functions. All patients had significant expression of CD61 (14/14) and CD34 (13/13) antigens, and a high frequency of expression of CD13 (9/12), CD33 (10/12) and CD11b (9/11). The BC in 6/15 patients was presented with thrombocytosis, 7/15 had a normal platelet count and two patients had thrombocytopenia. A grade IV myelofibrosis was present in 8/10 patients. Six patients evolved additional karyotypic abnormalities. Two patients had extramedullary BC. The serum activity of LDH (med. 1095.6) was elevated in all patients. A platelet dysfunction was documented in 4/5 patient tested. There are no clinical and hematological characteristics specific for Mk BC of CML. Normal or elevated platelet count (med. 427.4 x 109/L) in BC of CML with prominent expression of CD34 and CD61 antigens, and significant myelofibrosis (grade IV) are the most consistent clinical findings.

[Immunophenotyping of leukemic cells in the diagnosis of hairy cell leukemia]. / Znacaj imunofenotipizacije leukemijskih celija za dijagnostiku leukemije vlasastih (hairy) celija.

INTRODUCTION: It is established that immunophenotyping constitutes a useful method in the diagnosis of hairy cell leukaemia. However, no single marker is specific for hairy cell leukaemia. Two-color-flow cytometry can aid in the diagnosis by showing coexpression of CD11c, HC2 or CD25 with pan B cell markers. Recently, Matutes E. et al. [17] proposed a scoring system of immunophenotypic markers, which could be used to evaluate the diagnosis of hairy cell leukaemia. AIM OF THE STUDY: The aim of the study was to: a) confirm previous observations of immunophenotypic characteristics of hairy cell leukaemia; b) identify antibody combinations of two-color immunofluorescence staining that are most useful in the diagnosis of hairy cell leukaemia; c) examine the value of a scoring system of immunophenotypic markers in the diagnosis of hairy cell leukaemia. METHODS: We analyzed peripheral blood of 46 patients with hairy cell leukaemia using indirect immunofluorescence flow cytometry (EPICS-Coulter) with an extended panel of monoclonal antibodies: CD19, CD2O, CD22, CD24, CD10, HLA-DR, CD11c, CD25, HC2, Slg, kappa and lambda light chains. The diagnosis of hairy cell leukaemia in all patients was made using conventional criteria based on cell morphology, TRAP cytochemistry and bone marrow histology. One fourth of patients were also analyzed using two-color flow cytometry with three antibody combinations as follows: CD19 + CD11c, CD19 + CD25 and CD19 + HC2. RESULTS: Our results showed that hairy cells of our patients had a uniform and unique immunophenotype with expression of the following markers: CD19, CD22, CD2O, CD11c and HLA-DR in 100% of patients, CD24 in 93%, CD25 in 88%, Slg in 82%, HC2 in 67%, CD1O in 50%, kappa light chains in 38% and lambda light chains in 35% of patients (Table 1). The level of detectable circulating hairy cells varied widely, from 2% to 93% of total lymphocytes, and 12 patients (26%) with less than 5% of detectable hairy cells were excluded from analysis. Two-color cytometry showed that antibody combination CD19 + CD11c was coexpressed in 100% of patients, CD19 + CD25 in 78% of patients and CD19 + HC2 in 57% of patients (Table 2). Only patients with 5% or more double colored hairy cells for one antibody combination, were included in the analysis. On the basis of our results of in immunophenotyping of hairy cell leukaemia patients and results of other authors (17), we made our scoring system which considers the reactivity with the following markers: CD19, CD11c, CD25 and HC2. Each marker gives 1 point if positive and 0 point if negative. Score 4 had 83% of patients, score 3 had 14% of patients, score 2 had 3% of patients and no patient had score 1 or 0 (Table 3). CONCLUSION: Our results demonstrated that immunophenotyping with a selective panel of monoclonal antibodies could significantly increase the accuracy in diagnosis of hairy cell leukaemia. Two-color flow cytometry with antibody combination CD19 + CD11c showed coexpression of hairy cells in 100% of our patients. The scoring system for hairy cell leukaemia used in our patients showed that high scores 3 and 4 had 97% of patients.

Characterization of CD13 and CD33 surface antigen-negative acute myeloid leukemia.

From a cohort of 220 adults with newly diagnosed acute myeloid leukemia (AML), 8 (3.6%) exhibited a rare variant of aberrant membrane phenotype. It was characterized with typical myeloid morphologic and cytochemical patterns and absence of myeloid associated antigens (CD13, CD33, CD14, glycophorin A, CD61). According to the French-American-British criteria, disease in 5 patients was classified as M1 and in 3 patients as M2. CD34, CD38, HLA-DR, and CD45 were strongly expressed in 4 of 5, 3 of 3, 8 of 8, and 3 of 3 analyzed cases, respectively. CD7 antigen was strongly expressed in 4 of 6 patients. Except for predominance of male sex and high frequency of CD7 antigen expression, no other remarkable clinical or biologic characteristics were noted. Detected variant of AML with the unusual membrane phenotype (CD34+, HLA-DR-positive, CD38+, CD45+, CD7+) might represent an example of extreme asynchrony in sequences of morphologic and immunologic maturation or abnormal epitope expression on leukemic cell membrane molecules CD13 and CD33. Although the clinical significance of this AML variant is unclear, the existence of such cases demonstrates the continued need for simultaneous cytochemical and immunologic studies in the evaluation of acute leukemias.

Severe hypercalcaemia and extensive osteolytic lesions in an adult patient with T cell acute lymphoblastic leukaemia.

Hypercalcaemia is a rare feature of acute lymphoblastic leukaemia (ALL) in adults, particularly of the T cell type. We report on a 24-year-old patient with T-ALL, who presented with symptoms of hypercalcaemia (vomitus, acute renal failure), bone pain, extensive osteolytic lesions and normal white cell count without circulating blasts. An increased serum tumor necrosis factor (TNF-alpha) concentration of 35 pg/ml was found; it remained elevated at 52 pg/ml four weeks later, after having achieved haematological remission. Serum concentrations of IL-1beta, IL-6 and IL-2 were within the control range. The pathophysiology of hypercalcaemia in malignancy and possible mediators of bone resorption, in particular TNF-alpha, are discussed.

[Acute megakarioblastic leukemia: de novo and transformation of chronic myeloid leukemia]. / Prikaz bolesnika s akutnom megakarioblastnom leukemijom.

Acute megakarioblastic leukemia (AMKL) is a rare myeloproliferative syndrome with a fulminant clinical course characterized by progressive pancytopenia, palior, weakness and severe haemorrhage. Two cases of AMKL are presented: a 18-year old male with pancytopenia and massive haemorrhage, lymphadenopathy, organomegaly and mediastinal tumour. The diagnosis of AMKL was established by cytological and immunocytochemical analyses of peripherial blood cells (blasts were GPIIIa and GPIb postitive), by histological analysis or the bone marrow and lymph node, and immunohistochemical analysis of lymph node. The second case had megakarioblastic transformation of HGL which was confirmed by cytomorphological and immunophenotypical analyses. In spite of therapy, the patients died soon after the first signs of the disease.