Extramedullary hand mirror cells in pathologic conditions of lymphoid tissue

The hand mirror (HM) cell phenomenon, which usually affects pathologic cells of lymphoid tissue in bone marrow, especially in acute lymphoblastic leukemia (ALL), was seen in B-cell lymphoma cells in cerebrospinal fluid (CSF). It was subsequently detected not only in the bone marrow, but also in extramedullary sites of the lymphoma. Subsequent phase-contrast microscopic study of 45 consecutive specimens of CSF revealed lymphoid cells with HM features in a case of ALL (in which HM cells were subsequently found in the bone marrow) and in a case of acute viral meningoencephalitis. These observations demonstrate that the HM cell phenomenon, which is considered to be a cellular alteration resulting from incorporation in the cell of antigen-antibody complexes, is not unique to bone marrow. It can be present in extramedullary sites and can be seen in exfoliated cells in CSF, where its detection is facilitated by the use of phase-contrast microscopy.     

Hybrid acute leukemia: therapeutical implications of immunological phenotyping.

We phenotyped blood and bone marrow cells from a patient with acute Ph1+ acute leukemia longitudinally during the four months he received intensive chemotherapy. At presentation this case of biphenotypic acute leukemia had two immunologically different types of blast cells, one expressed CD10 (CALLA), CD13 (MY7) and CD33 (MY9) but lacked CD20 (B1), the other type expressed no CD10 or CD33. The phenotype, during AML induction therapy, changed to a more CD10+, CD20+ ALL one. ALL therapy based on these findings induced improvement in bone marrow function but the patient died of septicemia at day 134. The use of concomitant immunophenotyping (IP) and cell cycle analysis had shown proliferation advantage of the more lymphoid malignant cells. These results suggest that it is possible to induce lineage-associated changes in the phenotype of hybrid malignant cells and that these leukemias might be treated best according to longitudinal immunophenotyping of the blast cells.     

Growth of normal versus leukemic bone marrow cells in long term culture from acute lymphoblastic and myeloblastic leukemias

The ability of the in vitro long-term bone marrow culture (LTBMC) system to impair the survival of leukemic cells and to enhance the growth of normal progenitors has been studied. Bone marrow cells from 19 acute lymphoblastic leukemia (ALL) and 30 acute myeloid leukemia (AML) patients at diagnosis were grown in LTBMC for 4-10 weeks. In half of the cases the leukemic population declined down to undetectable levels and was replaced by putative normal hemopoietic precursors, both in ALL and in AML. In the remaining cases, leukemic cells persisted throughout the culture time and few if any normal hemopoietic cells were detected. These data led us to extend to the lymphoid compartment the previous observation of decreasing leukemic myeloid blasts in LTBMC. The potential of such cultures as an in vitro purging system for autologous bone marrow transplantation in selected poor-prognosis lymphoid malignancies should be explored, as has been done for acute and chronic myeloid leukemias.     

Daxx在急性白血病的表达及意义

目的研究Daxx在急性白血病(AL)中的表达及其与AL的分型、临床特征、疗效及预后的关系.方法应用免疫组织化学方法检测88例初治AL骨髓细胞Daxx蛋白的表达情况,分析其与FAB分型、临床特征、疗效及预后的关系.结果急性髓细胞白血病(AML)骨髓细胞中Daxx的表达显著高于正常对照组(P<0.05)与急性淋巴细胞白血病(ALL)组(P<0.05).在AML亚型中,Daxx在M3中的表达显著高于M1,M2,M3,M4和M5.采用逐步回归法分析,校正其它参数,Daxx的表达与初诊时的白细胞数及骨髓原始细胞数目呈正相关,与疗效呈负相关. 结论 Daxx在急性髓细胞白血病中广泛表达,其表达的紊乱可能在AML的发病中起作用,还与AML的某些临床特征、疗效及预后密切相关.     

Advanced lymphoblastic clones detection in T-cell leukemia

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignant neoplasm of the lymphocyte precursors that suffered malignant transformation arresting the lymphoid cell differentiation. Clinical studies revealed monoor, more rarely, oligoclonal nature of the disease. A precise identification of malignant clone markers is both the crucial stage of early diagnostics and the essential prognostic factor for therapeutic treatment. Here we present an improved system for unbiased detection of lymphoblastic clones in bone marrow aspirates of T-ALL patients. The system based on multiplex PCR of rearranged T-cell receptor locus (TRB) and straightforward sequencing of the resulted PCR fragments. Testing of the system on genomic DNA from Jurkat cell line and four clinical bone marrow aspirates revealed a set of unique TRB rearrangements that precisely characterize each of tested samples. Therefore, the outcome of the system produces highly informative molecular genetic markers for further monitoring of minimal residual disease in T-ALL patients.     

Acute lymphoblastic leukemia (ALL) antigens detected with antisera to E rosette-froming and non-E rosette-forming ALL blasts.

Based on the presence or absence of erythrocyte receptors(E) a T cell marker, acute lymphocytic leukemia (ALL), can be divided into E+ALL and E-ALL. We studied cell surface antigens on blasts from 12 children with untreated ALL: eight with E-ALL and four with E+ALL. Heterologous antisera were raised against thymus cells, E+ and E-ALL blasts, appropriately absorbed and tested by immunofluorescence and a radiolabeled antibody assay with normal and leukemic lymphoid cells. By both methods, anti-thymus and anti-E+ALL sera reacted with human thymocytes. Specific binding of anti-E+ALL serum to T antigens was indicated by the fact that a single absorption with thymocytes abolished its binding to allogenic thymocytes, and the reactivity of anti-E+ALL serum with thymus, blood and bone marrow lymphocytes was similar to that of anti-thymus serum. After exhaustive absorption with blood leukocytes, anti-E+ALL and E-ALL sera were negative against normal lymphocytes and bone marrow cells from children with ALL in remission. Anti-thymus and anti-E+ALL sera reacted with blasts from patients with E+ALL, but not with E-ALL. In contrast, anti-E+ALL serum reacted with 40 to 96% of blasts from all children with E-ALL, whereas of the four patients with E+ALL, two were negative and two had the lowest percentage of immunofluorescent cells (10 to 22%). These results were confirmed with the radiolabeled antibody assay. Patients with active E-ALL had cells bearing E-ALL antigen(s) in the peripheral blood and bone marrow, but the number of immunofluorescent cells was lower in blood. Cells reactive with anti-E-ALL serum did not react with thymus cells, blood lymphocytes, remission bone marrow cells, Raji cells, PWM and PHA-induced blasts and CLL cells bearing mIg (uk). These data suggest that the antigen detected on E-ALL blasts by anti-E-ALL serum is neither a HLA-related nor a cell differentiation antigen. Thus, by using antiserum to E+ALL blasts, we have confirmed the presence of a T cell-specific antigen(s) on E+ALL cells. This antiserum did not recognize other leukemia-associated antigens common to E+ and E-ALL. We have also demonstrated an antigen(s) which is regularly expressed on E-ALL blasts and is either not detectable or is present in a lower proportion of E+ALL blasts.     

Cutting edge: Natural helper cells derive from lymphoid progenitors

Natural helper (NH) cells are recently discovered innate immune cells that confer protective type 2 immunity during helminth infection and mediate influenza-induced airway hypersensitivity. Little is known about the ontogeny of NH cells. We report in this study that NH cells derive from bone marrow lymphoid progenitors. Using RAG-1Cre/ROSA26(YFP) mice, we show that most NH cells are marked with a history of RAG-1 expression, implying lymphoid developmental origin. The development of NH cells depends on the cytokine receptor Flt3, which is required for the efficient generation of bone marrow lymphoid progenitors. Finally, we demonstrate that lymphoid progenitors, but not myeloid-erythroid progenitors, give rise to NH cells in vivo. This work therefore expands the lymphocyte family, currently comprising T, B, and NK cells, to include NH cells as another type of innate lymphocyte that derives from bone marrow lymphoid progenitors.     

Effect of thymic humoral factor in vitro on human bone marrow and blood cells from B-, T- and null-cell acute lymphatic leukemia.

The nature of null-cell acute lymphatic leukemia (ALL) was investigated with the aid of a thymic humoral factor (THF), bone marrow cells, and a local xenogeneic graft-versus-host reaction (GVHR). Lymphocytes obtained from the blood and bone marrow of six children with T-cell ALL, five with null-cell ALL, one with perinatal B-cell ALL, one with acute myelocytic leukemia, and one with erythroleukemia were tested for membrane surface markers (E, EAC, and SM Ig); functional activity of T cells was tested by a local GVHR. All of the specimens obtained at the initial presentation showed a lack of functional activity of the lymphocytes. Incubation of null cell and acute myelocytic leukemia (AML) bone marrow with THF led to the acquisition of the characteristics of functional, immunocompetent T cells. No such effect was seen when the bone marrow of T-cell ALL and peripheral blood lymphocytes of B-cell perinatal ALL were incubated with THF. This study demonstrates that the null cell in ALL bone marrow can be differentiated into a T cell whereas the stem cell in AML bone marrow constitutes a pluripotential undifferentiated cell which also can mature into a T cell.     

Quantitative cytology in leukemia research

A study was undertaken to determine the usefulness of flow cytometric analysis of bone marrow cells as an objective means for diagnosis, classification and prognosis in patients with leukemia. Abnormal DNA content as a marker of neoplastic disease was found in only 15% of 264 adult patients with acute leukemia (13% in AML, 26% in ALL/AUL). Alternative means of tumor cell detection in heterogeneous marrow samples include determination of nucleolar antigen density and double-stranded RNA content. Phenotypic characterization of leukemia subtypes can be afforded by RNA content analysis of acridine orange-stained cells, demonstrating significantly higher mean RNA content values in AML, compared to ALL/AUL. Cytokinetic parameters amenable to flow cytometric analysis include measurements of cell cycle compartment distribution by DNA content, of cycle traverse rate by BUdR-induced modification of fluorescence intensity of DNA specific dyes and of growth fraction employing the method of in situ DNA denaturation and subsequent acridine orange staining. Determination of cell cycle distribution and RNA content pretreatment and serially during remission induction in 82 patients demonstrated a significantly lower pretreatment biopsy S phase proportion in responding patients with AML compared to individuals failing treatment whereas an opposite trend was noted in patients with ALL/AUL. While of no prognostic impact pretreatment, serial determinations of the RNA content during the first chemotherapy induction course revealed significant differences between responding and failing patients with AML. Also, patients attaining remission demonstrated a rise in marrow biopsy S phase compartment size by day 10 to 14 of treatment, thus, predicting remission during marrow hypoplasia. We conclude that quantitative cytologic examination of marrow cells from patients with acute leukemia provides useful diagnostic and prognostic information that should aid in the stratification of patients with poor prognosis to receive new agents.     

Disruption of Annexin II /p11 Interaction Suppresses Leukemia Cell Binding,Homing and Engraftment,and Sensitizes the Leukemia Cells to Chemotherapy

The bone marrow microenvironment plays an important role in acute lymphoblastic leukemia (ALL) cell proliferation, maintenance, and resistance to chemotherapy. Annexin II (ANX2) is abundantly expressed on bone marrow cells and complexes with p11 to form ANX2/p11-hetero-tetramer (ANX2T). We present evidence that p11 is upregulated in refractory ALL cell lines and patient samples. A small molecule inhibitor that disrupts ANX2/p11 interaction (ANX2T inhibitor), an anti-ANX2 antibody, and knockdown of p11, abrogated ALL cell adhesion to osteoblasts, indicating that ANX2/p11 interaction facilitates binding and retention of ALL cells in the bone marrow. Furthermore, ANX2T inhibitor increased the sensitivity of primary ALL cells co-cultured with osteoblasts to dexamethasone and vincristine induced cell death. Finally, in an orthotopic leukemia xenograft mouse model, the number of ALL cells homing to the bone marrow was reduced by 40–50% in mice injected with anti-ANX2 antibody, anti-p11 antibody or ANX2T inhibitor compared to respective controls. In a long-term engraftment assay, the percentage of ALL cells in mouse blood, bone marrow and spleen was reduced in mice treated with agents that disrupt ANX2/p11 interaction. These data show that disruption of ANX2/p11 interaction results in reduced ALL cell adhesion to osteoblasts, increased ALL cell sensitization to chemotherapy, and suppression of ALL cell homing and engraftment.     

Leukaemic peripheral blood plasma and bone marrow plasma: comparison of influence on lymphocyte proliferation

Abstract. Peripheral blood plasma from some children with untreated acute lymphoblastic leukaemia (ALL) exerted an inhibitory effect in vitro on phytohaemagglutinininduced lymphocyte transformation of normal peripheral blood lymphocytes. This occurred at concentrations beyond that required for optimal response as judged by reduction of blast cell formation and tritiated thymidine and tritiated uridine incorporation into DNA and RNA, respectively. In contrast, bone marrow plasma from these patients was non-inhibitory or contained significantly less inhibitory activity. Bone marrow plasma from the majority of healthy controls was superior to their peripheral blood plasma in enhancing phytohaemagglutinin-induced mitogenesis. The difference between an individual's bone marrow- and peripheral blood-derived plasma in enhancing proliferation of patient and healthy control cells was significantly greater amongst the patients than the healthy control group; this was attributed mainly to the increased inhibitory activity of ALL peripheral blood plasma compared with normal plasma. Medium conditioned by phytohaemagglutinin-stimulated normal peripheral blood lymphocytes was effective in neutralizing the inhibitory activity of ALL peripheral blood plasma. Taken together, these in vitro results are at least suggestive that in vivo , in healthy subjects, the rapidly proliferating cells in the bone marrow and the 'resting' blood cells in the circulation may be under the influence of a fine balance of different types and/or levels of humoral growth stimulatory and inhibitory factors and that in ALL an unstable balance of these factors exists. The decreased proliferation of circulating blast cells compared with bone marrow blasts in ALL may be attributed, at least in part, to exposure to the different levels of inhibitor(s) in the circulation and bone marrow as demonstrated in vitro by our results.     

Granulocytopoiesis in children with acute lymphoblastic leukaemia.

Numbers, proliferative potential, and differentiative capacity of bone marrow granulocyte-macrophage precursor cells were studied in 130 children with acute lymphoblastic leukaemia (ALL), including 77 children in an acute phase of the disease and 53 in remission. Bone marrow samples from 65 children without haematopoietic abnormalities were used as controls. The numbers of clonogenic precursors were found to be below normal in all phases of ALL, particularly during the acute period when the bone marrow was heavily infiltrated with leukaemic cells. It is shown that the decreases in the numbers and proliferative potential of the precursor cells during the acute phases was associated with the effects of leukaemic blast cells, but that in remission the observed reduction in the precursor cell pool was due to the cytostatic effect of therapy. The differentiative capacity of clonogenic granulocyte and macrophage precursors was not altered in children with ALL.     

Bone Marrow Recovery by Morphometry during Induction Chemotherapy for Acute Lymphoblastic Leukemia in Children

Bone marrow architecture is grossly distorted at the diagnosis of ALL and details of the morphological changes that accompany response to Induction chemotherapy have not been reported before. While marrow aspirates are widely used to assess initial response to ALL therapy and provide some indications, we have enumerated marrow components using morphometric analysis of trephine samples with the aim of achieving a greater understanding of changes in bone marrow niches. Morphometric analyses were carried out in the bone marrow trephine samples of 44 children with ALL, using a NanoZoomer HT digital scanner. Diagnostic samples were compared to those of 32 control patients with solid tumors but without marrow involvement. Samples from patients with ALL had significantly increased fibrosis and the area occupied by bony trabeculae was lower than in controls. Cellularity was higher in ALL samples due to leukemic infiltration while the percentage of normal elements such as megakaryocytes, adipocytes, osteoblasts and osteoclasts were all significantly lower. During the course of Induction therapy, there was a decrease in the cellularity of ALL samples at day 15 of therapy with a further decrease at the end of Induction and an increase in the area occupied by adipocytes and the width of sinusoids. Reticulin fibrosis decreased throughout Induction. Megakaryocytes increased, osteoblasts and osteoclasts remained unchanged. No correlation was found between clinical presentation, early response to treatment and morphological changes. Our results provide a morphological background to further studies of bone marrow stroma in ALL.     

Prion Replication in the Hematopoietic Compartment Is Not Required for Neuroinvasion in Scrapie Mouse Model

Fatal neurodegenerative prion diseases are caused by the transmissible PrP^Sc prion agent whose initial replication after peripheral inoculation takes place in follicular dendritic cells present in germinal centers of lymphoid organs. However, prion replication also occurs in lymphoid cells. To assess the role of the hematopoietic compartment in neuroinvasion and prion replication, we generated chimeric mice, on a uniform congenic C57/BL6J background, by bone marrow replacement with hematopoietic cells expressing different levels of PrP protein. Nine different types of chimeric mice were inoculated intraperitoneally either with the lymphotropic Rocky Mountain Laboratory (RML) strain or the non lymphotropic ME-7 scrapie strain, at different doses. Here, we clearly demonstrate that overexpression of PrP by the hematopoietic system, or the lack of PrP expression by the bone marrow derived cells, does not change the incubation time period of the disease, even when the mice are infected at limiting doses. We conclude that the hematopoietic compartment is more or less permissive to prion replication, both for RML and ME-7, but does not play a role in neuroinvasion.     

Intestinal lymphoid tissue changes in the cortisone-treated Wistar rat

This paper continues the previous investigation of the Department on the lymphoid tissue of central and peripheral lymphoid organs under different experimental conditions. The morphological reactional modalities of the intestinal lymphoid tissue in the male Wistar rat were followed up under endocrine imbalance conditions following cortisone administration. Seven days after administration cortisone induced a hyperplasia of the intestinal lymphoid tissue in parallel with a depletion of the lymph node parenchyma and a hypercellularity of bone marrow. After a 6-week postcortisone interval, the lymphoid tissue showed changes corresponding to a cellular depletion in parallel with the restoration of the lymph node parenchyma and a normocellular bone marrow.     

Bone marrow transplantation: current results in leukemia

Bone marrow transplantation offers two potential therapeutic advantages over more conventional therapy of leukemia. It allows more intensive treatment to be given without regard to marrow toxicity and allows in the case of allogeneic marrow an additional immunotherapeutic effect through graft-versus-host disease (GVHD). Initially, allogeneic transplants in HLA matched sibling donors were only employed in end-stage patients. Although there were encouraging results in terms of long-term therapeutic effects, the overall mortality was prohibitive. Subsequently, patients were transplanted in remission with a marked improvement in overall survival in both acute lymphocytic leukemia and acute non-lymphocytic leukemia. The major obstacles to further improvement in the therapeutic effects of this procedure have been identified (i.e., GVHD, viral infection, and relapse in ALL) and are subject to intensive investigations that already show encouraging results. Syngeneic marrow transplantation is limited for obvious reasons, but early results have shown significant therapeutic effects, in particular, in chronic myelogenous leukemia. These results have encouraged others to use autologous bone marrow. Marrow contamination with unseen tumor cells is being approached by pharmacologic and immunologic techniques designed to "purge" marrow of tumor cells. Animal and initial clinical studies have been encouraging.     

Adolescents with acute lymphoblastic leukemia: a new "age"

Adolescents with acute lymphoblastic leukemia (ALL) have a higher incidence of T-cell immunophenotype, a higher incidence of Philadelphia chromosome positive ALL, a lower incidence of high hyperdiploidy and TEL-AML1 translocation, and a lower incidence of extramedullary bulk disease compared to younger patients. There appears to be little difference between 10-15 and 16-21 year old patients. Adolescents with ALL have a lower event free survival (EFS) compared to younger patients. Adolescents 16-21 years of age treated on pediatric ALL trials have a significantly better EFS than those treated on adult trials. Pediatric treatment protocols utilize more vincristine, steroid and L-asparaginase compared to adult trials. In a recently completed Children's Cancer Group trial, adolescents 16-21 years of age had a four-year EFS of 73.1%. Avascular necrosis of bone is an important complication of therapy in adolescents with ALL.     

Granulopoietic studies in acute lymphocytic leukemia of children.

Studies have been carried out on the levels of serum and urine colony stimulating activity (CSA) and peripheral blood and bone marrow colony forming cell numbers in children with acute lymphocytic leukemia (ALL) during various phases of their disease. These studies have suggested that serum and urine levels of colony stimulating factor are reduced during the inital or relapse phase of the disease compared to levels found during remission. It has also been found that the number of bone marrow colony forming cells is reduced in relapse or before treatment and elevated during remission while the number of peripheral blood colony forming cells is increased during relapse or before treatment and normal during remission. It has also been shown that mixing of serum or leukemic cells with normal human bone marrow cells inhibits colony formation.