The advances in recent molecular biology have led to the large-scale production of the recombinant hematopoietic growth factors and interleukin, i.e., GM-CSF, G-CSF, M-CSF, erythropoietin, IL-3, IL-4, IL-5, and IL-6. It is useful to study the role of these factors in regulating hematopoiesis in vivo and in vitro. The promising results of GM-CSF, G-CSF, erythropoietin in clinical therapeutic application were summarized : GM-CSF and G-CSF may be useful in shortening the period of aplasia associated with ablative chemotherapy for bone marrow transplantation; the results of erythropoietin for treatment of patients with anemia due to renal failure indicate that a rise in hematocrit can be achieved with elimination of the blood transfusion requirement. IL-3, IL-4, IL-5, and IL-6 have the ability to form hemopoietic colonies in vitro, but the effects of these factors on hematopoiesis in vivo remains unclear. Further studies are necessary for clarifying the role of interleukins in regulating hematopoiesis in vivo.

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We have analyzed the alteration of the erythrocyte-glutamic-oxaloacetic transaminase (E-GOT) activity in rat hemolytic anemia model. In the model, hemolysis was induced by the intraperitoneal injection of acetylphenylhydrazine (APH). This model is very useful in studying the sequence of biochemical parameters of erythropoiesis after APH injection. Peripheral blood was withdrawn from the abdominal aorta with heparin, and E-GOT activity was measured daily. The E-GOT activity showed a peak at 7 days after APH injection. This result suggested that the measurement of E-GOT activity can be used to estimate the erythropoietic maturation process. In this study we have analyzed the E-GOT activities in patients with various anemic diseases. In hemolytic anemia, E-GOT activities were elevated in the condition of hemolytic crisis and compensated state, whereas in the uncompensated state the activities were decreased. In remission state without hemolysis the activities were normal. In aplastic anemia, patients' E-GOT activities were higher in the active state compared to the mild and remission states. A patient with renal anemia diagnosed as membranous proliferative gromerular nephritis with chronic renal failure showed very low E-GOT activity. In acute lymphoblastic leukemia, E-GOT activities were decreased at the onset but gradually recovered during induction chemotherapy and reached a normal level in the complete remission state. We have already reported that the erythrocyte protoporphyrin (FEP) and the reticulocytes measurement are good parameters for understanding the erythropoietic process. This paper indicates E-GOT is another useful parameter. In conclusion, the elevation of the E-GOT activity accompanied by reticulocytosis indicates the enhanced state of erythropoiesis. On the other hand, the elevation of the E-GOT without reticulocytosis reflects the erythropoietic maturation arrest. The decreased level of E-GOT with depressed reticulocytosis indicates erythropoietic suppression.

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Hemostatic function was studied serially in 25 children with acute lymphoblastic leukemia (ALL) receiving vincristine (VCR : day 1, 8, 15, 22), prednisolone (PDN : day 1-28 with tapered reduction), and L-asparaginase (L-Asp : day 22-31) as initial induction chemotherapy. As a result, L-Asp administration was followed by prolonged prothrombin time and activated partial thromboplastin time, decreased plasma fibrinogen, antithrombin-III and plasminogen levels. The change in platelet counts and increase of fibrinogen degradation products in serum were not observed. These hemostatic abnormalities returned to normal within 2 weeks after completion of L-Asp. On the other hand, plasma fibrinogen levels decreased remarkably before L-Asp administration only with VCR and PDN treatment. Early in the induction chemotherapy, leukemic cell lysis was suspected to be related with the accelerated fibrinogen consumption and decrease in plasma fibrinogen level. But it is considered that VCR reduces the production of fibrinogen.

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Eight children with severe aplastic anemia or acute leukemia underwent bone marrow transplantation (BMT) from major ABO incompatible donors. Removal of red blood cells from donor marrow was performed by hydroxyethyl starch (HES) sedimentation following processing with HM30 (HV50) blood cell separator. Marrow infusion caused no adverse effect except transient mild hemoglobinuria in one patient. No remarkable hemolysis was noted during the erythroid recovery stage. Durable engraftment was obtained in all children, but marked delay of erythroid recovery was observed in one patient who had had a high titer of pre-transplant anti-red blood cell antibody. The other three children with minor ABO incompatibility underwent BMT after removal of plasma from donor marrow by centrifugation. Marrow infusion was achieved safely and no hemolytic episode occurred.

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Seven children with advanced solid tumors (3 malignant lymphoma, 1 hepatocellular carcinoma, 1 hepatoblastoma, 1 extraskeletal Ewing's sarcoma, and 1 sacrococcygeal malignant teratoma) and three children with acute leukemia (AL) received high-dose chemotherapy followed by autologous bone marrow transplantation (ABMT). Anti-cancer drugs of conditioning regimens were chosen ac-cording to the results of in vitro sensitivity test using MTT dye reduction methods. In AL cases, the residual leukemic cells were purged by 4-hydroperoxycyclophosphamide. In 7 patients with solid tumors, 3 patients achieved complete remission, two had partial remission, one relapsed, and one showed no response. In 3 AL, only one patient continued complete remission. Side effects of ABMT were few and tolerable. Clinical trials of intensive chemotherapy plus ABMT might produce encouraging results in terms of response rate and long-term survival for selected patients with poor prognostic malignant diseases which are resistant or refractory to conventional forms of cancer therapy.

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Hematopoietic stem cells in bone marrow in a matured teratoma were studied in an 8-month-old girl. Similar numbers of erythroid and granulocytes-macrophage derived colonies were recognized as compared with normal bone marrow in children.

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To evaluate the effects of M-CSF (P-100) on neutrophil function, 13 patients who had been treated by allogeneic bone marrow transplantation were enrolled in this study. M-CSF (2×10⁵ units/ kg/day) was administered from one to 14 days posttransplantation. Neutrophil function was examined by microcalorie production assay using a thermo-active cell analyzer (ESCO-3000) and Candida albicans as phagocytic stimulus. Heat production of neutrophils was increased within 25 days posttransplant in the M-CSF-treated group without Candida stimulus. The increased heat production seemed to be the result of increased cell proliferation by the effect of M-CSF. Moreover, heat production increment when Candida albicans was employed as the phagocytic stimulus was also greater than that of the MCSF-non-treated group within 35 days posttransplant except in a few instances. In conclusion, M-CSF not only has a proliferative effect on neutrophils, but also increases cytokilling ability of neutrophils in some cases after bone marrow transplantation, or at least has no adverse effects on neutrophil function.

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We studied to clarify the mechanism of hemolysis with the use of ⁵¹Cr-labeled red cells and ferrokinetics, in 7 patients with hereditary spherocytosis (HS) and in 3 patients with abnormal pyruvate kinase (PK) deficiency, in pre-and post-splenectomy. In HS patients it was observed that reticulocyte counts increased, the red cell life span was reduced, and that there was an accumulation of ⁵¹Cr-labeled red cells in the spleen before splenectomy. The red cell life span returned nearly to normal after splenectomy. HS patients had levels of hemoglobin and reticulocyte counts within normal range. But in PK-deficient patients it was found that reticulocyte counts increased, the red cell life span was short or subnormal, and that abnormal PK-deficient reticulocyte was sequestrated and destroyed in the spleen and the liver before splenectomy. The red cell life span was reduced and its survival curve was bimodal (that is, short-and long-lived erythrocytes) after splenectomy. Reticulocyte count conversely increased and the level of hemoglobin was higher 1 to 2 g/dl than pre-splenectomy level. It was thought that PK-deficient patients with splenectomy had short-lived red cells of two groups. In one group, when reticulocytes reach matured red cells, those cells lost mitochondria, could not produce ATP, and were sequestrated and destroyed regularly in the liver and the bone marrow. In the other group, at the reticulocyte stage it is sequestrated and destroyed at random in the liver and the bone marrow.

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Leukemic cells from three patients with congenital leukemia without Down's syndrome were examined by ultracytochemistry. Electron microscopy showed ALL-like lymphoblasts in Patient 1, 2 types of ALL-like cells in Patient 2, and signs of basophilic leukemia in Patient 3. The myeloperoxidase reaction was negative in all 3 patients, and platelet peroxidase reaction was negative in two patients (Patients 1 and 3) examined. In Patient 3, the cells were strongly positive for acid phosphatase but weakly positive for acid mucopolysaccharides and sulfated glycoconjugates by ultracytochemistry, indicating immature basophils. Since congenital leukemia often exhibits atypical cytological features, various ultracytochemical examinations are necessary for its diagnosis.

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The Bone Marrow Transplantation Committee of the Society has conducted an annual registry of bone marrow transplantation in children in Japan since 1983. As of June 30, 1988, 410 patients had received transplants and were registered from 47 institutions. Bone marrow transplantation was performed in 114 cases of acute lymphoblastic leukemia (45 alive), 83 cases of acute nonlymphocytic leukemia (54 alive), 16 cases of chronic myelocytic leukemia (13 alive), 4 cases of juvenile-type chronic myelocytic leukemia (4 alive), 29 cases of non-Hodgkin's lymphoma (17 alive), 72 cases of malignant solid tumors (28 alive), 57 cases of aplastic anemia (52 alive), 22 cases of severe combined immune deficiency (8 alive), and 13 other cases (11 alive). The details are reported in this paper.

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A three-year-old girl developed rapidly progressing respiratory distress associated with fever and cytopenia after the completion of induction chemotherapy for acute lymphoblastic leukemia. A chest X-ray film was typical of cytomegalovirus (CMV) pneumonitis and the serial serological evaluations confirmed the infection. The patient did not show any response to the conventional intravenous immunoglobulin therapy and the condition deteriorated. Remarkable response, however, followed the subsequent administration of CMV immune globulin preparation and the patient recovered completely from the disease. This case illustrates the clinical value of the selective use of hyperimmune globulin preparations in the treatment of severe infections in immunocompromised children.

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Surface markers on leukemic cells such as E-rosette formation, surface immunoglobulin, or common ALL antigen (CALLA) have been thought to be one of the prognostic factors of acute lympho-blastic leukemia (ALL) in children. Four patients presented here were considered as ALL morpho-logically and cytochemically, but in all cases myeloid antigen MY7 (CD 13) or MY9 (CD33) were detected on the surface of the blast at first or second relapse. All patients were males and mean age of the onset was 11-years-3-months old, mean white blood cell count was 19, 500/μl, and mean duration of first remission was 23.5 months. Myeloid antigen was identified in four of eight cases of ALL which relapsed during the time from Jun. 1986 to Dec. 1987 in our hospital. It should thus be determined whether the expression of myeloid antigens on the ALL blast is one of the poor prognostic factors or not.

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This report describes a 14-year-old-boy with hemophilia B. He had been treated with a large amount of Factor IX concentrate since 6 months of age and showed a very high level of Factor IX inhibitor (185 Bethesda U/ml). He was admitted because of intracranial bleeding. We treated him successfully with “bypassing therapy” using activated and non-activated Prothrombin Complex Concentrates (Autoplex® and Proplex® ST, respectively). Based on the data from this patient and a few cases reported previously, clinical courses and therapeutic protocols for the hemophilic patients with high Factor IX inhibitor were discussed.

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A case of paroxysmal cold hemoglobinuria (PCH) was described, occurring in a 26-month old boy without a history of any previous infection and serologically negative for syphilis. After the patient was brought to a physician because of a fever and dark urine after exposure to cold weather over again, his anemia and hemoglobinuria were pointed out. He was admitted to the Department of Pediatrics, Hirosaki University School of Medicine, for further examination. Laboratory evaluation on admission revealed anemia and a high value of LDH. Direct Coombs test was negative ; the cold agglutinin activity was of low titer. Sugar water test was negative. Since Donath-Landsteiner (DL) test had been positive, Ehrlich finger test positive, and DL antibody positive, the patient was diagnosed as having PCH. Fragility of the patient's erythrocytes after cooling was demonstrated using Coil Planet Centrifuge. After the admission, his hemolytic attack disappeared by keeping warm. He was discharged on the 48th day of illness and DL antibody became negative on the 137th day of illness.

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A 6-year-old boy with acute lymphocytic leukemia developed hyperemia in the left conjunctiva bulbi during intensification therapy with vincristine and prednisolone 13 months after diagnosis. Ophthalmologic examination revealed cells in the anterior chamber. Examination of cytocentrifuged smears confirmed the presence of leukemic cells in the anterior chamber fluid. Bone marrow and cerebrospinal fluid were free of leukemic cells. Isolated eye relapse was diagnosed, and radiotherapy of 2, 000 cGy to the left eye and systemic reinduction therapy were given with intrathecal chemotherapy. Ten months later, routine follow-up examination revealed the reappearance of cells in the left anterior chamber and the presence of cells in the contralateral anterior chamber as well. At this time bone marrow and cerebrospinal fluid were again free of leukemic cells. This case illustrates that the eyes are one of the sanctuary sites independent of the central nervous system, and that the treatment of eye relapse is very difficult.

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The designation lymphoma syndrome leukemia (LSL) is applied to childhood atypical acute lymphoblastic leukemia (ALL) associated with lymphoma features. There is much controversy about whether LSL and lymphoblastic lymphoma, another representative disease of T-lymphoid malignancy, constitute and identical clinicopathological entity or not. We described here clinical features of 3 cases with LSL. All of them initially presented with massive mediastinal mass and other lymphoma-like lesions but no anemia or thrombocytopenia. Despite the induction of complete remission by conventional chemotherapy, early relapses consistently occurred. These cases became resistant to further chemotherapies, and eventually died of diffuse extramedullary involvements within 36 months after the onset. Leukemic blasts from the 3 cases were classified into French-American-British (FAB) L2 subtype of ALL, but the appearance of nuclei, nuclear chromatin, and cytoplasm somehow differed from those of leukemic blasts from typical ALL cases. Immunophenotyping showed that the leukemic blasts from the 3 cases were of T-cell lineage. LSL seems to represent a distinct clinical entity of ALL in childhood with absolutely poor prognosis, suggesting that very intensive chemotherapy or bone marrow transplantation has to be introduced as the initial treatment for this condition.

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Two children with necrotizing lymphadenitis were reported. Both patients, a 16-year-old. female (case 1) and a 15-year-old male (case 2), were admitted to the hospital because of fever and lymphadenopathy. The diagnosis of necrotizing lymphadenitis was established on the basis of the pathological finding of a lymph node. In the first case, hemophagocytosis was found in the bone marrow. The culture study of CFU-C revealed a decreased number of colony; leukopenia in peripheral blood was observed. The relationship between necrotizing lymphadenitis and other disorders with hemophagocytosis was discussed.

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A 7-year-old girl with Hb Köln, who showed mild hepatosplenomegaly and severe anemia (RBC 216 × 10⁴/mm³, Hb 5.2 g/dl, Ht 19.1%, reticulocyte 1%0) associated with an apparently upper respiratory infection, was reported. Erythroblast hyperplasia was observed in bone marrow. Further examination of hemoglobin revealed an abnormal band between Hb A and Hb A₂ in cellulose acetate membrane electrophoresis (pH 8.6). In the analysis of β chain separated by reverse-phase high-performance liquid chromatography, a change of valine to methionine, consistent with Hb Köln, was recognized. No other member of her family (father, mother, an elder brother and an elder sister) had the abnormal hemoglobin.

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Eleven patients with abdominal non-Hodgkin's lymphoma, 9 boys and 2 girls, varying in age from 4 to 11 years, were studied. In the four with intestinal primary lesion presenting with intussusception, their tumors were resectable and three are alive. In the seven with non-intestinal primary lesion, of whom two are alive, one had the partial excision, four had the biopsy alone, and two were diagnosed without laparotomy. The non-intestinal lymphomas, of which the primary masses were larger than that of the intestinal lymphoma, had more advanced intra-abdominal metastases than those in the intestinal lymphomas. Among the seven with non-intestinal lymphoma, the extra-abdominal metastasis was seen both at the mediastinal lymph nodes and the pleural cavity in four, at the inguinal nodes in one, and at the cervical nodes in the other. In the pathologic diagnosis, all eleven had the diffuse type which consisted of seven undifferentiated types (Burkitt's, 4; non-Burkitt's, 3), three large-cell types, and one lymphoblastic type. Immunological origin of the ten tumours detected consisted of seven B-cell, one T-cell, and two unclear lineages. It is suggested that whole abdominal irradiation can be avoided when an effective chemotherapy is employed.

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We report a 3-year-old boy having acute non-lymphocytic leukemia (ANLL) with both granuloid and monocytic features in the esterase activity. He presented with mild anemia, bleeding tendency, hepatomegaly and lymphadenopathy. The white blood cell count was 17, 300/μl with 34% of leukemia cells. The bone marrow contained 96% of leukemic cells which had promyelocytoid morphology. The cells were strongly positive for peroxidase. They were positive for both naphthol-AS-D-chloroacetate and α-naphthyl butyrate esterases, and the latter esterase activity was inhibited by NaF. The chromosome analysis revealed an abnormal 45, X, -Y, t (1p^-; 14q⁺), 9q⁺, 12q^- pattern. The leukemia appears to be an M3 type of ANLL with unusual cytological features.

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A 4-year-old previously healthy boy developed spontaneous inhibitors to Factor IX and lupus-like anticoagulant. A brief illness of presumed viral origin preceded hemorrhagic symptoms. Cutaneous ecchymosis and nasal bleeding were the predominant clinical features in this case. Hemorrhagic abnormalities and bruising tendency disappeared simultaneously without any treatment. Inhibitor to factor IX, approximately 1.2 Bethesda units/ml, was detected. Hypoprothrombinemia and delayed recovery of APTT made us suspect the existence of lupus-like anticoagulant. The existence of lupus-like anticoagulant was unmasked by the use of dilute thromboplastin (Tissue thromboplastin inhibition test) and was neutralized by rabbit brain phospholipid (Rabbit brain neutralization), and was sensitive to dilution. From these results, coexistence of lupus-like anticoagulant along with Factor IX inhibitor, which is a very rare event, was suggested.

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