Fms-like tyrosine kinase 3 ligand stimulation induces MLL-rearranged leukemia cells into quiescence resistant to antileukemic agents.

Fms-like tyrosine kinase 3 (FLT3) is highly expressed in acute lymphoblastic leukemia with the mixed-lineage leukemia (MLL) gene rearrangement refractory to chemotherapy. We examined the biological effect of FLT3-ligand (FL) on 18 B-precursor leukemic cell lines with variable karyotypic abnormalities, and found that nine of nine MLL-rearranged cell lines with wild-type FLT3, in contrast to other leukemic cell lines, are significantly inhibited in their proliferation in a dose-dependent manner by FL. This inhibition was due to induction of the G0-G1 arrest. A marked up-regulation of p27 by suppression of its protein degradation and an abrogation of constitutive signal transducers and activators of transcription 5 phosphorylation were revealed in arrested leukemia cells after FL stimulation. Importantly, FL treatment rendered not only cell lines but also primary leukemia cells with MLL rearrangement resistant to chemotherapeutic agents. MLL-rearranged leukemia cells adhering to the bone marrow stromal cell line, which expresses FL as the membrane-bound form, were induced to quiescent state resistant to chemotherapeutic agents, but their chemosensitivity was significantly restored in the presence of neutralizing anti-FL antibody. The FL/FLT3 interaction between leukemia cells and bone marrow stromal cells expressing FL at high levels should contribute, at least in part, to persistent minimal-residual disease of MLL-rearranged leukemia in bone marrow.

Roles of Bim in apoptosis of normal and Bcr-Abl-expressing hematopoietic progenitors.

Bcr-Abl kinase is known to reverse apoptosis of cytokine-dependent cells due to cytokine deprivation, although it has been controversial whether chronic myeloid leukemia (CML) progenitors have the potential to survive under conditions in which there are limited amounts of cytokines. Here we demonstrate that early hematopoietic progenitors (Sca-1(+) c-Kit(+) Lin(-)) isolated from normal mice rapidly undergo apoptosis in the absence of cytokines. In these cells, the expression of Bim, a proapoptotic relative of Bcl-2 which plays a key role in the cytokine-mediated survival system, is induced. In contrast, those cells isolated from our previously established CML model mice resist apoptosis in cytokine-free medium without the induction of Bim expression, and these effects are reversed by the Abl-specific kinase inhibitor imatinib mesylate. In addition, the expression levels of Bim are uniformly low in cell lines established from patients in the blast crisis phase of CML, and imatinib induced Bim in these cells. Moreover, small interfering RNA that reduces the expression level of Bim effectively rescues CML cells from apoptosis caused by imatinib. These findings suggest that Bim plays an important role in the apoptosis of early hematopoietic progenitors and that Bcr-Abl supports cell survival in part through downregulation of this cell death activator.

Induction of impaired membrane phospholipid asymmetry in mature erythrocytes after chemotherapy.

Senescent erythrocytes undergo a loss of phospholipid asymmetry in the plasma membrane and are removed from the circulation by phagocytosis.To examine the loss of phospholipid asymmetry in mature erythrocytes after chemotherapy, we monitored phosphatidylserine (PS)-exposing erythrocytes by using flow cytometry to detect annexin V-bound erythrocytes in the circulation of acute lymphoblastic leukemia patients after consolidation chemotherapy. Both the percentage and the absolute number of annexin V-positive erythrocytes gradually increased immediately after chemotherapy. This result paralleled the change in the serum levels of bilirubin, suggesting a direct induction of PS-externalization in mature erythrocytes and a subsequent increase in erythrocyte clearance by splenic macrophages. The PS-exposing erythrocyte counts showed negative correlations to platelet and reticulocyte counts, which reflect the hematopoietic potential in the bone marrow. This result suggests that bone marrow suppression could be responsible for the enlarged senescent population in the circulation. Moreover, PS-exposing erythrocytes in the circulation remained at relatively high levels even after the serum level of bilirubin normalized, indicating that the decreased clearance of senescent erythrocytes as a result of impaired phagocytosis following bone marrow suppression might also be involved in the increase in PS-exposing erythrocytes in the circulation late after chemotherapy.

Sequential 3-D MRI frontal volume changes in subacute sclerosing panencephalitis.

In patients with subacute sclerosing panencephalitis, clinical stages defined according to Jabbour correlate strongly with frontal lobe dysfunctions. Non-quantitative radiologic investigations in these patients have confirmed cerebral atrophy without identifying predominantly affected regions. We addressed this issue over an 8-year-old boy's course using volumetry based on three-dimensional T1-weighted gradient echo magnetic resonance imaging. Seven normal 6-12-year-old subjects served as controls. Whole-brain volume declined as Jabbour stage advanced from I to III. Frontal lobe volume and frontal-to-whole-brain volume ratios fell significantly as clinical stage progressed. Thus, cerebral atrophy in this SSPE patient was predominantly frontal, and paralleled clinical progression.

Lateralization of the frontal lobe functions elicited by a cognitive bias task is a fundamental process. Lesion study.

Cognitive neuroscience researchers have hypothesized that context-dependent and context-independent response selection is associated with the left and right frontal lobe, respectively, in right-handed adult males. Patients with left frontal lobe lesions show context-independent reasoning in a cognitive bias task (CBT), while those with right frontal lesions show context-dependent reasoning. Young children show more context-independent responses in a modified CBT (mCBT), while adolescents and adults show more context-dependent responses. We investigated the cognitive bias of right-handed children with unilateral frontal lobe lesions/epileptic foci to explore the plasticity of lateralization in the frontal lobes. The study included eight children with left frontal lobe lesions/epileptic foci (LLF) and four children with right frontal lobe lesions/epileptic foci (RLF). Twenty-three right-handed age-matched males served as controls. A computer presented version of the original card-choice task that was simplified and modified for children was used (mCBT). Simple visual stimuli differed dichotomously in shape, color, number, and shading. A target object presented alone was followed by two choices from which subjects made selection based on preference. Considering all four characteristics, the degree of similarity between the target and the subjects' choice was scored for 30 trials. A high score indicated a context-dependent response selection bias and a low score indicated a context-independent bias. The RLF subjects had a higher converted score (mean: 26.8+/-2.2), while LLF subjects showed a lower converted score (mean: 7.75+/-6.3). There were highly significant differences between LLF subjects and the other groups (P<0.001 vs. controls or RLF subjects). No significant correlations were observed between the converted scores and the age at onset, time since insult, or IQ in either LLF or RLF subjects. These findings suggest that the lateralization of frontal lobe function elicited by mCBT is fundamental and independent of language lateralization, rather than secondary to it. Furthermore, these findings also indicate that the timetable for the development of lateralized frontal lobe functions depends upon biologic factors.

[Study of saccadic eye movement in children with attention deficit/hyperactivity disorder].

Recent studies have postulated that impairment of behavior inhibition, leading to executive dysfunction in future, is the fundamental pathophysiology of attention deficit/hyperactivity disorder (AD/HD). To evaluate this, we studied two saccadic eye movement tasks: visual- and memory-guided, in 8 children with AD/HD (combined type;mean age, 9.1 years, range 6 approximately 11 years) and 16 healthy children (mean age, 9.2 years, range 6 approximately 12 years). In the memory-guided saccade task, reflexive anticipatory errors were more frequently observed in AD/HD than in controls. The children with AD/HD showed longer response latency in the memory-guided saccade task, and greater coefficient of variation in both the latency and amplitude in both the tasks. These results suggested that saccade tasks, especially the memory-guided task, may be useful in clarifying pathophysiology of AD/HD.

Effects of sodium in hydration solution on plasma methotrexate concentrations following high-dose methotrexate in children with acute lymphoblastic leukemia.

PURPOSE: To test whether a higher sodium dose in the hydration solution may facilitate faster methotrexate (MTX) elimination as compared with a lower sodium dose following high-dose MTX (HDMTX) treatment. METHODS: Intravenous solutions with alternate doses of sodium (regimen A 70 mEq/l, regimen B 100 mEq/l) were given to 30 children with acute lymphoblastic leukemia in two courses of HDMTX in a randomized crossover fashion. The plasma MTX concentrations every 24 h from the beginning of MTX administration and the adverse events associated with HDMTX were compared between the two hydration regimens. RESULTS: The plasma MTX concentrations were similar in the two hydration regimens at 24 h (A 50.9+/-7.4 vs B 40.9+/-5.4 microM, means+/- SE, P=0.17), but was significantly lower in regimen B at 48 and 72 h (A 0.65+/-0.17 vs B 0.27+/-0.03 microM, P=0.04; and A 0.14+/-0.03 vs B 0.05+/-0.01 microM, P=0.003). The time during which MTX plasma concentrations exceeded 0.1 microM was significantly longer in regimen A than in regimen B (A 3.83+/-0.18 vs B 3.13+/-0.06 days, P=0.001). The incidences of adverse events were similar between the two regimens ( P=0.78), and severe adverse events were not seen in either regimen. CONCLUSIONS: Hydration with a higher sodium dose facilitated faster MTX elimination following HDMTX. Sodium may have a beneficial effect on MTX-induced nephrotoxicity.

Nuclear accumulation of beta-catenin without an additional somatic mutation in coding region of the APC gene in hepatoblastoma from a familial adenomatous polyposis patient.

The APC (adenomatous polyposis coli) gene status in a familial adenomatous polyposis (FAP) patient who developed hepatoblastoma was analyzed by the yeast color assay. Although a single base insertion at codon 462 resulting in truncation of its product was documented in hepatoblastoma cells, no additional somatic mutation was detectable in the whole coding sequence of the APC gene. The nuclear accumulation of beta-catenin without mutation in the exons 2-4 of the beta-catenin gene, however, was observed in the tumor cells by immunohistochemistry. The similar nuclear accumulation of beta-catenin without an additional somatic mutation in its gene, in the absence of somatic mutation in cluster region of the APC gene, has been previously reported in the single FAP case. Moreover, review in the hepatoblastoma cases in the FAP families showed a relatively later onset of the disease when compared with the sporadic cases. These observations suggest that accumulation of beta-catenin without an additional somatic mutation in the APC gene might be a possible mechanism for tumorigenesis of hepatoblastoma in the FAP families.

The association of Ala45Thr polymorphism in NeuroD with child-onset Type 1a diabetes in Japanese.

Recently Iwata et al. reported that the polymorphism in NeuroD exon 2(Ala45Thr) was associated with adult-onset Type 1 diabetes in Japanese. Furthermore, the mutations in the NeuroD as a regulator of insulin transcription have been reported to result in Type 2 diabetes. We, therefore, aimed to clarify the role of this Ala45Thr polymorphism in the susceptibility to Type 1a, immune-mediated, diabetes of child-onset Japanese patients. Eighty patients with child-onset Type 1 diabetes were examined along with 121 non-diabetic subjects as the controls. The polymorphism in Ala45Thr was defined using the PCR-RFLP method. The GAD Ab, IA-2 Ab, HLA-DRB1 genotypes and residual beta-cell function at 3 years from onset were evaluated in relation to the difference in this polymorphism. The frequency of the Ala45Thr heterozygotes was significantly higher in the Type 1 diabetic patients than in the controls (21.3 versus 9.9%, P=0.0252). The frequency of loss of beta-cell function was higher in heterozygotes patients than in wild type homozygotes patients (P=0.0112). Type 1 diabetic patients with DRB1*0901 allele showed a significantly higher frequency, 27.9%, of the Ala45Thr variant than the controls (P=0.0041). In conclusion, the Ala45Thr polymorphism contributes to the risk of development of, and to the early deterioration of beta-cell function, in Type 1a diabetes among the Japanese population.

Age shifts frontal cortical control in a cognitive bias task from right to left: part I. Neuropsychological study.

Two functionally and neurally distinct cognitive selection mechanisms involve the prefrontal lobes: those based on internal representations (context dependent) and those involving exploratory processing of novel situations (context independent). We used a cognitive bias task (CBT) representing contextual reasoning to correlate lateralization with age in the frontal lobes. Subjects included 37 healthy right-handed male children and adolescents (age range, 5-18 years). Controls were 19 right-handed men from 20 to 30 years old. A computer-presented version of the original card-choice task simplified, modified for children was used (modified CBT; mCBT). Simple visual stimuli differed dichotomously in shape, color, number, and shading. A target object presented alone was followed by two choices from which subjects selected according to preference. Considering all four characteristics, similarity between target and subject choice was scored for 30 trials. A high score implied a context-dependent response selection bias and a low score, a context-independent bias. Similarity increased significantly with age. The youngest children (5-7 years) scored lower than ages from 11 years to adulthood. Between 7 and 9 years, scores began to increase with age to reach an adult level by age 13-16. Young children showed context-independent responses representing right frontal lobe function, while adolescents and adults showed context-dependent responses implicating left frontal lobe function. The locus of frontal cortical control in right-handed male subjects thus shifts from right to left as cognitive contextual reasoning develops.

Development of the prefrontal lobe in infants and children: a three-dimensional magnetic resonance volumetric study.

Relatively little is known about normal prefrontal lobe development. We used three-dimensional magnetic resonance imaging (MRI)-based brain volumetry to characterize developmental changes in prefrontal lobe volumes in infants and children. Prefrontal volumes were determined in 30 subjects aged 5 months to 18 years (221 months) and 3 adults aged 28-39 years (324-468 months). Images were acquired on a 1.5-T MRI system using T1-weighted gradient-echo sequences. Volumes of the frontal and prefrontal lobes were determined using a workstation, and the prefrontal-to-frontal volume ratio was calculated. Prefrontal lobe volume increased slowly until 8 years (96 months) of age, contrasting sharply with rapid growth between 8 and 14 years (96 and 168 months). The prefrontal-to-frontal volume ratio increased with age as a sigmoid growth curve. A prefrontal growth spurt occurs in late childhood. Knowledge of prefrontal lobe development is essential for understanding cognitive development and dysfunction.

Context-dependent reasoning in a cognitive bias task Part II. SPECT activation study.

A cognitive bias task (CBT) delineates two different cognitive selection mechanisms in the prefrontal cortex. To identify functional anatomy of context-dependent reasoning, we used technetium-99mhexamethyl- propyleneamine oxime (99mTc HM-PAO) single photon emission computed tomography (SPECT) and statistical parametric mapping. Twelve right-handed men 20-24 years old were instructed to look at a target card and then select the choice card (among two) that they preferred (modified CBT; mCBT). They also selected a choice card 2 weeks later without prior presentation of a target card (control task). In both tasks, 99mTc HM-PAO was injected intravenously about 15 s after initiation of the mCBT or control task. Brain images were obtained using a gamma camera and reconstructed by a UNIX-based workstation. Statistical analysis compared all activated images to control images. Results associated with P values of less than 0.01 (Z score > 2.36) were depicted on T1-weighted magnetic resonance images. All subjects preferred choices more similar to the target. SPECT activation occurred bilaterally in the dorsolateral prefrontal cortices and middle temporal gyri during performance of the CBT. Additionally, the left inferior prefrontal cortex and left fusiform gyrus showed significant activation compared with the control task. A neural network linking the temporal and prefrontal cortices prominently seen in the left hemisphere participates in context-dependent reasoning. Knowledge of such neural systems is essential for understanding prefrontal lobe function and dysfunction.

A variant of Fine-Lubinsky syndrome: a Japanese boy with profound deafness, cataracts, mental retardation, and brachycephaly without craniosynostosis.

A Japanese boy with a hearing deficit, cataracts, mental retardation, and brachycephaly without craniosynostosis is described. We believe that this patient represents a variant of the Fine-Lubinsky syndrome, and is the first report from a racial group other than Caucasian.

[NSE-positive lymphoblastic lymphoma in a boy with cutaneous involvement, giant splenomegaly, and hyper-gamma globulinemia].

We report a 6-year-old boy who was diagnosed as having neuron-specific enolase (NSE)-positive pro-T cell type lymphoblastic lymphoma preceded with a variety of symptoms such as skin rash, giant splenomegaly, and hyper-gamma globulinemia. He first showed cervical lymphadenopathy in June 1999, followed by a fever of unknown origin with atypical erythema, hepatosplenomegaly, and a few lymphoblastoid cells present in the bone marrow in September. However, no specific treatments were started at this point because a cervical lymph node biopsy failed to show malignancy and the patient's signs and symptoms resolved spontaneously. Two months later, oral prednisolone therapy was started due to recurrence of the fever and erythema, but resulted in exacerbation of the skin lesions and generalized lymphadenopathy. A biopsy of the right inguinal lymph node performed in January 2000 revealed proliferation of lymphoblastic cells positive for CD3, CD5 and NSE with a rearrangement of T cell receptor gene Jdelta, leading to the diagnosis of lymphoblastic lymphoma. After intensified chemotherapy, he received an autologous peripheral blood stem cell transplantation and has been in complete remission for 4 years.

[Spontaneous recovery from pancytopenia in a young female patient with paroxysmal nocturnal hemoglobinuria(PNH): changes in the GPI-anchor expression on peripheral blood cells].

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hemolytic disorder, caused by impaired cell surface expression of GPI-anchors in hematopoietic cells as a result of somatic mutation in the PIG-A gene, and often progresses into bone marrow aplasia. We experienced a girl diagnosed as having PNH with spontaneous recovery from pancytopenia, and analyzed the GPI-anchor expression on peripheral blood cells. Thrombocytopenia was first determined when she was 5-years old, and Ham and sugar water tests were negative at the age of 6 years. Subsequently, the pancytopenia slowly progressed, and the diagnosis of PNH was made at the age of 8 years based on the positive Ham test. Frame-shift of the PIG-A gene in exon 2 was confirmed in the peripheral granulocytes at the age of 11 years. Pancytopenia spontaneously subsided over two years after diagnosis, and an almost normal hematogram except for mild thrombocytopenia has been maintained for the subsequent 4 years. In periodical flow cytometric analysis of the CD55 expression on granulocytes and erythrocytes and the CD48 expression on lymphocytes, the population of the PNH clone was almost unchanged during the first two years of spontaneous recovery, but that of CD55-negative erythrocytes gradually decreased, suggesting that regression of PNH clone might be unnecessary in transient improvement of pancytopenia.

[Development of severe aplastic anemia in a girl with Hashimoto's thyroiditis and papillary thyroid carcinoma].

A 14-year-old girl was admitted because of general fatigue and cervical lymphadenopathy. She showed bilateral struma (IInd degree) and enlargement of her left cervical lymph nodes. Laboratory data revealed neutropenia (219/microliter) and thrombocytopenia (Plt 5.1 x 10(4)/microliter) with mild anemia (Hb 11.1 g/dl), and the bone marrow aspirate and biopsy specimens showed hypocellularity. In addition, auto-antibodies against thyroid peroxidase (TPO) and thyroglobulin (TG) were highly elevated. Computed tomography of the neck showed a nodule in the left thyroid lobe with marked lymphadenopathy, and fine needle aspiration biopsy demonstrated papillary thyroid carcinoma with Hashimoto's thyroiditis and metastasis to the lymph nodes. One month after left thyroid lobectomy and cervical lymphadenectomy, the patient's condition progressed to very severe aplastic anemia, and she received immunosuppressive therapy consisting of cyclosporin A and anti-thymocyte globulin. Hematologically, partial and complete responses were obtained three and six months later, respectively. Of interest, anti-TPO and TG antibody titers remarkably decreased after immunosuppressive therapy. The patient had HLA-DR 2(DRB 1*1501) and DR 8(DRB 1*0802). The former is frequently found in patients with cyclosporin A-dependent aplastic anemia, and the latter is frequently found in Asian patients with Hashimoto's thyroiditis, suggesting an underlying autoimmune background for the simultaneous outbreak of aplastic anemia and Hashimoto's thyroiditis complicated by thyroid carcinoma.

[Prevention of hepatic veno-occlusive disease by a combination of heparin and prostaglandin E1 in children undergoing hematopoietic stem cell transplantation].

The effect and safety of combination prophylaxis for hepatic veno-occlusive disease (VOD) following stem cell transplantation (SCT) was retrospectively evaluated in a total of 53 children who survived until day 30. Prophylaxis was started on the day before conditioning, and heparin (10 unit/kg/hr) alone was used in 6 patients, PGE1 (5 ng/kg/min) plus heparin plus PTX (200 mg/day) in 17 patients, lipo-PGE1 (0.5 ng/kg/min) plus heparin plus PTX in 7 patients, and lipo-PGE1 plus heparin in 23 patients. Diagnosis of VOD was made based on McDonald's criteria in 5 cases (9.4%), but not on Jones' criteria. No statistically significant difference was observed in the incidence of VOD among each prophylaxis group. The degree of VOD was mild in all of 5 cases, and all recovered with continuation of the prophylactic procedure. Each prophylactic procedure was performed without any significant adverse effect except for seizure induced by lipo-PGE1 in one patient with Lennox syndrome. Since both the incidence and fatality rate of VOD in children undergoing SCT are approximately 20% according to most of the previous reports, the present study suggests the effectiveness of combination prophylaxis.

[Measurement of the frontal and prefrontal lobe volumes in children with malnutrition by three dimensional magnetic resonance imaging scan].

To evaluate the effects of malnutrition in early life on the growth of the frontal and prefrontal lobes, we quantitatively measured the volumes of the frontal and prefrontal lobes by three dimensional (3-D) MRI in three children (1 year 2 months to 2 years 5 months) with malnutrition. The 3-D MRI data were acquired by the fast spoiled gradient recalled (SPGR) sequence using a 1.5T MR imager. The frontal and prefrontal lobe volumes were measured by the volume measurement function of the Workstation. The data obtained were compared with those of 16 normal subjects (13 children aged 5 months to 14 years, and 3 adults aged 27 to 39 years). The volumes of the frontal and prefrontal lobes in the subjects were smaller compared with age matched controls. The results suggest that malnutrition in early life affects the growth of the frontal and prefrontal lobes.

[Measurements of the frontal and prefrontal lobe volumes by three dimensional magnetic resonance imaging scan--III. Analysis of sex differences with advanced age].

To determine whether there is sex difference in the growth of the frontal and prefrontal lobes, we quantitatively measured the volume of these lobes by three dimensional (3-D) MRI in healthy 12 males (5 months to 39 years) and six females (1 year 11 months to 27 years). The left and right lobes were studied separately. The 3-D MRI data were acquired by the fast spoiled gradient recalled (SPGR) sequence using a 1.5T MR imager. The frontal and prefrontal lobe volumes were measured by the volume measurement function of the Workstation. In males, the left to right ratio (L/R ratio) of the frontal and prefrontal lobes increased with age. On the contrary, in females, L/R ratio of the frontal and prefrontal lobes showed no significant change with advancing age. These results highlighted sex-specific maturational changes of the frontal and prefrontal lobes and suggested that quantitative data on the frontal and prefrontal lobe are important in interpreting brain abnormalities in children with developmental disorders.

[Sympathetic skin response and emotional changes of visual stimuli].

We studied sympathetic skin response (SSR) to visual emotional stimuli in 11 normal children and 13 normal adults. The arousal and valence levels of original pictures were previously assessed by 20 normal children and adults. SSR appearance ratio tended to be high for pictures with high arousal and those with either high or low valence, and was higher in children than in adults. The response for pictures with high arousal and low valence was different between children and adults. These pictures were classified into 2 groups; one consisted of disgusting creatures such as worms and roaches, and another of violent scenes, symbolizing physiological and social unpleasure, respectively. In children, SSRs were evoked with the same ratio for these two groups of pictures. In adults, however, SSRs for social unpleasure pictures were evoked with a significantly higher ratio compared to those for physiological ones. These results indicate that children are more susceptible to emotional stimuli and that this susceptibility diminishes with development, suggesting the association between emotional signal and moral development.