Transient and mild reduction of consciousness during febrile illness in children.

We have studied the clinical and neuroimaging characteristics of transient and mild reduction of consciousness during febrile illness in children. We retrospectively evaluated 58 children admitted with mild reduction of consciousness within 12 h during febrile illness. 53 patients (91%) had delirious behavior, and 5 (9%) had no delirious behavior. We also compared the clinical characteristics, brain magnetic resonance imaging (MRI) findings, and electroencephalography (EEG) findings between patients with and without delirious behavior, and no statistically significant differences were observed in any of them between the 2 patient groups (P≥0.05). MRI was performed 0-4 days after onset in 23 patients. Reversible splenial or callosal and white matter lesions were observed in 2 of 3 patients without delirious behavior vs. 4 of 20 patients with delirious behavior on diffusion-weighted images. EEG was performed 0-3 days after onset in 29 patients. Transient abnormal findings were observed in 3 of 4 patients without delirious behavior vs. 11 of 25 patients with delirious behavior. In conclusion, we consider that transient and mild reduction of consciousness during febrile illness is a unique clinical group that is constituted by children both with and without delirious behavior.

Lyn is an important component of the signal transduction pathway specific to FLT3/ITD and can be a therapeutic target in the treatment of AML with FLT3/ITD.

Fms-like tyrosine kinase 3 (FLT3) is expressed in hematopoietic progenitor cells. An internal tandem duplication (ITD) of FLT3 (FLT3/ITD) is the most frequent mutation in human adult acute myeloid leukemia (AML). FLT3/ITD contributes to the constitutive activation of FLT3 itself and its downstream signal components, mitogen-activated protein kinase and signal transducers and activators of transcription 5 (STAT5), and enables interleukin (IL)-3-dependent cell lines to grow autonomously. In the present study, we showed the specific association of FLT3/ITD with Lyn, which led to the phosphorylation of Lyn in vivo. We also demonstrated that FLT3/ITD receptors displayed a higher affinity to bind to Lyn than wild-type FLT3 receptors in vitro and that this affinity was relative to the intensity of tyrosil phosphorylation of the receptor. Both treatment with small interfering RNA (siRNA) targeting Lyn and the Src family kinase inhibitor PP2 suppressed the IL-3-independent growth of FLT3/ITD-expressing 32D cells (FLT3/ITD-32D), reducing the constitutive phosphorylation of Lyn and STAT5. PP2 treatment of mice transplanted with FLT3/ITD-32D cells blocked the onset of tumors and decreased the size of established tumors. These results demonstrate that Lyn is an important component of the signal transduction pathway specific to FLT3/ITD and can be a therapeutic target in the treatment of AML with FLT3/ITD.

High-dose methotrexate therapy significantly improved survival of adult acute lymphoblastic leukemia: a phase III study by JALSG.

High-dose methotrexate (Hd-MTX) therapy has recently been applied to the treatment of adult acute lymphoblastic leukemia (ALL) based on pediatric protocols; however, its effectiveness for adult ALL has not yet been confirmed in a rigorous manner. We herein conducted a randomized phase III trial comparing Hd-MTX therapy with intermediate-dose (Id)-MTX therapy. This study was registered at UMIN-CTR (ID: C000000063). Philadelphia chromosome (Ph)-negative ALL patients aged between 25 and 64 years of age were enrolled. Patients who achieved complete remission (CR) were randomly assigned to receive therapy containing Hd-MTX (3 g/m2) or Id-MTX (0.5 g/m2). A total of 360 patients were enrolled. The CR rate was 86%. A total of 115 and 114 patients were assigned to the Hd-MTX and Id-MTX groups, respectively. The estimated 5-year disease-free survival rate of the Hd-MTX group was 58%, which was significantly better than that of the Id-MTX group at 32% (P=0.0218). The frequencies of severe adverse events were not significantly different. We herein demonstrated the effectiveness and safety of Hd-MTX therapy for adult Ph-negative ALL. Our results provide a strong rationale for protocols containing Hd-MTX therapy being applied to the treatment of adult ALL.

Irradiation inactivation of Listeria monocytogenes in low-fat ground pork at freezing and refrigeration temperatures.

Gamma radiation effectively controls Listeria monocytogenes in uncooked and in ready-to-eat foods. This study was conducted to determine if gamma radiation could be used to control L. monocytogenes in ground pork. Ground pork was contaminated with L. monocytogenes, kept at refrigeration (4 degrees C), chilling (0 degrees C), and freezing (-18 degrees C) temperatures overnight, exposed to gamma radiation and stored at 4 degrees C for 7 days, and at 0 and -18 degrees C for 60 days. Following irradiation, the meat was assayed for L. monocytogenes viable counts and lipid oxidation. A triangle test was performed to determine if sausage made from the irradiated and nonirradiated ground pork differed in sensory quality. It was observed that a 5-log reduction of L. monocytogenes viable counts would require a 3.0-kGy radiation dose. The results of a 60-day storage study of ground pork inoculated with 10(5) to 10(6) CFU of L. monocytogenes per gram indicated that counts for nonirradiated meat remained fairly constant at refrigeration, chilling, and freezing temperatures. However, irradiation of ground pork at 3.0 kGy could inactivate L. monocytogenes totally in ground pork subsequently held at all the temperatures used in this study. Lipid oxidation measurements, as determined by the thiobarbituric acid-reactive substance assay, ranged from 0.16 nmol/g for nonirradiated ground pork and 0.20 nmol/g for meat irradiated at 3.0 kGy. Sensory panelists could distinguish between irradiated and nonirradiated sausage but were divided on whether irradiation adversely affected the sausage quality. Our results suggest that gamma radiation could be useful to control L. monocytogenes in ground pork and improve the safety of ground pork products.

Development and analysis of patient-derived xenograft mouse models in intravascular large B-cell lymphoma.

Intravascular large B-cell lymphoma (IVLBCL) is a distinct disease entity with the peculiar characteristic that tumor cells proliferate within vessels. Despite recent advances in understanding the disease from clinical aspects, the underlying pathogenesis remains unknown. Here we demonstrate analyses of IVLBCL biology using four xenograft mouse models established from primary IVLBCL samples. In all four models, the main characteristic of IVLBCL tumor cell proliferation within vessels was retained. Time-lapse engraftment analyses revealed that the tumor cells initially engrafted and proliferated in the sinusoids and vessels in the liver and then engrafted and proliferated in multiple organs. Intriguingly, serial passage of tumor cells from the adrenal gland of a transplanted mouse developed from primary patient bone marrow cells into a second mouse showed that the tumor cells mainly distributed into the adrenal gland in the second mouse, implying the existence of clonal selection and/or evolution at engraftment of a specific organ. Gene expression profiling analyses demonstrated that the gene set associated with cell migration was enriched for normal peripheral blood B cells, indicating that inhibition of cell migration might be involved in IVLBCL pathogenesis. In conclusion, the mouse xenograft models described here are essential tools for uncovering IVLBCL biology.

Tandem-duplicated Flt3 constitutively activates STAT5 and MAP kinase and introduces autonomous cell growth in IL-3-dependent cell lines.

We have recently identified an internal tandem duplication of the human Flt3 gene in approximately 20% of acute myeloid leukemia (AML) cases. In the present study, the wild-type and the mutant Flt3 genes were transfected into two IL-3-dependent cell lines, 32D and BA/F3 cells. Mutant Flt3-transfected cells exhibited autonomous growth while wild-type Flt3-transfected cells with the continuous stimulation of Flt3 ligand exhibited a minimal proliferation. Cells expressing mutant Flt3 showed constitutive activation of STAT5 and MAP kinase. In contrast, Flt3 ligand stimulation caused rapid activation of MAP kinase but not STAT5 in cells expressing wild-type Flt3. Finally, we found constitutive activation of MAP kinase and STAT5 in all clinical samples of AML patients with mutant Flt3. Our study shows the significance of internal tandem duplication of Flt3 receptors for leukemia cell expansion.

c-Myb acetylation at the carboxyl-terminal conserved domain by transcriptional co-activator p300.

Transcription factor c-Myb plays important roles in cell survival and differentiation in immature hematopoietic cells. Here we demonstrate that c-Myb is acetylated at the carboxyl-terminal conserved domain by histone acetyltransferase p300 both in vitro and in vivo. The acetylation sites in vivo have been located at the lysine residues of the conserved domain (K471, K480, K485) by the use of the mutant Myb (Myb-KAmut), in which all three lysine residues are substituted into alanine. Electrophoretic mobility shift assay reveals that Myb-KAmut shows higher DNA binding activity than wild type c-Myb and that acetylation of c-Myb in vitro by p300 causes dramatic increase in DNA binding activity. Accordingly, transactivation activity of both mim-1 and CD34 promoters by Myb-KAmut is higher than that driven by wild type c-Myb. Furthermore, the bromodomain of p300, in addition to the histone acetyltransferase (HAT) domain, is required for effective acetylation of c-Myb, and hGCN5 is revealed to be a factor acetyl-transferase for c-Myb in vitro. We present a new manner of post-translational modification of the c-Myb protein and the potential significance of the acetylation in c-Myb.

DNA cleavage reaction and linoleic acid peroxidation induced by tea catechins in the presence of cupric ion.

Tea catechins with cupric ion promoted extensive DNA cleavage and fatty acid peroxidation in vitro under aerobic conditions. Neither cupric ions nor polyphenolic compounds including catechins alone induced DNA cleavage. While catalase significantly inhibited the DNA cleavage induced by catechins-Cu2+, superoxide dismutase (SOD) did not, indicating that H2O2 is probably involved in the DNA cleavage. These results suggest that the pro-oxidant property of catechins, which are generally considered to be anti-oxidants and anticarcinogens, is responsible for the O2 reducing ability of catechins catalyzed by cupric ion.

Determination of timing of brain injury in preterm infants with periventricular leukomalacia with serial neonatal electroencephalography.

OBJECTIVE: The aim of this study was to determine the timing of brain injury in infants with periventricular leukomalacia (PVL) with serial electroencephalography (EEG) recordings during the neonatal period. PATIENTS AND METHODS: We evaluated 172 preterm infants having a gestational age <33 weeks and weighing <2000 g. Initial EEG was recorded within 72 hours of life and then recorded once every 1 to 4 weeks. Serial cranial ultrasonography was performed and cystic PVL was diagnosed when multiple cystic formations of >3 mm in diameter were observed. RESULTS: Of the 172 infants studied, 26 were diagnosed as having cystic PVL by ultrasonography. EEG abnormalities were observed in 25 of 26 infants with PVL, although EEG abnormalities were seen in 20 of 146 infants without PVL. The initial EEG recordings were normal in 7 infants, but EEG abnormalities were observed later in 6 of these infants. In these 6 infants, the timing of injury was presumed to be postpartum. Only acute stage abnormalities were observed on initial EEG recording in 14 infants, and the timing of injury was presumed to be just before or around birth. Chronic stage abnormalities were recognized already on initial EEG recordings in the other 5 infants, and the timing of injury was presumed to be some time before birth. CONCLUSIONS: Our study indicates that it may be possible to determine the timing of injury in infants with PVL by serial EEG recordings.

Constitutively active STAT5A and STAT5B in vitro and in vivo: mutation of STAT5 is not a frequent cause of leukemogenesis.

We recently identified several constitutively active forms of signal transducers and activators of transcription 5 (STAT5) using polymerase chain reaction-driven random mutagenesis followed by retrovirus-mediated expression screening. All constitutively active STAT5 showed constitutive phosphorylation on their tyrosine residues and induced factor-independent growth in a mouse interleukin-3-dependent cell line, Ba/F3. Sequence analysis of these active STAT5 revealed two important mutations: S710F and N642H. The N642H mutation localized in the SH2 domain was able to induce autonomous growth of Ba/F3 cells by itself, whereas S710F in the effector domain was able to induce autonomous growth of Ba/F3 cells in concert with a second mutation including H298R and E150G. Recently, constitutive activation of STAT5 has been reported in patients' leukemic cells and is implicated in leukemogenesis. We attempted to clarify whether leukemic cells harbored activating mutations primarily in STAT5 proteins, and analyzed the sequence of STAT5 derived from 49 leukemic patients. No mutations were found, however, in the regions surrounding S710 and N642 of STAT5A and corresponding residues of STAT5B. We also cloned full-length cDNAs for STAT5s from three patients whose leukemic cells exhibited constitutive tyrosine phosphorylation of the STAT5 protein and expressed the derived STAT5 proteins in Ba/F3 cells. However, none of these clones exhibited constitutive tyrosine phosphorylation or gave rise to FI proliferation of Ba/F3 cells. These results indicate that constitutive activation of STAT5 is a secondary event in most leukemias.

Development of the monosynaptic reflex pathway in the human spinal cord.

Development of the monosynaptic reflex pathway of the spinal cord was investigated in 96 neurologically normal infants with ages ranging from 25 weeks in postconceptional age (PCA) to 24 months after full-term delivery (PDA) by examining H-reflexes from the triceps surae and hypothenar muscles in terms of their incidence, latency and maximal size in reference to the maximal M-wave. The triceps H-reflex was evoked in all cases, and the latency was longest (26 ms) in the youngest case of 25 weeks (PCA). It gradually shortened until full-term gestation, reaching the shortest value of 17 ms (mean). The H-reflex size initially increased until full-term gestation, reaching the maximum value of 70% and then reducing gradually to the plateau level of about 30% at 12 months (PDA). The hypothenar H-reflex could not be elicited until 32 weeks (PCA). The time course of changes in its latency and size was similar to those of the triceps H-reflex, except that it could not be elicited after 12 months (PDA). Thus, the monosynaptic reflex pathway is already functioning at the age of 25 weeks (PCA) in man. The significance of the systematic change in latency and excitability of the H-reflex with age is discussed.

Excretion of urinary epidermal growth factor in non-insulin dependent diabetes mellitus.

Morning urine samples were assayed for human EGF (hEGF) in 137 non-insulin dependent diabetic patients with normal serum creatinine and beta 2-microglobulin levels. Serving as controls, 80 age- and sex-matched healthy subjects were also examined. A significant positive correlation between hEGF excretion and the level of hemoglobin A1C (HbA1c) was present in those patients with a HbA1c value exceeding 8% (r = 0.37; p = 0.003) but not in the overall patients. The urinary hEGF level did not correlate with the concentration of glucose in urine or plasma. The mean urinary hEGF level of diabetic patients was significantly lower than that of healthy subjects. The mean urinary hEGF level was significantly lower in the patients with a diabetic history that exceeded five years as compared with those with a history below five years. The mean urinary hEGF level was significantly lower in the patients with retinopathy vs. those without retinopathy. In the patients with HbA1c value below 8%, the mean urinary hEGF level was lower in the patients whose urinary albumin level exceeded 1.7 mg/mmol.creatinine as compared with those whose urinary albumin excretion was below 1.7 mg/mmol.creatinine. These findings suggest that urinary hEGF excretion may decrease with the progression of diabetic complications in the patients with well-controlled glycemia, and that inadequate glycemic control may lead to an increased excretion of urinary hEGF in the early disease stage.

Effect of ascorbic acid on pituitary prolactin secretion in the non-ascorbate synthesizing Osteogenic Disorder Shionogi (ODS) rat.

We examined the secretion of prolactin (PRL) in the hereditary scurvy-prone Osteogenic Disorder Shionogi (ODS) rat to investigate the role of ascorbic acid (AsA) in pituitary lactotroph and hypothalamic function. Plasma PRL concentrations of conscious AsA-deficient or control ODS rats were measured before and after the administration of metoclopramide (MCP), 5-hydroxy-L-tryptophan (5-HTP) or saline. The basal plasma PRL levels did not differ between the two groups. However, the AsA-deficient rats exhibited significantly larger changes in plasma prolactin concentration in response to MCP or 5-HTP than the control ODS rats. Thus, AsA deficiency enhanced the stimulated, but not the basal, secretion of PRL in ODS rats. Our findings suggest that AsA may have an inhibitory effect on PRL secretion.

Background electroencephalographic (EEG) activities of very preterm infants born at less than 27 weeks gestation: a study on the degree of continuity.

AIMS: To clarify the features of the background electroencephalographic (EEG) activities in clinically well preterm infants born at less than 27 weeks gestation and to outline their chronological changes with increasing postconceptional age (PCA). METHODS: EEGs of clinically well premature infants born at less than 27 weeks gestation were recorded during the early postnatal period. The infants were separated into three groups according to their PCA at the time of EEG recording (21-22 weeks PCA, 23-24 weeks PCA, and 25-26 weeks PCA). The mean and maximum duration of interburst intervals (IBIs), the mean duration of bursts, and the percentage of continuous and discontinuous patterns in each PCA group were evaluated. RESULTS: There were three infants at 21-22 weeks PCA, seven at 23-24 weeks PCA, and five at 25-26 weeks PCA. Eighteen EEG recordings were obtained. The mean and maximum IBI duration decreased with increasing PCA. The percentage of continuous patterns increased with increasing PCA. Conversely, the percentage of discontinuous patterns decreased with increasing PCA. CONCLUSIONS: In premature infants born at less than 27 weeks gestation, the characteristics of the background EEG activities were similar to those of older premature infants. These changes reflect the development of the central nervous system in this period.

Neonatal EEG: a powerful tool in the assessment of brain damage in preterm infants.

Serial EEG recordings beginning immediately after birth are not only of great diagnostic and prognostic value but also useful to elucidate the timing and the mode of brain injuries in the preterm newborn. It is extremely useful to distinguish between acute stage and chronic stage EEG abnormalities. The former is characterized by findings of acute depression such as increased discontinuity, decreased faster frequency activities, and lowered amplitudes. The latter mainly includes dysmature patterns and disorganized patterns. The timing of brain insult can be assessed by considering EEG findings in relation to the time of birth. Different modes of brain injury are associated with different types of EEG abnormalities and different types of neurological outcome. Sudden strong brain insults are usually associated with findings of severe depression followed by disorganized pattern and later cerebral palsy, while persistent mild insults are usually associated with prolonged mild depression followed by dysmature pattern and later mental retardation. Routine serial EEG studies in preterm infants demonstrated that one fourth of cerebral palsies in these infants were of antenatal origin, two thirds of perinatal origin and postnatal injuries played the least role. Periventricular leucomalacia (PVL) manifesting itself on the ultrasound in the late neonatal period and suggesting postnatal origin was often found to be of antenatal origin with an EEG soon after birth. PVL without apparent causes was often associated with abnormal fetal heart rate patterns and early neonatal EEG abnormalities, and considered to have originated in the antepartum period.

Neurophysiological and neuroradiological features preceding infantile spasms.

Neonatal EEG, cerebral evoked potentials, CT scan findings and their evolution were investigated in 42 neonates who eventually developed the West syndrome. In infants with perinatal hypoxia, the EEG displayed most often marked or maximal depression in the first week of life. The highly abnormal initial EEG improved progressively with time. The most frequent findings in early infancy was discontinuous tracing and persistent alternating tracing at 1-2 months, and absent sigma rhythms at 3-4 months. The VEP, AEP and CT scan also showed findings suggesting severe degree of brain damage. In infants with meningitis, the initial EEG was less depressed but the EEG abnormality persisted or worsened. In infants with prenatal causes the neonatal EEG was variable, ranging from a normal background EEG to specific abnormalities. The evolution of paroxysmal abnormalities was similar in all groups. There were no epileptiform discharges in early months. The appearance of hypsarrhythmia was preceded by focal and then multifocal spike or sharp wave discharges.

Dysmature EEG pattern in EEGs of preterm infants with cognitive impairment: maturation arrest caused by prolonged mild CNS depression.

The significance of mild sustained brain injury in the pathogenesis of perinatal brain damage was investigated using serial EEG recordings performed on 102 early preterm infants surviving beyond 2 years of age. Sixteen infants (16%) elicited mild depression of background EEG activities in the neonatal period. Of nine infants with mild depression of prolonged duration (more than 3 weeks), five (56%) were diagnosed as having cognitive impairment in the follow up study. Four showed no signs of cerebral palsy, while one had cerebral palsy. The infants with cognitive impairment showed mild prolonged depressions in background EEG activities in the early neonatal period and dysmature EEG patterns in the late neonatal period. They also showed maturation arrest in EEG patterns during prolonged mild depression of background EEG activities. In addition to strong sudden depression of CNS causing deep white matter injury and motor impairment, prolonged mild depression is another mode of brain injury in early preterm infants which can induce future cognitive impairment.

Correlation between the serum level of endotoxin and periventricular leukomalacia in preterm infants.

The objective of our study was to determine the relation between the serum level of endotoxin at birth and the development of periventricular leukomalacia (PVL) in preterm infants. We studied 68 preterm infants whose gestational ages ranged between 27 and 33 weeks, and birthweights between 1000 and 2000 g. The serum endotoxin level was measured in the blood immediately after birth by means of a conventional chromogenic Limulus test. Serum endotoxin was taken as present when it was >5 pg/ml (indicated by ENDO(+)). Premature rupture of membranes (PROM), the mode of delivery, EEG findings and the development of PVL were investigated. Six infants were diagnosed as having cystic PVL on ultrasonography. Endotoxin was detected in 28 infants. PROM was observed in 18 ENDO(+) infants and 16 ENDO(-) infants (P = 0.085). Caesarean section was performed in nine ENDO(+) infants and 14 ENDO(-) infants (P = 0.29). PVL was observed in two ENDO(+) infants and four ENDO(-) infants (P > 0.99). EEG abnormalities were recognized in five ENDO(+) infants and six ENDO(-) infants (P > 0.99). Between ENDO(+) infants and ENDO(-) infants, no significant difference was observed in any of the four parameters. These results may indicate that endotoxin itself does not directly cause PVL.

Epilepsy in patients with spastic cerebral palsy: correlation with MRI findings at 5 years of age.

The aim of this study was to determine the relationship between epilepsy and the magnetic resonance imaging (MRI) findings in patients with spastic cerebral palsy at five years of age. We studied 14 patients with congenital anomaly and 116 with perinatal injury. The patients with perinatal injury were subdivided into two groups; those with preterm type injury alone (group P), and those with term type injury with or without preterm type injury (group T). Epilepsy was found in 37 of the 130 patients. The initial type of seizures was partial in 12 patients. infantile spasms in 20 and generalized in five. Kaplan-Meier analysis demonstrated that patients with congenital anomaly had a higher incidence and an earlier onset of epilepsy than those with perinatal injury. Of the patients with perinatal injury, group T patients showed a higher incidence and a later onset of epilepsy than group P patients.

Relation between the date of cyst formation observable on ultrasonography and the timing of injury determined by serial electroencephalography in preterm infants with periventricular leukomalacia.

The aim of this study was to clarify the relation between the date of cyst formation and the timing of injury in preterm infants with periventricular leukomalacia in order to address the issue of whether or not we can determine the timing of injury on the basis of ultrasonographic findings. We studied 33 preterm infants with cystic periventricular leukomalacia, the gestational ages being 32 weeks or less. As 27 of them exhibited either acute or chronic stage abnormalities in the initial electroencephalogram, the timing of injury was presumed to be before or around birth. For the remaining six infants, the timing of injury was considered to be postnatal in two infants, and was not determined in another four. The median date of cyst formation was 18 days of age (range, 10-39 days) in the 27 infants with abnormal initial electroencephalograms. For these 27 infants, the gestational age did not influence the date of cyst formation. In contrast, the date of cyst formation was significantly earlier in infants with widespread cysts than in those with localized cysts. In conclusion, it is difficult to determine the timing of injury from ultrasonographic findings, because the range of the date of cyst formation on ultrasonography was very wide among infants whose timing of injury was not greatly different.