Preventing Brain Damage from Hypoxic-Ischemic Encephalopathy in Neonates: Update on Mesenchymal Stromal Cells and Umbilical Cord Blood Cells.

Neonatal hypoxic-ischemic encephalopathy (HIE) causes permanent motor deficit "cerebral palsy (CP)," and may result in significant disability and death. Therapeutic hypothermia (TH) had been established as the first effective therapy for neonates with HIE; however, TH must be initiated within the first 6 hours after birth, and the number needed to treat is from 9 to 11 to prevent brain damage from HIE. Therefore, additional therapies for HIE are highly needed. In this review, we provide an introduction on the mechanisms of HIE cascade and how TH and cell therapies such as umbilical cord blood cells and mesenchymal stromal cells (MSCs), especially umbilical cord-derived MSCs (UC-MSCs), may protect the brain in newborns, and discuss recent progress in regenerative therapies using UC-MSCs for neurological disorders.The brain damage process "HIE cascade" was divided into six stages: (1) energy depletion, (2) impairment of microglia, (3) inflammation, (4) excitotoxity, (5) oxidative stress, and (6) apoptosis in capillary, glia, synapse and/or neuron. The authors showed recent 13 clinical trials using UC-MSCs for neurological disorders.The authors suggest that the next step will include reaching a consensus on cell therapies for HIE and establishment of effective protocols for cell therapy for HIE. KEY POINTS: · This study includes new insights about cell therapy for neonatal HIE and CP in schema.. · This study shows precise mechanism of neonatal HIE cascade.. · The mechanism of cell therapy by comparing umbilical cord blood stem cell with MSC is shown.. · The review of recent clinical trials of UC-MSC is shown..

Metabolomic analysis and mass spectrometry imaging after neonatal stroke and cell therapies in mouse brains.

Ischemic brain injury provokes complex, time-dependent downstream pathways that ultimately lead to cell death. We aimed to demonstrate the levels of a wide range of metabolites in brain lysates and their on-tissue distribution following neonatal stroke and cell therapies. Postnatal day 12 mice underwent middle cerebral artery occlusion (MCAO) and were administered 1 × 105 cells after 48 h. Metabolomic analysis of the injured hemisphere demonstrated that a variety of amino acids were significantly increased and that tricarboxylic acid cycle intermediates and some related amino acids, such as glutamate, were decreased. With the exception of the changes in citric acid, neither mesenchymal stem/stromal cells nor CD34+ cells ameliorated these changes. On-tissue visualization with matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) imaging revealed that the signal intensity of glutamate was significantly decreased in the infarct area, consistent with the metabolomic analysis, while its intensity was significantly increased in the peri-infarct area after MCAO. Although cell therapies did not ameliorate the changes in metabolites in the infarct area, mesenchymal stem cells ameliorated the increased levels of glutamate and carnitine in the peri-infarct area. MALDI-MS imaging showed the location-specific effect of cell therapies even in this subacute setting after MCAO. These methodologies may be useful for further investigation of possible treatments for ischemic brain injury.

Autologous cord blood cell therapy for neonatal hypoxic-ischaemic encephalopathy: a pilot study for feasibility and safety.

Neonatal hypoxic-ischaemic encephalopathy (HIE) is a serious condition; many survivors develop neurological impairments, including cerebral palsy and intellectual disability. Preclinical studies show that the systemic administration of umbilical cord blood cells (UCBCs) is beneficial for neonatal HIE. We conducted a single-arm clinical study to examine the feasibility and safety of intravenous infusion of autologous UCBCs for newborns with HIE. When a neonate was born with severe asphyxia, the UCB was collected, volume-reduced, and divided into three doses. The processed UCB was infused at 12-24, 36-48, and 60-72 hours after the birth. The designed enrolment was six newborns. All six newborns received UCBC therapy strictly adhering to the study protocol together with therapeutic hypothermia. The physiological parameters and peripheral blood parameters did not change much between pre- and postinfusion. There were no serious adverse events that might be related to cell therapy. At 30 days of age, the six infants survived without circulatory or respiratory support. At 18 months of age, neurofunctional development was normal without any impairment in four infants and delayed with cerebral palsy in two infants. This pilot study shows that autologous UCBC therapy is feasible and safe.

Newborn screening for mucopolysaccharidoses: Measurement of glycosaminoglycans by LC-MS/MS.

Mucopolysaccharidoses (MPS) are a family of lysosomal storage disorders which can lead to degenerative and irreversible skeletal, cardiovascular, pulmonary, and neurological damage. Current treatments, including hematopoietic stem cell transplantation and enzyme replacement therapy, have been found most effective if administered before clinical symptoms are present, highlighting the urgent need for the development of newborn screening. This study analyzed 18,222 dried blood spot samples from newborns for both enzyme activity and glycosaminoglycan (GAG) concentration levels. GAG levels were measured using liquid chromatography tandem mass spectrometry. Results were compared to our previously established cutoff values for three subtypes of GAGs: dermatan sulfate (DS) and heparan sulfate (HS0S and HSNS). Samples that were high for two of the three GAGs were identified and screened a second time. Samples were also measured for iduronate-2-sulfatase and alfa-L-iduronidase activity. A total of 300 samples were above the established cutoff values for at least two of the three GAGs after the first screening. One sample was determined through clinical and genetic testing to be a true positive for MPS II. The false positive rate after the first GAG screening was 1.64%. A Cochran's formula test showed that the samples available for the second screening were representative samples (p = .0000601). False positive rate after second GAG screening, extrapolated from the representative sample was 0.4%. False positive rate after enzyme activity assay by fluorimetry for IDUA and IDS enzymes was 0.21% and 0.18%. A combination of GAG and enzyme assays provided no false positive and false negative samples. Two-tier screening involving a combination of enzyme activity and multiple GAGs should be considered the gold standard for the diagnosis of MPS patients.

Autophagy in the Central Nervous System and Effects of Chloroquine in Mucopolysaccharidosis Type II Mice.

Mucopolysaccharidosis type II (MPS II) is a rare lysosomal storage disease (LSD) involving a genetic error in iduronic acid-2-sulfatase (IDS) metabolism that leads to accumulation of glycosaminoglycans within intracellular lysosomes. The primary treatment for MPS II, enzyme replacement therapy, is not effective for central nervous system (CNS) symptoms, such as intellectual disability, because the drugs do not cross the blood-brain barrier. Recently, autophagy has been associated with LSDs. In this study, we examined the morphologic relationship between neuronal damage and autophagy in IDS knockout mice using antibodies against subunit c of mitochondrial adenosine triphosphate (ATP) synthetase and p62. Immunohistological changes suggesting autophagy, such as vacuolation, were observed in neurons, microglia, and pericytes throughout the CNS, and the numbers increased over postnatal development. Oral administration of chloroquine, which inhibits autophagy, did not suppress damage to microglia and pericytes, but greatly reduced neuronal vacuolation and eliminated neuronal cells with abnormal inclusions. Thus, decreasing autophagy appears to prevent neuronal degeneration. These results suggest that an autophagy modulator could be used in addition to conventional enzyme replacement therapy to preserve the CNS in patients with MPS II.

Functional neuroimaging in Rasmussen syndrome.

PURPOSE: For a diagnosis of Rasmussen syndrome (RS), clinical course together with electroencephalography (EEG) and magnetic resonance imaging (MRI) findings are considered important, but there are few reports on functional neuroimaging. This study investigated cerebral blood flow (CBF)-single photon emission computed tomography (SPECT), central benzodiazepine receptor (BZR)-SPECT, and fluorine-18 fluorodeoxy glucose-positron emission tomography (FDG-PET) in RS patients, and correlated neuroimaging results with MRI and pathological findings. METHODS: Twenty-three patients diagnosed with RS according to Bien's (2005) diagnostic criteria (including 12 patients with a histological diagnosis) were studied. CBF-SPECT, BZR-SPECT and FDG-PET images were visually evaluated, and the findings correlated with MRI and histological findings. RESULTS: Hypoperfusion areas were observed in 16 of 22 patients by interictal CBF-SPECT. Hyperperfusion areas were observed in 10 of 12 patients by ictal CBF-SPECT, which correlated with ictal onset area by ictal EEG (IOAE). In the limited data of BZR-SPECT in nine patients, lowered uptake was detected in all nine patients, including two with no MRI abnormalities. Lowered glucose metabolism was observed in affected areas in all five patients by FDG-PET. Histological examination revealed findings of chronic encephalitis in all 12 patients examined, concomitant with focal cortical dysplasia in five patients. CONCLUSION: In RS patients, functional neuroimaging reveals clear abnormal findings, even before the appearance of MRI abnormalities. BZR-SPECT and FDG-PET could detect the IOAE efficiently even in the absence of MRI abnormalities, while interictal CBF-SPECT occasionally failed to detect IOAE if MRI was normal. Based on BZR-SPECT, refractory epileptic seizures in RS may suggest possible impairment of inhibitory neurons.

Future perspectives of cell therapy for neonatal hypoxic-ischemic encephalopathy.

Neonatal ischemic brain injury causes permanent motor-deficit cerebral palsy. Hypoxic-ischemic encephalopathy (HIE) is a very serious condition that can result in death and disability. In 1997, we reported that irreversible neuronal cell damage is induced by the elevation of intracellular Ca ion concentration that has occurred in sequence after excess accumulation of the excitatory neurotransmitter glutamate during ischemia. We also reported that hypothermia was effective in treating ischemic brain damage in rats by suppressing energy loss and raising intracellular Ca ion concentration. Following the 2010 revised International Liaison Committee on Resuscitation guideline, our group developed the Guideline for the treatment of Hypothermia in Japan, and we started online case registry in January 2012. However, therapeutic hypothermia must be initiated within the first 6 h after birth. By contrast, cell therapy may have a much longer therapeutic time window because it might reduce apoptosis/oxidative stress and enhance the regenerative process. In 2014, we administered autologous umbilical cord blood stem cell (UCBC) therapy for neonatal HIE, for the first time in Japan. We enrolled five full-term newborns with moderate-to-severe HIE. Our autologous UCBC therapy is leading to new protocols for the prevention of ischemic brain damage.

Hematopoietic Stem Cell Transplantation for Patients with Mucopolysaccharidosis II.

There is limited information regarding the long-term outcomes of hematopoietic stem cell transplantation (HSCT) for mucopolysaccharidosis II (MPS II). In this study, clinical, biochemical, and radiologic findings were assessed in patients who underwent HSCT and/or enzyme replacement therapy (ERT). Demographic data for 146 HSCT patients were collected from 27 new cases and 119 published cases and were compared with 51 ERT and 15 untreated cases. Glycosaminoglycan (GAG) levels were analyzed by liquid chromatography tandem mass spectrometry in blood samples from HSCT, ERT, and untreated patients as well as age-matched controls. Long-term magnetic resonance imaging (MRI) findings were investigated in 13 treated patients (6 ERT and 7 HSCT). Mean age at HSCT was 5.5 years (range, 2 to 21.4 years) in new patients and 5.5 years (range, 10 months to 19.8 years) in published cases. None of the 27 new patients died as a direct result of the HSCT procedure. Graft-versus-host disease occurred in 8 (9%) out of 85 published cases, and 9 (8%) patients died from transplantation-associated complications. Most HSCT patients showed greater improvement in somatic features, joint movements, and activity of daily living than the ERT patients. GAG levels in blood were significantly reduced by ERT and levels were even lower after HSCT. HSCT patients showed either improvement or no progression of abnormal findings in brain MRI while abnormal findings became more extensive after ERT. HSCT seems to be more effective than ERT for MPS II in a wide range of disease manifestations and could be considered as a treatment option for this condition.

Application of a diagnostic methodology by quantification of 26:0 lysophosphatidylcholine in dried blood spots for Japanese newborn screening of X-linked adrenoleukodystrophy.

X-linked adrenoleukodystrophy (X-ALD) is a rare inherited metabolic disease that results in the accumulation of very long chain fatty acids (VLCFA) in plasma and all tissues. Recent studies regarding cerebral X-ALD (CALD) treatment emphasize the importance of its early diagnosis. 26:0 lysophosphatidylcholine (LysoPC) is a sensitive biomarker for newborn screening of X-ALD, while its application for Japanese DBS is unclear. Therefore, we evaluated the feasibility of 20:0 LysoPC and 24:0 LysoPC along with 26:0 LysoPC for diagnosing X-ALD in a cohort of newborns (n = 604), healthy adults (n = 50) and patients (n = 4). Results indicated that 26:0 LysoPC had strong significance for discrimination of patients by the amounts of 2.0 to 4.0 and 0.1 to 1.9 pmol/punch for patients and newborns/healthy adults, respectively. Based on these values, we recommend that further diagnostic confirmation is essential if the amount of 26:0 LysoPC in DBS is above 1.7 pmol/punch.

Histologic Chorioamnionitis, Amniotic Fluid Interleukin 6, Krebs von den Lungen 6, and Transforming Growth Factor ß1 for the Development of Neonatal Bronchopulmonary Dysplasia.

BACKGROUND: Chorioamnionitis (CAM) is an important risk factor for the development of bronchopulmonary dysplasia (BPD) in preterm infants. OBJECTIVES: To evaluate the effects of CAM on the development of BPD using interleukin 6 (IL-6), Krebs von den Lungen 6 (KL-6), and transforming growth factor ß1 (TGF-ß1) in the amniotic fluid as markers for inflammation, lung injury, and fibrosis/remodeling, respectively. METHODS: Amniotic fluid concentrations of IL-6, KL-6, and TGF-ß1 were measured with enzyme-linked immunosorbent assay or electro-chemiluminescence immunoassay. RESULTS: Of the 36 preterm infants, 18 were exposed to histologically confirmed CAM. Of these, 12 were later diagnosed as having BPD. The IL-6, KL-6, and TGF-ß1 levels in the amniotic fluid significantly increased with increasing histologic severity of CAM. Moreover, these markers were higher in the BPD group with histologic CAM than those without. CONCLUSIONS: Our study suggests that CAM is likely to induce inflammatory, injury, and remodeling processes in the fetal lung.

Differences between Pediatricians and Internists in Advance Care Planning for Adolescents with Cancer.

OBJECTIVE: To evaluate differences between pediatricians and internists in the practice of and barriers to advance care planning (ACP) for adolescent patients with cancer. STUDY DESIGN: A self-reported questionnaire was administered to assess the practice of ACP, advance directives, and barriers to ACP for adolescent patients with cancer. All 3392 Japanese board-certified hematologists were surveyed, and 600 hematologists (227 pediatricians, 373 internists) who take care of adolescent patients with cancer with decision-making capacity were analyzed. RESULTS: If a patient's prognosis for survival was <3 months, pediatricians were significantly less likely to discuss ACP with their patients than internists, including discussions regarding the patient's medical condition (59% vs 70%), the patient's understanding of his/her medical condition (55% vs 66%), do not attempt resuscitation orders (17% vs 24%), and ventilator treatment if the patient's condition worsened (19% vs 25%). More than 75% of hematologists (both pediatricians and internists) discussed all ACP topics with patients' families. Similarly, with regard to advance directives, pediatricians were less likely than internists to discuss cardiopulmonary resuscitation (24% vs 47%) and the use of ventilators (31% vs 51%), vasopressors (24% vs 42%), and antibiotics (21% vs 31%) with their patients. Both pediatricians and internists discussed these issues more often with patients' families than with patients, especially cardiopulmonary resuscitation (98%) as well as the use of ventilators (98%) and vasopressors (91%). CONCLUSIONS: Pediatricians were less likely than internists to discuss ACP and advance directives with patients, and both pediatricians and internists tended to discuss ACP and advance directives more often with patients' families.

Double pancreatic tumors in an adolescent: Imaging features.

Insulinoma is generally identified as a single tumor and seldom occurs in children or adolescents. A 14-year-old girl with difficulty in waking was found to have hyperinsulinemic hypoglycemia. On abdominal ultrasonography two hypoechoic masses (8 and 12 mm in diameter) were seen in the pancreatic body: the larger mass was hypervascular, whereas the smaller one was hypovascular. Contrast-enhanced computed tomography showed enhancement of the larger mass, but did not delineate the smaller mass. On fat-suppressed T1-weighted magnetic resonance imaging, the larger mass was hypointense, but the smaller mass was hyperintense. Pathologically, the larger tumor was normal density, insulin positive, and rich in vascularity, whereas the smaller tumor was high density, insulin negative, and poor in vascularity. The present case suggests that difficulty waking should be considered as a potential etiology in insulinoma, and multiple suspected pancreatic insulinomas should be evaluated using a combination of imaging modalities to characterize each tumor.

Diabetic pregnancy activates the innate immune response through TLR5 or TLR1/2 on neonatal monocyte.

Diabetes mellitus (DM) during pregnancy causes congenital malformation, macrosomia, respiratory distress syndrome, and other abnormalities in neonates, but whether maternal DM affects the neonatal innate immune system is unknown. Therefore we aimed to reveal the influence of DM in pregnancy on the toll-like receptor (TLR)-mediated innate immune response in neonates. Cord blood was collected after full-term vaginal or cesarean delivery and classified into a DM group (n=8) and non-DM (control) group (n=7). Mononuclear cells were harvested from cord blood by using density gradient centrifugation, after which anti-CD14 magnetic beads were used to isolate monocytes from the mononuclear population. After monocytes were cultured with lipopolysaccharide (TLR4 ligand), flagellin (TLR5 ligand), Pam3CSK4 (TLR1/TLR2 ligand), zymosan (TLR2/TLR6 ligand), or macrophage-activating lipopeptide (TLR2/TLR6 ligand) for 12h, the cytokine levels (interleukin [IL]-8, IL-6, IL-1ß, IL-10, tumor necrosis factor alpha and IL-12) in the culture supernatants were measured. Compared with the control group, the DM group had higher concentrations of IL-8 (P=0.01) and tumor necrosis factor alpha (P=0.02) after monocyte cultures were stimulated with Pam3CSK4 and higher concentrations of IL-8 (P=0.01) after flagellin treatment. In contrast, stimulation with lipopolysaccharide, zymosan, or macrophage-activating lipopeptide did not lead to any difference in cytokine profiles between the two groups. These data indicate that maternal DM induces excessive inflammatory activation in neonates via a TLR5- or TLR1/2-mediated innate immune response.

Activity of daily living for Morquio A syndrome.

The aim of this study was to evaluate the activity of daily living (ADL) and surgical interventions in patients with mucopolysaccharidosis IVA (MPS IVA). The factor(s) that affect ADL are age, clinical phenotypes, surgical interventions, therapeutic effect, and body mass index. The ADL questionnaire comprises three domains: "Movement," "Movement with cognition," and "Cognition." Each domain has four subcategories rated on a 5-point scale based on the level of assistance. The questionnaire was collected from 145 healthy controls and 82 patients with MPS IVA. The patient cohort consisted of 63 severe and 17 attenuated phenotypes (2 were undefined); 4 patients treated with hematopoietic stem cell transplantation (HSCT), 33 patients treated with enzyme replacement therapy (ERT) for more than a year, and 45 untreated patients. MPS IVA patients show a decline in ADL scores after 10years of age. Patients with a severe phenotype have a lower ADL score than healthy control subjects, and lower scores than patients with an attenuated phenotype in domains of "Movement" and "Movement with cognition." Patients, who underwent HSCT and were followed up for over 10years, had higher ADL scores and fewer surgical interventions than untreated patients. ADL scores for ERT patients (2.5years follow-up on average) were similar with the-age-matched controls below 10years of age, but declined in older patients. Surgical frequency was higher for severe phenotypic patients than attenuated ones. Surgical frequency for patients treated with ERT was not decreased compared to untreated patients. In conclusion, we have shown the utility of the proposed ADL questionnaire and frequency of surgical interventions in patients with MPS IVA to evaluate the clinical severity and therapeutic efficacy compared with age-matched controls.

Glucocorticoids and erythropoietin in chronic lung disease of prematurity: Proliferative potential in lung fibroblast and epithelial cells exposed to tracheal aspirates.

BACKGROUND: We investigated the effects of glucocorticoids, erythropoietin (EPO) and spironolactone (SPL) n human fetal lung fibroblasts and human alveolar epithelial cells exposed to tracheal aspirate fluid (TAF) from extremely premature infants with chronic lung disease (CLD), characterized by fibrosis and changes in the alveolar epithelium. METHODS: Fibroblasts and epithelial cells (FHs 738Lu and A549, respectively) were treated with different concentrations of hydrocortisone (HDC), dexamethasone (DEX), betamethasone (BET), SPL, and EPO in the absence or presence of TAF from infants with CLD (gestational age, 25.3 ± 0.8 weeks; birthweight, 658 ± 77 g; postnatal age, 0-28 days) and assayed for proliferation. RESULTS: Exposure to TAF resulted in a concentration-dependent proliferation of fibroblasts and epithelial cells. Proliferation of TAF-exposed fibroblasts was suppressed most significantly by 100 µmol/L DEX (21%, P = 0.046) and 300 mIU/mL EPO (18%, P = 0.02) and promoted most significantly by 0.4 µmol/L HDC (10%, P = 0.04). Epithelial proliferation was promoted by 4 µmol/L HDC (15%, P = 0.04), 10 µmol/L DEX (53%, P < 0.01), 0.2 µmol/L BET (56%, P < 0.01), and 300 mIU/mL EPO (35%, P < 0.01) in the presence of TAF. Treatment with glucocorticoids alone did not significantly affect fibroblast proliferation. CONCLUSIONS: Glucocorticoids and EPO reduced fibroproliferation while promoting epithelial cell growth in vitro within certain dose ranges. Appropriate doses of glucocorticoids and EPO may be useful in the prevention and resolution of CLD in extremely premature infants.

Hematopoietic stem cell transplantation for Morquio A syndrome.

Morquio A syndrome features systemic skeletal dysplasia. To date, there has been no curative therapy for this skeletal dysplasia. No systemic report on a long-term effect of hematopoietic stem cell transplantation (HSCT) for Morquio A has been described. We conducted HSCT for 4 cases with Morquio A (age at HSCT: 4-15years, mean 10.5years) and followed them at least 10years (range 11-28years; mean 19years). Current age ranged between 25 and 36years of age (mean 29.5years). All cases had a successful full engraftment of allogeneic bone marrow transplantation without serious GVHD. Transplanted bone marrow derived from HLA-identical siblings (three cases) or HLA-identical unrelated donor. The levels of the enzyme activity in the recipient's lymphocytes reached the levels of donors' enzyme activities within two years after HSCT. For the successive over 10years post-BMT, GALNS activity in lymphocytes was maintained at the same level as the donors. Except one case who had osteotomy in both legs one year later post BMT, other three cases had no orthopedic surgical intervention. All cases remained ambulatory, and three of them could walk over 400m. Activity of daily living (ADL) in patients with HSCT was better than untreated patients. The patient who underwent HSCT at four years of age showed the best ADL score. In conclusion, the long-term study of HSCT has demonstrated therapeutic effect in amelioration of progression of the disease in respiratory function, ADL, and biochemical findings, suggesting that HSCT is a therapeutic option for patients with Morquio A.

Zymosan Induces Immune Responses Comparable with Those of Adults in Monocytes, Dendritic Cells, and Monocyte-Derived Dendritic Cells from Cord Blood.

OBJECTIVE: To investigate the differences in toll-like receptor (TLR)-mediated immune responses between human neonates and adults, focusing on the cytokine profiles of monocytes, dendritic cells (DCs), and monocyte-derived DCs (MoDCs) in cord and adult blood. STUDY DESIGN: Purified monocytes, DCs, and MoDCs were stimulated with the following TLR ligands: lipopolysaccharide (TLR4), Pam3CSK4 (TLR1/2), flagellin (TLR5), zymosan (TLR2), polyinosinic:polycytidylic acid (TLR3), imiquimod (TLR7), and CpG (TLR9). Interleukin (IL)-8, IL-6, tumor necrosis factor, IL-1ß, and IL-10 concentrations were analyzed in culture supernatants. RESULTS: Compared with the effects in adult blood, lipopolysaccharide-, Pam3CSK4-, flagellin-, and polyinosinic:polycytidylic acid-stimulated inflammatory cytokine production in cord blood was generally weak in monocytes, comparable in DCs, and elevated in MoDCs. CpG- and imiquimod-stimulated cytokine production in DCs was comparable in cord blood and adult blood, but cytokine production was almost absent in monocytes and MoDCs in both cord blood and adult blood. In contrast, zymosan stimulation produced comparable inflammatory cytokine profiles in the monocytes, DCs, and MoDCs of cord blood and adult blood. CONCLUSION: The immaturity of TLR-mediated innate immunity in neonates depends on monocytes rather than on DCs. Our results indicate that zymosan-mediated TLR2 signaling may be useful for developing a neonatal vaccine adjuvant.

Successful emergency combined therapy with partial splenic arterial embolization and endoscopic injection therapy against a bleeding duodenal varix in a child.

There is no consensus guidelines for treating duodenal variceal bleeding, which is a rare and life-threatening complication of portal hypertension. Here we report an exceedingly unusual case in a 9-year-old boy who had developed left-sided portal hypertension after surgical treatment for pancreatoblastoma followed by a duodenal variceal bleeding with massive melena, severe anemia (hemoglobin 4.5 g/dL) and hypovolemic shock. Emergency partial splenic arterial embolization (PSE) provided a reduction of variceal bleeding and improved blood pressure. Endoscopic injection sclerotherapy (EIS) was subsequently performed and stopped the duodenal variceal bleeding without the complication of portal vein thrombosis caused by injected sclerosant under hepatopetal flow. Our case demonstrates that emergency combined therapy with PSE and EIS can be considered as the therapeutic option for the management of left-sided portal hypertension-induced ectopic variceal bleedings in order to avoid the complication of portal embolization by EIS and provide effective hematostasis.

Hepatic focal nodular hyperplasia with congenital portosystemic shunt.

Hepatic focal nodular hyperplasia (FNH) is a rare benign tumor in children. Vascular anomalies have been identified as pathological features of FNH, but the etiology remains unclear. We describe a rare case including the time course of formation of hepatic FNH in response to congenital portosystemic shunt (PSS). A 4-month-old girl was identified on newborn mass screening to have hypergalactosemia, but no inherited deficiencies in galactose-metabolizing enzymes were found. Ultrasonography and per-rectal portal scintigraphy showed intrahepatic PSS of the right lobe as a cause of the hypergalactosemia. At age 12 months, the patient had elevated hepatic enzymes and small hypoechoic hepatic lesions around the shunt. On abdominal contrast-enhanced ultrasonography spoke-wheel sign and central stellate scar were seen, which are typical features of hepatic FNH without biopsy. Congenital intrahepatic PSS should be evaluated on abdominal contrast-enhanced ultrasonography and observed over time because of its potential to develop into hepatic FNH.

Biased hypermutation occurred frequently in a gene inserted into the IC323 recombinant measles virus during its persistence in the brains of nude mice.

Measles virus (MV) is the causative agent of measles and its neurological complications, subacute sclerosing panencephalitis (SSPE) and measles inclusion body encephalitis (MIBE). Biased hypermutation in the M gene is a characteristic feature of SSPE and MIBE. To determine whether the M gene is the preferred target of hypermutation, an additional transcriptional unit containing a humanized Renilla reniformis green fluorescent protein (hrGFP) gene was introduced into the IC323 MV genome, and nude mice were inoculated intracerebrally with the virus. Biased hypermutation occurred in the M gene and also in the hrGFP gene when it was inserted between the leader and the N gene, but not between the H and L gene. These results indicate that biased hypermutation is usually found in a gene whose function is not essential for viral proliferation in the brain and that the location of a gene in the MV genome can affect its mutational frequency.