Akinori Hoshika

Inducing proliferation of human amniotic epithelial (HAE) cells for cell therapy.

Probably because amnion is derived from the fetus and is exposed to the maternal immune system, human amniotic epithelial (HAE) cells do not express the HLA-A, -B, -C, or -DR antigens on their surfaces, suggesting that HAE cells do not induce rejection (immune reaction) after allotransplantation. And the amnion, like the placenta, is useless to the mother and child after birth. Therefore, HAE cells or tissues were expected to be suitable for allotransplantation. Because HAE cells produce large amounts of enzymes, amnion transplantation has been carried out in order to correct inborn errors of metabolism by supplementing lysosomal enzyme deficiencies. However, several problems remain before amnion allotransplantation can be accepted as effective. The HAE cell population is limited, because the maximum number of HAE cells obtainable from one donor is about 2 × 108 cells, and HAE cells proliferate poorly in in vitro culture. In this study, we aimed at increasing the HAE cell population in vitro. First, we investigated the effect of several cytokines on HAE cell proliferation and found that hepatocyte growth factor (HGF), epidermal growth factor (EGF), and transforming growth factor-β stimulated it, whereas IL-6 and LIF inhibited it. Second, we investigated the effects of amniotic fluid on HAE cell proliferation and observed that IL-6 in amniotic fluid inhibits it. Then, to inhibit the dying of cells, we attempted to inhibit apoptosis (one mode of cell death). Treatment with caspase III inhibitor increased the cell viability of HAE cells by 20%.

Umbilical Cord Milking Stabilizes Cerebral Oxygenation and Perfusion in Infants Born before 29 Weeks of Gestation

OBJECTIVE To investigate the effects of umbilical cord milking at birth on cerebral perfusion and systemic perfusion in very low birth weight (VLBW) infants. STUDY DESIGN Cerebral tissue oxygenation index and cerebral fractional tissue oxygen extraction were monitored in 50 stable VLBW infants (gestational age <29 weeks, birth weight <1250 g), with 26 allocated to the milked group and 24 to the control group. We used near-infrared spectroscopy 3-6, 12, 18, 24, 36, 48, and 72 hours after birth. Left ventricular end-diastolic dimension, left ventricular ejection fraction, left ventricle (LV) Tei index (measurement of left ventricular systolic and diastolic function), left ventricular cardiac output, and superior vena cava flow were measured concurrently using echocardiography. RESULTS There were no significant differences in gestational age and birth weight between the 2 groups. Hematocrit, left ventricular end-diastolic dimension, left ventricular cardiac output, and superior vena cava flow were higher in the milked group than in the control group, with improvement in the LV Tei index despite the absence of left ventricular ejection fraction changes within 24 hours after birth. Tissue oxygenation index increased and cerebral fractional tissue oxygen extraction decreased in the milked group within 24 hours after birth. CONCLUSION Umbilical cord milking stabilized cerebral oxygenation and perfusion in VLBW infants by improving LV diastolic function by increasing LV preload.

Noninvasive monitoring of deterioration in skeletal muscle function with forearm cast immobilization and the prevention of deterioration

BackgroundIn this research inactivity was simulated by immobilizing the forearm region in a plaster cast. Changes in skeletal muscle oxidative function were measured using near-infrared spectroscopy (NIRS), and the preventative effect of the training protocol on deterioration of skeletal muscle and the clinical utility of NIRS were examined.MethodsFourteen healthy adult men underwent immobilization of the forearm of the non-dominant arm by plaster cast for 21 days. Eight healthy adult subjects were designated as the immobilization group (IMM) and six were designated as the immobilization + training group (IMM+TRN). Grip strength, forearm circumference and dynamic handgrip exercise endurance were measured before and after the 21-day immobilization period. Using NIRS, changes in oxidative function of skeletal muscles were also evaluated. Muscle oxygen consumption recovery was recorded after the completion of 60 seconds of 40% maximum voluntary contraction (MVC) dynamic handgrip exercise 1 repetition per 4 seconds and the recovery time constant (TcVO2mus) was calculated.ResultsTcVO2mus for the IMM was 59.7 ± 5.5 seconds (average ± standard error) before immobilization and lengthened significantly to 70.4 ± 5.4 seconds after immobilization (p < 0.05). For the IMM+TRN, TcVO2mus was 78.3 ± 6.2 seconds before immobilization and training and shortened significantly to 63.1 ± 5.6 seconds after immobilization and training (p < 0.05).ConclusionsThe training program used in this experiment was effective in preventing declines in muscle oxidative function and endurance due to immobilization. The experimental results suggest that non-invasive monitoring of skeletal muscle function by NIRS would be possible in a clinical setting.

Diagnostic and predictive value of CSF d-ROM level in influenza virus-associated encephalopathy.

The aim of this study was to assess the validity of serum and CSF oxidative status of patients with IE in their initial stage through the d-ROM (Diacron-Reactive Oxygen Metabolites, Italy) test, compared to those with other neurological diseases. The study was conducted on the following four groups: (1) influenza virus-associated encephalopathy (IE, n = 8), including four patients showing neurological sequelae or mortal; (2) influenza virus-associated febrile seizures (IFS, n = 11); (3) febrile convulsion (FC, n = 10): (4) enterovirus-associated encephalopathy (EE, n = 4), including one patient with neurological sequelae. The CSF d-ROM levels in the IE group were significantly higher than those in the IFS and the FC groups but not in the EE group. In addition, general laboratory findings such as leukocytes, platelets, C-reactive protein, aspartate aminotransferase, creatinine, creatinine kinase and LDH, including interleukin-6 (IL-6), were analyzed in each group. The CSF d-ROM levels in the IE group were significantly higher than those in the IFS and FC groups but not in the EE group. As for the serum d-ROM levels and general laboratory findings, with the exception of CSF IL-6 levels in IE, no significant differences were detected compared with the other groups. In patients with IE, the CSF d-ROM levels could be a valid predictive biomarker of the severity, and oxidative stress may be related to the pathogenesis of IE.

Combined Treatment of Steroids and Cyclosporine in Kimura Disease

Kimura disease is a rare but distinctive chronic eosinophilic inflammatory disorder that is characterized by tumor-like lesions in the soft tissue and lymph nodes of the head and neck or parotid gland. Recently, many immunopathogenetic features of underlying T lymphocytes and related cytokines have been noted in Kimura disease. However, few previous studies have investigated the serial levels of cytokines in children. In this report we describe an 11-year-old Japanese boy with relapsing Kimura disease. Before the diagnosis of Kimura disease, the patient had a swelling on his left neck. Steroids were effective, but the tumor relapsed within a few months as the steroids were tapered. He was treated with steroids and cyclosporine. This treatment was done by measuring serial levels of serum soluble interleukin-2 receptor, interleukin-4, interleukin-5, and eosinophil cationic protein. These results suggest the activation of T-helper cells and T-helper 2 cytokines, that after activated B cells and eosinophilic infiltration play an important role in Kimura disease, and that cyclosporine suppresses the activity of this disease.

Methylation status of the p15 and p16 genes in paediatric myelodysplastic syndrome and juvenile myelomonocytic leukaemia

Aberrant DNA methylation is frequently observed in adults with myelodysplastic syndrome (MDS), and is recognized as a critical event in the diseases pathogenesis and progression. This is the first report to investigate the methylation status of p15 and p16, cell cycle regulatory genes, in children with MDS (n = 9) and juvenile myelomonocytic leukaemia (JMML; n = 18) by using a methylation‐specific polymerase chain reaction. The frequency of p15 hypermethylation in paediatric MDS was 78% (7/9), which was comparable to that in adult MDS. In contrast, p15 hypermethylation in JMML was a rare event (17%; 3/18). In JMML, clinical and laboratory characteristics including PTPN11 mutations and aberrant colony formation were not different between the three patients with hypermethylated p15 and the others. Aberrant methylation of p16 was not detected in children with either MDS or JMML. Since p15 and p16 genes were unmethylated in two children with JMML, in whom the disease had progressed with an increased number of blasts, a condition referred to as blastic crisis, we infer that the aberrant methylation of these genes is not responsible for the progression of JMML. The results suggest that demethylating agents may be effective in most children with MDS and a few patients with JMML.

Inflammatory cytokines as predictors of resistance to intravenous immunoglobulin therapy in Kawasaki disease patients.

Kawasaki disease (KD) is an acute systemic vasculitis. Activation of the immune system is a central feature of KD. Some KD patients are resistant to initial high‐dose intravenous immunoglobulin (IVIG) treatment. The study aimed to determine the predictors of IVIG resistance.

Changes in cerebral perfusion in extremely LBW infants during the first 72 h after birth.

Cerebral perfusion and its relation with systemic circulation in extremely LBW (ELBW) infants in the early neonatal period are not well understood. The cerebral tissue oxygenation index (TOI) and cerebral fractional tissue oxygen extraction (FTOE) were monitored in stable 16 ELBW infants (GA <29 wk) using near-infrared spectroscopy (NIRS) at 3–6, 12, 18, 24, 36, 48, and 72 h after birth. The left ventricular end-systolic wall stress (ESWS), left ventricular ejection fraction (LVEF), left ventricular cardiac output (LVCO), and superior vena cava (SVC) flow were also measured simultaneously using echocardiography. The ESWS increased till 18 h and then decreased; LVEF, LVCO, and SVC flow decreased till 12 h and increased thereafter. The TOI decreased till 12 h and correlated with SVC flow; FTOE increased until 12 h and then decreased. These changes in variables of NIRS and echocardiographic measurements contrasted to changes in mean arterial blood pressure (MABP), which showed trends of continuous and gradual increase after birth. We conclude that even stable ELBW infants undergo evident transitional changes in cerebral oxygenation and perfusion in the early postnatal period, which may reflect changes in cardiac function and cardiac output.

Low-volume muscular endurance and strength training during 3-week forearm immobilization was effective in preventing functional deterioration

PurposeThe purpose of this study was to determine whether endurance and strength hand grip exercises during 3-week upper limb immobilization preserve muscle oxidative capacity, endurance performance and strength.MethodsTen healthy adult men underwent non-dominant forearm immobilization by plaster cast for 21 days. Five healthy adult subjects were designated as the immobilization (IMM) group and five were designated as the immobilization + training (IMM+TRN) group. Grip strength, forearm circumference, dynamic handgrip endurance and muscle oxygenation response were measured before and after the 21 day immobilization period. Using near-infrared spectroscopy (NIRS), muscle oxygen consumption recovery (VO2mus) was recorded after a submaximal exercise and the recovery time constant (TcVO2mus) was calculated. Reactive hyperemic oxygenation recovery was evaluated after 5 minutes ischemia. Two training programs were performed by the IMM+TRN group twice a week. One exercise involved a handgrip exercise at 30% maximum voluntary contraction (MVC) at a rate of 1 repetition per 1 second until exhaustion (about 60 seconds). The other involved a handgrip exercise at 70% MVC for 2 seconds with a 2 second rest interval, repeated 10 times (40 seconds).ResultsThere was a significant group-by-time interaction between the IMM and IMM+TRN groups in the TcVO2mus (p = 0.032, F = 6.711). A significant group-by-time interaction was observed between the IMM and IMM+TRN groups in the MVC (p = 0.001, F = 30.415) and in grip endurance (p = 0.014, F = 9.791). No significant group-by-time interaction was seen in forearm circumference and reactive hyperemic oxygenation response either in IMM or IMM+TRN group.ConclusionThe training programs during immobilization period used in this experiment were effective in preventing a decline in muscle oxidative function, endurance and strength.

Different effects between 7-nitroindazole and L-NAME on cerebral hemodynamics and hippocampal lesions during kainic acid-induced seizures in newborn rabbits.

We examined the effects of 7-nitroindazole (7-NI) and N-omega-nitro-L-arginine methyl ester (L-NAME) on the endogenous nitric oxide (NO) production in vivo, cerebral hemodynamics, and hippocampal lesions to investigate the different roles between endothelial NOS (eNOS) and neuronal NOS (nNOS) during kainic acid (KA)-induced seizures in newborn rabbits. After a pre-treatment with 7-NI (50 mg/kg, i.p.), L-NAME (20 mg/kg, i.v.) or saline (1 ml, i.v.), KA (12 mg/kg, i.v.) was administered. NO production in the brain, regional cerebral blood flow (rCBF), cerebral oxygenation (concentrations of oxyhemoglobin (HbO2), deoxyhemoglobin (HbR), and total hemoglobin (tHb) in the brain tissue), and electroencephalography (EEG) were continuously monitored throughout the experiment lasting at least 60 min after the KA administration. There was a significant increase in NO generation in the brain during KA-induced seizures, which was inhibited by a pre-treatment with 7-NI or L-NAME. KA-induced seizures also increased rCBF significantly, which was inhibited not by 7-NI but by L-NAME. L-NAME pre-treatment caused a significant decrease in HbO2 and a significant increase in HbR during KA-induced seizures, compared with 7-NI and saline pre-treatment. EEG abnormalities and Neuronal damages in the hippocampus were significantly lower in 7-NI- and L-NAME-treated animals respectively, than in saline-treated animals. The present data demonstrated that the selective nNOS inhibitor, 7-NI, attenuated neither rCBF nor cerebral oxygenation during the seizures, while the non-selective NOS (nNOS and eNOS) inhibitor, L-NAME, attenuated both. These findings suggest that NO, probably originating from eNOS, may play an important role in the cerebral circulation. Both 7-NI and L-NAME inhibited the NO production and EEG abnormalities during the seizures that led to less damage to the hippocampus.