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To determine the neuronal function of genes in vivo, the neuron-specific deletion of a target gene in animals is required. Tau, a microtubule-associated protein, is expressed abundantly in neurons but scarcely in glias and other tissues. Therefore, to generate mice that express Cre recombinase in neurons, we inserted Cre recombinase into the tau locus. By crossing these tau-Cre mice with ROSA26 lacZ reporter mice, we observed Cre recombinase activity in the neurons from most of the central nervous system, but not in glias nor in non-neuronal tissues. This neuronal-specific activity appeared during embryogenesis. We further crossed tau-Cre mice with rab8 'floxed' mice, and showed that the recombination was nearly complete in the brain, but incomplete or non-detectable in other tissues. Thus, tau-Cre knockin mouse is a useful tool for studying the neuronal function of a gene in vivo.

We present a case of pemphigus vulgaris (PV) accompanied with protein-losing gastroenteropathy (PLE). A 9-year-old girl developed multiple oral ulcerations and erosions. She was first treated with oral antibiotics and a topical steroid without improvement. Laboratory data showed eosinophilia (absolute eosinophil count 1.08 x 10(9)/L) and hypoproteinemia (total serum protein 3.9 g/dL, albumin 2.2 g/dL). A biopsy specimen from the ileum showed intense eosinophil infiltration and albumin scintigraphy demonstrated protein exduation from the same site. Endoscopic examination of the oesophagus showed multiple ulcerations and erosions, and biopsy specimen showed eosinophilic spongiosis and immunohistologic staining demonstrated deposits of IgG and C3 in the intercellular space. Antidesmoglein-3 antibody elevated, she was diagnosed as PV complicated with PLE. Immunofluorescence study of a biopsy specimen from the terminal ileum showed no significant immunoglobulin or complement deposition, and autoantibody against intestinal mucosa was unclear in this case. Gastrointestinal evaluations should be considered in patients with hypoproteinemia associated with PV.

Background: Bronchopulmonary dysplasia (BPD) is thought to be one form of developmental arrest of the lung. Hepatocyte growth factor (HGF) participates in normal lung growth and in lung regeneration. Objectives: The purpose of this study is to investigate whether HGF can improve alveolarization and attenuates functional abnormalities of a murine model of BPD induced by hyperoxia. Methods: Three-day-old CD-1 mice were exposed to 90% of oxygen or room air (control group) for 7 days. These animals were then kept in room air for the next 7 days. Recombinant human (rh) HGF (100 μg/g b.w., divided 3 times, rhHGF group) or vehicle (vehicle group) was administered intraperitoneally during hyperoxia. On day 17, the pulmonary function test and bronchial hyperresponsiveness (BHR) were examined. Mean linear intercepts (MLI) were measured as parameters of alveolarization. Cell renewal (on day 10) and vascularization of the lung were also evaluated. Results: Exposure to hyperoxia induced increased airway resistance and BHR. These animals showed a severely simplified alveolar structure, increased MLI, decreased cell renewal (16.1 ± 2.4 vs. 29.6 ± 2.4%, p < 0.05), and decreased vascularization (15.1 ± 0.3 vs. 18.4 ± 1.5 vessels/hpf, p < 0.05, vehicle vs. control group, respectively). rhHGF treatment during exposure to hyperoxia significantly reduced all of these changes (27.9 ± 1.7%, 18.2 ± 0.5 vessels/hpf for cell renewal and vascularization, respectively; all values are p < 0.05 against vehicle animals). Conclusion: HGF partially protects against the inhibition of alveolarization and improves functional abnormality in the hyperoxia-induced neonatal mice model of BPD.

Intravenous immunoglobulin is effective to resolve inflammation of Kawasaki disease and reduce the incidence of coronary artery abnormalities. However approximately 20% of patients with Kawasaki disease had persistent or recurrent fevers after intravenous immunoglobulin and are considered to have a high risk for coronary artery abnormalities. Recently, we developed a new risk score that resistance to intravenous immunoglobulin could be identified with high sensitivity and specificity in advance using seven laboratory and demographic variables available before initiation of primary therapy. In this article, we will review prediction of intravenous immunoglobulin unresponsiveness. In addition, we will focus on the risk stratification of primary therapy using the risk score.

Lung fibroblasts are a major source of several cytokines including CC chemokine eotaxin. We aimed to study the regulation of eotaxin-1/CCL11 production by dexamethasone and analyze its molecular mechanisms in human lung fibroblasts. Normal human lung fibroblast cells were exposed to IL-4 (40 ng/ml) and/or dexamethasone (10(-6)-10(-9) M), and eotaxin mRNA expression and production was evaluated. Mechanisms of transcriptional regulation were assessed by Western blotting and dual luciferase assay for eotaxin promoter. The effects of dexamethasone on suppressor of cytokine signaling (SOCS)-1 and eotaxin mRNA expression in the cells transfected with expression vector (pAcGFP1-C1) or short interfering RNA (siRNA) for SOCS-1 were also investigated. Within 24 hours, dexamethasone inhibited IL-4-induced eotaxin mRNA expression and protein production, while eotaxin production was markedly increased at 48 and 72 hours after coincubation with IL-4 and dexamethasone. IL-4-induced eotaxin promoter activity was inhibited by dexamethasone at 8 hours, but enhanced at 48 hours after coincubation. Dexamethasone suppressed SOCS-1 mRNA expression but enhanced IL-4-induced STAT6 phosphorylation at 36 to 48 hours after coincubation. Enhanced expression of eotaxin mRNA by dexamethasone 48 hours after coincubation was completely diminished in the cells transfected with either expression vector or siRNA for SOCS-1. These results indicated that dexamethasone, depending on the exposure duration, can either inhibit or enhance IL-4-induced expression and production of eotaxin in the lung fibroblasts. The mechanisms of later enhanced production may depend on the prolonged transcriptional activity of the eotaxin gene, in part due to inhibition of SOCS-1 expression.

Leukotrienes (LTs) are one of the most important mediators in the pathophysiology of asthma. We measured the intracellular amounts of reduced glutathione (GSH) and oxidized glutathione (GSSG) in cultured human airway epithelial cells. LTC 4 affects the GSH/GSSG ratio by activating signals to increase interleukin-8 (IL-8) production. Pretreatment with a reducing agent, glutathione monochrome ester (GSH-OEt), and with a leukotriene receptor antagonist, montelukast, significantly suppressed LTC(4)-induced time-dependent changes in the intracellular redox state, and also suppressed upregulation of IL-8 production by suppressing NF-kappaB activation. Our observations led to the hypothesis that LTC(4)-induced oxidative stress is likely to contribute to amplification of airway inflammation.

We studied the effects of azelastine and disodium cromoglycate on bronchial responsiveness in 26 asthmatic children, using histamine inhalation challenge.We divided the subjects into three groups. For 12 weeks, the 1st group of patients inhaled 20mg of disodium cromoglycate 4 times daily, the 2nd group took oral administrated 2mg of Azelastine twice daily and the 3rd group had no antiallergic drugs. Histamine inhalation challenges were performed before and at the end of treatment period. There was no statistical difference between the results obtained before and at the end of treatment in the group which didn't have antiallergic drugs. But the threshold of histamine had significantly increased after treatment of DSCG and Azelastine (p

Cellular adhesion is crucial roles for eosinophil effector functions such as degranulation. Here, we focused on the role of a β2 integrin, αMβ2, in intracellular signaling pathways of human eosinophil activation. We found that the ligation of αMβ2 triggers the activation of an intracellular signaling cascade, including protein tyrosine phosphorylation and phosphoinositide turnover, and subsequent cellular degranulation in human eosinophils. These signaling events may play important roles in adhesion-dependent cellular functions of eosinophils.

Object The authors report the case of a boy with PFIC type 2 or BRIC type 2 who suffered from liver dysfunction at 2 months after birth. Methods and results A liver biopsy specimen revealed mild liver cirrhosis, and the findings resembled those observed in Byler disease. Genetic examination revealed a normal familial intrahepatic cholestasis-1 gene, but a heterozygous mutation for the ABCB11, C1620A (F540L), was observed. Therefore, the patient was initially diagnosed with PFIC type 2. For 3 years after the diagnosis, he had severe pruritus, an increased serum bile acid, and normal serum values of gamma-glutamyl transaminase. At the age of 2, treatment with administration of ursodeoxycholic acid was started; subsequently, a gradual improvement in his liver function was observed. At the age of 3, he suffered from massive intestinal and pulmonary hemorrhage, which improved immediately after the administration of vitamin K. He was then admitted to our hospital for liver transplantation. At 1 month after the admission, his liver dysfunction showed further improvement, except for a mild increase in the serum bile acid level. This condition did not show any change during the 5-year follow-up period. In addition, the patient showed severe growth failure and was diagnosed with growth hormone deficiency. Hence, he receives growth hormone administration. Conclusion The patient could be genetically diagnosed with bile salt export pump disease of PFIC type 2 or BRIC type 2. Various clinical features are observed in PFIC or BRIC patients with ABCB11 mutation.

A cross-sectional survey, Asthma Insights and Reality in Japan (AIRJ) conducted in 2000 demonstrated that the level of asthma control and inhaled corticosteroid (ICS) use in Japanese asthmatics were low compared to the average in European countries. The purpose of the AIRJ survey in 2005 was to assess changes in the burden of asthma and asthma management in Japan since 2000 AIRJ survey.We used identical methods to AIRJ 2000, including random-digit dialing to identify current asthmatics throughout Japan in 2005. A detailed telephone survey which included items on the burden of asthma and clinical management was conducted in 800 households (400 adults, 400 children) from the identified asthmatics. We compared AIRJ 2005 results with those of AIRJ 2000 separately for adults and children with statistical adjustment for differences in population characteristics between AIRJ 2000 and AIRJ 2005.Participants in 2005 reported significantly lower prevalence of symptoms, including a decrease in daytime symptoms in adults (52% to 46%, p=0.046) and a decrease in night-time awaking in children (40% to 33%, p=0.039). The number of patients who experienced one or more asthma episodes in the past year (ER visits, hospitalization, or unscheduled doctor visits) decreased significantly from 2000 in adults (41% to 31%, p0.01) and children (62% to 49%, p0.001). ICS use rate increased significantly in adults (12% to 18%, p=0.019) and had a tendency to increase in children (5% to 8%, p=0.059).There have been improvements in the burden of asthma patients and clinical disease management as well as more use of ICS in Japan since 2000. Nevertheless, in most patients, ICS use still remained low, and asthma management fell short of the goals of treatment outlined in the Japanese guidelines. There is a need to increase awareness of and compliance with the asthma guidelines and to popularize standardized therapy in Japan.

The present study aimed to examine whether and how colonic motility is affected by mild ischemia-induced intestinal injury in conscious rats through in vivo monitoring of colonic contractions, specifically with regard to the interstitial cells of Cajal (ICC) and the effect of nitric oxide (NO).Using miniature strain-gauge transducers, colonic motility with or without ischemia was recorded in conscious rats on the 4th, 7th, and 14th days after surgery. Histological examination for c-kit-positive cells was performed.In control nonischemic rats, the number and duration of contractions (NC and DC, respectively) decreased gradually, but the mean amplitude of contractions (MC) and motility index (MI) did not change. On the 7th day, the NC in the ischemic group increased significantly when compared with that in the control group (P = 0.037). The DC in the ischemic group was lower than that in the control group; the difference was significant on the 4th day (P = 0.008). The MIs in the ischemic group were lower than those in the control group. In both groups, administration of NGnitro-L: -arginine methyl ester on the 7th day increased only the resting cecal motility. Pathological examinations revealed c-kit-positive cells in both groups.Changes such as increased NC with shortened DC accompanied with decreased MI must have occurred at the ischemic site and might have been induced by an ischemic event. However, there exists a possibility that ICC and NO do not play a role in mild ischemia-induced dysmotility.

In Japan, the prevalence of food allergy has been increasing and a variety of problems have emerged regarding what should be considered a food allergy. A treatment regimen consists of avoiding the offending food (elimination diet therapy) and receiving nourishment from alternative foods (substitutional diet therapy). There is a growing concern that confusion has resulted from the lack of a consensus on the procedures for diagnosing and treating food allergies. The Food Allergy Committee of the Japanese Society of Pediatric Allergy and Clinical Immunology established the “Guidelines for Diagnosis and Management of Pediatric Food Allergy.” Definition, classification, pathophysiology, clinical disorders and management of food allergy are discussed and determined.

Aim: Asthmatic children are more likely to outgrow their symptoms than adult patients. Thus, we wanted to know whether there were any age-related differences in the time course of the allergic airway inflammation. Methods: BALB/C mice at different ages (young: 3 days after birth, and mature: 8 weeks of age) were sensitized with ovalbumin (OVA). Subsequently, animals were challenged with aerosolized OVA during 1, 2, 4 or 8 consecutive weeks. Bronchial hyperresponsiveness (BHR), serum IgE levels, the degrees of inflammatory cell infiltration (ICI) and goblet cell metaplasia (GCM) in the airways, and the number of eosinophils and cytokine levels in bronchoalveolar lavage fluid (BALF) were examined. Results: At 1 week, airway inflammation and BHR occurred similarly between young and mature mice. However, BHR disappeared at 4 weeks in young, whereas it persisted even at 8 weeks in mature mice. GCM, ICI and eosinophilia in BALF attenuated with time, with more remarkable reduction in young mice. The BALF IL-4 level was high during the first 2 weeks in both groups, while the IL-2 level was significantly increased at 2 weeks solely in young mice. Conclusion: Different time courses in airway inflammation and in BHR may relate to the different prognoses between childhood and adult asthma. The understanding of the mechanisms underlying this age-related differences may be helpful to induce remission in asthmatic patients.

We identified and analyzed novel thyrotropin (TSH) receptor mutations in three Japanese families with resistance to TSH.The TSH receptor gene was sequenced and the mutations were determined. The mutant TSH receptors were transfected into COS-7 cells, and their functions were analyzed.The patients were compound-heterozygotes for the R450H mutation and novel mutations in the TSH receptor gene. The first patient was a compound-heterozygote for R450H and V473I. The second sibling possessed R450H and R519C. The third sibling had R450H and R519G.The R450H mutant exhibited moderately impaired receptor functions and a moderately decreased cell surface expression in agreement with previous results. The V473I mutant exhibited an almost normal TSH binding, a slightly decreased cyclic adenosine monophosphate (cAMP) response, a moderately decreased inositolphosphate (IP) response, and an almost normal cell surface expression. TSH binding and TSH stimulation of cAMP and IPs were markedly decreased in the R519C and R519G mutants. Cell surface expression was decreased in the R519C mutant and negligible in the R519G mutant. All of these mutants showed normal intracellular synthesis of TSH receptors.These novel inactivating mutations contribute to understanding of the structure-function relationship of the TSH receptor. To date, all of the patients with TSH resistance resulting from TSH receptor mutations identified in Japan possessed the R450H mutation at least in one allele. These observations suggest that the R450H mutation is a commonly observed TSH receptor mutation in patients with TSH resistance in Japan.

This Asthma InsightsReality in Japan survey was conducted to access the achievement of asthma control by patients and current status of asthma treatment in 2005.This was cross-sectional, randomized-digit-dial household telephone survey, conducted between September and December 2005.A total 400 adults (mean 52.2 years) and 400 parents of children (mean 8.2 years) with asthma were interviewed by telephone. More than 40% of these patients reported having daytime asthma symptoms and over 30% in nighttime symptoms with awakening. Asthma episodes, i.e. hospitalization, emergency room visit, unscheduled visit, work/school absence were observed in 7, 4, 33, 29% of total patients during the past year, respectively. Eighteen percent of adult and 8% of children used inhaled corticosteroid. Awareness of peak-flow meter and experience of lung function test were low.The rate of patients who experienced asthma symptoms or episodes tended to decrease, and use of inhaled corticosteroids seemed to be increased since 2002 when a similar survey took place. However, asthma remains poorly controlled and the current level is still far from achieving the goal of asthma management objected by the Japanese guidelines. Further dissemination of standardize therapy in compliance with the guideline is required.

In contrast to an abundance of data concerning age-related changes of bronchial sensitivity, the relationship between age and rapidity of bronchoconstriction (bronchial reactivity) remains unclear. We studied age and bronchial reactivity in children with asthma. Enrolled in this study were 511 asthmatic subjects and 115 age-matched control subjects 1 to 16 years of age. Bronchial reactivity was represented by the slope of the methacholine transcutaneous oxygen pressure dose-response curve (SPO(2)) in younger children and the slope of the respiratory resistance dose-response curve (SRrs) in older children. Overall, SPO(2) and SRrs were higher in asthmatic than control subjects. SPO(2) increased significantly from 1 to 6 years in asthmatic subjects, reaching a plateau after age 7. This age-related change in SPO(2) also was seen in controls. SRrs in asthmatic subjects decreased after age 13, while SRrs in controls showed no significant change between age 7 and 16. Age-related change in bronchial reactivity occurs during childhood, possibly reflecting early changes in airway smooth muscle maturity and later changes in airway wall rigidity.

The success of allogeneic bone marrow transplantation (allo-BMT) in children with myelodysplastic syndrome (MDS) is mainly affected by relapse. The role of additional immunotherapy for pediatric patients with MDS relapsing after allo-BMT remains to be established. Because immunotherapy is generally more effective for patients who bear a low tumor burden, monitoring of minimal residual diseases (MRD) is essential for early diagnosis at molecular relapse. We report here that a patient with relapsed MDS after allo-BMT was successfully treated by the rapid discontinuation of immunosuppressive therapy at molecular relapse while monitoring Wilms tumor (WT1) gene expression levels. Quantitative analysis of WT1 gene expression appeared to be a useful tool to monitor MRD in the present case.

Background: It has been reported that histamine stimulates ion channels on airway epithelial cells and induces changes in osmolarity and the ion composition of the periciliary field of airway epithelia. Objective:To investigate the effect of inhaled histamine on epithelial cell swelling, we studied the role of airway epithelial cells under histamine inhalation challenge in an animal model of airway inflammation using ozone exposure. Method: After exposure to 3.0 ppm ozone for 2 h, guinea pigs were anesthetized and tracheostomized, and then, lung resistance (RL) was measured. Histamine inhalation challenge and histological examination were performed. Result:The values of RL before histamine inhalation in the control group and the ozone-exposed group were 0.26 ± 0.11 and 0.45 ± 0.34 cm H2O/s, respectively. RL increased significantly after histamine inhalation both in the control and the ozone-exposed groups. The threshold of histamine (PC200) in the ozone-exposed group was significantly lower than that in the control group. A significant swelling of the epithelial cells after histamine inhalation was observed both in the control and the ozone-exposed groups, with a greater increase in the ozone-exposed group compared with the control group. However, no change in wall thickness was observed in the histamine/antihistamine or the ozone/histamine/antihistamine group. Conclusion: Our results suggest the possibility that the airway epithelial cell swelling plays a role in the increase in RL after histamine inhalation, especially in the presence of airway inflammation.

The treatment of massive osteolysis with lymphangioma and/or hemangioma (Gorham-Stout syndrome) has been controversial. The authors report on a patient with multiple massive osteolyses and extensive lymph-hemangiomatosis whose lesions were reduced by interferon alfa therapy. A 2-year-old girl had complained of left chylothorax. Thoracoscopy showed an increase in small lymphatic vessels in the chest wall. The chylothorax was improved by coagulation of the lymphatic vessels. Later, multiple massive osteolyses appeared in the left 11th and 12th ribs, the TH10-L3 vertebrae, and the right femur. There were also hemangiomas in the liver and spleen, a tumor lesion in the left lower chest wall, and hemangiomatous change on the skin surface of the left back. The left lung had only a minimal air content. After OK-432 was injected into the femur and chest wall lesions, the femur lesion disappeared. Then, as right chylothorax appeared, OK-432 was injected into the right pulmonary cavity. The chylothorax disappeared, but pericardial effusion appeared. After steroid pulse therapy, pericardial effusion disappeared. During these treatments, the 7th to 10th ribs disappeared from the x-ray and scoliosis developed. One month later, a cloudy fluid collection in the right lung was found on computed tomography. Interferon alfa and steroid pulse therapy were started. Interferon alfa (1,500,000 units) was subcutaneously administered daily for 2 months and was gradually reduced and maintained at 1,500,000 unit/wk. Steroids were also reduced and maintained at 5 mg/d of predonine. Later, the progress of osteolysis and the extension of lymph-hemangiomatosis stopped. Ten months later, hemangioma in the back disappeared, and the 7th to 10th ribs, which had disappeared, reappeared. The interferon alfa therapy was stopped 14 months after it was administered. The patient's condition has been stable for 10 months since then. At this time, computed tomography shows regression of the hemangiomatous lesion in the back. The authors clinically diagnosed the patient as having Gorham-Stout syndrome with extension of lymph-hemangiomatosis. Interferon alfa with or without steroid therapy should be a choice for patients with extension lesions.

Continuous-wave 456-nm blue light generation is reported from a periodically polled MgO:LiNbO3 crystal by single-pass frequency doubling of a 912-nm Nd:GdVO4 laser. A maximum blue power of 167 mW with 8.3% infrared-to-blue conversion efficiency and 4.2%/W normalized conversion efficiency resulted from a 10-mm-long nonlinear device with 4.19-μm domain period and that operated at 44.5°C temperature.

Positron emission tomography (PET) with [18F]fluoro-D-deoxyglucose (FDG) has been used to detect seizure foci and evaluate surgical resection with localization related epilepsies. However, few investigations have focused on generalized epilepsy in children. To reveal the pathophysiology of generalized epilepsy, we studied 11 patients with generalized epilepsy except West syndrome, and 11 patients with localization related epilepsy without organic disease. The FDG PET was performed by simultaneous emission and transmission scanning. We placed regions of interest (ROI) on bilateral frontal lobe, parietal lobe, occipital lobe, temporal lobe, basal ganglia, thalamus and cerebellum. Standardized uptake value (SUV) was measured and normalized to SUV of ipsilateral cerebellum. Then, we compared the data of generalized epilepsy to those of localization related epilepsy. FDG PET revealed significant interictal glucose hypometabolism in bilateral basal ganglia in generalized epilepsy compared to that in localization related epilepsy (right side: p = 0.0095, left side: p = 0.0256, Mann-Whitney test). No other region showed any significant difference (p > 0.05) between the two groups. These findings indicate that the basal ganglia is involved in the outbreak of generalized seizures or is affected secondarily by the epileptogenicity itself.

Adrenomedullin is a vasodilator peptide that displays a variety of effects, such as hypotension and vasodilatation. The aim of this study was to test the effect of intravenous adrenomedullin on the motility pattern of the small intestine, and the functional involvement of adrenomedullin in endotoxin-induced small-intestinal motility disturbance.Jejunal motility was recorded in fasted conscious rats, using miniature strain-gauge force transducers sutured to the serosal surface of the small-intestinal wall.Intravenous administration of adrenomedullin at doses of 3, 6, and 10 microg/kg per min over 30 min disrupted phase 3 of the migrating motor complex, with the disruption lasting for 61.9 +/- 5.1, 52.2 +/- 10.6, and 74.1 +/- 25.2 min, respectively. The interval from drug administration to the onset of disruption decreased as the dose of adrenomedullin increased to 41.5 +/- 11.0, 11.6 +/- 3.4, and 0 min, respectively (P0.05). An increase in the motility index was also dose-dependent (P0.05) at these doses of adrenomedullin. Lipopolysaccharide (50 microg/kg) induced disruption of phase 3, which lasted for 138.7 +/- 5.4 min. Previous administration of the putative adrenomedullin-receptor antagonist, AM (22-52), at a 50 microg/kg dose, attenuated the disruption induced by lipopolysaccharide to 74.4 +/- 3.5 min (P0.01).Our findings (1) suggest that intravenous adrenomedullin causes small-intestinal motility disturbances, and (2) support the hypothesis that adrenomedullin overproduction plays an important role in lipopolysaccharide-induced disruption of the motility pattern in the small intestine in rats.

Intracellular oxidation and reduction (redox state) correspond closely to the surrounding environment. Most environmental factors affecting this balances such as oxidants, ultraviolet light, radioactive emissions, infections, and allergic reactions represent oxidative stress upon cells. We examined intracellular redox state after oxidative stress upon cultured human airway epithelial cells (Calu-3) by measuring reduced (GSH) or oxidized (GSSG) glutathione. We studied cytokine production, which is related to glutathione redox regulation, in response to ozone and also evaluated the effect of pretreatment with an ethyl ester of reduced glutathione (GSH-OEt) on cytokines. Ozone exposure (3.0 ppm, 3 min) time-dependently changed the redox state, while increasing production of interleukin(IL)-8 and IL-6, mRNA and protein. Treatment with GSH-OEt before ozone suppressed IL-8, but stimulated IL-6 production. Thus, oxidative stress affects intracellular glutathione redox state, in airway epithelial cells, activating signals to increase production of cytokine, modulation that may exacerbate respiratory symptoms.

Background: The pathophysiology of infantile asthma may differ from that in older children or in adults, partly because of the different immune response depending upon maturation. In adult mice, the sensitizing dose of antigen is known to be critical to the polarized development of helper T cell subsets and allergic airway inflammation. We wanted to know the characteristics of allergic airway inflammation of infantile asthma by developing a murine model. Methods: BALB/C mice at different stages of maturation (juvenile: 3 days after birth; adult: 8 weeks of age) were sensitized with 10 or 1,000 µg ovalbumin (OVA) or vehicle. The animals were then challenged with aerosolized OVA or saline once a day during 6 consecutive days. After the final challenge, bronchial hyperresponsiveness (BHR), bronchoalveolar lavage fluid (BALF), histological changes in the airways and immunological status were examined. Results: In both juvenile and adult animals, sensitization with 10 µg OVA induced the T helper 2 response (elevated IL-4 and decreased IFN-γ levels). BHR, airway eosinophilia, the inflammatory cell infiltration, goblet cell metaplasia (GCM), and IgE antibody production were more prominent in animals given this dose than 1,000 µg OVA. Among these responses, GCM as well as BALF IL-4, and BHR were comparable between juvenile and adult animals, whereas other parameters were lower in juvenile animals, especially in those given 1,000 µg OVA. Conclusion: GCM and, consequently, airway mucus hypersecretion may be an important component of allergic airway inflammation in infantile asthma.

Background/Purpose The authors studied gastrointestinal motility in a patient with total intestinal aganglionosis (TIA) and the effect of octreotide (OCT) on ileal motility in this patient. Methods The 3200-g girl received ileostomy at 50 cm proximal to the ileocecal site and jejunostomy at 15 cm distal to the ligament of Treitz because of severe ileus owing to TIA. Histology of the intestines, including jejunostomy, showed no ganglion cells. Gastro-duodeno-jejunal and distal ileal manometries were done 8 months after birth. Results In upper gastrointestinal manometry, sporadic contractions and clusters consisting of 3 to 5 contractions were observed in the duodenum and jejunum, but no typical phase 3 was observed during the 3-hour recording period. In ileal manometry, long-lasting repetitive contractions were recorded at 2 distal sites. In the most proximal ileum, the frequency of contractions was less than in the 2 distal sites. OCT administration induced a decrease in the amplitude of contractions during the first 20 minutes. The amplitude increased thereafter and reached a level higher than that before OCT administration. Conclusions In this patient, the predominant manometry finding was the remarkable hypermotility of the ileum. OCT induced a transient decrease in ileal motility and an increase in motility thereafter.

Hepatitis C virus (HCV) causes chronic hepatitis, liver cirrhosis and hepatocellular carcinoma in addition to acute hepatitis. The HCV genome encodes two envelope glycoproteins, E1 and E2. To investigate the role of E1 and E2 in HCV infection, we used a recombinant vesicular stomatitis virus (VSV), VSVdeltaG*, harboring the green fluorescent protein gene instead of the VSV G envelope protein gene. It was complemented with the native form of E1 and E2, or E1 or E2 alone, to make HCV pseudotypes VSVdeltaG*(HCV), VSVdeltaG*(E1), and VSVdeltaG*(E2). Neither E1 nor E2 expression was detected on the cell surface, as reported. Unlike previous reports, infectious activities of VSVdeltaG*(HCV), VSVdeltaG*(E1) and VSVdeltaG*(E2) pseudotypes were detected under conditions where VSV was completely neutralized by anti-VSV. We could enhance the infectious titers 100-fold by sonication upon virus harvest. Bovine lactoferrin efficiently inhibited infection by VSVdeltaG*(HCV) as well as VSVdeltaG*(E2), as the interaction between E2 and lactoferrin has been thought to contribute to the inhibition of HCV infectivity. VSVdeltaG*(HCV) infected many adherent cell lines, including hepatic cell lines, but not most hematopoietic cell lines. Treatment of cells with trypsin, tunicamycin, or sulfated polysaccharides before infection reduced the infectivity of VSVdeltaG*(HCV) by about 90%, suggesting that a cell surface protein(s) with sugar chains plays an important role in HCV infection. The VSV pseudotypes developed here would be useful for analyzing the early stages of HCV infection.

Objectives To evaluate the efficacy of anti-regurgitant milk (AR milk) with reduced concentration of locust bean gum (LBG) compared with the usual commercially available concentration of this thickener. Methods Thirty infants with daily regurgitation but no other medical problems were randomly assigned to one of two groups. Infants in group A (n = 16) were fed either HL-450, an AR milk thickened with a commonly used concentration of LBG (0.45 g/100 mL) or control milk (HL-00; no LBG) in a crossover manner for periods of 1 week. The order of milk was randomly chosen for each subject. Infants in group B (n = 14) were fed HL-350, an AR milk with a reduced LBG concentration (0.35 g/100 mL), or HL-00 in the same crossover fashion. The number of episodes of regurgitation, feeding time, and body weight gain were recorded. Three infants in group B did not complete the protocol and were excluded. Results Both AR formulas decreased the number of regurgitation episodes by approximately 50% compared with control. Five mothers who gave their infants HL-450 and no mothers who fed their children HL-350 reported that the infants had difficulty sucking the formula through the nipple. Thirteen (81.3%) mothers who used HL-450 and 9 (81.8%) mothers who used HL-350 preferred the AR milk to the control milk. Conclusions An AR milk with reduced LBG was as effective in reducing regurgitation as one with the usually available concentration of LBG.

Background . Procalcitonin (PCT) is a new parameter of inflammation, the clinical usefulness of which is currently being evaluated. Materials and methods . We determined simultaneously the serum concentrations of PCT and C-reactive protein (CRP) as well as the white blood cell (WBC) count in 25 patients with Kawasaki disease (KD), 17 with bacterial infections, 10 with systemic autoimmune diseases, 17 with viral infections and 18 healthy children. The optimal cut-off value of each parameter for predicting coronary aneurysms was determined using receiver operating characteristic curves. Results . Significantly higher serum concentrations of PCT were observed in patients with KD (2.3±3.0 ng/ml) and bacterial infections (2.2±2.9 ng/ml) than in patients with autoimmune diseases (0.4±0.4 ng/ml) or viral infections (0.4±0.3 ng/ml), or in healthy children (0.2±0.1 ng/ml). The serum PCT but not the WBC count or CRP, differentiated the KD patients from the patients with autoimmune diseases. The optimal cut-off value of 3.0 ng/ml of PCT increased the prediction rate of coronary aneurysms that subsequently occurred in 4 (16%) patients with KD. Conclusions . The serum PCT may be clinically useful for determining the severity of KD and for narrowing the differential diagnosis of patients with inflammatory diseases.

Ultraviolet radiation, the third-harmonic of a Nd:GdVO4 laser operating at 1.06-microns wavelength, is reported from a single-pass cascaded first-order periodically poled MgO:LiNbO3 crystals. Continuous-wave 47 mW at 354 nm with 3.8%/Wcm normalized conversion efficiency resulted.

Continuous-wave power of 1.18 W at 531 nm with 16.8% conversion efficiency is reported at room temperature from a 2-mm-thick, 25-mm long uncoated periodically polled MgO:LiNbO3 by single-pass frequency doubling of a Nd:GdVO4 laser.

Objective The aim of this multicenter prospective and randomized study was to determine the effect of adding corticosteroids to intravenous gamma globulin (IVGG) therapy on serum cytokine levels, as well as to see its effect on the clinical course in children in the acute phase of Kawasaki disease (KD). Study design Patients with KD (n = 32) were randomized to receive either IVGG alone (G group) or IVGG plus corticosteroids (G + S group). The clinical course and cytokine responses between groups were compared. Results The pretreatment serum levels of interleukin (IL)-2, IL-6, IL-8, and IL-10 were significantly higher in patients with KD than in healthy controls. Although IVGG alone failed to reduce cytokine concentrations within 24 hours of IVGG administration, corticosteroids plus IVGG reduced IL-2, IL-6, IL-8, and IL-10 levels. The levels of IL-2, IL-6, IL-8, and IL-10 within 24 hours after initiating IVGG therapy were significantly lower in the G + S group than in the G group. The duration of fever was shorter, and the C-reactive protein concentration decreased more quickly in the G + S group than in the G group. Conclusions These findings suggest that corticosteroids rapidly ameliorate symptoms by reducing cytokine levels in children with KD.

Eosinophils produce cysteinyl leukotrienes such as leukotriene C4 and D4 upon stimulation by platelet-activating factor or other mediators, and these cells themselves express cysteinyl leukotriene receptors. Pranlukast, a compound developed in Japan, antagonizes cysteinyl leukotriene receptors and inhibits contraction of airway smooth muscle, microvascular leakage into airways, and eosinophil infiltration. This agent can decrease symptoms of bronchial asthma, but its specific influences on effector functions of eosinophils important to the pathogenesis and exacerbation of asthma remain unknown. In the present study, we investigated the effect of pranlukast on human eosinophil functions. Eosinophils obtained from peripheral blood of normal volunteers were stimulated by platelet-activating factor, leukotriene D4, or phorbol ester. Superoxide anion generation was measured by reduction of cytochrome c. Expression of alphaMbeta2 was analyzed by flow cytometry. To evaluate eosinophil degranulation, eosinophil protein X, a toxic granule constituent, was measured by radioimmunoassay in sample supernatants. Pranlukast partially inhibited major eosinophil effector functions of superoxide anion generation and degranulation induced by platelet-activating factor, although at concentrations tested pranlukast failed to significantly reduce platelet-activating factor-induced alphaMbeta2 expression. Pranlukast completely inhibited leukotriene D4-induced superoxide generation and alphaMbeta2 expression. In contrast, pranlukast at 10(-6)M did not affect phorbol ester-induced superoxide generation at 120 minutes, degranulation, or alphaMbeta2 expression. The results suggested that inhibition by pranlukast of platelet-activating, factor-induced eosinophil effector functions such as superoxide generation and degranulation might result at least partly from antagonism of autocrine mechanisms involving cysteinyl leukotrienes produced in response to platelet-activating factor.

Background: Cellular adhesion is crucial for eosinophil effector functions. Objective: We sought to elucidate the role of the actin cytoskeleton in cellular adhesion and superoxide anion generation by human eosinophils. Methods: Eosinophils were stimulated with platelet-activating factor (PAF) or complement component 5a on human serum albumin-coated plates with or without an actin-polymerization inhibitor, cytochalasin B (CB), or cytochalasin D (CD). Superoxide anion generation was measured on the basis of reduction of absorbance associated with cytochrome c.2 Eosinophil adhesion was assessed on the basis of eosinophil protein X content in adherent cells. Transient stimulus-induced increase of intracellular calcium and translocation of protein kinase C (PKC) βII, PKC δ, PKC ζ, and p47 phagocyte oxidase (a component of nicotinamide adenine dinucleotide phosphate oxidase) were also investigated. Results: CB, CD, or antibodies against CD18 (the β2 chain of integrin, αMβ2) inhibited stimulus-induced eosinophil superoxide anion generation. Stimulus-induced eosinophil adhesion was unaltered by CB, whereas it was significantly suppressed by CD or anti-CD18 antibodies. Transient PAF-induced intracellular calcium increase was also unaffected by CB or CD, but stimulus-induced eosinophil shape changes and translocation of PKCs and p47 phagocyte oxidase to the cell membrane region were completely inhibited by CB. PAF-induced eosinophil degranulation was inhibited by CB, CD, or anti-CD18 antibodies, whereas complement component 5-induced degranulation was not suppressed by CB. Conclusion: By itself, β2 integrin-dependent cellular adhesion is not sufficient for promoting eosinophil effector function. Adequate actin assembly is required for eosinophil adhesion and also for full superoxide anion generation in eosinophils. (J Allergy Clin Immunol 2003;112:126-33.)

Ultraviolet second-harmonic generation in first-order periodically poled MgO:LiNbO3 is presented. Using a high-voltage multipulse application method, we fabricated a first-order nonlinear grating with a period of 1.8 microm and depth of approximately 150 microm over 10-mm interaction length in a 1-mm-thick MgO:LiNbO3 substrate. In a single-pass configuration, continuous-wave 30-mW UV light at 362.5 nm was generated for a fundamental power of 522 mW, corresponding to a normalized conversion efficiency of 11%/W.

An outbreak of methicillin-resistant Staphylococcus aureus (MRSA) colonization occurred from November 2001 in the neonatal intensive care unit (NICU) of our hospital. Since the establishment of our NICU in 1991, some MRSA has been detected in NICU patients. For MRSA infection preventive measures, utilization of the following items was implemented: mupirocin ointment, diluted povidone iodine, methylrosaniline chloride, and disposable rubber gloves. Patients in whom MRSA was detected received intranasal administration of the mupirocin ointment three times daily and were bathed in, or their entire body was wiped with diluted povidone iodine once daily for the first 3 days in each week. In addition, they received an intraoral application of methylrosaniline chloride daily. All therapy was done until MRSA strains were undetectable for 3 continuous weeks. Genotypes of 13 MRSA strains isolated from eight inpatients and one mother were analyzed by pulsed-field gel electrophoresis (PFGE). All PFGE patterns were identical, except for one, which had one distinct migrating fragment. These data suggested that this MRSA outbreak was caused by the same strain, which was derived from the mother of a low-birth-weight infant born on October 30, 2001. Gradually, the number of inpatients carrying MRSA decreased, until finally MRSA was no longer observed, in April 2002. Fortunately, we controlled the MRSA outbreak immediately, and none of the inpatients developed severe MRSA infection. We think that in our NICU, which is isolated from other hospital wards, it is important to prevent the entrance of MRSA-carrying mothers.

To determine whether the inhaled diuretic, amiloride, can modify bronchoconstriction induced by ultrasonically nebulized distilled water (UNDW) in asthmatic children, a double-blind, randomized, placebo-controlled study was done. The UNDW inhalation challenge was performed in 12 asthmatic children (nine boys and three girls; mean age +/- SEM, 11.2 +/- 0.75 yr), who had at least a 20% fall in FEV1 after UNDW inhalation. On separate days, these subjects underwent UNDW inhalation challenge after inhalation of amiloride (0.3 mg/m2 body surface area) or placebo (0.9% saline). Neither bronchodilation nor bronchoconstriction after amiloride inhalation was observed. The mean value and SEM of the UNDW PD20 after placebo inhalation was 4.55 +/- 0.80 ml. After inhalation of amiloride, PD20 increased to 7.93 +/- 0.75 ml (p < 0.01). We concluded that inhaled amiloride exerted a protective effect against UNDW-induced bronchoconstriction in asthmatic children.

SummaryAgonists at β2 adrenoceptors are used widely as bronchodilators in treating bronchial asthma. These agents also may have important anti-inflammatory effects on eosinophils in asthma. We examined whether widely prescribed β2-adrenoceptor agonists differ in ability to suppress stimulus-induced eosinophil effector functions such as superoxide anion (O2–) generation and degranulation. To examine involvement of cellular adhesion in such responses, we also investigated effects of β2 agonists on cellular adhesion and on CD11b expression by human eosinophils. O2– was measured using chemiluminescence. Eosinophil degranulation and adhesion were assessed by a radioimmunoassay for eosinophil protein X (EPX). CD11b expression was measured by flow cytometry. Fenoterol inhibited platelet-activating factor (PAF)-induced O2– generation by eosinophils significantly more than salbutamol or procaterol. Fenoterol partially inhibited PAF-induced degranulation by eosinophils similarly to salbutamol or procaterol. Fenoterol inhibited phorbol myristate acetate (PMA)-induced O2– generation and degranulation by eosinophils, while salbutamol or procaterol did not. Fenoterol inhibition of PMA-induced O2– generation was not reversed by ICI-118551, a selective β2-adrenoceptor antagonist. Fenoterol, but not salbutamol or procaterol, significantly inhibited PAF-induced eosinophil adhesion. Fenoterol inhibited O2– generation and degranulation more effectively than salbutamol or procaterol; these effects may include a component involving cellular adhesion. Inhibition also might include a component not mediated via β2 adrenoceptors.

Background: Prostaglandins, synthesized by cyclo-oxygenase (COX), regulate renal hemodynamics and also epithelial water and solute transport. Developmental changes occur in COX-2 mRNA expression and its response to lipopolysaccharide stimulation in rats. We examined age-related changes in COX mRNA expression induced by hypoxia in the renal cortex and medulla of developing rats. Methods: Total RNA was extracted from 1- and 4-week-old male Wistar rats exposed to one or 4 h of hypoxia (8% O2). Cyclo-oxygenase mRNA was quantitatively analyzed using a real-time polymerase chain reaction with dual-labeled fluorogenic probes. Results: Expression of COX-1 mRNA did not change in response to hypoxia in the cortex or medulla in either infantile or adult rats. In infantile rats, COX-2 mRNA expression was not induced by one or 4 h of hypoxia. In adults, 1- and 4-h exposures to hypoxia induced COX-2 mRNA in the renal cortex, and 1-h of exposure induced COX-2 mRNA in the medulla. Conclusions: Response of expression of COX-2 mRNA in rats exposed to acute hypoxia show age-related variability treated by acute hypoxia. Cyclo-oxygenase-2 might not play a major role in the hypoxic infantile rat kidney.

To study the effect of maturation on abilities of superoxide radicals (O-2) generation in the airways, we compared stimuli-induced O-2 generation by alveolar macrophages in immature (aged 10+/-2 days) and adult (aged 90+/-2 days) guinea-pigs. The production of O-2 was assayed by chemiluminescence method, using a Cypridina luciferin analog as a highly sensitive and specific probe for O-2. Whereas no significant difference in cell components of bronchoalveolar lavage fluid was observed between immature and adult animals, O-2 generation induced by stimulation of alveolar macrophages was greater in immature than in adult animals, with significant differences observed after platelet-activating factor (100 nM) or phorbol myristate acetate (0.5 micro g/ml). The results suggest that alveolar macrophages from immature animals are far more potent O-2 generators than the same cells of adult animals.

Insulin-like growth factor-I activates a calcium-permeable cation channel GRC (growth factor-regulated channel). In the present study, we investigated the immunohistochemical localization of GRC in human tissues using a polyclonal anti-GRC antibody. Immunoreactive GRC was detected in the stomach, duodenum, large intestine and prostate. In these tissues, GRC-expressing cells were distributed solitarily in the epithelium and coexpressed chromogranin-A, a marker of neuroendocrine cells. GRC was also expressed in the epithelium of the pancreatic duct, mammary gland, parotid gland, and submandibular gland. Epithelial cells of the renal tubule and the tracheal gland were also stained with anti-GRC antibody. In the lung, alveolar macrophages expressed GRC. In the brain, Purkinje cells of the cerebellum and arachnoid cells of the meningitis expressed GRC. These results indicate that GRC is expressed in restricted types of cells in particular tissues, and that GRC may modulate calcium entry in these cells.

Background: Platelet-activating factor (PAF) is a potent stimulator of human eosinophils involved in the pathogenesis of allergic diseases. However, intracellular signaling mechanisms in eosinophils involving the PAF receptor are incompletely understood. Objective: We sought to determine the roles of protein kinase C (PKC) and cyclic AMP-dependent protein kinase (protein kinase A [PKA]) in signaling pathways of human eosinophils stimulated with PAF. Methods: After pretreatment with a PKC inhibitor, bisindolylmaleimide I, or a PKA inhibitor, H89, we investigated PAF-evoked functions, such as CD11b expression, cellular adhesion, superoxide anion generation, and degranulation in human eosinophils. Results: Preincubation of eosinophils with bisindolylmaleimide I resulted in enhancement of upregulated CD11b expression and adhesion induced by PAF. H89 pretreatment also enhanced PAF-induced cellular adhesion. Superoxide anion generation and degranulation were suppressed by means of inhibition of either PKC or PKA. Conclusion: PKC and PKA negatively regulate PAF-induced CD11b upregulation and cellular adhesion but promote eosinophil effector functions, such as superoxide anion generation and degranulation. PKC and PKA modulate PAF-evoked intracellular signaling of the eosinophil function in distinct ways. (J Allergy Clin Immunol 2002;110:241-8.)