Search

A 77-year-old man was referred to our hospital for abnormal thoracic radiographs. Computed tomography (CT) revealed a 20-mm subpleural ground-glass opacity in the right S

High-throughput, high-accuracy detection of emerging viruses allows for pandemic prevention and control. Currently, reverse transcription-polymerase chain reaction (RT-PCR) is used to diagnose the presence of SARS-CoV-2. The principle of the test is to detect RNA in the virus using a pair of primers that specifically binds to the base sequence of the viral RNA. However, RT-PCR is a sophisticated technique requiring a time-consuming pretreatment procedure for extracting viral RNA from clinical specimens and to obtain high sensitivity. Here, we report a method for detecting novel coronaviruses with high sensitivity using artificial intelligent nanopores utilizing a simple procedure that does not require RNA extraction. Artificial intelligent nanopore platform consists of machine learning software on the servers, portable high-speed and high-precision current measuring instrument, and scalable, cost-effective semiconducting nanopore modules. Here we show that the artificial intelligent nanopores are successful in accurate identification of four types of coronaviruses, HCoV-229E, SARS-CoV, MERS-CoV, and SARS-CoV-2, which are usually extremely difficult to detect. The positive/negative diagnostics of the new coronavirus is achieved with a sensitivity of 95 % and specificity of 92 % with a 5-minute diagnosis. The platform enables high throughput diagnostics with low false negatives for the novel coronavirus.

BACKGROUND: The efficacy and safety of benralizumab, an interleukin-5 receptor alpha-directed cytolytic monoclonal antibody, for the prevention of exacerbations in patients with moderate to very severe chronic obstructive pulmonary disease (COPD) are not known.METHODS: In the GALATHEA and TERRANOVA trials, we enrolled patients with COPD (at a ratio of approximately 2:1 on the basis of eosinophil count [≥220 per cubic millimeter vs. RESULTS: In GALATHEA, the estimates of the annualized exacerbation rate were 1.19 per year (95% confidence interval [CI], 1.04 to 1.36) in the 30-mg benralizumab group, 1.03 per year (95% CI, 0.90 to 1.19) in the 100-mg benralizumab group, and 1.24 per year (95% CI, 1.08 to 1.42) in the placebo group; the rate ratio as compared with placebo was 0.96 for 30 mg of benralizumab (P = 0.65) and 0.83 for 100 mg of benralizumab (P = 0.05). In TERRANOVA, the estimates of the annualized exacerbation rate for 10 mg, 30 mg, and 100 mg of benralizumab and for placebo were 0.99 per year (95% CI, 0.87 to 1.13), 1.21 per year (95% CI, 1.08 to 1.37), 1.09 per year (95% CI, 0.96 to 1.23), and 1.17 per year (95% CI, 1.04 to 1.32), respectively; the corresponding rate ratios were 0.85 (P = 0.06), 1.04 (P = 0.66), and 0.93 (P = 0.40). At 56 weeks, none of the annualized COPD exacerbation rate ratios for any dose of benralizumab as compared with placebo reached significance in either trial. Types and frequencies of adverse events were similar with benralizumab and placebo.CONCLUSIONS: Add-on benralizumab was not associated with a lower annualized rate of COPD exacerbations than placebo among patients with moderate to very severe COPD, a history of frequent moderate or severe exacerbations, and blood eosinophil counts of 220 per cubic millimeter or greater (Funded by AstraZeneca [GALATHEA and TERRANOVA] and Kyowa Hakko Kirin [GALATHEA]; GALATHEA and TERRANOVA ClinicalTrials.gov numbers, NCT02138916 and NCT02155660.).

Isovaleric acidemia (IVA) is an organic acid disease caused by a deficiency of isovaleryl-CoA dehydrogenase. Deficiency of this enzyme leads to accumulation of organic acids, such as isovalerylcarnitine and isovalerylglycine. The proposed IVA treatments include leucine restriction and l-carnitine and/or glycine supplementation, which convert isovaleric acid into non-toxic isovalerylcarnitine and isovalerylglycine, respectively. We examined the therapeutic response using the leucine load test and performed a 10-year follow-up in the patient. Methods:We evaluated the patient with IVA beginning at 5 years of age, when he presented with a mild to intermediate metabolic phenotype. Ammonia, free carnitine, isovalerylcarnitine, and isovalerylglycine were analyzed in the urine and blood after a meal consisting of 1600 mg leucine with glycine alone (250 mg/kg/day), l-carnitine alone (100 mg/kg/day), or both glycine and l-carnitine for four days each. Results:(Leucine load test) Three hours after the meal, serum ammonia levels increased most dramatically with glycine treatment alone, then with both in combination, and least with l-carnitine alone. Urinary isovalerylglycine levels increased 2-fold more with glycine supplementation than those following supplementation with both agents or with l-carnitine alone. Treatment with both agents resulted in a gradual increase in urinary acylcarnitine levels during the 6-h period following the leucine load, reaching concentrations comparable to those observed with l-carnitine alone. (Clinical course) After initiation of both glycine (200 mg/kg/day) and l-carnitine (100 mg/kg/day) supplementation at 5 years of age, doses were gradually reduced to 111.7 mg/kg/day and 55.8 mg/kg/day, respectively, at 15 years of age. His mind and body had developed without any sequelae. Discussion:We concluded that l-carnitine conjugated isovaleric acid earlier than glycine. Additionally, during the 10-year follow-up period, the patient displayed no clinical deterioration., 論文

Identification of the genes responsible for adult-onset type II citrullinemia (CTLN2) and citrin protein function have enhanced our understanding of citrin deficiency. Citrin deficiency is characterized by 1) neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD); 2) adaptation/compensation stage with unique food preference from childhood to adulthood; and 3) CTLN2. The treatment of NICCD aims to prevent the progression of cholestasis, and it includes medium chain triglycerides (MCT) milk and lactose-free milk, in addition to medications (e.g., vitamin K2, lipid-soluble vitamins and ursodeoxycholic acid). Spontaneous remission around the age of one is common in NICCD, though prolonged cholestasis can lead to irreversible liver failure and may require liver transplantation. The adaptation/compensation stage (after one year of age) is characterized by the various signs and symptoms such as hypoglycemia, fatty liver, easy fatigability, weight loss, and neuropsychiatric symptoms. Some poorly-controlled patients show failure to thrive and dyslipidemia caused by citrin deficiency (FTTDCD). Diet therapy is the key in the adaptation/compensation stage. Protein- and fat-rich diet with a protein: fat: carbohydrate ratio being 15-25%: 40-50%: 30-40% along with the appropriate energy intake is recommended. The use of MCT oil and sodium pyruvate is also effective. The toxicity of carbohydrate is well known in the progression to CTLN2 if the consumption is over a long term or intense. Alcohol can also trigger CTLN2. Continuous intravenous hyperalimentation with high glucose concentration needs to be avoided. Administration of Glyceol® (an osmotic agent containing glycerol and fructose) is contraindicated. Because the intense treatment such as liver transplantation may become necessary to cure CTLN2, the effective preventative treatment during the adaptation/compensation stage is very important. At present, there is no report of a case with patients reported having the onset of CTLN2 who are on the diet therapy and under the appropriate medical support during the adaptation/compensation stage.

Galactosemia is caused by metabolic disturbances at various stages of galactose metabolism, including deficiencies in enzymes involved in the Leloir pathway (GALT, GALK1, and GALE). Nevertheless, the etiology of galactosemia has not been identified in a subset of patients. This study aimed to explore the causes of unexplained galactosemia. Trio-based exome sequencing and/or Sanger sequencing was performed in eight patients with unexplained congenital galactosemia. In vitro enzymatic assays and immunoblot assays were performed to confirm the pathogenicity of the variants. The highest blood galactose levels observed in each patient were 17.3–41.9 mg/dl. Bilateral cataracts were observed in two patients. In all eight patients, we identified biallelic variants (p.Arg82*, p.Ile99Leufs*46, p.Gly142Arg, p.Arg267Gly, and p.Trp311*) in the GALM encoding galactose mutarotase, which catalyzes epimerization between β- and α-D-galactose in the first step of the Leloir pathway. GALM enzyme activities were undetectable in lymphoblastoid cell lines established from two patients. Immunoblot analysis showed the absence of the GALM protein in the patients’ peripheral blood mononuclear cells. In vitro GALM expression and protein stability assays revealed altered stabilities of the variant GALM proteins. Biallelic GALM pathogenic variants cause galactosemia, suggesting the existence of type IV galactosemia.

Primary lung tumor tissue exhibiting numerous multinucleated osteoclast-like giant cells (OGCs) is an extremely rare histological type. In this study, we describe the case of an 82-year-old woman. Chest CT demonstrated an extensive crazy-paving appearance in both of the lung fields and a solid round mass in the right lower lobe. A pathological examination of the tumor revealed sarcomatoid carcinoma containing OGCs combined with diffused, invasive lepidic predominant adenocarcinoma. We herein present an extremely rare lung cancer, invasive lepidic predominant adenocarcinoma extensively involving the lung with areas of sarcomatoid carcinoma containing OGCs.

The mitochondrial aspartate-glutamate carrier isoform 2 (citrin) and mitochondrial glycerol-3-phosphate dehydrogenase (mGPD) double-knockout mouse has been a useful model of human citrin deficiency. One of the most prominent findings has been markedly increased hepatic glycerol 3-phosphate (G3P) following oral administration of a sucrose solution. We aimed to investigate whether this change is detectable outside of the liver, and to explore the mechanism underlying the increased hepatic G3P in these mice. We measured G3P and its metabolite glycerol in plasma and urine of the mice under various conditions. Glycerol synthesis from fructose was also studied using the liver perfusion system. The citrin/mGPD double-knockout mice showed increased urine G3P and glycerol under normal, fed conditions. We also found increased plasma glycerol under fasted conditions, while oral administration of different carbohydrates or ethanol led to substantially increased plasma glycerol. Fructose infusion to the perfused liver of the double-knockout mice augmented hepatic glycerol synthesis, and was accompanied by a concomitant increase in the lactate/pyruvate (L/P) ratio. Co-infusion of either pyruvate or phenazine methosulfate, a cytosolic oxidant, with fructose corrected the high L/P ratio, leading to reduced glycerol synthesis. Overall, these findings suggest that hepatic glycerol synthesis is cytosolic NADH/NAD+ ratio-dependent and reveal a likely regulatory mechanism for hepatic glycerol synthesis following a high carbohydrate load in citrin-deficient patients. Therefore, urine G3P and glycerol may represent potential diagnostic markers for human citrin deficiency.

Isovaleric acidemia (IVA) is an autosomal recessive inborn error affecting leucine metabolism. It is caused by a deficiency in isovaleryl-CoA dehydrogenase (IVD), a mitochondrial matrix enzyme that catalyzes the oxidation of isovaleryl-CoA to 3-methylcrotonyl-CoA. IVD is a FAD-containing enzyme, consisting of four identical subunits. Clinical features of IVA include poor feeding, vomiting, lethargy, developmental delay, metabolic acidosis, and a characteristic "sweaty foot" odor. IVA is one of the target disorders for newborn screening by tandem mass spectrometry (MS/MS). The human IVD gene is located on chromosome 15q. To date, over 50 disease-causing mutations have been reported worldwide. In this study, we searched for IVD mutations in five Japanese patients with IVA (neonatal type, two patients; chronic intermittent type, two patients; and mild biochemical type, one patient). The diagnosis of IVA was confirmed by urinary organic acid analysis using gas chromatography and mass spectrometry. All coding exons and the flanking introns in the IVD gene were amplified by PCR and were directly sequenced. We thus identified six hitherto unknown mutations (p.G94D, p.E116K, p.M167T, p.L243P, p.L246P, and c.696+1G>T) and four previously reported (p.R53P, p.R395C, p.Y403C, and p.E411K) pathogenic mutations. All patients were compound heterozygotes, and each mutation was identified in a single patient. Pathogenicity of newly identified mutations was validated using computational programs. Among them, the p.M167T is believed to influence FAD binding, as the position 167 is present in one of the FAD-binding sites. Our results have illustrated the heterogeneous mutation spectrum and clinical presentation of IVA in the Japanese patients.

Mutations in the X-linked gene CDKL5 cause early-onset epileptic encephalopathy and severe developmental delay. Because this disorder predominantly affects females, the full clinical spectrum of male patients remains elusive. We herein report a 16-year-old boy, who suffered from intractable seizures 20 days after birth. Serial electroencephalograms detected recurrent focal epileptiform discharges from age 4 months, which evolved to hypsarrhythmia later in infancy. Mass-spectrometric analyses revealed increase in urinary excretion of methylmalonic acid without perturbed concentrations of propionic acid, homocystein and methionine. Whole-exome sequencing identified a de novo, truncating mutation in CDKL5 (NM_003159.2:c.419dupA, p.Asn140Lysfs*8). Targeted sequencing excluded concomitant mutations in methylmalonic academia-associated genes. No methylmalonic acidemia has been reported in children with CDKL5 disorder. Extensive analyses on organic acid metabolism for males with CDKL5 mutations will gain more insight into their biochemical profiles in infancy.

Propionic acidemia is an autosomal recessive disorder that is due to deficiency in the enzyme propionyl-CoA carboxylase. Cardiomyopathy is a well-known phenomenon in propionic acidemia that it may rapidly progress to death. Here we describe a case of propionic acidemia in a 27-year-old man who developed adult-onset secondary dilated cardiomyopathy. In early infancy he was diagnosed with propionic acidemia and was later noted to have mild mental retardation, mild renal failure, and optic nerve atrophy. Although he was in good energy status with a low-protein diet and carnitine supplementation, he was admitted to our university hospital with decompensate heart failure, which resulted in low-output cardiac syndrome with massive mitral regurgitation and left ventricular dyssynchrony. Cardiac resynchronization therapy (CRT) and continuous hemodiafiltration followed by hemodialysis (HD) dramatically improved his clinical status.

Kocuria kristinae is a catalase-positive, coagulase-negative, Gram-positive coccus found in the environment and in normal skin and mucosa in humans; however, it is rarely isolated from clinical specimens and is considered a nonpathogenic bacterium. We describe a case of catheter-related bacteremia due to K. kristinae in a young adult with propionic acidemia undergoing periodic hemodialysis. The patient had a central venous catheter implanted for total parenteral nutrition approximately 6 months prior to the onset of symptoms because of repeated acute pancreatitis. K. kristinae was isolated from two sets of blood cultures collected from the catheter. Vancomycin followed by cefazolin for 16 days and 5-day ethanol lock therapy successfully eradicated the K. kristinae bacteremia. Although human infections with this organism appear to be rare and are sometimes considered to result from contamination, physicians should not underestimate its significance when it is isolated in clinical specimens.

The male patient is the third child of unrelated Japanese parents. His older sister had tachypnea and feeding difficulties, and died at 5 days of age. The patient was delivered at term (birthweight, 3.8 kg), following an unremarkable pregnancy. He presented with tachypnea, metabolic acidosis, and hyperammonemia (944 μmol · L−1) at 6 days. ### Questions for consideration: 1. What is the differential for infantile presentation of hyperammonemia in the neonatal period? 2. What laboratory tests would you pursue? GO TO SECTION 2 Hyperammonemia occurs in urea cycle disorders (e.g., ornithine transcarbamylase deficiency) and organic acidurias (e.g., methylmalonic aciduria, propionic aciduria, and isovaleric aciduria) and fatty acid oxidation defects (e.g., multiple acyl-CoA dehydrogenase deficiency). The existence of acidosis with ketosis indicates organic aciduria, whereas respiratory alkalosis is observed in urea cycle defects. Diagnosis is based on quantitative assay of amino acids and acylcarnitines from dried blood and organic acids in urine samples. ### Case: part 2. Elevated levels of 2-methylcitric acid and 3-hydroxypropionic acid were found in the urine. The plasma propionic acid concentration was increased (4.5 mg · dL−1), and propionyl-CoA …

Background: Serum levels of pneumocyte biomarkers KL-6 and surfactant protein D (SP-D) are useful diagnostic markers for interstitial lung diseases. However, associations of serum KL-6 and SP-D with radiologic findings in nonspecific interstitial pneumonia (NSIP) remain unclear. Objectives: To determine whether serum levels of KL-6 and SP-D reflect fibrotic and/or inflammatory processes in NSIP, we investigated the correlation between high-resolution computed tomography (HRCT) findings and serum KL-6 and SP-D levels. Methods: Serum levels of KL-6 and SP-D were measured in 21 patients with biopsy-confirmed NSIP. The radiographic extent of 6 HRCT patterns and total HRCT score, defined as the scored fibrotic index, were assessed. Changes in the levels of serum markers and CT findings during follow-up were also monitored. Results: Serum levels of KL-6 in NSIP positively correlated with the total HRCT score and overall extent of interstitial disease. Serum levels of SP-D in NSIP showed a positive correlation with the area of ground-glass attenuation without traction bronchiectasis and the inflammatory CT pattern, but the levels were inversely correlated with the area of ground-glass attenuation with traction bronchiectasis and the fibrotic CT pattern. The follow-up CT and serum marker changes after treatment showed that percent change of disease extent was reflected in both markers, especially KL-6. Further, the decreased fibrotic pattern correlated with both biomarkers. Conclusions: The results indicate that serum levels of KL-6 in NSIP reflect the overall extent of interstitial lesions, which include both inflammatory and fibrotic lesions, while the levels of SP-D mainly reflect the extent of inflammatory lesions.

Isovaleric acidemia (IVA) is an autosomal recessive inborn error affecting leucine metabolism. It is caused by a deficiency in isovaleryl-CoA dehydrogenase (IVD), a mitochondrial matrix enzyme that catalyzes the oxidation of isovaleryl-CoA to 3-methylcrotonyl-CoA. IVD is a FAD-containing enzyme, consisting of four identical subunits. Clinical features of IVA include poor feeding, vomiting, lethargy, developmental delay, metabolic acidosis, and a characteristic "sweaty foot" odor. IVA is one of the target disorders for newborn screening by tandem mass spectrometry (MS/MS). The human IVD gene is located on chromosome 15q. To date, over 50 disease-causing mutations have been reported worldwide. In this study, we searched for IVD mutations in five Japanese patients with IVA (neonatal type, two patients; chronic intermittent type, two patients; and mild biochemical type, one patient). The diagnosis of IVA was confirmed by urinary organic acid analysis using gas chromatography and mass spectrometry. All coding exons and the flanking introns in the IVD gene were amplified by PCR and were directly sequenced. We thus identified six hitherto unknown mutations (p.G94D, p.E116K, p.M167T, p.L243P, p.L246P, and c.696+1GT) and four previously reported (p.R53P, p.R395C, p.Y403C, and p.E411K) pathogenic mutations. All patients were compound heterozygotes, and each mutation was identified in a single patient. Pathogenicity of newly identified mutations was validated using computational programs. Among them, the p.M167T is believed to influence FAD binding, as the position 167 is present in one of the FAD-binding sites. Our results have illustrated the heterogeneous mutation spectrum and clinical presentation of IVA in the Japanese patients.

We report herein a 1.5-year-old girl with methylmalonic acidemia (MMA) in whom two missense mutations were found: a novel I739T mutation located in exon 13 and the L494X mutation in exon 8. The results of organic acid test showed a pronounced increase in methylmalonate excretion with increased methylcitrate and 3-OH-propionate excretion, leading to a diagnosis of MMA, and Vitamin B12 administration was started. Analysis of the mut gene confirmed a T-to-A substitution at nucleotide position 1481 in exon 8 and a T-to-C substitution at nucleotide position 2216 in exon 13, leading to the amino acid isoleucine at position 739 being changed to threonine, resulting in c.2216T > C (p.I739T). The patient has now been on high-dose oral administration of Vitamin B12 and carnitine therapy (900 mg of levocarnitine chloride) for 5 years without experiencing further attacks, and her cognitive and motor development is normal. Further tests on residual enzyme activity, as well as experience with more cases, may shed light on the relationship between gene mutations and phenotypes in MMA.

An end-tidal CO2 (ETCO2) monitor (capnometer) is used most often as a noninvasive substitute for P aCO2 in anesthesia, anesthetic recovery and intensive care. However, the utility and accuracy of the portable capnometer in spontaneously breathing patients with or without chronic pulmonary diseases has received little recognition. To determine the utility of the portable capnometer in general wards and in in-home care, we examined the correlation between ETCO2 measured by a portable capnometer and simultaneous P aCO2 measured in 41 spontaneously breathing patients. TV-ETCO2 (ETCO2 measured by tidal volume maneuver) was lower than P aCO2 by an average of 9.0 mmHg and VC-ETCO2 (ETCO2 measured by vital capacity maneuver) was lower than P aCO2 by an average of 0.5 mmHg. The mean difference between P aCO2 and VC-ETCO2 was not statistically significant. Regression analysis showed a close correlation between VC-ETCO2 and P aCO2 (r = 0.91, P < 0.0001). Thus, VC-ETCO2 was highly correlated with P aCO2. Furthermore, a close correlation between VC-ETCO2 and P aCO2 was also observed in patients with compromised pulmonary function (r = 0.88, P < 0.0001 in patients with below 70% of FEV1.0%; r = 0.89, P < 0.0001 in patients with below 80% of %VC). Our studies show that VC-ETCO2 measured by the portable capnometer gives a reliable point estimate ofP aCO2, and can be useful to evaluate the respiratory condition of spontaneously breathing patients in general wards and in in-home care.

Here, we report a case of systemic lupus erythematosus (SLE) complicated by cytomegalovirus (CMV)-induced hemophagocytic syndrome (HPS) and colitis. A 44-year-old woman with SLE was treated with corticosteroid and cyclophosphamide for lupus nephritis. Although her lupus nephritis improved, fever, progressive pancytopenia and intestinal bleeding were observed. A bone marrow aspiration showed an increase in mature histiocytes with hemophagocytosis. In addition, a colonoscopy showed hemorrhagic colitis with ulcer and the biopsy specimen from the colon revealed typical CMV cells with CMV inclusions confirmed by immunohistochemistry. Furthermore, a large number of CMV antigen-positive leukocytes was detected, suggesting an active CMV infection. CMV infection is serious in compromised hosts. Therefore clinicians should be aware of the clinical settings in which this infection can arise and the target organs potentially affected in order to initiate the appropriate intervention.

Here, we report two rare female cases of Sjogren's syndrome with multiple bullae, involving a 66- and a 51-year-old. Neither had any obvious pulmonary complaint. Chest radiographs and high-resolution CT (HRCT) scans showed interstitial linear and nodular opacities and multiple bullae. In the first case spirometry indicated an obstructive change judged by FEV1.0, and ?? 50/ ?? 25. In both cases, histologic examination of the lung revealed thickening of alveolar septa and interstitial mononuclear cell infiltration. In the first case the bullae decreased in size with corticosteroid treatment. Airway narrowing due to peribronchiolar mononuclear cell infiltration causes a check-valve mechanism, which may lead to bullae formation. Although a rare occurrence, it is important to recognize that cystic or bullous lung disease can accompany Sjogren's syndrome.(Intenal Medicine 41:124-128, 2002)

To determine prognostic factors of nosocomial pneumonia in general wards, we performed prospective clinical study using multivariate statistical analysis. Eighty patients with nosocomial pneumonia in our units were enrolled in the study between December, 1996 and January 1998. Clinical setting and severity of pneumonia were evaluated, and laboratory data were collected at the occurrence of nosocomial pneumonia. Death due to nosocomial pneumonia occurred in 29 of 80 patients (mortality rate 36%). Univariate analysis showed the following factors associated with mortality: the presence of an ultimately or rapidly fatal underlying condition, prior antibiotics use, use of antacids, presence of 'high-risk' micro-organisms, sepsis, respiratory failure, multiple organ failure, bilateral chest X-ray infiltrates, a Simplified Acute Physiology Score (SAPS) indexor = 11, albumin3.0 g dl(-1), and lactate dehydrogenase (LDH)or = 796 IUI(-1). Furthermore, multivariate analysis identified three factors significantly associated with mortality: the presence of an ultimately or rapidly fatal underlying condition [odds ratio (OR)=7.0; 95% confidence interval (CI)=1.2-41.1; P=0.03]; SAPS indexor = 11 (OR=7.6; 95% CI=1.1-51.9, P=0.04); LDHor = 796 IUI(-1) (OR=28.2; 95% CI=2.0-406, P=0.01). Our study indicates that host factors and disease severity factors are important prognostic factors of nosocomial pneumonia in general wards.

West syndrome (WS), an intractable epileptic encephalopathy of infancy, is refractory to many antiepileptic drugs; however, adrenocorticotropic hormone (ACTH) is an effective treatment for WS. The mechanism behind the efficacy of ACTH is mediated by biochemical processes that remain unknown. We examined the effects of ACTH therapy with tetracosactide (TCS), a synthetic ACTH analogue, on brain metabolism in patients with WS, using (1)H magnetic resonance spectroscopy (¹H-MRS). In six patients with cryptogenic WS, we performed single-voxel ¹H-MRS at the occipital lobe cortex. Measurements were taken prior to TCS treatment, a few days after therapy, and several months after therapy. Data were also compared with subjects having only mild psychomotor delays. The metabolites measured were glutamine plus glutamate (Glx), N-acetylaspartate (NAA), choline (Cho), and myoinositol (mI); each was expressed as a ratio with creatine plus phosphocreatine (total creatine: tCr). The Glx/tCr ratio was significantly reduced after the TCS treatment. The NAA/tCr ratio was also significantly reduced after the treatment compared with the control group, although the change in NAA signal was heterogeneous among patients, correlating with respective outcomes. The Cho/tCr and mI/tCr ratios were not affected by TCS treatment. The reduction in Glx suggests a decrease in the glutamate-glutamine cycle, which plays a pivotal role in synthesizing neurotransmitters such as glutamate and GABA. TCS-induced Glx reduction may induce changes in synaptic signal transduction, thereby accounting for the effect of TCS on WS. The change in NAA indicates altered neuronal activity, which may be correlated with outcome in WS patients.

Discoidin domain receptor 1 (DDR1) tyrosine kinases constitute a novel family of receptors characterized by a unique structure in the ectodomain (discoidin-I domain). The DDR1 ligand is the extracellular matrix protein collagen.To identify receptor tyrosine kinases (RTKs) involved in control of growth and differentiation of human bronchial epithelial (HBE) cells, a polymerase chain reaction-based search for RTKs in HBE cells was performed. DDR1 was the most abundant clone identified. Northern analysis detected a 3.6 kb DDR1 messenger ribonucleic acid (mRNA) expressed in HBE cells and transformed HBE lines, BET-1A and BEAS-2B. In addition, fluorescence-activated cell sorter (FACS) analyses using an anti-DDR1 antibody showed that DDR1 was expressed on HBE cells and two HBE lines.Immunohistochemical staining using human bronchial tissue demonstrated that DDR1 was mainly expressed at the basolateral cell surface of the bronchial epithelium. Furthermore, immunostaining of type IV collagen, a major component of the basement membrane, clearly showed that the basement membrane was closely attached to the basal surface of the bronchial epithelium.Since collagen binds to and activates discoidin domain receptor 1 tyrosine kinase, colocalization of discoidin domain receptor 1 and its ligand type IV collagen demonstrates a potential interaction of discoidin domain receptor 1 on the bronchial epithelium with type IV collagen. Further study of this interaction may define the functional significance of the collagen-discoidin domain receptor 1 signalling pathway in health and in disease.

Holocarboxylase synthetase (HCS) deficiency is a rare autosomal recessive disorder of biotin metabolism. Including three new Japanese patients we diagnosed in this study, ten Japanese families have, so far, been accumulated. In these families, the mutations 237Leu > Pro (sevenalleles) and 1067delG (five alleles) were predominant; 508Arg > Trp and 550Val > Met mutations were identified in three families in the heterozygous form and in one patient in the homozygous form, respectively. To determine the origin of these mutations, we identified new polymorphic microsatellite markers in the HCS gene and analyzed the haplotypes of the patients. All the 237Leu > Pro and the 1067delG alleles were associated with haplotype 2-2. This finding is consistent with the notion that these mutations are founder mutations in the Japanese population. Three Japanese 508Arg > Trp alleles were associated with several haplotypes, including 2-3 and 1-4. The haplotype of a Taiwanese patient homozygous for the 508Arg > Trp mutation was 2-3/2-3. The haplotype of one Japanese patient homozygous for the 550Val > Met mutation was 1-4/1-4, whereas that of a Jewish patient with the same homozygous mutation was 2-3/2-3. Both mutations were associated with at least two haplotypes and were found in several ethnic groups. The changes 508Arg > Trp and 550Val > Met occurred at CpG dinucleotide. The data suggest that these two mutations represent a mutational hot-spot.

We recently showed that macrophage-stimulating protein (MSP), a serum protein homologous to hepatocyte growth factor, promotes ciliary motility by activating its receptor, RON, on the airway ciliated epithelium. To investigate the functional involvement of MSP and RON in bronchiectasis, in which mucociliary clearance (MCC) is impaired, first we examined RON expression on the bronchial ciliated epithelium of patients with bronchiectasis. We confirmed RON expression at the apical surface of bronchial ciliated epithelium of patients with bronchiectasis as well as those of normal human bronchus. Next, we examined whether MSP is present in sputum of patients with bronchiectasis and normal control subjects. By Western blotting, we found that half of the MSP in sputum is present as a biologically active alpha/beta chain heterodimer (mature MSP). In addition, we found that the MSP concentrations in sputum were significantly elevated in patients with bronchiectasis (n=8; 16.8+/-3.0 ng ml(-1)) compared with normal controls (n=9; 8.4+/-2.4 ng ml(-1); P

The P-channel DINOR flash memory, which uses the band-to-band tunneling induced hot electron (BBHE) program method having the advantages of high scalability, high efficiency, and high oxide reliability, was fabricated by 0.35-/spl mu/m-rule CMOS process and was investigated in detail. An ultra-high programming throughput of less than 8 ns/byte (=4 /spl mu/s/512 byte) and a low current consumption of less than 250 /spl mu/A were achieved by utilizing 512-byte parallel programming. Furthermore, we investigated its endurance characteristics up to 10/sup 6/ program/erase cycles, and window narrowing and G/sub m/ degradation were found to be very small even after 10/sup 6/ cycles. It is thought that the BBHE injection point contributes to the G/sub m/ stability and the oxide-damage-reduced operation contributes to the good window narrowing characteristics. The P-channel DINOR flash memory realizing high programming throughput with low power consumption is one of the strongest candidates for the next generation of high-performance, low-voltage flash memories.

Holocarboxylase synthetase (HCS) is an essential enzyme for the biotinylation of several mammalian carboxylases. A deficiency of HCS is accountable for early onset biotin-responsive multiple carboxylase deficiency. To address the mechanism of biotin responsiveness, we analyzed the kinetic properties of the previously identified mutant, L237P, and another mutant, V550M, described in this report. The V550M mutant contains a G to A transition at position 1935, which is within the putative biotin binding site, whereas the mutation in L237P occurs outside the biotin binding site. Km and Vmax values for the mutant proteins were determined by overexpressing cDNAs encoding the mutants in transformed fibroblasts from an HCS-deficient patient. Enzyme activity assays were performed using apo-carboxyl carrier protein as a substrate. A Km for biotin that was larger than the value found for the wild-type cDNA was observed in fibroblasts transfected with the V550M cDNA, but not the L237P cDNA. The Vmax for the expressed L237P cDNA was 4.3% of that observed for the wild-type cDNA. Biotin-responsiveness in the patient with the L237P mutation was neither due to an increased affinity for biotin nor a restoration of stability of the mutant by biotin treatment. A new mechanism of biotin responsiveness in HCS deficiency is presented.

Macrophage-stimulating protein (MSP) is an 80-kD serum protein with homology to hepatocyte growth factor (HGF). Its receptor, RON tyrosine kinase, is a new member of the HGF receptor family. The MSP-RON signaling pathway has been implicated in the functional regulation of mononuclear phagocytes. However, the function of this pathway in other types of cells has not been elucidated. Here we show that in contrast to the HGF receptor, which was expressed at the basolateral surface, RON was localized at the apical surface of ciliated epithelia in the airways and oviduct. In addition, MSP was found in the bronchoalveolar space at biologically significant concentrations. MSP bound to RON on normal human bronchial epithelial cells with a high affinity (Kd = 0.5 nM) and induced autophosphorylation of RON. Activation of RON by MSP led to a significant increase in ciliary beat frequency of human nasal cilia. These findings indicate that the ciliated epithelium of the mucociliary transport apparatus is a novel target of MSP. Ciliary motility is critical for mucociliary transport. Our findings suggest that the MSP-RON signaling pathway is a novel regulatory system of mucociliary function and might be involved in the host defense and fertilization.

Holocarboxylase synthetase (HCS) deficiency is an inherited disease of biotin metabolism characterized by a unique pattern of organic aciduria, metabolic acidosis, and skin lesions. By analysis of five patients in four unrelated families, two mutations were identified: a transition from T to C which causes an amino-acid substitution of proline for leucine at position 237 (L237P) and a single deletion of guanine (delG 1067) followed by premature termination. One patient was homozygous for the L237P mutation, three patients in two families were compound heterozygotes of the missense and deletion alleles, and the other patient was heterozygous for the L237P mutation. Inheritance was successfully demonstrated in all of the patients' families by a modified PCR followed by restriction enzyme digestion. The two mutations accounted for seven of eight mutant alleles, while neither mutation was detected in 108 normal healthy Japanese children (216 alleles). Transient expression in cultured fibroblasts from a patient showed that the L237P mutation was responsible for decreased HCS activity. These results suggest that the L237P and delG1067 mutations are frequent disease-causing mutations in Japanese patients with HCS deficiency. This PCR-based technique may therefore be useful for detecting mutations among Japanese patients.

Interleukin-13 (IL-13) is a pleiotropic cytokine that inhibits the production of inflammatory cytokines of monocytes. We investigated the effects of IL-13 on the clonal growth of human hematopoietic progenitors. IL-13 alone did not support any colony formation. IL-13 markedly suppressed macrophage colonies that were formed in the presence of IL-3 and erythropoietin, granulocyte-macrophage colony-stimulating factor, or macrophage colony-stimulating factor. Macrophage colony cells showed dendritic cell-line morphology and cellular aggregates. IL-13 did not affect granulocyte colony and erythroid burst formation. Delayed addition of IL-13 and replating onto the culture dishes with IL-13 showed that macrophage colony formation was suppressed during days 8 and 14 of culture. These results indicate that IL-13 affects the growth of the late stage of committed macrophage progenitors. Single-cell culture of isolated CD34+CD33+ cells with IL-13 confirmed that macrophage colony formation was significantly suppressed. These results show that IL-13 directly suppresses the proliferation of differentiating macrophages. In addition, these suppressive effects of IL-13 were synergistic with IL-4. Furthermore, in the liquid culture of bone marrow cells in the presence of IL-13, the number of CD14 (monocyte-macrophage antigen)-positive cells decreased and CD18 (LFA-1 beta)-positive cells increased. It is concluded that IL-13 affects the growth of the late stage of macrophage precursors as well as mature monocytes. Induction of differentiation of human monocytes may be correlated with the suppression of their progenitors.

Intestinal lactase is required for the hydrolysis of lactose that is the most essential carbohydrate in milk and the primary diet source of newborn. Congenital lactase deficiency [CLD (MIM 223000)] is a severe gastrointestinal disorder and is characterized by watery diarrhea due to an extremely low or the lack of lactase activity in the intestinal wall from birth. CLD is a rare disease and occurs more frequently in Finland. Recent studies have shown that mutations in the coding region of the lactase (LCT) gene underlie CLD in patients from Finland and other European countries. Here, we report two novel mutations in the LCT gene in a Japanese female infant with clinical features consistent with those of CLD. She suffered from severe watery diarrhea from the age of 2 days on breast milk/lactose containing cow's milk formula. With the lactose-free hydrolyzed cow's milk formula, diarrhea was stopped, and she has now developed well on a lactose-free diet. She shows a lactose-intolerance pattern on the lactose challenge test. Sequence analysis revealed the two mutations in her LCT gene: c.4419C>G (p.Y1473X) in exon 10 transmitted from her mother and c.5387delA (p.D1796fs) in exon 16 transmitted from her father. Both mutations cause premature truncation of lactase polypeptide and are supposed to be responsible for CLD. To our knowledge, this is the first report on mutations in the LCT gene in Japan. We suggest that an increased awareness is required regarding CLD.

We present the first mutation proven case of Fanconi-Bickel syndrome, a rare type of glycogen storage disease, from India. A four-year-old girl presented with severe growth retardation, genu varum and hepatomegaly. Investigations confirmed severe hypophosphatemic rickets and Fanconi syndrome. Molecular analysis confirmed a homozygous deletion insertion mutation in Glut 2 gene.

Fanconi-Bickel syndrome is a rare inherited disorder of carbohydrate metabolism. The disease is characterized by the association of a massive hepatomegaly due to glycogen accumulation, severe hypophosphatemic rickets and marked growth retardation due to proximal renal tubular dysfunction. Fanconi-Bickel syndrome is a single gene disease and is caused by defects in the facilitative glucose transporter 2 (GLUT2) gene (SLC2A2) on chromosome 3q26.1-26.3, which encodes for the glucose transporter protein 2 expressed in hepatocytes, pancreatic beta-cells, enterocytes, and renal tubular cells. Several mutations in a gene encoding a glucose transporter have been reported in patients with Fanconi-Bickel syndrome. Here we report a Turkish child who had a novel mutation that has not been described before and we discuss the knowledge regarding genetic mutations in this rare disease.

The conventional method used for glass melting is air-fuel firing, which is inefficient, energy-intensive and time-consuming. In this study, an innovative in-flight melting technology was developed and applied to glass production for the purposes of energy conservation and environmental protection. Three types of heating sources, radio-frequency (RF) plasma, a 12-phase alternating current (ac) arc and an oxygen burner, were used to investigate the in-flight melting behavior of granulated powders. Results show that the melted particles are spherical with a smooth surface and compact structure. The diameter of the melted particles is about 50% of that of the original powders. The decomposition and vitrification degrees of the prepared powders decrease in the order of powders prepared by RF plasma, the 12-phase ac arc and the oxygen burner. The largest heat transfer is from RF plasma to particles, which results in the highest particle temperature (1810 °C) and the greatest vitrification degree of the raw material. The high decomposition and vitrification degrees, which are achieved in milliseconds, shorten the melting and fining times of the glass considerably. Our results indicate that the proposed in-flight melting technology is a promising method for use in the glass industry.

Isolated 3-methylcrotonyl-CoA carboxylase (MCC) deficiency appears to be the most frequent organic aciduria detected in tandem mass spectrometry (MS/MS) screening programs in the United States, Australia, and Europe. A pilot study of newborn screening using MS/MS has recently been commenced in Japan. Our group detected two asymptomatic MCC deficiency patients by the pilot screening and collected data on another three MCC deficiency patients to study the molecular bases of the MCC deficiency in Japan. Molecular analyses revealed novel mutations in one of the causative genes, MCCA or MCCB, in all five of the patients: nonsense and frameshift mutations in MCCA (c.1750C > T/c.901_902delAA) in patient 1, nonsense and frameshift mutations in MCCB (c.1054_1055delGG/c.592C > T) in patient 2, frameshift and missense mutations in MCCB (c.1625_1626insGG/c.653_654CA > TT) in patient 3, a homozygous missense mutation in MCCA (c.1380T > G/ 1380T > G) in patient 4, and compound heterozygous missense mutations in MCCB (c.569A > G/ c.838G > T) in patient 5. No obvious clinical symptoms were observed in patients 1, 2, and 3. Patient 4 had severe neurological impairment and patient 5 developed Reye-like syndrome. The increasing use of MS/MS newborn screening in Japan will further clarify the clinical and genetic heterogeneity among patients with MCC deficiency in the Japanese population.

Methylmalonic acidemia (MMA) is caused by a deficiency in the activity of l-methylmalonyl-CoA mutase (MCM), a vitamin B12 (or cobalamin, Cbl)-dependent enzyme. Apoenzyme-deficient MMA (mut MMA) results from mutations in the nuclear gene MUT. Most of the MUT mutations are thought to be private or restricted to only a few pedigrees. Our group elucidated the spectrum of mutations of Japanese mut MMA patients by performing mutation and haplotype analyses in 29 patients with mut MMA. A sequence analysis identified mutations in 95% (55/58) of the disease alleles. Five mutations were relatively frequent (p.E117X, c.385 + 5G > A, p.R369H, p.L494X, and p.R727X) and four were novel (p.M1V, c.753_753 + 5delGGTATA, c.1560G > C, and c.2098_2099delAT). Haplotype analysis suggested that all of the frequent mutations, with the exception of p.R369H, were spread by the founder effect. Among 46 Japanese patients investigated in the present and previous studies, 76% (70/92) of the mutations were located in exons 2, 6, 8, and 13. This finding – that a limited number of mutations account for most of the mutations in Japanese mut MMA patients – is in contrast with results of a previous study in Caucasian patients.