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Impaired response to erythropoiesis-stimulating agents (ESAs) is associated with increased mortality in patients with end-stage kidney disease. However, the underlying mechanisms are not fully elucidated. Accumulating data reveal that selenium (Se), a trace element, plays a key role in stress erythropoiesis and erythrocyte homeostasis. We evaluated the relationship between serum Se levels and the response to ESAs in hemodialysis patients.In this cross-sectional study, we determined serum Se levels in 173 hemodialysis patients. We analyzed the association of serum Se with ESA responsiveness, as defined by ESA resistance index.Of the study participants, 50% had lower Se levels than the population-based reference values. We found that serum Se levels were significantly and inversely correlated with erythropoiesis resistance index (ERI) but not transferrin saturation (TSAT) or ferritin levels. Multiple regression analyses confirmed the association between Se levels and ESA hyporesponsiveness, independently of other known factors, such as iron status, being female, and dialysis vintage (β = -0.11,Serum Se levels are associated with the response to ESAs and can predict ESA resistance independently of iron status in Japanese hemodialysis patients. These data open the possibility to test whether Se supplementation reduces ESA demand.

BACKGROUND Percutaneous endoscopic gastrostomy with jejunal extension (PEG-J) is often used to treat patients with neurological impairment and difficulty in swallowing. However, these patients often develop copper deficiency. This report describes a case of isolated neutropenia, which is a rare manifestation of copper deficiency. CASE SUMMARY Our patient was a 19-year-old boy with neurological impairment and gastroesophageal reflux. He received PEG-J feeding, including an enteral supplement containing copper and zinc. However, as his serum zinc level was low (53 μg/dL) at the age of 19 years and 2 mo, we changed to a zinc-rich supplement containing 22 mg/d of zinc and 1.0 mg/d of copper. The supplement comprised a mixture of isocal 1.0 junior (5 packs/d), Tezon [2 packs (250 mL)/d], and cocoa powder. Seven months later, he had neutropenia (606/mm3) with a serum copper level of 16 μg/dL. There were no other manifestations of copper deficiency, including anemia. Copper deficiency and neutropenia both improved following the administration of cocoa powder and Tezon. CONCLUSION In patients receiving long-term PEG-J feeds, white blood cell counts, hemoglobin, and serum levels of copper and zinc should be regularly monitored.

Menkes disease is a rare hereditary disease in which systemic deficiency of copper due to mutation of the ATP7A gene causes severe neurodegenerative disorders. The present parenteral drugs have limited efficacy, so there is a need for an efficacious drug that can be administered orally. This study focused on glyoxal-bis (N(4)-methylthiosemicarbazonato)-copper(II (CuGTSM), which has shown efficacy in macular mice, a murine model of Menkes disease, and examined its pharmacokinetics. In addition, nanosized CuGTSM (nCuGTSM) was prepared, and the effects of nanosizing on CuGTSM pharmacokinetics were investigated. CuGTSM or nCuGTSM (10 mg/kg) was administered orally to male macular mice or C3H/HeNCrl mice (control), and plasma was obtained by serial blood sampling. Plasma concentrations of CuGTSM and GTSM were measured by LC-MS/MS and pharmacokinetic parameters were calculated. When CuGTSM was administered orally, CuGTSM and GTSM were both detected in the plasma of both mouse strains. When nCuGTSM was administered, the Cmax was markedly higher, and the mean residence time was longer than when CuGTSM was administered for both CuGTSM and GTSM in both mouse strains. With macular mice, the AUC ratio (GTSM/CuGTSM) was markedly higher and the plasma CuGTSM concentration was lower than with C3H/HeNCrl mice when either CuGTSM or nCuGTSM was administered. Absorption of orally administered CuGTSM was confirmed in macular mice, and the nano-formulation improved the absorption and retention of CuGTSM in the body. However, the plasma concentration of CuGTSM was lower in macular mice than in control mice, suggesting easier dissociation of CuGTSM.

BACKGROUND Many trace elements are essential for infant growth and development during early life. Their concentrations in breast milk vary depending on social and economic factors. Nonetheless, the present available values in Japan were derived from lactating mothers approximately 15 years ago. METHODS Healthy mothers who gave birth to a single infant after 37 weeks of gestation at Teikyo University Hospital were recruited between July 2016 and December 2017. They were encouraged to collect breast milk samples and a self-administered food frequency questionnaire at 1 and 3 months postpartum. Anthropometric data for the mothers and their infants were also collected. Overall, 79 subjects were analyzed after excluding subjects with inadequate data in the food frequency questionnaire, insufficient breast milk samples, and medication that could affect dietary intakes. Trace element concentrations were determined by inductively coupled plasma mass spectrometry, and their correlation with several factors were investigated. RESULTS Trace element concentrations were widely distributed as previously reported. Median concentrations of Cr, Mn, Fe, Cu, Zn, Se, and Mo were 0.8, 0.8, 98, 50, 272, 2.2, and 0.7 µg/dL at 1 month postpartum and 0.6, 0.7, 55, 33, 177, 2.1, and 0.7 µg/dL at 3 months postpartum, respectively. There were no correlations between trace element concentrations and either mothers' intakes or infants' growth. In contrast, there were significant correlations between several trace elements and macronutrients in addition to inter-element correlations among almost all trace elements. CONCLUSIONS Trace element concentrations in mature breast milk were determined from recently lactating mothers in Japan.

Introduction: Transient neonatal diabetes mellitus (TNDM) is a rare condition that is characterized by the presence of diabetes mellitus during the first 6 months of life and remission by 18 months of age. It usually relapses at a median age of 14 years. Hyperinsulinaemic hypoglycaemia is a relatively common complication during remission. Although β-cell function is reported to be impaired at relapse, the clinical course of glycaemic profiles during remission in patients with TNDM remains largely unknown. Case Presentation: Longitudinal glycaemic profiles were investigated annually from remission (185 days) to relapse (14.5 years) in a patient with TNDM due to paternal 6q24 duplication using the oral glucose tolerance test (glucose intake: 1.75 g/kg to a maximum of 75 g). The patient’s β-cell function and insulin sensitivity were assessed by calculating the insulinogenic index, homeostasis model assessment of β-cell function (HOMA-β), homeostasis model assessment of insulin resistance (HOMA-IR), quantitative insulin sensitivity check index, and Matsuda index. Early insulin response to glucose intake was impaired throughout remission, whereas fasting insulin and β-cell function by HOMA-β gradually increased in the first few years since remission, followed by a gradual decline in function. In contrast, HOMA-IR fluctuated and peaked at 6.5 years of age. Conclusion: This is the first report of annual longitudinal glycaemic profiles in a patient with 6q24-related TNDM during remission. We identified fluctuations in β-cell function and insulin resistance during remission.

For preterm and very low birthweight infants, the mother’s own milk is the best nutrition. Based on the latest information for mothers who give birth to preterm and very low birthweight infants, medical staff should encourage and assist mothers to pump or express and provide their own milk whenever possible.If the supply of maternal milk is insufficient even though they receive adequate support, or the mother’s own milk cannot be given to her infant for any reason, donor human milk should be used.Donors who donate their breast milk need to meet the Guideline of the Japan Human Milk Bank Association.Donor human milk should be provided according to the medical needs of preterm and very low birthweight infants, regardless of their family’s financial status.In the future, it will be necessary to create a system to supply an exclusive human milk‐based diet (EHMD), consisting of human milk with the addition of a human milk‐derived human milk fortifier, to preterm and very low birthweight infants.

Background: Body image self-discrepancy reflects a preference for weight loss regardless of normal body size and is a distorted cognition that may be a precursor to eating disorders. The aim of this study was to investigate factors associated with body image self-discrepancy among healthy junior high school students in Japan.Method: This cross-sectional study was conducted at one junior high school in Saitama, Japan, in December 2016. After excluding obese participants (defined as 20% above their ideal weight), 304 students (mean age, 13.9years; n=181 girls, 59.5%) who fell into underweight (n=22, 7.2%) and normal weight categories were selected. Body image self-discrepancy was measured using the Contour Drawing Rating Scale which includes eight separate figures representing body sizes. We then calculated the difference by subtracting ideal from current body sizes and defined body image self-discrepancy if the difference >1.Results: Girls constituted 92% (n=49) of the 53 students with body image self-discrepancy. In all students, multivariable stepwise models demonstrated that female gender (OR, 6.92, 95% CI: 2.33–20.51), a calorie-restricted diet (OR, 5.18, 95% CI: 2.22–12.05), and psychological symptoms (OR, 1.47, 95% CI: 1.15–1.87) were significantly associated with an increased risk of body image self-discrepancy. Specifically for girls, an increased risk of body image self-discrepancy was associated with calorie-restricted suppers and psychological symptoms.Conclusion: Body image self-discrepancy among healthy adolescents in Japan was found to be closely linked to being a girl, having a calorie-restricted diet, and having psychological symptoms.

The essential trace element zinc maintains liver functions. Liver diseases can alter overall zinc concentrations, and hypozincemia is associated with various hepatic pathologies. Modulating systemic zinc through dietary supplementation is potentially useful for liver diseases. We evaluated the usefulness of zinc (NPC-02; acetate formulation) supplementation. We conducted two NPC-02 studies on zinc-deficient patients (serum zinc

Families are involved in decision-making regarding end-of-life (EOL) care in Japan. However, how support from medical professionals toward families' decision-making affects families' satisfaction with EOL care has not been adequately studied. We aimed to examine the impact of support from medical professionals considering the care recipients' preferences on families' overall satisfaction with EOL care and the mediating effect of fulfilling care recipients' preferences.We administered self-report questionnaires through home-nursing providers to bereaved families (n=753), who lost loved ones between April 2015 and March 2016. Analyses were conducted with 237 of these bereaved families whose loved ones had been ≥65 years old, and had no missing values in key variables. We asked whether the families had received any support from medical professionals in determining the care recipients' EOL preferences, in discussing how to honor the care recipients' own interests, and in supporting the families' decision-making. We also collected data measuring the overall satisfaction with EOL care, families' perceptions that the care recipients' preferences were honored during EOL care, and demographic characteristics of care recipients and caregivers.Data from 58 male and 179 female family members were analyzed. The average age was 65.8 (standard deviation [SD]=11.9) years. Care recipients were 113 men and 124 women, and their average age was 83.0 (SD=9.1) years old at the time of death. A path analysis revealed that support for families from medical professionals was related to families' satisfaction with EOL care through the mediating factor of fulfilling care recipients' preferences.Support from medical professionals considering care recipients' preferences will help families' involvement in EOL decision-making.

Menkes disease is a copper metabolism disorder caused by mutations in ATP7A, a copper-transporting P-type ATPase. In this study, oral copper supplementation via glyoxal-bis(N(4)-methylthiosemicarbazonato)-copper(II) (CuGTSM), a lipophilic copper complex, was investigated in male hemizygous macular (MoMl/y) mice, a mouse model of Menkes disease. CuGTSM was administered by oral gavage on postnatal days 5, 8, 11, 17, 23, and 32. The copper levels in the organs and serum, copper-dependent enzyme activities in the brain, and ceruloplasmin (Cp) activity in the serum were measured at 15 days and 3 and 8 months of age. Histological analysis of the intestines and the rotarod test were also performed. CuGTSM treatment extended the lifespan of MoMl/y mice and partly restored the copper concentrations and cytochrome oxidase and DBH activities in the brain; however, the rotarod test showed impaired motor performance. The treatment also increased copper concentrations and Cp activity in the serum. In suckling MoMl/y mice, CuGTSM treatment transiently induced diarrhea accompanied by copper accumulation and altered villus morphology in the ileum. Oral administration of CuGTSM extended the lifespan of MoMl/y mice. Oral administration is attractive, but pharmaceutical studies are needed to reduce the adverse enteral effects.

According to the recent 2015 Nutrition Survey, the prevalence of being underweight (Body Mass Index, BMI

Background and Aim Few studies of zinc monotherapy for presymptomatic Wilson disease have focused on young children. We therefore evaluated long-term efficacy and safety of zinc monotherapy for such children, and established benchmarks for maintenance therapy. Methods We retrospectively and prospectively examined children under 10 years old with presymptomatic Wilson disease who received zinc monotherapy from time of diagnosis at 12 participating pediatric centers in Japan. Results Twenty-four patients met entry criteria. Aspartate aminotransferase and alanine aminotransferase decreased significantly beginning 1 month after initiation of treatment and usually remained under 50 U/L from 1 to 8 years of treatment. Twenty four-hour urinary copper decreased significantly at 6 months, and usually remained under 75 µg/day and between 1 and 3 µg/kg/day for the remainder of the study. All patients continued to take zinc, and none became symptomatic. In patients under 6 years old who received 50 mg/day of zinc as an initial dose, aspartate aminotransferase and alanine aminotransferase significantly decreased at 1 month after initiation of treatment, as did γ-glutamyltransferase and 24-hour urinary copper at 6 months. Conclusions To our knowledge, this is the first multicenter study of zinc monotherapy for young children with presymptomatic Wilson disease. Such monotherapy proved highly effective and safe. Maintaining normal transaminase values (or values under 50 U/L when normalization is difficult) and 24-hour urinary copper excretion between 1 and 3 µg/kg/day and under 75 µg/day is a reasonable goal. An initial dose of 50 mg/day is appropriate for patients under 6 years old.

Menkes disease (MD) is a lethal infantile neurodegenerative disorder with X-linked inheritance, characterized by progressive neurodegenerative symptoms caused by pathogenic variants in the ATP7A. Early diagnosis and treatment are important, although the diagnosis is difficult prior to 2 months of age. We present an unusually severe case of MD with skull fractures at the birth and repeated fractures during the neonatal period, with further examinations leading to diagnosis. The patient died due to hemorrhagic shock, due to multiple arterial occlusion despite initiation of copper-histidine therapy in early infancy. Bone fracture at birth and multiple arterial occlusion are very rare findings in MD. This unusual and severe presentation emphasizes the importance of early diagnosis and treatment. A congenital bone fracture should be considered as a possible presentation of MD, especially in cases without birth complications.

This study examined whether maternal nutritional intake and breast milk macronutrient content influence the weight of breastfed infants. We investigated 129 healthy mothers with singleton babies born from July 2016 to December 2017 in a university hospital in Tokyo, Japan. Information was obtained by a self-administered food frequency questionnaire at 1 (valid response n = 92, mean age, 34 years) and 3 (n = 57) months after delivery. Breast milk was sampled at 1 and 3 months and the macronutrient contents were analyzed. The average pre-pregnancy body mass index and weight gain during pregnancy were 20.7 &plusmn, 2.6 kg/m2 and 9.6 &plusmn, 3.7 kg, respectively. At 1 month, average maternal calorie intake was 1993 &plusmn, 417 kcal/day, which was lower than the intake recommended by Japanese Dietary Reference Intakes for breastfeeding mothers. There were no significant differences with regard to maternal calorie and protein intake, and breast milk macronutrient content between breastfed infants with weight above and below the 25th percentile of its distribution at both 1 and 3 months. This study suggests that suboptimal calorie intake by breastfeeding mothers and breast milk macronutrient content were not associated with weight of their infants at 1 and 3 months after delivery.

The objective of the study was to investigate whether an infant formula supplemented with galacto-oligosaccharides (GOS; OM55N) was able to stimulate the growth of indigenous bifidobacteria and to establish microbiota similar to that of breastfed infants. A randomised, double-blind, placebo-controlled trial was performed using 35 healthy term infants (31-54 days of age; 42±6 days) to determine whether infant formula with 0.3 g/dl GOS (OM55N) stimulated the growth of bifidobacteria in the infants’ guts. At the trial onset and 2 weeks after, the infants’ faecal samples were examined for microbiota composition (bacterial abundance and α-diversity) and faecal characteristics. Among the 35 infants, 5 were withdrawn and 8 were excluded from the final evaluation before breaking the blinding since the indigenous bifidobacteria were not detected at the trial onset. After 2 weeks, the abundance of Bifidobacteriaceae was significantly increased in the GOS feeding group compared to the control (+11.6±24.1% vs -3.9±13.0%; P=0.043). The Shannon index, which accounts for both abundance and evenness of the present species, was significantly decreased with GOS supplementation (-0.1±0.4 vs +0.4±0.4; P=0.014). Faecal characteristics such as pH and organic acids were similar in both groups, with no statistical differences. No adverse side effects related to the formula consumption were reported. Although the concentration of GOS was relatively low, the infant formula with GOS increased the abundance of bifidobacteria and resulted in a reduced α-diversity of the microbiota.

Background Biotin deficiency has been reported in Japanese infants fed special formulas for medical reasons, including those with milk allergy and congenital metabolic diseases, because these formulas contain little biotin. Serum biotin measurement is useful for diagnosing biotin deficiency. We applied a simple and rapid method to analyze serum biotin, and established normal ranges for children and adults. Methods Serum biotin in 188 healthy Japanese children aged 0–4 years and in 25 healthy adults was analyzed using a Biotin ELISA Kit (immundiagnostik). The effects of various conditions on the measurement of serum biotin were also examined. Results Median biotin in children aged 0–4 years was 10.4 ng/dL (IQR, 7.9–13.4 ng/dL), and that in adults was 12.9 ng/dL (IQR, 10.8–15.8 ng/dL). Normal range was 4.7–22.0 ng/dL in children and 8.4–20.5 ng/dL in adults (calculated using two-sided 95%CI). Measurements obtained with this method were not affected by frozen storage, freeze–thaw, or hemolysis, indicating that serum biotin can be analyzed accurately under these conditions, with a possible application to plasma samples. Conclusions Serum biotin was significantly lower in children than in adults, with the normal range being 4.7–22.0 ng/dL in children and 8.4–20.5 ng/dL in adults. This simple and accurate enzyme-linked immunosorbent assay method is useful for diagnosing biotin deficiency.

A 7-month-old girl was brought to hospital due to vomiting. Upon admission, she was in a convulsive state and stupor with extremely low blood glucose. Head computed tomography showed brain edema, and comprehensive treatment for acute encephalopathy was initiated immediately. Severe hypoglycemia, metabolic acidosis, elevation of ammonia and serum transaminases and creatine kinase suggested metabolic decompensation. Infusion of a high-glucose solution containing vitamins, biotin, and l-carnitine resolved the metabolic crisis quickly, but brain damage was irreversible. She was found to have been fed exclusively on a hypoallergenic formula (HF) for 7 months, although she was found later to be non-allergic. Evidence of inborn metabolic diseases was absent, therefore biotin deficiency and carnitine deficiency were concluded to be a consequence of reliance on a HF for a prolonged period. Health-care professionals should warn parents of the consequences of using HF.

BACKGROUND Given that nutritional biotin deficiency in Japanese infants has been reported, a straightforward method for estimating biotin level is needed. The biotin content in infant formula, breast milk, and the sera of infants fed with various types of formula were measured using avidin-binding assay. METHODS A commercially available ELISA kit was used for the measurement of biotin in 54 types of formula, including hydrolysate formulas for milk allergy, as well as in breast milk and in the sera of 27 infants fed with these formulas. RESULTS The biotin content reached the recommended value in only five formulas. All of the hydrolysate formulas and more than half of the special formulas contained biotin

Background Cyclic vomiting syndrome is characterized by recurrent vomiting that is associated with increased adrenocorticotropic hormone and antidiuretic hormone levels during cyclic vomiting syndrome attacks. However, both prognosis and treatment remain unclear. We therefore evaluated the clinical features, prognosis, and effectiveness of the prophylaxis of cyclic vomiting syndrome as well as the relationship between symptoms and adrenocorticotropic hormone/antidiuretic hormone levels. Methods We included 31 patients with cyclic vomiting syndrome who were admitted to Teikyo University between 1996 and 2008. All patients were diagnosed with cyclic vomiting syndrome based on the criteria of the second edition of the International Headache Classification. The patients (25 of 31) were followed until 2013. Results The median overall duration of the disorder was 66 (3-179) months. Follow-up was completed for 25 patients with cyclic vomiting syndrome, of whom 44% (n = 11) developed migraine. Valproic acid, valproic acid with phenobarbital, phenobarbital, and amitriptyline were effective in nine, four, three, and one patients, respectively. Abnormally high adrenocorticotropic hormone (n = 17) and antidiuretic hormone (n = 18) levels were found among the 25 patients for whom follow-up data were available. The following correlations were significant: attack duration and adrenocorticotropic hormone levels (correlation coefficient: 0.5153, P = 0.0084) and attack duration and antidiuretic hormone levels (correlation coefficient: 0.5666, P = 0.0031). Antidiuretic hormone levels in patients with bilious vomiting were higher than in those without bilious vomiting ( P = 0.048). Conclusions Most patients with cyclic vomiting syndrome recovered completely and benefited from prophylactic therapy, although half of them developed migraines.

Zinc deficiency is common in Japan, yet awareness on this disorder is lacking. The Japanese Society of Clinical Nutrition recently issued the Japan’s Practical Guideline for Zinc Deficiency 2018 setting forth criteria for diagnosing zinc deficiency, i.e., (a) one or more symptoms of zinc deficiency or low serum alkaline phosphatase, (b) ruling out other diseases, (c) low serum zinc, and (d) alleviation of symptoms upon zinc administration. Serum zinc

Menkes disease (MD) is an X-linked recessive disorder caused by dysfunction of a copper-transporting protein, leading to severe neurodegeneration in early childhood. We investigated whether a lipophilic copper chelator, disulfiram, could enhance copper absorption from the intestine and transport copper across the blood–brain barrier in MD model mice. Wild type and MD model mice were pretreated with disulfiram for 30 min before oral administration of 64CuCl2. Each organ was sequentially analyzed for radioactivity with γ counting. Copper uptake into the brain parenchyma was assessed by ex vivo autoradiography. In wild type mice, orally administered copper was initially detected in the intestine within 2 h, reaching a maximum level in the liver (19.6 ± 3.8 percentage injected dose per gram [%ID/g]) at 6 h. In MD model mice, the copper reached the maximum level in the liver (5.3 ± 1.5 %ID/g) at 4 h, which was lower than that of wild type mice (19.0 ± 7.4 %ID/g) (P

The deficiency or excess intake of trace elements, including zinc, copper, selenium and iodine, has often been reported. Zinc deficiency is often observed in infants fed breast milk with low zinc concentration, individuals administered chelating medicines, athletes and patients with diabetes mellitus, hepatic cirrhosis or nephrosis syndrome. Menkes disease is associated with severe copper deficiency, and there is no effective treatment. Deficiencies of selenium and iodine are observed in patients who receive special formulas of milk and enteral formula with low selenium and iodine concentrations, respectively. In contrast, neonatal transient hypothyroidism due to excess intake of iodine in pregnant women has also reported in Japan. It is expected that collaborative studies by researchers and clinicians will contribute to clarify the detail mechanism, diagnosis and treatment of these abnormalities.

Menkes disease (MD) is a neurodegenerative disorder characterized by copper deficiency. It is caused by defective intestinal absorption of copper resulting from a deficiency of a copper-transporting ATPase, ATP7A. We investigated the effects of combination therapy with copper and disulfiram, a known lipophilic chelator. We synthesized a copper-disulfiram complex (Cu-DSF) and determined its crystal structure by X-ray crystallographic analysis. Unfortunately, Cu-DSF was not orally bioavailable due to its lipophilicity. We therefore planned to use cyclodextrin as a solubilizing agent to increase the water solubility of Cu-DSF. After comparisons of the effects of cyclodextrins (α, β, γ), it was found that addition of β-cyclodextrin (β-CyD) increased the solubility of Cu-DSF. Moreover, the modified β-CyD, hydroxypropyl-β-cyclodextrin, was yet more effective as a solubilizing agent. For the development of a convenient method to determine the concentration of Cu-DSF included by β-cyclodextrins, a standard curve based on UV-visible(VIS) absorption was derived.

Background: Menkes disease is a lethal disorder associated with copper metabolism. Although early treatment with copper-histidine injections can improve outcomes, early diagnosis is difficult because the clinical features of Menkes disease are subtle or do not manifest in affected neonates. Previous report stated that the low activity of dopamine β-hydroxylase, a copper-dependent enzyme, leads to increases in the urine homovanillic acid/vanillylmandelic acid (HVA/VMA) ratios in patients with Menkes disease, and indicated that a urine HVA/VMA ratio cut-off value of >4 is useful in screening for Menkes disease. Methods : We examined the standard values of the urine HVA/VMA ratio in unaffected neonates and assessed its use as a screening parameter for Menkes disease among neonates. In total, 112 neonates, aged between 1 and 6days, were enrolled in the study and were classified into 2 groups based on their urine HVA/VMA ratios: high (>4) and low (⩽4). Results : Multivariate logistic analysis revealed that mechanical ventilation was an independent risk factor for a high urine HVA/VMA ratio (odds ratio: 21.94; 95% confidence interval: 2.82–247.03; p =0.004). The mean urine HVA/VMA ratio was 2.47±0.67 among 92 neonates who did not receive mechanical ventilation. Conclusion : This study established standard values for the urine HVA/VMA ratio in newborn babies that could be useful in screening for Menkes disease among neonates.

Background Menkes disease (MNK; MIN 309400) is an X-linked recessive lethal disorder of copper metabolism caused by mutations in ATP7A (MIM 300011), which encodes a transmembrane copper-transporting P-type ATPase. This study assessed mutations in ATP7A in Japanese patients with MNK and their families using gene analysis. Methods A total of 66 patients with MNK born between 1975 and 2013 in Japan were investigated in this study. Definite diagnosis of MNK was carried out on polymerase chain reaction (PCR) amplification and direct sequencing of each exon. Genetic analysis was also performed on 39 women for carrier diagnosis, and on nine fetuses and 10 neonates for the diagnosis of MNK. Results We detected 55 different mutations, of which 20 were de novo mutations. The mutations were located around the six copper binding sites, first to third and six transmembrane domains, and the ATP binding site. Of 30 mothers, 23 (76.7%) were carriers. Approximately half of the male siblings of patients with MNK were also diagnosed with MNK. Conclusion Mutations in ATP7A varied widely across patients, although approximately half of the mutations were located in exons 4, 9, 10, and 15. Approximately 23% of patients did not inherit the mutations from their mothers, but had de novo mutations. An early definite diagnosis is necessary for the early treatment of MNK, and gene analysis serves as an effective method for detecting mutations in ATP7A.

Few studies of zinc monotherapy for presymptomatic Wilson disease have focused on young children. We therefore evaluated long-term efficacy and safety of zinc monotherapy for such children and established benchmarks for maintenance therapy.We retrospectively and prospectively examined children under 10 years old with presymptomatic Wilson disease who received zinc monotherapy from time of diagnosis at 12 participating pediatric centers in Japan.Twenty-four patients met entry criteria. Aspartate aminotransferase and alanine aminotransferase decreased significantly beginning 1 month after initiation of treatment and usually remained under 50 U/L from 1 to 8 years of treatment. Twenty four-hour urinary copper decreased significantly at 6 months and usually remained under 75 μg/day and between 1 and 3 μg/kg/day for the remainder of the study. All patients continued to take zinc, and none became symptomatic. In patients under 6 years old who received 50 mg/day of zinc as an initial dose, aspartate aminotransferase and alanine aminotransferase significantly decreased at 1 month after initiation of treatment, as did γ-glutamyltransferase and 24-h urinary copper at 6 months.To our knowledge, this is the first multicenter study of zinc monotherapy for young children with presymptomatic Wilson disease. Such monotherapy proved highly effective and safe. Maintaining normal transaminase values (or values under 50 U/L when normalization is difficult) and 24-h urinary copper excretion between 1 and 3 μg/kg/day and under 75 μg/day is a reasonable goal. An initial dose of 50 mg/day is appropriate for patients under 6 years old.

A supernumerary ovary is an exceedingly rare disorder, in which the structure containing ovarian tissue is located at some distance from the normally placed ovary. 16 cases of endometriosis or tumors originating in a supernumerary ovary have been published in the English literature, but no case of coexisting endometriosis and a tumor has been published. We present the case of a 40-year-old female with cystic endometrioma with coexisting fibroma originating in a supernumerary ovary in the rectovaginal pouch. The present case is the first to be reported with coexisting endometriosis and a tumor originating in a supernumerary ovary. Our experience with this case and the results of our previous studies of rectovaginal endometriosis indicated that the possibility of originating in a supernumerary ovary shall be examined in cases of cystic endometrioma in the rectovaginal pouch.

We have developed an easy and specific enzyme-linked immunoassay (ELISA) for the simultaneous determination of serum metallothinein-1 (MT-1) and 2 (MT-2) in both humans and experimental animals. A competitive ELISA was established using a specific polyclonal antibody against rat MT-2. The antibody used for this ELISA had exhibited the same cross-reactivity with MT in humans and experimental animals. The NH2 terminal peptide of MT containing acetylated methionine was shown to be the epitope of this antibody. The reactivity of this ELISA system with the liver, kidney and brain in MT1/2 knock-out mice was significantly low, but was normal in an MT-3 knock-out mouse. The lowest detection limit of this ELISA was 0.6ng/ml and the spiked MT-1was fully recovered from the plasma. We investigated the normal range of MT1/2 (25-75%tile) in 200 healthy human serum and found it to be 27-48ng/ml, and this was compared with the serum levels in various liver diseases. The serum MT1/2 levels in chronic hepatitis C (HCV) patients were significantly lower than healthy controls and also other liver diseases. In the chronic hepatitis cases, the MT1/I2 levels increased gradually, followed by the progression of the disease to liver cirrhosis and hepatocellular carcinoma. In particular, we found significantly elevated MT1/2 plasma levels in Wilson's disease patients, levels which were very similar to those in the Long-Evans Cinnamon (LEC) rat (model animal of Wilson's disease). Furthermore, a significantly elevated MT1/2 level was found in patients with Menkes disease, an inborn error of copper metabolism such as Wilson's disease.

Menkes disease (MD), an X-linked recessive disorder of copper metabolism caused by mutations in the copper-transporting ATP7A gene, results in growth failure and severe neurodegeneration in early childhood. Subcutaneous copper-histidine injection is the standard treatment for MD, but it has limited clinical efficacy. Furthermore, long-term copper injection causes excess copper accumulation in the kidneys, resulting in renal dysfunction. To attempt to resolve this issue, we used PET imaging with 64Cu to investigate the effects of disulfiram on copper biodistribution in living mice serving as an animal model for MD (MD model mice). Methods: Macular mice were used as MD model mice, and C3H/He mice were used as wild-type mice. Mice were pretreated with 2 types of chelators (disulfiram, a lipophilic chelator, and d-penicillamine, a hydrophilic chelator) 30 min before 64CuCl2 injection. After 64CuCl2 injection, emission scans covering the whole body were performed for 4 h. After the PET scans, the brain and kidneys were analyzed for radioactivity with γ counting and autoradiography. Results: After copper injection alone, marked accumulation of radioactivity (64Cu) in the liver was demonstrated in wild-type mice, whereas in MD model mice, copper was preferentially accumulated in the kidneys (25.56 ± 3.01 percentage injected dose per gram [%ID/g]) and was detected to a lesser extent in the liver (13.83 ± 0.26 %ID/g) and brain (0.96 ± 0.08 %ID/g). Copper injection with disulfiram reduced excess copper accumulation in the kidneys (14.54 ± 2.68 %ID/g) and increased copper transport into the liver (29.42 ± 0.98 %ID/g) and brain (5.12 ± 0.95 %ID/g) of MD model mice. Copper injection with d-penicillamine enhanced urinary copper excretion and reduced copper accumulation in most organs in both mouse groups. Autoradiography demonstrated that disulfiram pretreatment induced copper transport into the brain parenchyma and reduced copper accumulation in the renal medulla. Conclusion: PET studies with 64Cu revealed that disulfiram had significant effects on the copper biodistribution of MD. Disulfiram increased copper transport into the brain and reduced copper uptake in the kidneys of MD model mice. The application of 64Cu PET for the treatment of MD and other copper-related disorders may be useful in clinical settings.

In our previous report, an 80% ethanol bitter gourd seed extract (BGSE) was found to suppress proliferation of adult T-cell leukemia (ATL) cell lines. The present study aimed to identify the bioactive compounds from BGSE specific against ATL. From the result of an HPLC-MS analysis, α-eleostearic acid (α-ESA) was present in BGSE at 0.68% ± 0.0022% (±SD, n = 5). In the cell proliferation test, α-ESA potently suppressed proliferation of two ATL cell lines (ED and Su9T01; IC50 = 8.9 and 29.3 µM, respectively) more than several other octadecanoic acids. However, α-ESA moderately inhibited phytohemagglutinin-activated human peripheral blood mononuclear cells (PBMC; IC50 = 31.0 µM). These results suggest that BGSE-derived α-ESA has potential as a functional food constituent because of its activity against ATL, particularly against ED cells. Moreover, α-ESA might be effective for the prevention of moderate adverse effects of ATL on normal T cells.

Recent studies have demonstrated that gut microbiota development influences infants' health and subsequent host physiology. However, the factors shaping the development of the microbiota remain poorly understood, and the mechanisms through which these factors affect gut metabolite profiles have not been extensively investigated. Here we analyse gut microbiota development of 27 infants during the first month of life. We find three distinct clusters that transition towards Bifidobacteriaceae-dominant microbiota. We observe considerable differences in human milk oligosaccharide utilization among infant bifidobacteria. Colonization of fucosyllactose (FL)-utilizing bifidobacteria is associated with altered metabolite profiles and microbiota compositions, which have been previously shown to affect infant health. Genome analysis of infants' bifidobacteria reveals an ABC transporter as a key genetic factor for FL utilization. Thus, the ability of bifidobacteria to utilize FL and the presence of FL in breast milk may affect the development of the gut microbiota in infants, and might ultimately have therapeutic implications., The factors shaping the development of infants' gut microbiota are poorly understood. Here, the authors show that alterations in gut microbiota development in infants are associated with the presence of bifidobacteria having a gene for utilisation of human milk oligosaccharides.

Infants are vulnerable to zinc deficiency. Thus, abnormally low breast milk zinc levels cause transient neonatal zinc deficiency (TNZD) in breast-fed infants. TNZD has been considered to be rare because of a paucity of citations in the published literature. However, recent studies of affected mothers identified four missense mutations in the solute carrier family 30 member 2 gene (SLC30A2), which encodes the zinc transporter, ZnT2. Genetic analyses of SLC30A2/ZnT2 in three Japanese mothers secreting low-zinc milk (whose infants developed TNZD) were performed. The effects of identified mutations were examined in a cell-based assay. Furthermore, 31 single-nucleotide polymorphisms (SNPs) in SLC30A2/ZnT2 were evaluated for their potential involvement in low-zinc levels in milk. Each mother had a different novel heterozygous mutation in SLC30A2/ZnT2. One mutation reduced splicing efficiency of the SLC30A2/ZnT2 transcript, and all ZnT2 mutants were defective in zinc transport and were unstable in cells. Moreover, four SNPs caused a significant loss of zinc-transport activity, similar to that in disease-causing ZnT2 mutants. Our results indicate that many SLC30A2/ZnT2 mutations cause or potentially cause TNZD. Genetic information concerning TNZD pathogenesis is limited, and our results suggest that the TNZD frequency may be higher than previously thought.

Menkes disease (MNK) is an X-linked recessive disorder. Incidence of live-born infants with MNK is 2.8 per million live births in Japan. The aim of this study was to observe congenital malformations (CMs) in MNK patients. Subjects comprised 35 Japanese male patients with classical MNK who received copper histidine treatment. Patient clinical data were obtained anonymously from medical records or medical record summaries by pediatrician’s retrospective review through a survey. We observed 21 different CMs in 14 patients. Eight of these had a single CM, while six had multiple CMs. The most frequent CM was higher arched palate with other CMs found in five patients. There was no relationship between CMs and mutations in the ATP7A gene. Using Mann–Whitney U tests, age at death was also significantly lower in MNK patients with CMs (P < 0.05), compared to those without CMs, even though there was no significant difference of age onset, age at diagnosis and age at start of treatment with copper histidine between both groups of patients. Sudden death occurred in three MNK patients with CMs only: two with congenital heart disease, and one with microphallus.

Menkes disease (MD) is a genetic neurodegenerative disorder characterized by copper deficiency due to a defect in ATP7A. Standard treatment involves parenteral copper-histidine administration. However, the treatment is ineffective if initiated after two months of age, because the administered copper accumulates in the blood-brain barrier and is not transported to neurons. To resolve this issue, we investigated the effects of a combination therapy comprising copper and disulfiram, a lipophilic chelator, in the macular mouse, an animal model of MD. Seven-day-old macular mice treated subcutaneously with 50 μg of CuCl(2) on postnatal day 4 were used. The mice were given a subcutaneous injection of CuCl(2) (10 μg) with oral administration of disulfiram (0.3mg/g body weight) twice a week until eight weeks of age, and then sacrificed. Copper concentrations in the cerebellum, liver, and serum of treated macular mice were significantly higher than those of control macular mice, which received only copper. Mice treated with the combination therapy exhibited higher cytochrome c oxidase activity in the brain. The ratios of noradrenaline and adrenaline to dopamine in the brain were also increased by the treatment, suggesting that dopamine β-hydroxylase activity was improved by the combination therapy. Liver and renal functions were almost normal, although renal copper concentration was higher in treated macular mice than in controls. These results suggest that disulfiram facilitates the passage of copper across the blood-brain barrier and that copper-disulfiram combination therapy may be an effective treatment for MD patients.