The immunoreactive signature of monocyte-derived dendritic cells from patients with Down syndrome.

The clinical spectrum of Down syndrome (DS) ranges from congenital malformations to premature aging and early-onset senescence. Excessive immunoreactivity and oxidative stress are thought to accelerate the pace of aging in DS patients; however, the immunological profile remains elusive. We investigated whether peripheral blood monocyte-derived dendritic cells (MoDCs) in DS patients respond to lipopolysaccharide (LPS) distinctly from non-DS control MoDCs. Eighteen DS patients (age 2~47 years, 12 males) and 22 controls (age 4~40 years, 15 males) were enrolled. CD14-positive monocytes were immunopurified and cultured for 7 days in the presence of granulocyte-macrophage colony-stimulating factor and IL-4, yielding MoDCs in vitro. After the LPS-stimulation for 48 hours from days 7 to 9, culture supernatant cytokines were measured by multiplex cytokine bead assays, and bulk-prepared RNA from the cells was used for transcriptomic analyses. MoDCs from DS patients produced cytokines/chemokines (IL-6, IL-8, TNF-α, MCP-1, and IP-10) at significantly higher levels than those from controls in response to LPS. RNA sequencing revealed that DS-derived MoDCs differentially expressed 137 genes (74 upregulated and 63 downregulated) compared with controls. A gene enrichment analysis identified 5 genes associated with Toll-like receptor signaling (KEGG: hsa04620, p = 0.00731) and oxidative phosphorylation (hsa00190, p = 0.0173) pathways. MoDCs obtained from DS patients showed higher cytokine or chemokine responses to LPS than did control MoDCs. Gene expression profiles suggest that hyperactive Toll-like receptor and mitochondrial oxidative phosphorylation pathways configure the immunoreactive signature of MoDCs in DS patients.

High-Titer Anti-ZSCAN1 Antibodies in a Toddler Clinically Diagnosed with Apparent Rapid-Onset Obesity with Hypothalamic Dysfunction, Hypoventilation, and Autonomic Dysregulation Syndrome.

Severe obesity in young children prompts for a differential diagnosis that includes syndromic conditions. Rapid-Onset Obesity with Hypothalamic Dysfunction, Hypoventilation, and Autonomic Dysregulation (ROHHAD) syndrome is a potentially fatal disorder characterized by rapid-onset obesity associated with hypoventilation, neural crest tumors, and endocrine and behavioral abnormalities. The etiology of ROHHAD syndrome remains to be established, but recent research has been focusing on autoimmunity. We report on a 2-year-old girl with rapid-onset obesity during the first year of life who progressed to hypoventilation and encephalitis in less than four months since the start of accelerated weight gain. The patient had a high titer of anti-ZSCAN1 antibodies (348; reference range < 40), and the increased values did not decline after acute phase treatment. Other encephalitis-related antibodies, such as the anti-NDMA antibody, were not detected. The rapid progression from obesity onset to central hypoventilation with encephalitis warns about the severe consequences of early-onset ROHHAD syndrome. These data indicate that serial measurements of anti-ZSCAN1 antibodies might be useful for the diagnosis and estimation of disease severity. Further research is needed to determine whether it can predict the clinical course of ROHHAD syndrome and whether there is any difference in antibody production between patients with and without tumors.

BCS1L mutations produce Fanconi syndrome with developmental disability.

Fanconi syndrome is a functional disorder of the proximal tubule, characterized by pan-aminoaciduria, glucosuria, hypophosphatemia, and metabolic acidosis. With the advancements in gene analysis technologies, several causative genes are identified for Fanconi syndrome. Several mitochondrial diseases cause Fanconi syndrome and various systemic symptoms; however, it is rare that the main clinical symptoms in such disorders are Fanconi syndrome without systematic active diseases like encephalomyopathy or cardiomyopathy. In this study, we analyzed two families exhibiting Fanconi syndrome, developmental disability and mildly elevated liver enzyme levels. Whole-exome sequencing (WES) detected compound heterozygous known and novel BCS1L mutations, which affect the assembly of mitochondrial respiratory chain complex III, in both cases. The pathogenicity of these mutations has been established in several mitochondria-related functional analyses in this study. Mitochondrial diseases with isolated renal symptoms are uncommon; however, this study indicates that mitochondrial respiratory chain complex III deficiency due to BCS1L mutations cause Fanconi syndrome with developmental disability as the primary indications.

The earliest enzyme replacement for infantile-onset Pompe disease in Japan.

BACKGROUND: Infantile-onset Pompe disease (IOPD) is the most severe phenotype of a lysosomal storage disorder caused by acid alpha-glucosidase (GAA) deficiency. An enzymatic newborn screening (NBS) program started regionally in Japan in 2013 for early enzyme replacement therapy (ERT). We report the ERT responses of the first NBS-identified Japanese IOPD case and of another case diagnosed prior to NBS, to discuss the problems of promptly starting ERT in Japan. METHODS: Acid alpha-glucosidase activity was measured by fluorometric assay in both patients. The diagnosis of IOPD was confirmed by next-generation followed by Sanger-method sequencing (patient 1) or direct sequencing of polymerase chain reaction (PCR)-amplified products (patient 2) of the GAA gene. RESULTS: A female infant identified by NBS had a novel out-of-frame (p.F181Dfs*6) variant and a reported pathogenic (p.R600C) variant, along with two pseudodeficiency variants. Enzyme replacement therapy was started at age 58 days when the infant had increased serum levels of creatine kinase and slight myocardial hypertrophy. Clinical and biochemical markers improved promptly. She has been alive and well without delayed development at age 14 months. Patient 2, a Japanese male, received a diagnosis of IOPD at age 5 months before the NBS era. He had a homozygotic variant of GAA (p.R608X), later registered as a cross-reactive immunological material (CRIM)-negative genotype, and developed a high titer of anti-rhGAA antibodies. The patient has survived myocardial hypertrophy with continuous respiratory support for 12 years of ERT. CONCLUSIONS: Enzyme replacement therapy should not be delayed over the age of 2 months for reversible cardiac function, although CRIM-negative cases may hamper turnaround time reduction.

Rapid and long-lasting efficacy of high-dose ambroxol therapy for neuronopathic Gaucher disease: A case report and literature review.

Gaucher disease (GD) is a lysosomal storage disorder caused by a deficiency in the GBA1-encoded enzyme, ß-glucocerebrosidase. Enzyme replacement therapy is ineffective for neuronopathic Gaucher disease (nGD). High-dose ambroxol has been administered as an alternative treatment for a group of patients with nGD. However, little is known about the clinical indication and the long-term outcome of patients after ambroxol therapy. We herein report a case of a female patient who presented with a progressive disease of GD type 2 from 11 months of age and had the pathogenic variants of p.L483P (formerly defined as p.L444P) and p.R502H (p.R463H) in GBA1. A combined treatment of imiglucerase with ambroxol started improving the patient's motor activity in 1 week, while it kept the long-lasting effect of preventing the deteriorating phenotype for 30 months. A literature review identified 40 patients with nGD, who had received high-dose ambroxol therapy. More than 65% of these patients favorably responded to the molecular chaperone therapy, irrespective of p.L483P homozygous, heterozygous or the other genotypes. These results highlight the long-lasting effect of ambroxol-based chaperone therapy for patients with an expanding spectrum of mutations in GBA1.

Genotype-Phenotype Correlations in Thirty Japanese Patients with Congenital Hypothyroidism Attributable to TG Defects.

CONTEXT: Thyroglobulin (Tg), encoded by TG, is essential for thyroid hormone synthesis. TG defects result in congenital hypothyroidism (CH). Most reported patients were born before the introduction of newborn screening (NBS). OBJECTIVE: We aimed to clarify the phenotypic features of patients with TG defects diagnosed and treated since the neonatal period. SUBJECTS AND METHODS: We screened 1061 patients with CH for thirteen CH-related genes and identified thirty patients with TG defects. One patient was diagnosed due to hypothyroidism-related symptoms and the rest were diagnosed via NBS. Patients were divided into two groups according to their genotypes, and clinical characteristics were compared. We evaluated the functionality of the seven missense variants using HEK293 cells. RESULTS: Twenty-seven rare TG variants were detected, including fifteen nonsense, three frameshift, two splice-site, and seven missense variants. Patients were divided into two groups: thirteen patients with biallelic truncating variants and seventeen patients with monoallelic/biallelic missense variants. Patients with missense variants were more likely to develop thyroid enlargement with TSH stimulation than patients with biallelic truncating variants. Patients with biallelic truncating variants invariably required full hormone replacement, whereas patients with missense variants required variable doses of levothyroxine. Loss of function of the seven missense variants was confirmed in vitro. CONCLUSION: To our knowledge, this is the largest investigation on the clinical presentation of TG defects diagnosed in the neonatal period. Patients with missense variants showed relatively mild hypothyroidism with compensative goiter. Patients with only truncating variants showed minimal or no compensative goiter and required full hormone replacement.

Guidelines for Newborn Screening of Congenital Hypothyroidism (2021 Revision).

Purpose of developing the guidelines: Newborn screening (NBS) for congenital hypothyroidism (CH) was started in 1979 in Japan, and early diagnosis and treatment improved the intelligence prognosis of CH patients. The incidence of CH was once about one in 5,000-8,000 births, but has been increased with diagnosis of subclinical CH. The disease requires continuous treatment and specialized medical facilities should conduct differential diagnosis and treatment in patients who are positive by NBS to avoid unnecessary treatment. The Guidelines for Mass Screening of Congenital Hypothyroidism (1998 version) were developed by the Mass Screening Committee of the Japanese Society for Pediatric Endocrinology in 1998. Subsequently, the guidelines were revised in 2014. Here, we have added minor revisions to the 2014 version to include the most recent findings. Target disease/conditions: Primary congenital hypothyroidism. Users of the Guidelines: Physician specialists in pediatric endocrinology, pediatric specialists, physicians referring pediatric practitioners, general physicians, laboratory technicians in charge of mass screening, and patients.

Critical diseases in neonates after discharge home from birth hospital: A retrospective study from a tertiary hospital in Japan.

INTRODUCTION: To establish actionable neonatal screening during the first month of life, we investigated critical diseases in seemingly healthy newborns discharged from birth hospitals. METHODS: This retrospective study enrolled previously healthy full-term infants who visited our hospital, a tertiary hospital in Japan, from home between 5 and 28 days after birth from 2009 to 2018. Infants with known perinatal or congenital diseases, positive newborn screening results, or accidental injuries were excluded. Data were collected from electronic medical records, including principal diagnosis, clinical details, and prognosis at 18 months of age. RESULTS: Ninety-seven (58 %) of 168 eligible neonates were admitted to the hospital, and 71 (42 %) were not. The median admission rate in patients with disease onset at ≤14 days after birth (80 %) was significantly higher than that in patients with disease onset at ≥15 days (42 %). Among 45 patients who received intensive medical care, 5 died and 10 developed neurodevelopmental sequelae. Four of 5 patients died by 100 days. Among 25 diseases treated in intensive care unit, 17 (68 %) diseases had a prevalence of <1 per 2000 live births. The commonly used diagnostic methods were imaging (n = 58, 35 %) and physical examination (n = 34, 20 %). CONCLUSION: Critical diseases due to rare and heterogeneous causes in ostensibly healthy newborns occurred predominantly in the first two weeks of life. Optimal newborn screening and health check-up protocols may benefit from the wide spectrum of life-threatening diseases occurring in home after birth.

Clinical and molecular aspects of Japanese children with medium chain acyl-CoA dehydrogenase deficiency.

We report the outcome of 16 Japanese patients with medium chain acyl-CoA dehydrogenase deficiency. Of them, 7 patients were diagnosed after metabolic crisis, while 9 were detected in the asymptomatic condition. Of the 7 symptomatic cases, 1 died suddenly, and 4 cases had delayed development. All 9 patients identified by neonatal or sibling screening remained healthy. Of 14 mutations identified, 10 were unique for Japanese, and 4 were previously reported in other nationalities. Presymptomatic detection including neonatal screening obviously improves quality of life of Japanese patients, probably regardless of the genotypes.

Bezafibrate can be a new treatment option for mitochondrial fatty acid oxidation disorders: evaluation by in vitro probe acylcarnitine assay.

BACKGROUND: The number of patients with mitochondrial fatty acid oxidation (FAO) disorders is recently becoming larger with the spread of newborn mass screening. Despite the advances in metabolic and molecular characterization of FAO disorders, the therapeutic studies are still limited. It was reported recently that bezafibrate (BEZ), an agonist of peroxisome proliferating activator receptor (PPAR), can restore FAO activity in cells from carnitine palmitoyltransferase-2 (CPT2) and very-long-chain acyl-CoA dehydrogenase (VLCAD) deficiencies as well as clinical symptoms in the adult patients. METHODS: In this study, the therapeutic effect of BEZ was determined by in vitro probe acylcarnitine (IVP) assay using cultured fibroblasts and tandem mass spectrometry on various FAO disorders. The clinical trial of BEZ treatment for a boy with the intermediate form of glutaric acidemia type 2 (GA2) was also performed. RESULTS: The effect of BEZ was proven in cells from various FAO disorders including GA2, deficiencies of VLCAD, medium-chain acyl-CoA dehydrogenase, CPT2, carnitine acylcarnitine translocase and trifunctional protein, by the IVP assay. The aberrantly elevated long- or medium-chain acylcarnitines that are characteristic for each FAO disorder were clearly corrected by the presence of BEZ (0.4 mmol/L) in culture medium. Moreover, daily administration of BEZ in a 2-year-old boy with GA2 dramatically improved his motor and cognitive skills, accompanied by sustained reduction of C4, C8, C10 and C12 acylcarnitines in blood, and normalized the urinary organic acid profile. No major adverse effects have been observed. CONCLUSION: These results indicate that BEZ could be a new treatment option for FAO disorders.

Anesthesia management in living-donor liver transplantation in a patient with carbamoyl phosphate synthetase deficiency: a case report.

BACKGROUND: Carbamoyl phosphate synthetase deficiency (CPS1D) is a urea-cycle disorder (UCD). We report successful perioperative management of pediatric living donor liver transplantation (LDLT) in a CPS1D patient. CASE PRESENTATION: A 10-year-old female patient with CPS1D underwent LDLT. Proper administration of dextrose 50% and 60 kcal/kg/day with L-arginine and L-carnitine resulted in the avoidance of intraoperative hyperammonemia induced by hypercatabolism. Serum ammonia level transiently increased to 61 mmol/L in the anhepatic phase and decreased to 44 mmol/L after reperfusion. CONCLUSIONS: We suggest anesthesia management with administration of dextrose to avoid hyperammonemia during LDLT in patients with CPS1D.

Clinical and molecular investigation of 37 Japanese patients with multiple acyl-CoA dehydrogenase deficiency: p.Y507D in ETFDH, a common Japanese variant, causes a mortal phenotype.

Multiple acyl-CoA dehydrogenase deficiency (MADD) is an inherited metabolic disease caused by a defect in electron transfer flavoprotein alpha (ETFA), ETF beta (ETFB), or ETF dehydrogenase (ETFDH), and riboflavin metabolism disorders have recently been reported to present as mimicking MADD. MADD is roughly classified into neonatal (type 1 or 2) and later-onset (type 3) forms. To identify clinicogenetic characteristics in Japan, we investigated 37 Japanese patients with MADD diagnosed from 1997 to 2020. The causes of MADD were ETFDH deficiency in 26 patients, ETFA deficiency in four, ETFB deficiency in six, and riboflavin metabolism disorder in one. All 15 patients with the neonatal-onset type died by 2 years of age, while five of 22 patients with the later-onset form died by 3 years of age. Furthermore, 8 of 15 patients with the later-onset form of ETFDH deficiency treated with riboflavin were riboflavin non-responders. p.Y507D in ETFDH was identified as the most common variant (9 of 48 alleles, 18.8%). Of two patients with a homozygous p.Y507D variant, one experienced disease onset and died in the neonatal period, while the other experienced disease onset at two months of age and died at two years old, suggesting that the p.Y507D variant results in fatal outcomes. Our study concluded that more than half of Japanese patients with MADD died by three years old, and more than half of patients with the later-onset form had poor responsiveness to riboflavin, partly due to the unique Japanese p.Y507D variant in ETFDH.

Clinical and molecular investigation of 19 Japanese cases of glutaric acidemia type 1.

Glutaric acidemia type 1 (GA1) is a metabolic disease caused by a deficiency of glutaryl-CoA dehydrogenase (GCDH). Untreated patients mostly develop severe striatal degeneration. More than 200 mutations have been reported in the GCDH gene, and common R402W and IVS10-2A>C were found in Caucasian and Chinese/Taiwanese, respectively. However, in Japan, genetic mutations have only been reported in a few cases. Herein, we report the clinical and molecular basis of GA1 in 19 Japanese patients, including six previously reported patients. All cases showed high urinary glutaric acid excretion. Eleven patients were severely impaired (three patients died), three had mild impairment, and five showed normal development. Four of 5 patients that developed normally were detected in the presymptomatic stage by neonatal or sibling screening. Nineteen mutations in 26 alleles were identified, and eight of them (89 or 90delC, Y155C, IVS4+2T＞C, G244S, Q352X, G354A, K361E, and 1144-1145delGC) were novel. S305L (12.1%, 4/34 alleles) was found in several cases, suggesting that this mutation is a common mutation. In contrast, R402W was not identified and IVS10-2A>C was only found in one allele, suggesting that Japanese patients with GA1 show allelic heterogeneity and have a different genetic background to patients from other countries. One of a pair of sisters with the same mutations (M339V/S305L) lacking residual activity was severely retarded, whereas the older girl remains asymptomatic at 22 years of age, indicating that genotype does not necessarily predict GA1 phenotype. We consistently found that there was no association between genotype and phenotype. However, children with mild impairment were diagnosed and treated earlier than severely impaired cases {4.7±2.5 months (range: 2-8 months) vs. 11.6±12.7 months (range: 4-51 months)}. Our results suggest that early detection and treatment but not genotype are associated with better patient outcome, reinforcing the importance of neonatal screening.

Fully Automated Quantitative Measurement of Serum Organic Acids via LC-MS/MS for the Diagnosis of Organic Acidemias: Establishment of an Automation System and a Proof-of-Concept Validation.

Gas chromatography-mass spectrometry has been widely used to analyze hundreds of organic acids in urine to provide a diagnostic basis for organic acidemia. However, it is difficult to operate in clinical laboratories on a daily basis due to sample pretreatment processing. Therefore, we aimed to develop a fully automated system for quantifying serum organic acids using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The pretreatment CLAM-2030 device was connected to an LC-MS/MS system for processing serum under optimized conditions, which included derivatizing serum organic acids using 3-Nitrophenylhydrazine. The derivatized organic acids were separated on a reverse-phase Sceptor HD-C column and detected using negative-ion electrospray ionization multiple reaction monitoring MS. The automated pretreatment-LC-MS/MS system processed serum in less than 1 h and analyzed 19 serum organic acids, which are used to detect organic acidemias. The system exhibited high quantitative sensitivity ranging from approximately 2 to 100 µM with a measurement reproducibility of 10.4% CV. Moreover, a proof-of-concept validation of the system was performed using sera from patients with propionic acidemia (n = 5), methylmalonic acidemia (n = 2), and 3-methylcrotonylglycinuria (n = 1). The levels of marker organic acids specific to each disease were significantly elevated in the sera of the patients compared to those in control samples. The automated pretreatment-LC-MS/MS system can be used as a rapid in-hospital system to measure organic acid levels in serum for the diagnosis of organic acidemias.

A case report with functional characterization of a HNF1B mutation (p.Leu168Pro) causing MODY5.

We previously performed next-generation sequencing-based genetic screening in patients with autoantibody-negative type 1 diabetes, and identified the p.Leu168Pro mutation in HNF1B. Here,we report the clinical course of the patient and the results of functional characterization of this mutation. The proband had bilateral renal hypodysplasia and developed insulin-dependent diabetes during childhood. The pathogenicity of Leu168Pro-HNF1B was evaluated with three-dimensional structure modeling, Western blotting, immunofluorescence analysis and luciferase reporter assays using human embryonic kidney 293 cells. Three-dimensional structure modeling predicted that the Leu168 residue is buried in the DNA-binding Pit-Oct-Unc-specific (POUS) domain and forms a hydrophobic core. Western blotting showed that the protein expression level of Leu168Pro-HNF1B was lower than that of wild-type (WT) HNF1B. Immunofluorescence staining showed that both WT- and Leu168Pro-HNF1B were normally localized in the nucleus. The cells transfected with WT-HNF1B exhibited 5-fold higher luciferase reporter activity than cells transfected with an empty vector. The luciferase activities were comparable between WT-HNF1B/Leu168Pro-HNF1B and WT-HNF1B/empty vector co-transfection. In conclusion, Leu168Pro is a protein-destabilizing HNF1B mutation, and the destabilization is likely due to the structural changes involving the hydrophobic core of POUS. The disease-causing Leu168Pro HNF1B mutation is a loss-of-function mutation without a dominant-negative effect.

Brain-sparing cord blood transplantation for the borderline stage of adrenoleukodystrophy.

BACKGROUND: Adrenoleukodystrophy (ALD) is an X-linked disorder characterized by rapidly progressive deterioration of neurocognitive functions and premature death. In addition to the difficulty in identifying the earliest signs of ALD, treatment-associated exacerbation of neurological symptoms has been an obstacle to achieve successful hematopoietic cell transplantation (HCT) for affected children. CASE REPORT: We report a 9-year-boy with ALD. He presented with impairment in social skills compatible to the diagnosis of autism spectrum disorder from 3 years of age. He showed progressive strabismus, slurred speech and dysmetria at 6 years of age. The head MRI showed symmetrical T2-hyperintense lesions in the occipital white matters with a gadolinium enhancement, which extended to the internal capsules. The Loes score was thus calculated as 13. Very-long-chain-fatty-acids were increased to 1.800 (C24:0/C22:0) and 0.077 (C26:0/C22:0) in leukocytes. Sanger sequencing confirmed the pathogenic variant in ABCD1 (NM_000033.4:p.Gly512Ser). After multidisciplinary discussions over the treatment options, we performed a cord blood HCT with a reduced intensity conditioning (fludarabine, melphalan and brain-sparing total body irradiation). He was fully recovered with >90% chimerism of donor leukocytes at 55 days after HCT. He experienced three times of generalized seizures after discharge, that has been well controlled for 2 years without other complications or neurocognitive deteriorations. CONCLUSION: For patients with ALD on a borderline indication for HCT, brain-sparing irradiation might be an alternative option in reduced intensity conditioning. Careful decision-making process and tailored conditioning are critical for the successful outcome of HCT for children with ALD.

Physical, cognitive, and social status of patients with urea cycle disorders in Japan.

Urea cycle disorders (UCDs) are inherited metabolic diseases that lead to hyperammonemia. Severe hyperammonemia adversely affects the brain. Therefore, we conducted a nationwide study between January 2000 and March 2018 to understand the present status of UCD patients in Japan regarding diagnosis, treatments, and outcomes. A total of 229 patients with UCDs (126 patients: ornithine transcarbamylase deficiency [OTCD]; 33: carbamoyl phosphate synthetase 1 deficiency [CPS1D]; 48: argininosuccinate synthetase deficiency [ASSD]; 14: argininosuccinate lyase deficiency [ASLD]; and 8: arginase 1 deficiency [ARG1D]) were enrolled in the present study. Although growth impairment is common in patients with UCDs, we discovered that Japanese patients with UCDs were only slightly shorter than the mean height of the general adult population in Japan. Patients with neonatal-onset UCDs are more likely to experience difficulty finding employment and a spouse; however, some patients with late-onset UCDs were employed and married. Additionally, intellectual and developmental disabilities, such as attention deficit hyperactivity disorder (ADHD) and autism, hinder patients with UCDs from achieving a healthy social life. Moreover, we identified that it is vital for patients with UCDs presenting with mild to moderate intellectual disabilities to receive social support. Therefore, we believe the more robust social support system for patients with UCDs may enable them to actively participate in society.

The expanding phenotype of hypokalemic periodic paralysis in a Japanese family with p.Val876Glu mutation in CACNA1S.

BACKGROUND: Hypokalemic periodic paralysis (HypoPP) is an autosomal dominant disease characterized by the episodic weakness of skeletal muscles and hypokalemia. More than half patients with HypoPP carry mutations in CACNA1S, encoding alpha-1 subunit of calcium channel. Few reports have documented the non-neuromuscular phenotypes of HypoPP. METHODS: The proband is a Japanese woman who developed HypoPP at 6 years of age. An excessive insulin secretion with the oral glucose tolerance test rationalized that she had experienced frequent attacks of paralysis on high-carbohydrate diets. RESULTS: Voglibose and acetazolamide effectively controlled her paralytic episodes. Her 8-year-old son and 2-year-old daughter started showing the paralytic symptoms from 4 and 2 years of age, respectively. Laboratory tests revealed high concentrations of creatinine kinase in serum and elevated renin activities in plasma of these children. The targeted sequencing confirmed that these three patients had an identical heterozygous mutation (p.V876E) in CACNA1S. CONCLUSION: Our data indicate that the p.V876E mutation in CACNA1S contributes to the early onset of neuromuscular symptoms and unusual clinical phenotypes of HypoPP.

A novel molecular aspect of Japanese patients with medium-chain acyl-CoA dehydrogenase deficiency (MCADD): c.449-452delCTGA is a common mutation in Japanese patients with MCADD.

We studied 11 Japanese patients with medium-chain acyl-CoA dehydrogenase deficiency (MCADD) and found a common mutation, c.449-452delCTGA, which accounted for 45% of the mutations. Seven of 10 independent patients carried at least one copy of this mutation. Phenotypes of homozygous patients with the c.449-452delCTGA mutation varied from asymptomatic to life-threatening metabolic decompensation in Japanese patients with MCADD, similar to the phenotypic variations in Caucasians. This study suggests the genotypic difference between those of Caucasians and Japanese regarding MCADD.

Clinical and molecular aspects of Japanese patients with mitochondrial trifunctional protein deficiency.

Mitochondrial trifunctional protein (MTP) deficiency is a rare inherited metabolic disorder of mitochondrial fatty acid oxidation. We newly characterized three novel mutations in 2 Japanese patients with MTP deficiency, and investigated the clinical and molecular aspects of 5 Japanese patients including 3 previously reported cases. Herein, we describe the characterization of four missense mutations, R214C, H346R, R411K, and V422G, in the HADHB gene, which have been identified in Japanese patients, employing a newly developed, sensitive transient expression analysis. Co-transfection of wild-type HADHA and HADHB cDNAs in SV40-transfected fibroblasts from a MTP-deficient patient yielded sufficient enzyme activity to evaluate low-level residual enzyme activity, using two incubation temperatures of 30 degrees C and 37 degrees C. At 30 degrees C, residual enzyme activity was higher than that at 37 degrees C in V422G, R214C, and R411K. However, H346R, which was seen in the most severe case, showed no enzyme activity at both temperatures. Our results demonstrate that a defect of HADHB in MTP deficiency is rather common in Japanese patients, and the mutational spectrum is heterogeneous. The present findings showed that all missense mutations in this study were disease-causing. Although the number of patients is still limited, it is suggested that the phenotype is correlated with the genotype and a combination of two mutant alleles of the HADHB gene in MTP deficiency.