Synthase Deficiency Research Articles

Nutrient-dependent phosphorylation channels lipid synthesis to regulate PPARα

Peroxisome proliferator-activated receptor (PPAR)α is a nuclear receptor that coordinates liver metabolism during fasting. Fatty acid synthase (FAS) is an enzyme that stores excess calories as fat during feeding, but it also activates hepatic PPARα by promoting synthesis of an endogenous ligand. Here we show that the mechanism underlying this paradoxical relationship involves the differential regulation of FAS in at least two distinct subcellular pools: cytoplasmic and membrane-associated. In mouse liver and cultured hepatoma cells, the ratio of cytoplasmic to membrane FAS-specific activity was increased with fasting, indicating higher cytoplasmic FAS activity under conditions associated with PPARα activation. This effect was due to a nutrient-dependent and compartment-selective covalent modification of FAS. Cytoplasmic FAS was preferentially phosphorylated during feeding or insulin treatment at Thr-1029 and Thr-1033, which flank a dehydratase domain catalytic residue. Mutating these sites to alanines promoted PPARα target gene expression. Rapamycin-induced inhibition of mammalian/mechanistic target of rapamycin complex 1 (mTORC1), a mediator of the feeding/insulin signal to induce lipogenesis, reduced FAS phosphorylation, increased cytoplasmic FAS enzyme activity, and increased PPARα target gene expression. Rapamycin-mediated induction of the same gene was abrogated with FAS knockdown. These findings suggest that hepatic FAS channels lipid synthesis through specific subcellular compartments that allow differential gene expression based on nutritional status.

Neonatal hyperphenylalaninemia screening from 1985 to 2012 in Nanjing: a retrospective analysis of 27 years

Objective To explore the incidence,classification,treatment and prognosis of neonatal hyperphenylalaninemia(HPA) in Nanjing.Methods Infants born from September 1st,1985 to September 1st,2012 in Nanjing were investigated.Heel blood samples of neonates were collected at 72 hours after birth,and the concentration of blood phenylalanine was determined by fluorescent quantitative method in Nanjing Neonatal Screening Center within 20 days after birth.The deferential diagnosis was performed in all 81 cases diagnosed as HPA by tetrahydrobiopterin loading test,urinary pterin analysis and dihydropteridine reductase activity determination.The blood phenylalanine concentration and mental development were followed up regularly on infants with HPA.Mutations of phenylalanine hydroxylase (PAH) gene were analyzed by gene sequencing in 20 patients with PAH deficiency.The relationship between phenylalanine concentration and mental development was analyzed by variance analysis,LSD and Pearson correlation analysis.Results (1) The incidence of HPA in neonates in Nanjing was 1/8800(81/712 818).Among 81 confirmed cases,78 cases (96.3％) suffered from PAH deficiency,including 37 classical phenylketonurias (PKU),23 mild PKU and 18 mild HPA; the three patients (3.7％) with tetrahydrobiopterin deficiency were all 6-pyruvoyl tetrahydropterin synthase deficiency.(2) Among these 81 cases,eight refused to any treatment (two of them died),six had mental retardation (one with epilepsy),and six were lost in follow-up.For the rest 67 patients who received medical treatment,the ages at the last visit were three months to 26 years old.Among the 55 patients who received mental development evaluation,five of them were abnormal [intelligence quotient (IQ)＜75].(3) Negative correlation was observed between IQ and the average concentration of phenylalanine(r=-0.53,P＜0.05),while no correlation between IQ and the initial concentration of phenylalanine was observed (r =-0.39,P＞0.05).(4)Among the 20 patients with PAH deficiency who were tested for gene sequencing,18 mutations of PAHgenewere detected.The most common mutation were EX6-96A＞G and R241C(34.28％).Conclusions The incidence of HPA in Nanjing was higher than the average national level.With early diagnosis and standard treatment,over 90％ of HPA patients could have normal mental development.The mutation spectrum of PAH gene of Nanjing population is different from other areas,and the most common mutations of PAH gene are EX6-96A＞G and R241C. Key words: Phenylketonurias; phenylalanine hydroxylase; Mutation; Neonatal screening; Retrospective studies

New case of mitochondrial HMG-CoA synthase deficiency. Functional analysis of eight mutations

Mitochondrial HMG-CoA synthase deficiency is a rare inherited metabolic disorder that affects ketone-body synthesis. Acute episodes include vomiting, lethargy, hepatomegaly, hypoglycaemia, dicarboxylic aciduria, and in severe cases, coma. This deficiency may have been under-diagnosed owing to the absence of specific clinical and biochemical markers, limitations in liver biopsy and the lack of an effective method of expression and enzyme assay for verifying the mutations found. To date, eight patients have been reported with nine allelic variants of the HMGCS2 gene. We present a new method of enzyme expression and a modification of the activity assay that allows, for first time, the functional study of missense mutations found in patients with this deficiency. Four of the missense mutations (p.V54M, p.R188H, p.G212R and p.G388R) did not produce proteins that could have been detected in soluble form by western blot; three produced a total loss of activity (p.Y167C, p.M307T and p.R500H) and one, variant p.F174L, gave an enzyme with a catalytic efficiency of 11.5%. This indicates that the deficiency may occur with partial loss of activity of enzyme. In addition, we describe a new patient with this deficiency, in which we detected the missense allelic variant, c.1162G>A (p.G388R) and the nonsense variant c.1270C>T (p.R424X).

Cutaneous dyspigmentation in patients with ganglioside GM3 synthase deficiency

Ganglioside GM3 synthase deficiency is a rare autosomal recessive metabolic disorder characterized by infantile onset of severe irritability and epilepsy, failure to thrive, developmental stagnation, and cortical blindness. Because of the lack of easily recognizable dysmorphism and specific neurologic manifestations, identification of patients with this condition is extremely challenging. Here we report on previously undescribed pigmentary abnormalities in 20 of 38 patients with GM3 synthase deficiency. All 20 of the patients showed freckle-like hyperpigmented macules, ranging in size from 2 to 5 mm in diameter and usually found bilaterally on the extremities, especially the dorsal aspects of the hands and feet. Seven of these patients also had depigmented macules and patches, especially on the face and extremities. These cutaneous changes were asymptomatic, and were not associated with the severity or particular phenotype of the neurologic disease. They became visible only after the first years of life with an increased incidence with advancing age. These distinct pigmentary features are not identified in 54 normal siblings, and may provide a useful clue in identifying patients with ganglioside metabolic disorders.

Recurrent Encephalopathy: NAGS (N-Acetylglutamate Synthase) Deficiency in Adults

N-acetyl-glutamate synthase (NAGS) deficiency is a rare autosomal recessive urea cycle disorder (UCD) that uncommonly presents in adulthood. Adult presentations of UCDs include; confusional episodes, neuropsychiatric symptoms and encephalopathy. To date, there have been no detailed neurological descriptions of an adult onset presentation of NAGS deficiency. In this review we examine the clinical presentation and management of UCDs with an emphasis on NAGS deficiency. An illustrative case is provided. Plasma ammonia levels should be measured in all adult patients with unexplained encephalopathy, as treatment can be potentially life-saving. Availability of N-carbamylglutamate (NCG; carglumic acid) has made protein restriction largely unnecessary in treatment regimens currently employed. Genetic counselling remains an essential component of management of NAGS.

Two novel mutations of the CYP11B2 gene in a Japanese patient with aldosterone deficiency type 1

Isolated hypoaldosteronism is a rare and occasionally life-threatening cause of salt wasting in infancy. A 2-month-old Japanese boy of unrelated parents was examined for failure to thrive and poor weight gain. Laboratory findings were hyponatremia, hyperkalemia, high plasma renin and low aldosterone levels. Spot urine analysis by gas chromatography-mass spectrometry (GC-MS) showed that urinary excretion of corticosterone metabolites was elevated. Whereas excretion of 18-hydroxycortricosterone metabolites was within the normal range, excretion of aldosterone metabolites was undetectable. The patient was therefore suspected to have aldosterone synthase deficiency type 1. Sequence analysis of CYP11B2, the gene encoding aldosterone synthase (CYP11B2), showed that the patient was a compound heterozygote for c.168G>A, p.W56X in exon 1 and c.1149C>T, p.R384X in exon 7. p.W56X was inherited from his mother and p.R384X was from his father. Since both alleles contain nonsense mutations, a lack of CYP11B2 activity was speculated to cause his condition. To our knowledge, this is the first Japanese patient in which the molecular basis of aldosterone synthase deficiency type 1 has been clarified. This case also indicates that spot urinary steroid analysis is useful for diagnosis.

A novel homozygous TMEM70 mutation results in congenital cataract and neonatal mitochondrial encephalo-cardiomyopathy

Mutations in the TMEM70 gene are the most common cause of nuclear encoded ATP synthase deficiency resulting in a syndrome characterized by neonatal lactic acidosis, cardiomyopathy, and encephalomyopathy. Here we report on the first Turkish patient who presented after birth with lactic acidemia, severe hpotonia, hypertrophic cardiomyopathy and bilateral congenital cataract. TMEM70 genetic analysis revealed the causative homozygous c.535C>T novel mutation that result in substitution of a highly conserved tyrosine into histidine at position 179. In this report we focused on a detailed description of the clinical features of this syndrome with special emphasis on the typical facial dysmorphic features. Our report underscores TMEM70 deficiency as a pan-ethnic well defined phenotype. In cases with high suspicion sequencing of TMEM70 should be performed even before the traditional invasive muscle biopsy to confirm the diagnosis.

Demographics, diagnosis and treatment of 256 patients with tetrahydrobiopterin deficiency in mainland China: results of a retrospective, multicentre study

National coverage of neonatal screening for hyperphenylalaninaemia (HPA) in China is still low and tests to differentiate causes of HPA are not performed in many centres. This study aimed to describe the demographics, geographic distribution, diagnosis, treatment and clinical outcomes of treatment, including intellectual development, in patients with tetrahydrobiopterin (BH4) deficiency in mainland China. This was a retrospective, multicentre, chart review in patients with BH4 deficiency across mainland China born 1985-2010. Two hundred fifty six patients were included; 59.9% (267/446) of parents were from eastern China. Median (interquartile range) age at diagnosis decreased from 12.0 (5.5, 102.0) months to 2.0 (1.0, 3.5) months in patients born 1985-1999 (n = 28) and 2005-2010 (n = 152), respectively. 6-Pyruvoyl-tetrahydropterin synthase (PTPS) deficiency was the primary cause of BH4 deficiency (96.0%); four hotspot mutations accounted for 76.6% of PTS gene mutations; two novel variants in the QDPR gene were identified. Most patients (83.6%) received treatment with BH4, L-dopa, 5-hydroxytryptophan and/or diet therapy. Target blood Phe concentration was confirmed at 88.9% of visits; median (Q1, Q3) blood Phe concentration was 106.8 (73.0, 120.0) μmol/L during therapy and 117.0 (67.1, 120.0) μmol/L at last visit. Median (Q1, Q3) WISC IQ score was 80.0 (69.0, 90.0) in 33 patients. DQ scores were within normal range (≥85) for 37/59 (62.7%) patients. Physical development indicators were within normal ranges. Treatment-related adverse events, reported in 20/256 (7.8%) patients, were mild-to-moderate in severity. This study provides valuable information on the current and historical situation of BH4 deficiency in mainland China.

Déficit en aldostérone-synthase : 4 observations pédiatriques

Neonatal salt-wasting syndromes are rare but potentially serious conditions. Isolated hypoaldosteronism is an autosomal recessive inherited disorder of terminal aldosterone synthesis, leading to selective aldosterone deficiency. Two different biochemical forms of this disease have been described, called aldosterone synthase deficiency or corticosterone methyl oxydase, types I and II. In type I, there is no aldosterone synthase activity and the 18 hydroxycorticosterone (18 OHB) level is low, whereas in type II, a residual activity of aldosterone synthase persists and 18 OHB is overproduced. We report on four patients with isolated hypoaldosteronism. In 2 of them, who were recently diagnosed with aldosterone synthase deficit, we discuss the symptoms and treatment. The 2 other patients are now adults. We discuss the long-term outcome, the quality of adult life, aldosterone synthase deficits, as well as the pathophysiology and molecular analysis.

Homozygosity for a mutation in the CYP11B2 gene in an infant with congenital corticosterone methyl oxidase deficiency type II

Isolated aldosterone synthase deficiency can be the source of life-threatening salt wasting and failure to thrive in infancy. We studied an infant with failure to thrive and persistent hyponatremia despite oral sodium supplementation. Initial analyses revealed highly elevated plasma renin but normal values of plasma aldosterone. The biochemical diagnosis of corticosterone methyl oxidase deficiency type II was established by multisteroid analysis, revealing a pathognomonic pattern with a highly elevated ratio of 18-OH-corticosterone to aldosterone. This reflects an enzymatic defect in the aldosterone synthase that is responsible for the terminal steps in the aldosterone biosynthesis. Molecular genetic analysis supported the diagnosis revealing homozygosity for a pathogenic c.554C>T (p.T185I) variation in exon 3 of the CYP11B2 gene encoding aldosterone synthase. Homozygosity for two other polymorphic variations c.504C>T (p.F168F) and c.518A>G (p.K173R) were identified as well. Treatment with fludrocortisone resulted in catch-up growth. Discontinuation of treatment at the age of 9 years was later possible without any clinical or biochemical deterioration. Isolated deficiency in aldosterone biosynthesis should be considered in neonates and infants with failure to thrive and salt wasting. Normal levels of plasma aldosterone compared with highly elevated levels of plasma renin indicate an impaired aldosterone biosynthesis and suggest the disorder. Recognition of its existence is important as fludrocortisone replacement therapy effectively normalizes sodium balance and growth.

ENOS Deficiency Predisposes Podocytes to Injury in Diabetes

Endothelial nitric oxide synthase (eNOS) deficiency may contribute to the pathogenesis of diabetic nephropathy in both experimental models and humans, but the underlying mechanism is not fully understood. Here, we studied two common sequelae of endothelial dysfunction in diabetes: glomerular capillary growth and effects on neighboring podocytes. Streptozotocin-induced diabetes increased glomerular capillary volume in both C57BL/6 and eNOS(-/-) mice. Inhibiting the vascular endothelial growth factor receptor attenuated albuminuria in diabetic C57BL/6 mice but not in diabetic eNOS(-/-) mice, even though it inhibited glomerular capillary enlargement in both. In eNOS(-/-) mice, an acute podocytopathy and heavy albuminuria occurred as early as 2 weeks after inducing diabetes, but treatment with either captopril or losartan prevented these effects. In vitro, serum derived from diabetic eNOS(-/-) mice augmented actin filament rearrangement in cultured podocytes. Furthermore, conditioned medium derived from eNOS(-/-) glomerular endothelial cells exposed to both high glucose and angiotensin II activated podocyte RhoA. Taken together, these results suggest that the combined effects of eNOS deficiency and hyperglycemia contribute to podocyte injury, highlighting the importance of communication between endothelial cells and podocytes in diabetes. Identifying mediators of this communication may lead to the future development of therapies targeting endothelial dysfunction in albuminuric individuals with diabetes.

Refractory epilepsy and mitochondrial dysfunction due to GM3 synthase deficiency

We report two children, born from consanguineous parents, who presented with early-onset refractory epilepsy associated with psychomotor delay, failure to thrive, blindness and deafness. Polarographic and spectrophotometric analyses in fibroblasts and liver revealed a respiratory chain (RC) dysfunction. Surprisingly, we identified a homozygous nonsense mutation in the GM3 synthase gene by using exome sequencing. GM3 synthase catalyzes the formation of GM3 ganglioside from lactosylceramide, which is the first step in the synthesis of complex ganglioside species. Mass spectrometry analysis revealed that the complete absence of GM3 ganglioside and its biosynthetic derivatives was associated with an upregulation of the alternative globoside pathway in fibroblasts. The accumulation of Gb3 and Gb4 globosides likely has a role in RC dysfunction and in the decrease of mitochondrial membrane potential leading to apoptosis, which we observed in fibroblasts. We show for the first time that GM3 synthase deficiency, responsible for early-onset epilepsy syndrome, leads to a secondary RC dysfunction. Our study highlights the role of secondary mitochondrial disorders that can interfere with the diagnosis and the evolution of other metabolic diseases.

TMEM70: a mutational hot spot in nuclear ATP synthase deficiency with a pivotal role in complex V biogenesis

Mammalian complex V (F1F0-ATP synthase or ATPase) uses the proton gradient to generate ATP during oxidative phosphorylation and requires several helper proteins, including TMEM70, to form the holoenzyme in a stepwise process in which nuclear DNA is combined with mitochondrial DNA-encoded subunits. We report the clinical and molecular findings in three patients presenting lactic acidosis, 3-methylglutaconic aciduria, and hypertrophic cardiomyopathy. All three showed an isolated defect of fully assembled ATP synthase in association with a "common" (c.317-2A > G) and a new (c.628A > C/p.T210P) variant in TMEM70. Interestingly, one of the patients also showed nitric oxide-responsive pulmonary arterial hypertension, a finding never before associated with TMEM70 deficiency. In addition to widening the clinical and mutational spectrum of defective ATP synthase, our study also suggests that mutant TMEM70 associates in high molecular weight complexes (470-550kDa) when expressed in Hela cells and exerts a direct action in ATP synthase biogenesis and assembly, mediating the incorporation of F1 moieties.

Hypotensions d’origine endocrinienne

Hypotension is defined by a low blood pressure either permanently or only in upright posture (orthostatic hypotension). In contrast to hypertension, there is no threshold defining hypotension. The occurrence of symptoms for systolic and diastolic measurements respectively below 90 and 60 mm Hg establishes the diagnosis. Every acute hypotensive event should suggest shock, adrenal failure or an iatrogenic cause. Chronic hypotension from endocrine origin may be linked to adrenal failure from adrenal or central origin, isolated hypoaldosteronism, pseudohypoaldosteronism, pheochromocytoma, neuro-endocrine tumors (carcinoïd syndrome) or diabetic dysautonomia. Hypotension related to hypoaldosteronism associates low blood sodium and above all high blood potassium levels. They are generally classified according to their primary (hyperreninism) or secondary (hyporeninism) adrenal origin. Isolated primary hypoaldosteronisms are rare in adults (intensive care unit, selective injury of the glomerulosa area) and in children (aldosterone synthase deficiency). Isolated secondary hypoaldosteronism is related to mellitus diabetes complicated with dysautonomia, kidney failure, age, iatrogenic factors, and HIV infections. In both cases, they can be associated to glucocorticoid insufficiency from primary adrenal origin (adrenal failure of various origins with hyperreninism, among which congenital 21 hydroxylase deficiency with salt loss) or from central origin (hypopituitarism with hypo-reninism). Pseudohypoaldosteronisms are linked to congenital (type 1 pseudohypoaldosteronism) or acquired states of resistance to aldosterone. Acquired salt losses from enteric (total colectomy with ileostomy) or renal (interstitial nephropathy, Bartter and Gitelman syndromes…) origin might be responsible for hypotension and are associated with hyperreninism-hyperaldosteronism. Hypotension is a rare manifestation of pheochromocytomas, especially during surgical removal when the patient has not been prepared with calcium inhibitors. Every flush with hypotension should suggest a carcinoid crisis, which is very sensitive to subcutaneous somatostatin analog. An accurate etiological diagnosis should allow treat efficiently endocrine hypotension without inducing hypertension in supine posture.

D.P.16 Metabolic and hemodynamic alterations in the mdx skeletal muscle revisited using multi-parametric functional NMR

The mdx mouse remains a popular model of the Duchenne muscular dystrophy. While criticized for the mild impairment of muscle function and structure, the mdx remains highly relevant in terms of energy and microvascular control disturbances associated to muscle dystrophy. In this study, a non-invasive multi-parametric functional NMR approach (mpf) was used to revisit these anomalies. This NMR approach interleaved arterial spin labeled imaging and 31P spectroscopy to measure perfusion, capillary oxygenation and muscle energetics of the gastrocnemius of C57Bl/10wt and mdx mice (between 10 and 65weeks of age). Hind-limb ischemia (30min) or repeated bouts of exercises (2min) and recovery were investigated. After ischemia, phosphocreatine (PCr) depleted more in 28-week-old mdx vs wt mice (ΔPCr: 82±6% vs 62±10%). Resynthesis of PCr at reactive hyperemia appeared prolonged (TCr: 88.7 ±21.5s vs 53±23.3s). In parallel, vasodilatation lasted less at reactive hyperemia in mdx mice compared to wt mice. Similarly, a shorter reperfusion profile was visible in 65-week old mdx mice following exercise, confirming a default in maintaining vasodilatation in old mdx vs old wt mice but also compared to young mdx mice. Moreover capillary oxygenation at reactive hyperemia showed reduced O2 extraction in mdx mice. In this study, the mpf-NMR protocol proved again its capacity to characterize in vivo functional anomalies, this time in dystrophic muscle. The underlying mechanisms require further investigation, with neuronal NO synthase (nNOS) deficiency most likely playing a direct role in the perfusion profile alterations. The defective nNOS might also contribute indirectly, through PFK inhibition, to the faster PCr depletion during ischemia, which could be indicative of impaired glycolysis. On the other hand, the slower PCr resynthesis and lower oxygen extraction help to appreciate and quantify the real impact in vivo of mitochondrial dysfunction in mdx.

Dyrk1A, a Serine/Threonine Kinase, is Involved in ERK and Akt Activation in the Brain of Hyperhomocysteinemic Mice

Hyperhomocysteinemia due to cystathionine beta synthase (CBS) deficiency is associated with diverse brain disease. Whereas the biological actions linking hyperhomocysteinemia to the cognitive dysfunction are not well understood, we tried to establish relationships between hyperhomocysteinemia and alterations of signaling pathways. In the brain of CBS-deficient mice, a murine model of hyperhomocysteinemia, we previously found an activation of extracellular signal-regulated kinase (ERK) pathway and an increase of Dyrk1A, a serine/threonine kinase involved in diverse functions ranging from development and growth to apoptosis. We then investigated the relationship between Dyrk1A and the signaling pathways initiated by receptor tyrosine kinases (RTK), the ERK and PI3K/Akt pathways. We found a significant increase of phospho-ERK, phospho-MEK, and phospho-Akt in the brain of CBS-deficient and Dyrk1a-overexpressing mice. This increase was abolished when CBS-deficient and Dyrk1A-transgenic mice were treated with harmine, an inhibitor of Dyrk1A kinase activity, which emphasizes the role of Dyrk1A activity on ERK and Akt activation. Sprouty 2 protein level, a negative feedback loop modulator that limits the intensity and duration of RTK activation, is decreased in the brain of CBS-deficient mice, but not in the brain of Dyrk1A transgenic mice. Furthermore, a reduced Dyrk1A and Grb2 binding on sprouty 2 and an increased interaction of Dyrk1A with Grb2 were found in the brain of Dyrk1A transgenic mice. The consequence of Dyrk1A overexpression on RTK activation seems to be a decreased interaction of sprouty 2/Grb2. These observations demonstrate ERK and Akt activation induced by Dyrk1A in the brain of hyperhomocysteinemic mice and open new perspectives to understand the basis of the cognitive defects in hyperhomocysteinemia.

Abstract 39: Organic Nitrates Improve Calcium Handling and Myofilament Sensitivity Impaired by Nitroso-Redox Disequilibrium

Oxidative stress is a key factor in the dysfunctional calcium (Ca 2+ ) handling and contractile performance in heart disease. Organic nitrates are used for the treatment of cardiovascular afflictions, however endogenous nitric oxide (NO) has a broad spectrum of actions and the mechanisms by which it regulates Ca 2+ cycling and contractility is poorly understood. Neuronal NO synthase (nNOS) deficiency induces a nitroso-redox disequilibrium characterized by oxidative stress and altered contractility and calcium handling. We hypothesize that treatment of nNOS −/− mice with organic nitrates such as nitroglycerin (TNG) or isosorbide dinitrate (ISDN) reduces cytosolic calcium levels and restores myofilament responsiveness to calcium with no changes in contractility. Cardiomyocytes (CMs) were isolated from nNOS −/− (N=7) and wild type (WT) mice (N=3). Cells were loaded with fura-2 and then electrically evoked intracellular Ca 2+ and sarcomere length were simultaneously measured in an IonOptix system. It has been shown that L-type Ca 2+ current is exacerbated in nNOS −/− cardiomyocytes. In consequence, at 1 Hz pacing rate, Ca 2+ peak and transient amplitude were increased in nNOS −/− (Peak Ca 2+ : 571 ± 38 nM) compared to WT (401 ± 32 nM; p = 0.012) cardiomyocytes, which was reduced toward normal by 10 nmol/L TNG (453 ± 55 nM; p = 0.11) as well as 10 nmol/L ISDN (394 ± 65; p = 0.035). Phospholamban phosphorylation has been shown to be reduced in nNOS −/− CMs, possibly due to the enhanced activity of protein phosphatases induced by oxidative stress. Thus, Ca 2+ decay and sarcomere relaxation were slower in nNOS −/− compared to WT CMs ( p < 0.0001). Treatment with TNG ( p < 0.0001 vs. NOS1 −/− ) or ISDN ( p < 0.0001 vs. NOS1 −/− ) accelerated Ca 2+ decay and relaxation. Ca 2+ sensitivity was impaired in nNOS −/− ( p < 0.021 vs. WT). Although organic nitrates either reduce or do not affect myofilament sensitivity in normal myocytes, TNG and ISDN increased their responsiveness to Ca 2+ in nNOS −/− . In conclusion, restoration of NO availability and subsequent attenuation of the nitroso-redox imbalance, improved excitation-contraction coupling in nNOS −/− CMs, decreased intracellular Ca 2+ which is counteracted by the improved myofilament sensitivity resulting in no net change in contractility.

Long-term follow-up of tetrahydrobiopterin therapy in patients with tetrahydrobiopterin deficiency in Japan

Tetrahydrobiopterin (BH4) deficiency is a rare, congenital and lethal condition resulting in phenylalanine build-up that can lead to mental retardation and developmental defects, unless properly treated. About 1 million newborn infants in Japan undergo neonatal PKU screening every year, of which about 1 in 2 million are diagnosed with the condition. In this post-marketing surveillance study, 19 patients with BH4 deficiency in whom BH4 supplementation with sapropterin dihydrochloride (Biopten®) (hereafter referred to as ‘BH4 therapy’) was initiated before the age of 4years, were followed up for ⩽28years. Patients who screened positive for BH4 deficiency were treated with supplemental BH4 plus L-dopa and 5-hydroxytryptophan. Data on the patients’ clinical courses were collected once yearly at 10 medical centers in Japan. Seventeen patients were diagnosed with 6-pyruvoyl tetrahydropterin synthase deficiency and two with dihydropteridine reductase deficiency at an average age of 3.6months; the mean age at end of follow-up was 14.6years. Average duration of BH4 therapy (mean dose, 5mg/kg per day) was 13.2years. Serum phenylalanine was reduced from more than 10mg/dL at the start of drug administration to less than 2mg/dL at end of follow-up. No abnormalities in height or weight were observed in any patients, except for one female patient with familial obesity. No unwarranted side effects were reported throughout the long-term course of treatment, even during pregnancy. BH4 therapy can effectively maintain serum phenylalanine levels within the normal range in patients with BH4 deficiency, and demonstrated excellent long-term safety, with no side effects.

Novel CYP11B2 mutation causing aldosterone synthase (P450c11AS) deficiency

Aldosterone synthase (P450c11AS) deficiency is a rare autosomal recessive disorder, presenting with severe salt-losing in early infancy. It is caused by inactivating mutations of the CYP11B2 gene. Here, we describe three unrelated Asian patients who have clinical and hormonal features compatible with aldosterone synthase deficiency and identify their CYP11B2 mutations. Patient 1 was a Thai female infant. Patient 2 was an Indian boy, and patient 3 was a Thai male infant. All subjects presented at the age of 1-2 months with diarrhea, failure to thrive, and severe dehydration. Their plasma electrolytes showed hyponatremia, hyperkalemia, and acidosis. All patients had normal cortisol response and had elevated plasma renin activity with low aldosterone levels. The entire coding regions of the CYP11B2 gene were amplified by polymerase chain reaction and sequenced. Patient 1 was homozygous for a previously described mutation, p.T318M. Patient 2 was homozygous for a novel c.666delC mutation inherited from both parents resulting in p.223F>Sfsx295. No CYP11B2 mutation was detected in patient 3. We report the first CYP11B2 defects in Southeast Asian families responsible for aldosterone synthase deficiency and identified a novel CYP11B2 mutation. However, the affected gene(s) responsible for primary hypoaldosteronism other than CYP11B2 remain to be determined.

Course of psychiatric disorders in a case of 6-Pyruvoyl-tetrahydropterin synthase deficiency

Course of psychiatric disorders in a case of 6-Pyruvoyl-tetrahydropterin synthase deficiency