Clinical and biochemical outcomes in cobalamin C deficiency with use of high-dose hydroxocobalamin in the early neonatal period.

This case report describes a patient with early-onset cobalamin C deficiency who was started on treatment with high-dose parenteral hydroxocobalamin after diagnosis at 13 days of life. Prior to diagnosis, initial presenting symptoms included poor feeding, lethargy, apneic episodes, hypothermia, and hypotonia; these symptoms resolved after initiation of medication. Methylmalonic acid and homocysteine levels were trended and significantly improved with treatment. She was maintained on 2 mg/kg/day dosing of hydroxocobalamin. No adverse effects to treatment were observed. At the time of this report, the patient was 19 months of age; she had not manifested common findings of early-onset cobalamin C deficiency, including microcephaly, poor feeding, growth abnormalities, hypotonia, seizures, maculopathy, or neurodevelopmental delay. This report suggests that early initiation of high-dose hydroxocobalamin is safe and effective.

Protective effects of L-carnitine on behavioral alterations and neuroinflammation in striatum of glutaryl-COA dehydrogenase deficient mice.

Glutaric acidemia type 1 (GA1) is caused by glutaryl-CoA dehydrogenase deficiency that leads to a blockage in the metabolic route of the amino acids lysine and tryptophan and subsequent accumulation of glutaric acid (GA), 3-hydroxyglutaric acids and glutarylcarnitine (C5DC). Patients predominantly manifest neurological symptoms, associated with acute striatal degeneration, as well as progressive cortical and striatum injury whose pathogenesis is not yet fully established. Current treatment includes protein/lysine restriction and l-carnitine supplementation of (L-car). The aim of this work was to evaluate behavior parameters and pro-inflammatory factors (cytokines IL-1ß, TNF-α and cathepsin-D levels), as well as the anti-inflammatory cytokine IL10 in striatum of knockout mice (Gcdh-/-) and wild type (WT) mice submitted to a normal or a high Lys diet. The potential protective effects of L-car treatment on these parameters were also evaluated. Gcdh-/- mice showed behavioral changes, including lower motor activity (decreased number of crossings) and exploratory activity (reduced number of rearings). Also, Gcdh-/- mice had significantly higher concentrations of glutarylcarnitine (C5DC) in blood and cathepsin-D (CATD), interleukin IL-1ß and tumor factor necrosis alpha (TNF-α) in striatum than WT mice. Noteworthy, L-car treatment prevented most behavioral alterations, normalized CATD levels and attenuated IL-1ß levels in striatum of Gcdh-/- mice. Finally, IL-1ß was positively correlated with CATD and C5DC levels and L-car was negatively correlated with CATD. Our results demonstrate behavioral changes and a pro-inflammatory status in striatum of the animal model of GA1 and, most importantly, L-car showed important protective effects on these alterations.

Identification of 2,2-Dimethylbutanoic Acid (HST5040), a Clinical Development Candidate for the Treatment of Propionic Acidemia and Methylmalonic Acidemia.

Propionic acidemia (PA) and methylmalonic acidemia (MMA) are rare autosomal recessive disorders of propionyl-CoA (P-CoA) catabolism, caused by a deficiency in the enzymes P-CoA carboxylase and methylmalonyl-CoA (M-CoA) mutase, respectively. PA and MMA are classified as intoxication-type inborn errors of metabolism because the intramitochondrial accumulation of P-CoA, M-CoA, and other metabolites results in secondary inhibition of multiple pathways of intermediary metabolism, leading to organ dysfunction and failure. Herein, we describe the structure-activity relationships of a series of short-chain carboxylic acids which reduce disease-related metabolites in PA and MMA primary hepatocyte disease models. These studies culminated in the identification of 2,2-dimethylbutanoic acid (10, HST5040) as a clinical candidate for the treatment of PA and MMA. Additionally, we describe the in vitro and in vivo absorption, distribution, metabolism, and excretion profile of HST5040, data from preclinical studies, and the synthesis of the sodium salt of HST5040 for clinical trials.

Dietary Treatment from Birth to Pregnancy in a Woman with Methylmalonic Aciduria.

Methylmalonic aciduria is treated with a natural protein-restricted diet with adequate energy intake to sustain metabolic balance. Natural protein is a source of methylmalonic acid precursors, and intake is individually modified according to the severity and clinical course of the disease. The experience and approach to MMA treatment in European centers is variable with different amounts of natural protein and precursor-free l-amino acids being prescribed, although the outcome appears independent of the use of precursor-free l-amino acids. Further long-term outcome data is necessary for early treated patients with MMA. This case study, a woman with MMA followed from birth to the age of 35 years, including pregnancy, illustrates the long-term course of the disease and lifetime changes in dietary treatment. A low natural protein diet (1.5 g-1.0 g/kg/day) was the foundation of treatment, but temporary supplementation with precursor-free l-amino acids, vitamin-mineral mixture, and energy supplements were necessary at different timepoints (in childhood, adolescence, adulthood and pregnancy). Childhood psychomotor development was slightly delayed but within the normal range in adulthood. There were few episodes of metabolic decompensation requiring IV glucose, but at age 27 years, she required intensive care following steroid treatment. In pregnancy, she remained stable but received intensive biochemical and medical follow-up. This successful long-term follow-up of a patient with MMA from childhood, throughout pregnancy, delivery, and postpartum confirms that careful clinical, biochemical, and dietetic monitoring is crucial to ensure a favourable outcomes in MMA. Personalized treatment is necessary according to the individual clinical course. Knowledge about long-term treatment and clinical outcome is important information to influence future MMA clinical guidelines.

Analysis of 70 patients with hydrocephalus due to cobalamin C deficiency.

OBJECTIVE: To analyze the clinical characteristics of patients with hydrocephalus secondary to cobalamin C (cblC) deficiency and to discuss the optimal strategies for assessing and treating such patients by performing clinical and laboratory studies in 70 patients. METHODS: A total of 1,211 patients were clinically diagnosed with methylmalonic acidemia (MMA) from 1998 to 2019. Among them, cblC deficiency was confirmed in 70 patients with hydrocephalus by brain imaging and biochemical and genetic analysis. RESULTS: Of the 70 patients, 67 (95.7%) had early-onset MMA and homocystinuria. The patients typically had high blood propionylcarnitine and total homocysteine, low methionine, and methylmalonic aciduria. Signs of intracranial hypertension were relatively rare. We measured ventricular dilatation early in the disease by cranial ultrasound and MRI and/or CT. Eighteen different MMACHC mutations, including 4 novel mutations (c.427C>T, c.568insT, c.599G>A, and c.615C>A), were identified biallelically in all 70 patients. c.609G>A was the most frequent mutation, followed by c.658_660del, c.217C>T, and c.567dupT. Three cases were diagnosed by postmortem study. Metabolic therapy, including cobalamin injections supplemented with oral l-carnitine and betaine, was administered in the remaining 67 cases. A ventriculoperitoneal shunt was performed in 36 cases. During the follow-up, psychomotor development, nystagmus, impaired vision, and sunset eyes improved gradually. CONCLUSION: Hydrocephalus is a severe condition with several different causes. In this study, ventriculomegaly was found in 70 patients with cblC deficiency. Early diagnosis, etiologic treatment, and prompt surgical intervention are crucial to improve the prognosis of patients.

Biomedical applications of microbial phenylalanine ammonia lyase: Current status and future prospects.

Phenylalanine ammonia lyase (PAL) has recently emerged as an important therapeutic enzyme with several biomedical applications. The enzyme catabolizes l-phenylalanine to trans-cinnamate and ammonia. PAL is widely distributed in higher plants, some algae, ferns, and microorganisms, but absent in animals. Although microbial PAL has been extensively exploited in the past for producing industrially important metabolites, its high substrate specificity and catalytic efficacy lately spurred interest in its biomedical applications. PEG-PAL drug named Palynziq™, isolated from Anabaena variabilis has been recently approved for the treatment of adult phenylketonuria (PKU) patients. Further, it has exhibited high potency in regressing tumors and treating tyrosine related metabolic abnormalities like tyrosinemia. Several therapeutically valuable metabolites have been biosynthesized via its catalytic action including dietary supplements, antimicrobial peptides, aspartame, amino-acids, and their derivatives. This review focuses on all the prospective biomedical applications of PAL. It also provides an overview of the structure, production parameters, and various strategies to improve the therapeutic potential of this enzyme. Engineered PAL with improved pharmacodynamic and pharmacokinetic properties will further establish this enzyme as a highly efficient biological drug.

Inducible Slc7a7 Knockout Mouse Model Recapitulates Lysinuric Protein Intolerance Disease.

Lysinuric protein intolerance (LPI) is a rare autosomal disease caused by defective cationic amino acid (CAA) transport due to mutations in SLC7A7, which encodes for the y+LAT1 transporter. LPI patients suffer from a wide variety of symptoms, which range from failure to thrive, hyperammonemia, and nephropathy to pulmonar alveolar proteinosis (PAP), a potentially life-threatening complication. Hyperammonemia is currently prevented by citrulline supplementation. However, the full impact of this treatment is not completely understood. In contrast, there is no defined therapy for the multiple reported complications of LPI, including PAP, for which bronchoalveolar lavages do not prevent progression of the disease. The lack of a viable LPI model prompted us to generate a tamoxifen-inducible Slc7a7 knockout mouse (Slc7a7-/-). The Slc7a7-/- model resembles the human LPI phenotype, including malabsorption and impaired reabsorption of CAA, hypoargininemia and hyperammonemia. Interestingly, the Slc7a7-/- mice also develops PAP and neurological impairment. We observed that citrulline treatment improves the metabolic derangement and survival. On the basis of our findings, the Slc7a7-/- model emerges as a promising tool to further study the complexity of LPI, including its immune-like complications, and to design evidence-based therapies to halt its progression.

Prevention by L-carnitine of DNA damage induced by 3-hydroxy-3-methylglutaric and 3-methylglutaric acids and experimental evidence of lipid and DNA damage in patients with 3-hydroxy-3-methylglutaric aciduria.

3-hydroxy-3-methylglutaric aciduria (HMGA) is an inherited disorder of the leucine catabolic pathway in which occurs a deficiency of the 3-hydroxy-3-methylglutaryl-CoA lyase enzyme. Therefore, the organic acids 3-hydroxy-3-methylglutaric (HMG) and 3-methylglutaric (MGA), mainly, accumulate in tissues of affected patients. Lately, much attention has been focused on free radicals as mediators of tissue damage in human diseases, causing lipid peroxidation, protein oxidation and DNA damage. The treatment of this disease is based in a restricted protein ingest and supplementation with l-carnitine (LC), an antioxidant and detoxifying agent. In the present work, we investigated the in vitro oxidative damage to DNA induced by the accumulation of organic acids and oxidative stress parameters in vivo of patients with 3-HMG, as well as the effect of the recommended therapy. The in vitro DNA damage was analyzed by the alkaline comet assay in leukocytes incubated with HMG and MGA (1 mM, 2.5 mM and 5 mM) and co-incubated with LC (90 µM and 150 µM). The in vivo urinary 15-F2t-isoprostane levels and urinary oxidized guanine species were measured by ELISA kits in patient's urine before and after the treatment with LC. HMG and MGA induced a DNA damage index (DI) significantly higher than that of the control group. The DI was significantly reduced in the presence of LC. It was also verified a significant increase of oxidized guanine species and urinary isoprostane levels, biomarker of oxidative DNA damage and lipid peroxidation respectively, in patients before treatment. After the treatment and supplementation with LC, patients presented significantly lower levels of those biomarkers. Analyzing the data together, we can conclude that HMGA patients present oxidative lipid and DNA damage, which is induced by HMG and MGA, and the antioxidant therapy with LC can prevent that kind of injuries.

The Role of Oxidative Stress and Bioenergetic Dysfunction in Sulfite Oxidase Deficiency: Insights from Animal Models.

Sulfite oxidase (SO) deficiency is an autosomal recessive inherited neurometabolic disease caused by deficient activity of SO. It is biochemically characterized by tissue accumulation and high urinary excretion of sulfite, thiosulfate, and S-sulfocysteine. Severe neurological symptoms, including neonatal seizures, encephalopathy, and psychomotor retardation, are commonly observed in the affected patients, but the pathogenesis of the neurologic dysfunction is still poorly understood. In this minireview, we will briefly summarize the knowledge obtained from in vivo and in vitro findings from animal studies indicating that oxidative stress and mitochondrial dysfunction are involved in the pathophysiology of the brain damage in this disease. Recent reports have shown that sulfite induces free radical generation, impairs brain antioxidant defenses, and disturbs mitochondrial energy metabolism and biogenesis. Moreover, it has been evidenced that free radical scavengers and the pan-PPAR agonist bezafibrate are able to prevent most deleterious effects elicited by sulfite on the brain. These promising data offer new perspectives for potential therapeutic strategies for this condition, which may include the early use of appropriate antioxidants and PPAR agonists in addition to the available treatment.

Fifteen-year follow-up of Italian families affected by arginine glycine amidinotransferase deficiency.

BACKGROUND: Arginine:glycine amidinotransferase deficiency (AGAT-d) is a very rare inborn error of creatine synthesis mainly characterized by absence of brain Creatine (Cr) peak, intellectual disability, severe language impairment and behavioural disorder and susceptible to supplementary Cr treatment per os. Serial examinations by magnetic resonance spectroscopy are required to evaluate Cr recovery in brain during treatment of high doses of Cr per os, which have been proved beneficial and effective in treating main clinical symptoms. A long term study with detailed reports on clinical, neurochemical and neuropsychological outcomes of the first Italian patients affected by AGAT-d here reported can represent a landmark in management of this disorder thus enhancing medical knowledge and clinical practice. RESULTS: We have evaluated the long term effects of Cr supplementation management in four Italian patients affected by AGAT-d, correlating specific treatments with serial clinical, biochemical and magnetic resonance spectroscopy examinations as well as the neuropsychological outcome by standardized developmental scales. Consecutive MRS examinations have confirmed that Cr depletion in AGAT-d patients is reversible under Cr supplementation. Cr treatment is considered safe and well tolerated but side effects, including weight gain and kidney stones, have been reported. CONCLUSIONS: Early treatment prevents adverse developmental outcome, while patients diagnosed and treated at an older age showed partial but significant cognitive recovery with clear improvements in adaptive functioning.

Proposed recommendations for diagnosing and managing individuals with glutaric aciduria type I: second revision.

Glutaric aciduria type I (GA-I; synonym, glutaric acidemia type I) is a rare inherited metabolic disease caused by deficiency of glutaryl-CoA dehydrogenase located in the catabolic pathways of L-lysine, L-hydroxylysine, and L-tryptophan. The enzymatic defect results in elevated concentrations of glutaric acid, 3-hydroxyglutaric acid, glutaconic acid, and glutaryl carnitine in body tissues, which can be reliably detected by gas chromatography/mass spectrometry (organic acids) and tandem mass spectrometry (acylcarnitines). Most untreated individuals with GA-I experience acute encephalopathic crises during the first 6 years of life that are triggered by infectious diseases, febrile reaction to vaccinations, and surgery. These crises result in striatal injury and consequent dystonic movement disorder; thus, significant mortality and morbidity results. In some patients, neurologic disease may also develop without clinically apparent crises at any age. Neonatal screening for GA-I us being used in a growing number of countries worldwide and is cost effective. Metabolic treatment, consisting of low lysine diet, carnitine supplementation, and intensified emergency treatment during catabolism, is effective treatment and improves neurologic outcome in those individuals diagnosed early; treatment after symptom onset, however, is less effective. Dietary treatment is relaxed after age 6 years and should be supervised by specialized metabolic centers. The major aim of this second revision of proposed recommendations is to re-evaluate the previous recommendations (Kölker et al. J Inherit Metab Dis 30:5-22, 2007b; J Inherit Metab Dis 34:677-694, 2011) and add new research findings, relevant clinical aspects, and the perspective of affected individuals.

Eye Findings on Vigabatrin and Taurine Treatment in Two Patients with Succinic Semialdehyde Dehydrogenase Deficiency.

We describe for the first time two patients with succinic semialdehyde dehydrogenase (SSADH) deficiency, who were found to have abnormal electroretinogram (ERG) examinations at baseline, or 6 months after vigabatrin treatment was started. This was somewhat reversible with L-taurine treatment, or minimally progressive. The mechanism of injury to the retina may be induced by elevations of γ-aminobutyric acid causing peripheral photoreceptor and ganglion cell damage, and this can be exacerbated by the use of vigabatrin. The use of taurine supplementation in tandem with vigabatrin may allow reversal of retinopathy and mitigate or slow down further deterioration. Further prospective clinical trials are required to evaluate this further. We recommend starting L-taurine therapy together with vigabatrin if a trial of vigabatrin is commenced in a patient with SSADH deficiency. Close monitoring of visual fields or ERG is also recommended at baseline and during vigabatrin therapy.

Novel c.2216T > C (p.I739T) mutation in exon 13 and c.1481T > A (p.L494X) mutation in exon 8 of MUT gene in a female with methylmalonic acidemia.

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

S-adenosylmethionine treatment in methionine adenosyltransferase deficiency, a case report.

Reported is a female patient with methionine adenosyltransferase I/III (MAT I/III) deficiency, who was found to have pronounced hypermethioninemia on newborn mass spectroscopy screening, and had two compound heterozygous missense mutations in the gene encoding human MAT1A protein. Hypermethioninemia persisted and her mental development was deficient. At 4 years and 8 months, we started with the supplementary treatment of S-adenosylmethionine, the metabolic product of methionine catalyzed by MAT, which was effective in her neurological development.

Diagnosis, treatment and outcome of glutaric aciduria type I in Zhejiang Province, China.

BACKGROUND: Glutaric aciduria type I (GA I; MIM 231670) is a rare autosomal recessive disorder resulting from glutaryl-CoA dehydrogenase deficiency. This article reports our experience in the diagnosis, treatment and outcome of GA I patients in Zhejiang Province, China. MATERIAL/METHODS: A total of 129,415 newborns (accounting for approximately one-tenth of the annual births in Zhejiang Province) and 9640 high-risk infants were screened for inborn errors of metabolism in the Neonatal Screening Center of Zhejiang Province during a 3-year period. Tandem mass spectrometry and gas chromatography-mass spectrometry were used for diagnosis of the patients. Dietary modification, carnitine supplementation and aggressive treatment of intercurrent illnesses were adapted for GA I patients. RESULTS: Three infants were diagnosed with GA I by high-risk screening (detection rate: 1/3,213) and 2 were diagnosed by newborn screening (incidence: 1/64,708). Four patients (3 by high-risk screening and 1 by neonatal screening) undergoing MRI examination showed remarkable changes on T2-weighted image. Four patients accepted timely treatment, and in the patient diagnosed by neonatal screening, treatment was delayed until hypotonia appeared 3 months later. Neuropsychological assessment showed mental and motor retardation in 3 patients after treatment, including the patient diagnosed by neonatal screening. CONCLUSIONS: Individualized timely treatment and close monitoring of GA I patients needs to be optimized in China. Appropriate communication with parents may help to achieve successful management of GA I patients.

l-arginine:glycine amidinotransferase (AGAT) deficiency: clinical presentation and response to treatment in two patients with a novel mutation.

Creatine and creatine phosphate provide storage and transmission of phosphate-bound energy in muscle and brain. Of the three inborn errors of creatine metabolism causing brain creatine depletion, l-arginine:glycine amidinotransferase (AGAT) deficiency has been described in only two families. We describe clinical and biochemical features, magnetic resonance spectroscopy (MRS) findings and response to creatine supplementation in two siblings with a novel mutation in the AGAT-encoding GATM gene. The sister and brother were evaluated at age 12 and 18years, respectively, because of mild mental retardation, muscle weakness and low weight. Extensive work-up had previously yielded negative results. Electron microscopy of the muscle revealed tubular aggregates and the activity of respiratory chain complexes was decreased in the muscle. Urine organic acid concentrations normalized to urine creatinine concentration were all increased, suggesting a creatine metabolism disorder. Brain MRS was remarkable for absence of creatine. Urine guanidinoacetate levels by tandem mass spectrometry were low, suggesting AGAT deficiency. GATM sequencing revealed a homozygous single nucleotide insertion 1111_1112insA, producing a frame-shift at Met-371 and premature termination at codon 376. Eleven months after commencing treatment with oral creatine monohydrate 100mg/kg/day, repeat MRI/MRS showed significantly increased brain creatine in the sister and a slight increase in the older brother. The parents' impression of improved strength and stamina was substantiated by increased post-treatment versus pre-treatment scores in the Vineland Adaptive Behavior Scale, straight-arm raising and timed up-and-go tests. Similarly, there was an apparent improvement in cognitive function, with significantly increased IQ-scores in the sister and marginal improvement in the brother.

Thermolability of mutant MMACHC protein in the vitamin B12-responsive cblC disorder.

Methylmalonic aciduria and homocystinuria, cblC type, is the most common inborn error of cellular vitamin B12 metabolism. We previously showed that the protein carrying the mutation responsible for late-onset cblC (MMACHC-R161Q), treatable with high dose OHCbl, is able to bind OHCbl with wild-type affinity, leaving undetermined the disease mechanism involved [Froese et al., Mechanism of responsiveness, Mol. Genet. Metab. (2009).]. To assess whether the mutation renders the protein unstable, we investigated the thermostability of the wild-type and mutant MMACHC proteins, either unbound or bound to different cobalamins (Cbl), using differential scanning fluorimetry. We found that MMACHC-wt and MMACHC-R161Q are both very thermolabile proteins in their apo forms, with melting temperatures (T(m)) of 39.3+/-1.0 and 37.1+/-0.7 degrees C, respectively; a difference confirmed by unfolding of MMACHC-R161Q but not MMACHC-wt by isothermal denaturation at 35 degrees C over 120 min. However, with the addition of OHCbl, MMACHC-wt becomes significantly stabilized (Delta T(m max)=8 degrees C, half-maximal effective ligand concentration, AC(50)=3 microM). We surveyed the effect of different cobalamins on the stabilization of the wild-type protein and found that AdoCbl was the most stabilizing, exerting a maximum increase in T(m) of approximately 16 degrees C, followed by MeCbl at approximately 13 degrees C, each evaluated at 50 microM cofactor. The other cobalamins stabilized in the order (CN)(2)Cbi>OHCbl>CNCbl. Interestingly, the AC(50)'s for AdoCbl, MeCbl, (CN)(2)Cbi and OHCbl were similar and ranged from 1-3 microM, which compares well with the K(d) of 6 microM for OHCbl [Froese et al., Mechanism of responsiveness, Mol. Genet. Metab. (2009).]. Unlike MMACHC-wt, the mutant protein MMACHC-R161Q is only moderately stabilized by OHCbl (Delta T(m max)=4 degrees C). The dose-response curve also shows a lower effectivity of OHCbl with respect to stabilization, with an AC(50) of 7 microM. MMACHC-R161Q showed the same order of stabilization as MMACHC-wt, but each cobalamin stabilized this mutant protein less than its wild-type counterpart. Additionally, MMACHC-R161Q had a higher AC(50) for each cobalamin form compared to MMACHC-wt. Finally, we show that MMACHC-R161Q is able to support the base-off transition for AdoCbl and CNCbl, indicating this mutant is not blocked in that respect. Taken together, our results suggest that protein stability, as well as propensity for ligand-induced stabilization, contributes to the disease mechanism in late-onset cblC disorder. Our results underscore the importance of cofactor stabilization of MMACHC and suggest that even small increases in the concentration of cobalamin complexed with MMACHC may have therapeutic benefit in children with the late-onset, vitamin responsive cblC disease.

Reduction of lipid and protein damage in patients with disorders of propionate metabolism under treatment: a possible protective role of L-carnitine supplementation.

Disorders of propionate metabolism are autosomal recessive diseases clinically characterized by acute metabolic crises in the neonatal period and long-term neurological deficits whose pathophysiology is not completely established. There are increasing evidences demonstrating antioxidant properties for L-carnitine, which is used in the treatment of propionic and methylmalonic acidemias to increase the excretion of organic acids accumulated in tissues and biological fluids of the affected patients. In this work we aimed to evaluate lipid (malondialdehyde content) and protein (carbonyl formation and sulfhydryl oxidation) oxidative damage in plasma from patients with propionic and methylmalonic acidemias at the moment of diagnosis and during treatment with L-carnitine. We also correlated the parameters of oxidative damage with plasma total, free and esterified L-carnitine levels. We found a significant increase of malondialdehyde and carbonyl groups, as well as a reduction of sulfhydryl groups in plasma of these patients at diagnosis compared to controls. Furthermore, patients under treatment presented a marked reduction of the content of protein carbonyl groups, similar to controls, and malondialdehyde content in relation to patients at diagnosis. In addition, plasma total and free L-carnitine concentrations were negatively correlated with malondialdehyde levels. Taken together, the present data indicate that treatment significantly reduces oxidative damage in patients affected by disorders of propionate metabolism and that l-carnitine supplementation may be involved in this protection.

New indications and controversies in arginine therapy.

Arginine is an important, versatile and a conditionally essential amino acid. Besides serving as a building block for tissue proteins, arginine plays a critical role in ammonia detoxification, and nitric oxide and creatine production. Arginine supplementation is an essential component for the treatment of urea cycle defects but recently some reservations have been raised with regards to the doses used in the treatment regimens of these disorders. In recent years, arginine supplementation or restriction has been proposed and trialled in several disorders, including vascular diseases and asthma, mitochondrial encephalopathy lactic acidosis and stroke-like episodes (MELAS), glutaric aciduria type I and disorders of creatine metabolism, both production and transportation into the central nervous system. Herein we present new therapeutic indications and controversies surrounding arginine supplementation or deprivation.

Diagnostic work-up and management of patients with isolated methylmalonic acidurias in European metabolic centres.

The long-term outcome of patients with methylmalonic aciduria (MMA) is still uncertain due to a high frequency of complications such as chronic renal failure and metabolic stroke. The understanding of this disease is hampered by a huge variation in the management of these patients. The major aim of this study was to evaluate the current practice in different European metabolic centres. A standardized questionnaire was sent to 20 metabolic centres asking for standard procedures for confirmation of diagnosis, testing cobalamin responsiveness, dietary treatment, pharmacotherapy, and biochemical and clinical monitoring. Sixteen of 20 metabolic centres (80%) returned questionnaires on 183 patients: 89 of the patients were classified as mut(0), 36 as mut(-), 13 as cblA, 7 as cblB, and 38 as cblA/B. (1) Confirmation of diagnosis: All centres investigate enzyme activity by propionate fixation in fibroblasts; six centres also perform mutation analysis. (2) Cobalamin response: Ten centres follow standardized protocols showing large variations. A reliable exclusion of nonspecific effects has not yet been achieved by these protocols. (3) Long-term treatment: In cobalamin-responsive patients, most centres use hydroxocobalamin (1-14 mg/week i.m. or 5-20 mg/week orally), while two centres use cyanocobalamin. All cobalamin-nonresponsive patients and most cobalamin-responsive patients are supplemented with L: -carnitine (50-100 mg/kg per day). Fourteen centres use intestinal decontamination by antibiotic therapy. Most centres follow D-A-CH (n = 6) or Dewey (n = 4) recommendations for protein requirements. Fourteen centres regularly use precursor-free amino acid supplements. Standardized monitoring protocols are available in seven centres, again showing high variability.