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Mucopolysaccharidosis type I (MPS I), a lysosomal storage disease caused by a deficiency of α-L-iduronidase, leads to storage of the glycosaminoglycans, dermatan sulfate and heparan sulfate. Available therapies include enzyme replacement and hematopoietic stem cell transplantation. In the last two decades, newborn screening (NBS) has focused on early identification of the disorder, allowing early intervention and avoiding irreversible manifestations. Techniques developed and optimized for MPS I NBS include tandem mass-spectrometry, digital microfluidics, and glycosaminoglycan quantification. Several pilot studies have been conducted and screening programs have been implemented worldwide. NBS for MPS I has been established in Taiwan, the United States, Brazil, Mexico, and several European countries. All these programs measure α-L-iduronidase enzyme activity in dried blood spots, although there are differences in the analytical strategies employed. Screening algorithms based on published studies are discussed. However, some limitations remain: one is the high rate of false-positive results due to frequent pseudodeficiency alleles, which has been partially solved using post-analytical tools and second-tier tests; another involves the management of infants with late-onset forms or variants of uncertain significance. Nonetheless, the risk-benefit ratio is favorable. Furthermore, long-term follow-up of patients detected by neonatal screening will improve our knowledge of the natural history of the disease and inform better management.

Classic infantile Pompe disease is characterized by a severe phenotype with cardiomyopathy and hypotonia. Cardiomyopathy is generally hypertrophic and rapidly regresses after enzyme replacement therapy. In this report, for the first time, we describe a patient with infantile Pompe disease and hypertrophic cardiomyopathy that evolved into non-compaction myocardium after treatment. The male newborn had suffered since birth with hypertrophic cardiomyopathy and heart failure. He was treated with standard enzyme replacement therapy (ERT) (alglucosidase alfa) and several immunomodulation cycles due to the development of anti-ERT antibodies, without resolution of the hypertrophic cardiomyopathy. At the age of 2.5 years, he was treated with a new combination of ERT therapy (cipaglucosidase alfa) and a chaperone (miglustat) for compassionate use. After 1 year, the cardiac hypertrophy was resolved, but it evolved into non-compaction myocardium. Non-compaction cardiomyopathy is often considered to be a congenital, primitive cardiomyopathy, due to an arrest of compaction of the myocardium wall during the embryonal development. Several genetic causes have been identified. We first describe cardiac remodeling from hypertrophic cardiomyopathy to a non-compaction form in a patient with infantile Pompe disease treated with a new ERT. This has important implications both for the monitoring of Pompe disease patients and for the understanding of the pathophysiological basis of non-compaction myocardium.

Lysosomal storage disorders (LSDs) are a group of clinically heterogeneous disorders affecting the function of lysosomes and are characterized by an accumulation of undigested substrates within several cell types. In recent years there have been substantial advances in supportive care and drug treatment for some LSDs, leading to improved patient survival, as seen in Gaucher, Pompe and Fabry disease and some Mucopolysaccharidoses; however, many symptoms still persist. Thus it is now even more important to improve patients' quality of life and reduce symptoms and comorbidities. One potential way of achieving this goal is through adjunct nutritional therapy, which is challenging as patients may be overweight with associated consequences, or malnourished, or underweight. Furthermore, drugs used to treat LSDs can modify the metabolic status and needs of patients. There are currently not enough data to make specific dietary recommendations for individual LSDs; however, suggestions can be made for managing clinical manifestations of the diseases, as well as treatment-associated adverse events. The metabolic and nutritional status of adult patients must be regularly assessed and individualized dietary plans may be created to cater to a patient's specific needs. Damage to the autophagic process is a common feature in LSDs that is potentially sensitive to dietary manipulation and needs to be assessed in clinical studies.

Several disorders should be considered in the case of newborns and infants experiencing acute or recurrent symptoms after food ingestion. Immune-mediated adverse food reactions are the most frequent and always to be considered. Nevertheless, in the extensive differential diagnosis, clinicians should also include inherited metabolic disorders (IMDs). This review reports clinical features and diagnostic aspects of the most common IMDs that may present with acute manifestations triggered by food intake. Major focus will be amino acid and protein metabolism defects and carbohydrate disorders. Nowadays, for many of these disorders the risk of an acute presentation triggered by food has been decreased by the introduction of expanded newborn screening (NBS). Nevertheless, clinical suspicion remains essential because some IMDs do not have still reliable markers for NBS and a false negative screening result may occur. The aim of this review is to help pediatricians to take these rare inherited disorders into account in the differential diagnosis of acute or recurrent gastrointestinal symptoms related to food intake, which may avoid delayed diagnosis and potentially life-threatening consequences.

Aicardi-Goutieres syndrome is an inflammatory disease occurring due to mutations in any of TREX1, RNASEH2A, RNASEH2B, RNASEH2C, SAMHD1, ADAR or IFIH1. We report on 374 patients from 299 families with mutations in these seven genes. Most patients conformed to one of two fairly stereotyped clinical profiles; either exhibiting an in utero disease-onset (74 patients; 22.8% of all patients where data were available), or a post-natal presentation, usually within the first year of life (223 patients; 68.6%), characterized by a sub-acute encephalopathy and a loss of previously acquired skills. Other clinically distinct phenotypes were also observed; particularly, bilateral striatal necrosis (13 patients; 3.6%) and non-syndromic spastic paraparesis (12 patients; 3.4%). We recorded 69 deaths (19.3% of patients with follow-up data). Of 285 patients for whom data were available, 210 (73.7%) were profoundly disabled, with no useful motor, speech and intellectual function. Chilblains, glaucoma, hypothyroidism, cardiomyopathy, intracerebral vasculitis, peripheral neuropathy, bowel inflammation and systemic lupus erythematosus were seen frequently enough to be confirmed as real associations with the Aicardi-Goutieres syndrome phenotype. We observed a robust relationship between mutations in all seven genes with increased type I interferon activity in cerebrospinal fluid and serum, and the increased expression of interferon-stimulated gene transcripts in peripheral blood. We recorded a positive correlation between the level of cerebrospinal fluid interferon activity assayed within one year of disease presentation and the degree of subsequent disability. Interferon-stimulated gene transcripts remained high in most patients, indicating an ongoing disease process. On the basis of substantial morbidity and mortality, our data highlight the urgent need to define coherent treatment strategies for the phenotypes associated with mutations in the Aicardi-Goutieres syndrome-related genes. Our findings also make it clear that a window of therapeutic opportunity exists relevant to the majority of affected patients and indicate that the assessment of type I interferon activity might serve as a useful biomarker in future clinical trials. (c) 2015 Wiley Periodicals, Inc

OBJECTIVE: It has been hypothesized that reduced dietary availability of tryptophan may be the cause of impaired serotonin activity in underweight anorexics. The study reported here evaluated the relationship between tryptophan availability in the blood and nutritional status in anorexia nervosa. METHODS: The total amount of tryptophan and the ratio between tryptophan and other large neutral amino acids (TRP/LNAA) were assessed in a sample of 16 starving anorexic patients. Body weight and composition and energy intake were evaluated in all patients. All subjects also completed self-reported questionnaires such as the Hopkins Symptom Checklist and Eating Disorders Inventory (EDI). RESULTS: The TRP/LNAA ratio seems to be higher in patients with a more severe catabolic status. It is, in fact, significantly inversely correlated with body mass index, body fat, muscle mass, daily energy intake, and daily tryptophan intake. The TRP/LNAA ratio also correlates with growth hormone and the EDI drive for thinness. Patients who exercise excessively had significantly higher TRP/LNAA ratios. CONCLUSIONS: In starving anorexic patients, the TRP/LNAA ratio does not seem to be determined by the content of tryptophan in the diet, but it correlates with measures of catabolism. The relationship of the TRP/LNAA ratio to excessive exercise and starvation indicates the importance of further investigations exploring the role of tryptophan availability in maintaining anorexia nervosa.

Background: Disorders of fatty acid oxidation (FAO) are difficult to diagnose, primarily because in many of the FAO disorders measurable biochemical intermediates accumulate in body fluids only during acute illness. Increased concentrations of 3-hydroxy-fatty acids (3-OH-FAs) in the blood are indicative of FAO disorders of the long- and short-chain 3-hydroxy-acyl-CoA dehydrogenases, LCHAD and SCHAD. We describe a serum/plasma assay for the measurement of 3-OH-FAs with carbon chain lengths from C6 to C16. Methods: We used stable isotope dilution gas chromatography-mass spectrometry (GC-MS) with electron impact ionization and selected ion monitoring. Natural and isotope-labeled compounds were synthesized for the assay. Results: The assay was linear from 0.2 to 50 μmol/L for all six 3-OH-FAs. CVs were 5–15% at concentrations near the upper limits seen in healthy subjects. In 43 subjects, the medians (and ranges) in μmol/L were as follows: 3-OH-C6, 0.8 (0.3–2.2); 3-OH-C8, 0.4 (0.2–1.0); 3-OH-C10, 0.3 (0.2–0.6); 3-OH-C12, 0.3 (0.2–0.6); 3-OH-C14, 0.2 (0.0–0.4); and 3-OH-C16, 0.2 (0.0–0.5). 3-OH-FAs were increased in infants receiving formula containing medium chain triglycerides. Two patients diagnosed with LCHAD deficiency showed marked increases in 3-OH-C14 and 3-OH-C16 concentrations. Two patients diagnosed with SCHAD deficiency showed increased shorter chain 3-OH-FAs but no increases in 3-OH-C14 to 3-OH-C16. Conclusion: Measuring blood concentrations of the 3-OH-FAs with this assay may be a valuable tool for helping to rapidly identify deficiencies in LCHAD and SCHAD and may also provide useful information about the status of the FAO pathway.

Adriana Lombard0 ‘, Giancarlo Goi a, Emma Guagnellini b, Alessandra Fabi a, Gianalfredo Sciorelli ‘, Albert0 B. Burlina d and Guido Tettamanti a** U Department of Biological Chemisrty, The Medical School, University of Milan, Milan h Clinical Chemistry Laboraiory, Bassini Hospital, Cinisello Balsamo, Milan, ’ Immunohaematologv Division, National Institute o/Cancer, Milan and’ Departmenr of Paediatrics. University of Verona, Verona (Italy)

β- d -N- Acetylglucosaminidase isozymes were separated and assayed in the plasma of control healthy individuals and pregnant women by an automated method consisting in chromatofocusing on polybuffer exchanger PBE-94 column, flow-through fluorimetric determination of activity and computer assisted quantification. Under the established optimal conditions the method fractionated β- d -N- acetylglucosaminidase into four isozymes, A, I2, I1 and B, with the analytical coefficients of variation of 1.8, 2.2, 6.4 and 4.1%, respectively. Duration of a single analysis was 25 min including washing, and 10–15 successive runs could be performed on the same column with good reproducibility. A linear activity response was observed from 1–5 μl of plasma (depending on the individual isozyme) to 50 μl, and the detection limit was 0.016 mUnits. Isozyme A was heat labile. Upon sialidase treatment, isozymes A, I2 and I1 released sialic acid and were eluted from the column at less acidic pHs. In healthy individuals isozymes A, I2, I1 and B covered about 62.8, 6.9, 15.0 and 15.1% of the total β- d -N- acetyl-glucosaminidase activity, respectively. During pregnancy the plasma concentration of all isozymes increased. Isozyme I2 showed the highest enhancement (30-fold), followed by I1 (8-fold), B (5.6-fold) and A (3-fold). Interruption of pregnancy by either physiological delivery or ambulatory abortion was followed by a sharp fall of the concentration of all isozymes reaching, in a few days, the control levels.

The following enzymes of lysosomal origin were fluorimetrically determined in maternal plasma from the second to the ninth month of pregnancy at 1-mth intervals: β- d -N- acetylglucosaminidase (EC 3.2.1.30), β- d -glucuronidase (EC 3.2.1.31), β- d -glucosidase (EC 3.2.1.21), β- d -galactosidase (EC 3.2.1.22), α- d -galactosidase (EC 3.2.1.23), α- l -fucosidase (EC 3.2.1.51) and α- d -mannosidase (EC 3.2.1.24) (pH 4.0). As reference microsomal α- d -mannosidase (pH 5.7) was also studied. Thirty-eight healthy women, aged 18–37 yr, who had a normal pregnancy followed by normal parturition, were studied. All enzymes, with the only exception of β-D-galactosidase, showed a progressive and statistically significant increase of activity throughout pregnancy. At the end of pregnancy, the increase ranged from a maximum of 5.6-fold for β- d -N- acetylglucosaminidase to a minimum of 0.55-fold for α- d -mannosidase, pH 5.7. In the case of β- d -N- acetylglucosaminidase , the level at the fifth month of pregnancy was significantly higher than that at the third month, and from the sixth to the ninth month each level significantly differed from that of the month immediately preceding. After interruption of pregnancy (at the third month) and after normal delivery, the plasma level of some lysosomal enzymes, particularly of β- d -N- acetylglucosaminidase , sharply decreased to almost normal values. Therefore, the determination of some lysosomal enzymes, especially β- d -N- acetylglucosaminidase , can be suggested as predictive of a regular course of pregnancy. Women who developed gestational diabetes displayed a marked increase of the plasma levels of some lysosomal enzymes, particularly of β- d -N- acetylglucosaminidase , without any significant change of glycosylated haemoglobin, HbA1c. Thus, the plasma assay of this enzyme may constitute a valuable laboratory tool for monitoring gestational diabetes.

Several lysosomal enzymes (beta-N-acetyl-D-glucosaminidase, beta-D-glucuronidase, alpha-D-galactosidase, beta-D-galactosidase, alpha-D-glucosidase, beta-D-glucosidase, alpha-L-fucosidase and alpha-D-mannosidase) were determined in the serum of 54 non-insulin-dependent diabetics with different degrees of metabolic control and without complications and in 18 non-insulin-dependent diabetics with complications. The serum levels of beta-N-acetyl-D-glucosaminidase, beta-D-glucuronidase, alpha-D-galactosidase, and alpha-D-mannosidase were significantly (p less than 0.01) higher in the diabetics without complications. The levels of beta-N-acetyl-D-glucosaminidase and beta-D-glucuronidase were inversely proportional to the degree of metabolic control, in a statistically significant manner. Moreover the levels of these enzymes decreased to normal values during a 2-month period of controlled oral hypoglycemic drug-diet therapy resulting in metabolic compensation. The presence of complications was indicated by a further increase of serum beta-N-acetyl-D-glucosaminidase and beta-D-glucuronidase; however the portion of lysosomal enzyme activities due to complications remained unchanged after controlled therapy aimed at compensating the metabolism. The conclusion is drawn that in non-insulin-dependent diabetics, as already shown for insulin dependent-diabetics, serum lysosomal enzymes, especially beta-N-acetyl-D-glucosaminidase and beta-D-glucuronidase, are good intraindividual indicators of the metabolic control of the disease.

Summary: A case of an infant suffering from progressive lethargy, sparse scalp hair, autistic-like behavior, myoclonias, and drug-resistant generalized seizures is reported. Laboratory investigations revealed, in the absence of metabolic acidosis, an increased urinary excretion of 2-ketoglutaric acid and a small peak of 3-hydroxyisovaleric acid. The serum biotinidase activity was 0.15 nmol min-1 ml“(normal range 5.2 ± 0.9) in the propositus and 0.310 and 0.420 in her father and mother, respectively. The interictal EEG showed multifocal abnormalities; numerous seizures were recorded, with the pattern of true tonic-clonic fits, exceptional in infancy. Also myoclonias, auditory myoclonus, and repetitive startles were documented. Because of dramatic improvement of ail symptoms and signs after starting biotin (5 mg twice daily), the authors suggest a therapeutical trial in all drug-resistant infantile seizures. RESUME Les auteurs rapportent un cas d'un nourrisson presentant une lethargie progressive, une chevelure clairsemee, un comporte- ment d'allure autistique, des myoclonies et des crises generalisees resistant aux traitements. Les examens de laboratoire ont mis en evidence, en l'absence d'acidose metabolique, une excretion urinaire augmentee d'acide de ceto-glutarique, et un petit pic d'acide 3 hydroxy-hydro-valerique. l'activite biotinidase du serum a eteevaluee a 0.15 nmol/mn/ml (valeurs normales (5.2 ± 0.9) chez la patiente, les chiffres etant respectivement de 0.31 et de 0.42 chez le pere et la mere. l'EEG intercritique a montre des anomalies multi-focales; de nombreuses crises ont ete enregis-trees, d'aspect lonico-clonique vrai, ce qui est exceptionnel chez le nourrisson. Les auteurs ont aussi enregistre des myoclonies, un myoclonus reflexe aux stimulations auditives, et des sursauts repetitifs. Tous les sympodmes ont ete ametioris de facon im-portante apres mise en route d'un traitement par biotine (5 mg, 2 fois par jour); les auteurs suggerent que ce type de traitement doit etre essaye dans toutes les crises infantiles rebelles aux traitements.

The serum levels of the two enzymes of lysosomal origin, beta-N-acetyl-D-glucosaminidase and beta-D-glucuronidase, and the isozyme pattern of the former, were determined in control infants and in infants of diabetic mothers (IDM) on the 1st and 5th day after birth. IDM were divided into three groups. Group 1: class A diabetic mothers treated dietetically; Groups 2 and 3: class A and classes B, C, D diabetic mothers, respectively, treated with insulin. All, but one, diabetic mothers were in excellent metabolic control. In the controls the serum levels of both enzymes were quite elevated on the 1st day after birth, reflecting the condition of the mothers at the end of pregnancy, and increased further on the 5th day, presumably as a result of the concurrent burst of antiinsulin hormones. In Group 1 IDM the serum levels of the two enzymes were higher than in controls, on the 1st day, probably reflecting the higher concentrations present in the mothers at the end of pregnancy than in controls, but equalling the condition of normal neonates on the 5th day. This indicates that IDM of this group had a normal post-natal response of the lysosomal apparatus to hormone stress. In Groups 2-3 IDM the enzyme levels on the 1st day could not be distinguished from those of controls, while on the 5th day a decrease was seen, suggesting reduced effect of the antiinsulin hormone burst on the lysosomal apparatus. The isozyme pattern of beta-N-acetyl-D-glucosaminidase in all IDM was similar to that of controls. The behavior of serum lysosomal enzymes of Groups 2-3 IDM is a further indication that the lysosomal apparatus is extremely sensitive to even small metabolic perturbations occurring in diabetic mothers during pregnancy.

The plasma levels of three lysosomal enzymes, β- D-N -acetylglucosaminidase, β- D -glucuronidase, and α- L -fucosidase, were fluorimetrically determined in seven insulin-dependent diabetic patients one day before, one day after, and during a two-day treatment with the artificial pancreas, at 4 to 5 h intervals. A statistically significant decrease of the plasma level of each enzymes was observed during artificial pancreas treatment. The extent of decrease was 30 to 35% for β- N -acetylglucosaminidase, 35 to 40% for β- D -glucuronidase, and 20 to 25% for α- L -fucosidase. The decrease occurred earlier (at the first day of treatment) for β- D-N -acetylglucosaminidase, and later (at the second day of treatment, and lasting to the first day after treatment) for the other two enzymes. These results suggest a direct connection between the lysosomal apparatus and insulin-controlled metabolic pathways, and a potential role for lysosomal enzymes as indicators of the metabolic compensation in diabetes.

The optimal assay conditions and the stability of the following enzymes of lysosomal origin in human cerebrospinal fluid (CSF) were studied: acid phosphatase, beta-D-N-acetylglucosaminidase, alpha-D-galactosidase, beta-D-galactosidase, alpha-D-glucosidase, beta-D-glucosidase, alpha-L-fucosidase, alpha-D-mannosidase, beta-D-glucuronidase. The microsomal alpha-D-mannosidase, pH 5.7, was used as a reference non-lysosomal glycohydrolase. All the examined enzymes, with the only exception of beta-D-glucuronidase, underwent a more or less rapid loss of activity upon CSF storage in the temperature range from 37 degrees C to -80 degrees C. Storage in liquid nitrogen (-196 degrees C) was the only condition in which full activity for all tested enzymes was maintained for at least 15 days. Addition of human serum albumin to CSF, immediately after withdrawal, had a double effect in favouring enzyme stabilization and causing enzyme activation in some cases, and enzyme inhibition in others. Using conditions warranting enzyme stability the fluorimetric methods for lysosomal enzymes determination in cerebrospinal fluid appear to be highly reproducible (CV less than 5%) and simple enough for routine use.

Background The goal of newborn screening is early identification of babies with a high risk for disorders that may not be clinically evident at birth, but have severe consequences if untreated. New insight into inherited diseases and the ability to test for numerous diseases using new technique such as tandem mass spectrometry have made it practical to greatly expand the number of conditions tested. The expanded neonatal screening is now available and relatively simple, but this represents only a part of the picture. Positive results require follow-up confirmation. Most disorders screened require confirmatory biochemical or genetic tests and specialist visits. An efficient system is needed for managing the care of affected newborns. Expanded newborn screening is not yet available in all Regions of Italy, but discussions aimed at organizing universal access are underway. If these are successful, the role of the pediatrician as the primary contact with the parents is expected to become even more important. Methods We have conducted a survey of Italian pediatricians to assess their familiarity and opinions on newborn screening in general and on expanded newborn screening. All members of the Italian Association of Pediatricians (n = 9000) were invited to compile a 10-item questionnaire online. Results The response rate was 10 %, corresponding to 605 of 6000 active members. Respondents were from all Regions of Italy, with the highest number of responses coming from Lombardy (138, 22.8 %), Campania and Puglia (n = 61; 10.1 %). Interestingly, expanded neonatal screening was not available in any of these Regions at the time of the survey. Regarding their understanding of neonatal screening in general, most respondents (n = 552; 91.1 %) considered that they had at least a sufficient level of knowledge; however, only 59.6 % thought they had sufficient knowledge of expanded newborn screening. Conclusions Successful implementation of a universal expanded NBS program will require efficient procedures for follow-up, diagnosis and treatment to prevent morbidity and mortality of infants and to reduce the period of uncertainty for unaffected families. Pediatricians may need additional training to allow them to fulfill their tasks of coordinating this process while keeping families informed and reassured. Electronic supplementary material The online version of this article (doi:10.1186/s13052-015-0147-1) contains supplementary material, which is available to authorized users.