Diagnosis Of Lysosomal Storage Diseases Research Articles

Hepatomegaly and Splenomegaly: An Approach to the Diagnosis of Lysosomal Storage Diseases.

Clinical findings of hepatomegaly and splenomegaly, the abnormal enlargement of the liver and spleen, respectively, should prompt a broad differential diagnosis that includes metabolic, congestive, neoplastic, infectious, toxic, and inflammatory conditions. Among the metabolic diseases, lysosomal storage diseases (LSDs) are a group of rare and ultrarare conditions with a collective incidence of 1 in 5000 live births. LSDs are caused by genetic variants affecting the lysosomal enzymes, transporters, or integral membrane proteins. As a result, abnormal metabolites accumulate in the organelle, leading to dysfunction. Therapeutic advances, including early diagnosis and disease-targeted management, have improved the life expectancy and quality of life of people affected by certain LSDs. To access these new interventions, LSDs must be considered in patients presenting with hepatomegaly and splenomegaly throughout the lifespan. This review article navigates the diagnostic approach for individuals with hepatosplenomegaly particularly focusing on LSDs. We provide hints in the history, physical exam, laboratories, and imaging that may identify LSDs. Additionally, we discuss molecular testing, arguably the preferred confirmatory test (over biopsy), accompanied by enzymatic testing when feasible.

Out-of-pocket health expenditures in patients living with ınborn errors of metabolism

AimThe implementation of newborn screening programs for inborn errors of metabolism has advanced the diagnosis and management of affected infants and undoubtedly improved their outcomes. We aimed to determine out-of-pocket health expenditures of patients with inborn errors of metabolism during follow-up and treatment processes and to determine the economic burden on the families.Materials and methodsA total of 232 patients who voluntarily agreed to participate in the study and were regularly followed up in the Department of Pediatric Metabolism with the diagnosis of Inborn Errors of Metabolism between April 2022 and July 2022 were included. Questionnaires were asked about the demographic characteristics of patients, use of health services, follow-up, treatment procedures, frequency of controls and health expenditures.ResultsThe average out-of-pocket expenditure of the households in the last month was 1039.22 ± 1030.08 (minimum: 20, maximum: 5000) Turkish Liras. When we consider the catastrophic health expenditure rate as expenditure exceeding 40% of household income, we found that 9.9% (23 people) of parents included in the study made catastrophic health expenditures. The rate of catastrophic expenditure of patients with a diagnosis of Amino Acid Metabolism Disorders was found to be higher than that of patients with a diagnosis of Vitamin and Cofactor Metabolism Disorders. Similarly, patients with a diagnosis of lysosomal storage diseases had more expenditures than patients with a diagnosis of vitamin and cofactor metabolism disorders. When we compared the rate of catastrophic health expenditure of the patients with urea cycle disorders and the patients with a diagnosis of vitamin and cofactor metabolism disorders, the former had more expenditure than the latter (p < 0.05). There was no significant difference between other disease groups in terms of catastrophic expenditure. The rate of catastrophic expenditures of the households living as large family type were higher than the families living as nuclear family type (p < 0.01). A statistically significant difference was found between the rates of catastrophic expenditures of the families living in Ankara and those who were admitted from other provinces for follow-up and treatment (p < 0.001). However, there was no difference between the rates of catastrophic expenditure of the patients who received any treatment and those who were followed up without treatment (p > 0.05).ConclusionDue to the high rate of consanguineous marriages in our country, the development of newborn screening programs, the increase in awareness about metabolic diseases and the improvement in diagnostic methods, the frequency of metabolic diseases is increasing, and mortality and morbidity rates are significantly reduced with early diagnosis and treatment opportunities. It is necessary to carry out more comprehensive studies to determine and prevent the socioeconomic effects of out-of-pocket health expenditures of patients living with Inborn Errors of Metabolism.

An artifiсial intelligence computer system for differential diagnosis of lysosomal storage diseases

Aim. To improve the efficiency of diagnosis of hereditary lysosomal storage diseases using an intelligent computerbased decision support system.Materials and methods. Descriptions of 35 clinical cases from the literature and depersonalized data of 52 patients from electronic health records were used as material for clinical testing of the computer diagnostic system. Knowledge engineering techniques have been used to extract, structure, and formalize knowledge from texts and experts. Literary sources included online databases and publications (in Russian and English). On this basis, for each clinical form of lysosomal diseases, textological cards were created, the information in which was corrected by experts. Then matrices were formed, including certainty factors (coefficients) for the manifestation, severity, and relevance of signs for each age group (up to 1 year, from 1 to 3 years inclusive, from 4 to 6 years inclusive, 7 years and older). The knowledge base of the expert system was implemented on the ontology network and included a disease model with reference variants of clinical forms. Decision making was carried out using production rules.Results. The expert computer system was developed to support clinical decision-making at the pre-laboratory stage of differential diagnosis of lysosomal storage diseases. The result of its operation was a ranked list of hypotheses, reflecting the degree of their compliance with reference descriptions of clinical disease forms in the knowledge base. Clinical testing was carried out on cases from literary sources and patient data from electronic health records. The criterion for assessing the effectiveness of disease recognition was inclusion of the verified diagnosis in the list of five hypotheses generated by the system. Based on the testing results, the accuracy was 87.4%.Conclusion. The expert system for the diagnosis of hereditary diseases has shown fairly high efficiency at the stage of compiling a differential diagnosis list at the pre-laboratory stage, which allows us to speak about the possibility of its use in clinical practice.

A Comprehensive, Targeted NGS Approach to Assessing Molecular Diagnosis of Lysosomal Storage Diseases.

With over 60 different disorders and a combined incidence occurring in 1:5000–7000 live births, lysosomal storage diseases (LSDs) represent a major public health problem and constitute an enormous burden for affected individuals and their families. Several reasons make the diagnosis of LSDs an arduous task for clinicians, including the phenotype and penetrance variability, the shared signs and symptoms, and the uncertainties related to biochemical enzymatic assay results. Developing a powerful diagnostic tool based on next generation sequencing (NGS) technology may help reduce the delayed diagnostic process for these families, leading to better outcomes for current therapies and providing the basis for more appropriate genetic counseling. Herein, we employed a targeted NGS-based panel to scan the coding regions of 65 LSD-causative genes. A reference group sample (n = 26) with previously known genetic mutations was used to test and validate the entire workflow. Our approach demonstrated elevated analytical accuracy, sensitivity, and specificity. We believe the adoption of comprehensive targeted sequencing strategies into a routine diagnostic route may accelerate both the identification and management of LSDs with overlapping clinical profiles, producing a significant reduction in delayed diagnostic response with beneficial results in the treatment outcome.

Differential diagnosis of hereditary metabolic diseases using the expert knowledge-based system

Aim. The aim of the study was to create a computer decision support system using expert knowledge for the diagnosis of rare hereditary diseases due to the difficulty of their identification at the pre-laboratory stage.Material and Methods. Descriptions of the clinical picture of lysosomal storage diseases from literature sources were used as the research material. The methods included knowledge extraction, expert assessments, quantization of age intervals, and applied intelligent services to form a knowledge base.Results. The results of the study include the construction of models for a complex assessment of a sign and an integral assessment of a disease, on the basis of which the comparative analysis algorithm is implemented to assess each of the hypotheses put forward by the system. The results of testing the prototype of the created expert system on a control sample of patients with mucopolysaccharidosis showed the efficiency of 90%. Discussion. In the discussion, several diagnostic systems are considered and their distinction from the system, presented in this work, is shown.Conclusion. The results of the development of intelligent system based on knowledge for the diagnosis of lysosomal storage diseases are summarized and the perspectives for its development are highlighted.

Assessing Lysosomal Disorders in the NGS Era: Identification of Novel Rare Variants

Lysosomal storage diseases (LSDs) are a heterogeneous group of genetic disorders with variable degrees of severity and a broad phenotypic spectrum, which may overlap with a number of other conditions. While individually rare, as a group LSDs affect a significant number of patients, placing an important burden on affected individuals and their families but also on national health care systems worldwide. Here, we present our results on the use of an in-house customized next-generation sequencing (NGS) panel of genes related to lysosome function as a first-line molecular test for the diagnosis of LSDs. Ultimately, our goal is to provide a fast and effective tool to screen for virtually all LSDs in a single run, thus contributing to decrease the diagnostic odyssey, accelerating the time to diagnosis. Our study enrolled a group of 23 patients with variable degrees of clinical and/or biochemical suspicion of LSD. Briefly, NGS analysis data workflow, followed by segregation analysis allowed the characterization of approximately 41% of the analyzed patients and the identification of 10 different pathogenic variants, underlying nine LSDs. Importantly, four of those variants were novel, and, when applicable, their effect over protein structure was evaluated through in silico analysis. One of the novel pathogenic variants was identified in the GM2A gene, which is associated with an ultra-rare (or misdiagnosed) LSD, the AB variant of GM2 Gangliosidosis. Overall, this case series highlights not only the major advantages of NGS-based diagnostic approaches but also, to some extent, its limitations ultimately promoting a reflection on the role of targeted panels as a primary tool for the prompt characterization of LSD patients.

Finding vacuolated lymphocytes in fetal effusions improves the prenatal diagnosis of lysosomal storage diseases

There are many causes of fetal effusions, including the rare lysosomal storage diseases (LSDs). Vacuolated lymphocytes (VLs) are found in the blood of infants with LSDs, and their presence in fetal effusion could increase the risk of underlying LSD. Between 2006 and 2018, all fetal effusions samples from 43 fetal multidisciplinary centers were referred to a single laboratory. Cells were counted, and, if observed, VLs were categorized and counted. Screening for LSDs was performed by metabolite analyses on amniotic fluid supernatant. The diagnosis of an LSD was confirmed by measuring the activity of the corresponding enzyme and/or mutation analysis. Our laboratory received 614 ascitic fluids and 280 pleural fluids sampled between 22 and 33 weeks of gestation. The final diagnosis was LSD in 16 cases (1.8%). VLs were reported in all these 16 cases, in a mix of lymphocytes with and without vacuoles. Vacuoles in VLs varied in size and number. In most cases, VLs were easy to recognize, with numerous, large, round, well-defined vacuoles, but in three cases of LSDs, VLs were atypical. The finding of VLs in fetal effusions is an inexpensive first-line test that may help to prioritize biochemical and genetic tests for LSDs.

Tandem mass spectrometry-based multiplex assays for α-mannosidosis and fucosidosis

Multiplex tandem mass spectrometry (MS/MS)-based enzyme activity assays for newborn screening (NBS) and diagnosis of lysosomal storage diseases (LSDs) in newborns, using dried blood spots (DBS) on newborn screening cards, have garnered much attention due to its sensitivity, high precision, and the capability to screen for an unprecedented number of diseases in a single assay. Herein we report the development of MS/MS-based enzyme assays for the diagnosis of α-mannosidosis and fucosidosis. These new protocols are able to distinguish untreated patients from random newborns, carriers and a post-bone marrow transplant patient. We have successfully multiplexed the α-mannosidosis assay with a multiplex MS/MS assay for the screening and diagnosis of other LSDs, namely Fabry, Pompe, MPS I, Gaucher, Niemann-Pick-A/B, and Krabbe diseases. Additionally, we also multiplexed the fucosidosis NBS assay with a 5-plex assay that tests for MPS-II, MPS-IIIB, MPS-IVA, MPS-VI and MPS-VII.

Quand penser à une maladie de surcharge lysosomale (Morquio inclus) ?

Lysosomal storage diseases (LSDs) are a group of over 70 diseases that are characterized by lysosomal dysfunction, caused by progressive accumulation of metabolites due to defects in lysosomal enzymes and degradation pathways, most of which are inherited as autosomal recessive traits. These disorders are individually rare but collectively affect more than 1 in 4000 live births. LSDs typically present in infancy and childhood, although adult- onset forms also occur. Lysosomal storage diseases (LSDs) are rare systemic and progressive genetic metabolic diseases, which present a wide heterogeneity (from severe to attenuated form) and a wide phenotypic spectrum of nonspecific manifestations, which may lead to considerable diagnostic delay. The precise diagnosis can be difficult or impossible, taking into account clinical variability and overlapping symptoms among LSD. The most of these disorders manifest with neurological, musculoskeletal and visceral signs. If severe subtypes are often readily diagnosed, the more attenuated subtypes are frequently missed or diagnosis is significantly delayed. Early diagnosis has become essential since effective disease-specific treatments, including enzyme replacement therapy and stem cell transplantation, have become available for certain LSDs, necessary to prevent the development of disabling and irreversible manifestations. Diagnosis of LSD is driven by clinical suspicion and several clinical approaches are possible: what presentations should prompt consideration of a LSD? Is it a neurological or a visceral phenotype? How thinking about a mucopolysaccharidoses, an oligosaccharidoses, a sphingolipidoses? How to discuss for each clinical sign or symptom the differential diagnosis of all LSD?

Performance of the Four-Plex Tandem Mass Spectrometry Lysosomal Storage Disease Newborn Screening Test: The Necessity of Adding a 2nd Tier Test for Pompe Disease.

Early diagnosis of lysosomal storage diseases (LSDs) through newborn screening (NBS) has been adapted widely. The National Taiwan University Hospital Newborn Screening Center launched the four-plex tandem mass spectrometry LSD newborn screening test in 2015. The test determined activities of acid α-glucosidase (GAA; Pompe), acid α-galactosidase (GLA; Fabry), acid β-glucocerebrosidase (ABG; Gaucher), and acid α-l-iduronidase (IDUA; MPS-I) in dried blood spots (DBS). Through 2017, 64,148 newborns were screened for these four LSDs. The screening algorithm includes enzyme activity/ratio as the cutoffs for the first screening test and a second-tier test for Pompe disease screening. The second-tier Pompe disease screening test measured activity inhibition by acarbose. Twenty-nine newborns required a confirmatory test; six were confirmed to have Pompe disease, and nine were confirmed to have Fabry disease. The screen-positive rate for Pompe disease was 0.031%. Therefore, in Pompe disease newborn screening, a validated 2nd tier test is necessary to decrease false positives.

Clinical implementation of gene panel testing for lysosomal storage diseases.

BackgroundThe diagnostic workup in patients with a clinical suspicion of lysosomal storage diseases (LSD) is often difficult due to the variability in the clinical phenotype. The gold standard for diagnosis of LSDs consists of enzymatic testing. However, due to the sequential nature of this methodology and inconsistent genotype–phenotype correlations of certain LSDs, finding a diagnosis can be challenging.MethodWe developed and clinically implemented a gene panel covering 50 genes known to cause LSDs when mutated. Over a period of 18 months, we analyzed 150 patients who were referred for LSD testing and compared these results with the data of patients who were previously enrolled in a scheme of classical biochemical testing.ResultsOur panel was able to determine the molecular cause of the disease in 22 cases (15%), representing an increase in diagnostic yield compared to biochemical tests developed for 21 LSDs (4.6%). We were furthermore able to redirect the diagnosis of a mucolipidosis patient who was initially suspected to be affected with galactosialidosis. Several patients were identified as being affected with neuronal ceroid lipofuscinosis, which cannot readily be detected by enzyme testing. Finally, several carriers of pathogenic mutations in LSD genes related to the disease phenotype were identified as well, thus potentially increasing the diagnostic yield of the panel as heterozygous deletions cannot be detected.ConclusionWe show that the implementation of a gene panel for LSD diagnostics results in an increased yield in comparison to classical biochemical testing. As the panel is able to cover a wider range of diseases, we propose to implement this methodology as a first‐tier test in cases of an aspecific LSD presentation, while enzymatic testing remains the first choice in patients with a more distinctive clinical presentation. Positive panel results should however still be enzymatically confirmed whenever possible.

Vacuolated Lymphocytes as a Clue for Diagnosis of Lysosomal Storage Disease like GM1 Gangliosidosis.

Vacuolated Lymphocytes as a Clue for Diagnosis of Lysosomal Storage Disease like GM1 Gangliosidosis.

False Positive Diagnosis of Lysosomal Storage Disease Based on Dried Blood Spot Sample; Leucocyte Number of a Challenging Factor

False Positive Diagnosis of Lysosomal Storage Disease Based on Dried Blood Spot Sample; Leucocyte Number of a Challenging Factor

Validation of a Multiplex Tandem Mass Spectrometry Method for the Detection of Selected Lysosomal Storage Diseases in Dried Blood Spots

Abstract Background: Interest in screening methods for lysosomal storage diseases (LSDs) has increased in recent years, since early diagnosis and treatment are essential to prevent or attenuate the onset of symptoms and the complications of these diseases. In the current work, we evaluated the performance of tandem mass spectrometry (MS/MS) for the detection of some LSDs, aiming the future use of this methodology for the screening of these disorders. Methods: Standard curves and quality control dried blood spots were assayed to evaluate the precision, linearity, and accuracy. A total of 150 controls were grouped according to age and subjected to measurement of lysosomal enzymes deficient in Niemann-Pick A/B, Krabbe, Gaucher, Fabry, Pompe, and Mucopolysaccharidosis type I diseases. Samples from 59 affected patients with a diagnosis of LSDs previously confirmed by fluorimetric methods were analyzed. Results: Data from standard calibration demonstrated good linearity and accuracy and the intra- and interassay precisions varied from 1.17% to 11.60% and 5.39% to 31.24%, respectively. Except for galactocerebrosidase and ?-l-iduronidase, enzyme activities were significantly higher in newborns compared to children and adult controls. Affected patients presented enzymatic activities significantly lower compared to all control participants. Conclusion: Our results show that MS/MS is a promising methodology, suitable for the screening of LSDs, but accurate diagnoses will depend on its correlation with other biochemical and/or molecular analyses.

Chitotriosidase activity as additional biomarker in the diagnosis of lysosomal storage diseases.

To date, several genetic variants that lead to a deficiency of chitotriosidase activity have been described. The duplication of 24 bp (dup24bp) in exon 10 of the CHIT1 gene, which causes a complete loss of enzymatic activity of the gene product, is the most common among the European population. The aim of the study was to evaluate the possibility of using chitotriosidase activity as an additional biomarker in diagnosis of lysosomal storage diseases (LSDs) in Ukraine, to determine this parameter in blood plasma of the patients with various lysosomal diseases and to assess the effect of the presence of dup24bp in the CHIT1 gene on this parameter. It has been shown that chitotriosidase activity in blood plasma is a convenient additional biochemical marker in the diagnosis of some LSDs, namely Gaucher disease, Niemann-Pick disease A, B, C and GM1-gangliosidosis. Reference ranges of the normal chitotriosidase activity were determined in blood plasma of Ukrainian population and found to be 8.0-53.1 nmol 4-methylumbelliferone/h·ml of plasma. The total allele frequency of the dup24bp in the CHIT1 gene in Ukrainian population was determined, which amounted to 0.26 (323/1244) that is higher than in European population. It was indicated that moleculargenetic screening of dup24bp in the CHIT1 gene is a necessary stage in a protocol for the laboratory diagnosis of Gaucher disease, Niemann-Pick disease A, B, C as well as GM1-gangliosidosis to avoid incorrect diagnosis.

Galactocerebrosidase assay on dried-leukocytes impregnated in filter paper for the detection of Krabbe disease

BackgroundKrabbe disease (KD) is an inherited lysosomal storage disease (LSD) caused by the deficiency of galactocerebrosidase (GALC) and is characterized by a severe and progressive leukodystrophy with death frequently before one year of life in the classical early-onset form. As a consequence of the enzyme defect, globoid cells containing undigested galactosylceramide are observed and are characteristic of the disease. Hematopoietic stem cell transplantation is the current treatment for this disease, with some success in the classical cases if performed very early in life. Definitive diagnosis of KD is generally accessed by determination of GALC in leukocytes or fibroblasts. For the last few years, dried-blood filter paper (DBFP) samples have been increasingly used for lysosomal enzyme assays. Originally, some lysosomal enzymes could not be tested in DBFP samples using fluorometric assays, including GALC, heparan-sulfamidase and a few others. Recently, we reported successful results using dried-leukocytes filter paper (DLFP) samples for heparan sulfamidase and β-galactosidase. Extending these studies, we present now a new GALC assay on these type of samples. MethodsAdapted leukocyte fluorometric assay was used for the evaluation of GALC in DLFP samples. ResultsOur results using this method showed a clear discrimination between GALC levels observed in KD patients and healthy controls. ConclusionsThe assay is robust and reliable and could be adopted by reference laboratories for diagnosis of LSDs. It is expected that the use of DLPF would make it possible to diagnose patients living in isolated areas, where liquid samples usually have to be transported over several days and sometimes across country borders before reaching reference laboratories.

Assessment of a targeted resequencing assay as a support tool in the diagnosis of lysosomal storage disorders.

BackgroundWith over 50 different disorders and a combined incidence of up to 1/3000 births, lysosomal storage diseases (LSDs) constitute a major public health problem and place an enormous burden on affected individuals and their families. Many factors make LSD diagnosis difficult, including phenotype and penetrance variability, shared signs and symptoms, and problems inherent to biochemical diagnosis. Developing a powerful diagnostic tool could mitigate the protracted diagnostic process for these families, lead to better outcomes for current and proposed therapies, and provide the basis for more appropriate genetic counseling.MethodsWe have designed a targeted resequencing assay for the simultaneous testing of 57 lysosomal genes, using in-solution capture as the enrichment method and two different sequencing platforms. A total of 84 patients with high to moderate-or low suspicion index for LSD were enrolled in different centers in Spain and Portugal, including 18 positive controls.ResultsWe correctly diagnosed 18 positive blinded controls, provided genetic diagnosis to 25 potential LSD patients, and ended with 18 diagnostic odysseys.ConclusionWe report the assessment of a next–generation-sequencing-based approach as an accessory tool in the diagnosis of LSDs, a group of disorders which have overlapping clinical profiles and genetic heterogeneity. We have also identified and quantified the strengths and limitations of next generation sequencing (NGS) technology applied to diagnosis.

Oligosaccharide Analysis in Urine by MALDI-TOF Mass Spectrometry for the Diagnosis of Lysosomal Storage Diseases

There are 45 known genetic diseases that impair the lysosomal degradation of macromolecules. The loss of a single lysosomal hydrolase leads to the accumulation of its undegraded substrates in tissues and increases of related glycoconjugates in urine, some of which can be detected by screening of free oligosaccharides (FOS) in urine. Traditional 1-dimensional TLC for urine oligosaccharide analysis has limited analytical specificity and sensitivity. We developed fast and robust urinary FOS and glycoaminoacid analyses by MALDI-time-of-flight/time-of-flight (MALDI-TOF/TOF) mass spectrometry for the diagnosis of oligosaccharidoses and other lysosomal storage diseases. The FOS in urine equivalent to 0.09 mg creatinine were purified through sequential passage over a Sep-Pak C18 column and a carbograph column and were then permethylated. MALDI-TOF/TOF was used to analyze the permethylated FOS. We studied urine samples from individuals in 7 different age groups ranging from 0-1 months to ≥ 17 years as well as urine from known patients with different lysosomal storage diseases. We identified diagnostic urinary FOS patterns for α-mannosidosis, galactosialidosis, mucolipidosis type II/III, sialidosis, α-fucosidosis, aspartylglucosaminuria (AGU), Pompe disease, Gaucher disease, and GM1 and GM2 gangliosidosis. Interestingly, the increase in urinary FOS characteristic of lysosomal storage diseases relative to normal FOS appeared to correlate with the disease severity. The analysis of urinary FOS by MALDI-TOF/TOF is a powerful tool for first-tier screening of oligosaccharidoses and lysosomal storage diseases.

Prenatal-Onset Niemann–Pick Type C Disease with Nonimmune Hydrops Fetalis

Niemann-Pick type C (NPC; OMIM 257219) disease is a neurodegenerative lysosomal storage disorder characterized by accumulation of unesterified cholesterol in the lysosomal/late endosomal system. This autosomal recessive disorder occurs in approximately 1/150,000 births. The broad clinical spectrum ranges from a prenatal severe presentation to an adult-onset chronic neurodegenerative disease. Data about prenatal presentation of NPC are limited. A female newborn was born at 34(2) weeks' gestation with a birth weight of 3070 g, and transferred to the Neonatal Intensive Care Unit because of nonimmune hydrops fetalis (NIHF) and respiratory distress. On admission, a physical examination revealed skin edema, mild respiratory distress, and abdominal distention due to massive ascites. Hepatosplenomegaly and cholestasis increased progressively and bleeding diathesis occurred. Results of an abdominal ultrasonography showed hepatosplenomegaly and segmental multicystic dysplastic left kidney. Foamy cells with a lysosomal phospholipid storage pattern compatible with NPC were found in the bone marrow smear. Cultured fibroblasts showed a strongly elevated filipin staining (classical NPC cellular phenotype), establishing the diagnosis of NPC. The infant died on the 52(nd) day of life because of respiratory distress due to lung involvement of NPC, massive ascites, and progressive liver failure. Results of an autopsy showed multiorgan storage disease involving the liver, spleen, lymph nodes, thymus, lungs, and brain. Here, we present a preterm infant with NIHF as a sign of severe prenatal-onset NPC and review the literature.

Enzymatic Screening and Diagnosis of Lysosomal Storage Diseases.

Lysosomal storage diseases (LSDs) are a group of more than 50 genetic disorders. Clinical symptoms are caused by the deficiency of specific enzyme (enzymes) function and resultant substrate accumulation in the lysosomes, which leads to impaired cellular function and progressive tissue and organ dysfunction. Measurement of lysosomal enzyme activity plays an important role in the clinical diagnosis of LSDs. The major enzymatic testing methods include fluorometric assays using artificial 4-methylumbelliferyl (4-MU) substrates, spectrophotometric assays and radioactive assays with radiolabeled natural substrates. As many effective treatment options have become available, presymptomatic diagnosis and early intervention are imperative. Many methods were developed in the past decade for newborn screening (NBS) of selective LSDs in dried blood spot (DBS) specimens. Modified fluorometric assays with 4-MU substrates, MS/MS or LC-MS/MS multiplex enzyme assays, digital microfluidic fluorometric assays, and immune-quantification assays for enzyme contents have been reported in NBS of LSDs, each with its own advantages and limitations. Active technical validation studies and pilot screening studies have been conducted or are ongoing. These studies have provided insight in the efficacy of various methodologies. In this review, technical aspects of the enzyme assays used in clinical diagnosis and NBS are summarized. The important findings from pilot NBS studies are also reviewed.