May 19, 2005 - Lance Wyble, MD. Dorothy K. Grange, MD. Enid Gilbert-Barness, MD .... transition at nucleotide 45 in Exon 1); and a splice site mutation.
Perinatal/Neonatal Case Presentation Sialidosis Presenting as Severe Nonimmune Fetal Hydrops is Associated with Two Novel Mutations in Lysosomal a-Neuraminidase David J. Loren, MD Yvan Campos, BS Alessandra d’Azzo, PhD Lance Wyble, MD Dorothy K. Grange, MD Enid Gilbert-Barness, MD Frances V. White, MD Aaron Hamvas, MD
Sialidosis is a lysosomal storage disease characterized by accumulation of sialylated oligosaccharides in tissues, blood and urine and is caused by mutations in the gene for lysosomal a-neuraminidase (NEU1). There is wide variability in the age of onset and severity of symptoms in sialidosis. We report here a case of sialidosis due to novel mutations in NEU1 presenting as severe nonimmune hydrops fetalis. Journal of Perinatology (2005) 25, 491–494. doi:10.1038/sj.jp.7211335 Published online 19 May 2005
INTRODUCTION The multiple and diverse etiologies of nonimmune hydrops fetalis create a diagnostic challenge in the evaluation of affected fetuses and newborns. In the 1970s, nearly 60% of these cases were characterized as idiopathic. However, with improving diagnostic capabilities, this number has decreased to approximately 20% in more recent surveys.1,2 Inborn errors of metabolism are now more
Edward Mallinckrodt Department of Pediatrics (D.J.L., D.K.G., A.H.), Washington University School of Medicine, St. Louis, MO, USA; Department of Genetics and Tumor Cell Biology (Y.C., A.A.), St. Jude Children’s Research Hospital, USA; Department of Anatomy and Neurobiology (Y.C., A.A.), University of Tennessee, Memphis, TN, USA; Department of Pediatrics (L.W.), University of South Florida, Tampa, FL, USA; Department of Pathology (E.G.B.), University of South Florida, Tampa, FL, USA; Department of Pathology (F.V.W.), Washington University School of Medicine, St. Louis, MO, USA.
Current affiliation: Division of Neonatology (D.J.L.), University of Washington, Seattle, WA, USA. Address correspondence and reprint requests to Aaron Hamvas, MD, Department of Pediatrics, Washington University, Newborn Medicine, St. Louis Children’s Hospital, One Children’s Place, St. Louis, MO 63110, USA.
frequently identified as causes of nonimmune hydrops fetalis. Among these increasingly recognized metabolic disorders are lysosomal storage diseases (e.g., mucopolysaccharidosis VII & IVA, Type II Gaucher, Fabry Disease, Mucoliposis II, Niemann-Pick Type C, Gm1 Gangliosidosis), which are characterized by intracellular accumulation of incompletely processed metabolites, resulting in cell and organ dysfunction.3,4 Sialidosis (MIM# 256550), a rare lysosomal storage disease occurring in 1 in 250,000 to 2,000,000 live births, is due to a deficiency of lysosomal a-neuraminidase (N-acetyl-neuraminic acid hydrolase, EC 3.2.1.18),5 Approximately 40 mutations have been described in NEU1, the gene that encodes a-neuraminidase, which disrupt enzyme expression and/or activity and result in intracellular accumulation, increased serum concentration, and increased urinary excretion of sialyloligosaccharides. Of the NEU1 mutations identified to date, fewer than one-fourth have been associated with hydrops fetalis.6 We report a case of sialidosis presenting as fetal hydrops in which two novel mutations in NEU1 were identified. CASE REPORT The case patient was the first pregnancy for healthy Caucasian parents. Routine ultrasound performed around 18 weeks gestation demonstrated polyhydramnios, fetal scalp edema and ascites. Prenatal evaluations for chromosomal abnormalities, transplacentally acquired infections, structural anomalies, fetal anemia and arrhythmias were all negative. There was no family history of inborn errors of metabolism, other genetic disorders or unexplained neonatal deaths. Peritoneal-amniotic and pleuro-amniotic shunts failed to relieve the fetal fluid collections. Cesarean section was performed at 34 weeks gestation for fetal well-being after spontaneous rupture of membranes. The physical exam was notable for anasarca and hydroceles; the patient required mechanical ventilation immediately and for the duration of his course. Postnatal evaluations for immune-mediated hemolysis and infection were again negative. The peripheral blood smear was remarkable for lymphopenia and ‘‘atypical lymphocytes,’’ which, upon further examination, were determined to be monocytes with vacuolated cytoplasm (Figure 1). Histologic examination of the placenta showed hydropic changes with foamy material within the cytoplasm of the trophoblasts suggestive of a lysosomal storage disorder. Radiographic and ultrasound evaluations revealed
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Figure 1. Photomicrograph of a vacuolated monocyte in the patient’s peripheral blood smear (Wright’s stain, 400).
hepatosplenomegaly, ascites and hydroceles with all other organs and vascular structures being normal. A skeletal survey was normal; specifically, there was no evidence of dysostosis multiplex. A lung biopsy showed diffuse alveolar septal widening with numerous histiocyte-like cells with cytoplasmic vacuoles. Electron microscopy of the lung showed extensive vacuolization of macrophages, epithelial cells and endothelial cells (Figure 2). Testing for a panel of lysosomal storage diseases known to be associated with nonimmune fetal hydrops was performed on lymphocytes, plasma and cultured skin fibroblasts (David A. Wenger, PhD, Thomas Jefferson University). Bound sialic acid in lymphocytes was twice the normal concentration (39.3 nmol/mg with normal less than 20 nmol/mg). In skin fibroblasts, the sialic acid content was elevated and a-neuraminidase activity was A transition at nucleotide 45 in Exon 1); and a splice site mutation (G>A transition at nucleotide 1022 in Exon 5). The parents were each found to be heterozygous for one of the mutations. These substitutions have not been observed in controls indicating that they are not common polymorphisms. DISCUSSION The features in this case that initially suggested the presence of a storage disorder included hepatosplenomegaly and massive hydroceles along with foamy inclusions on placental 492
Figure 2. Electron microscopy of lung. Numerous cytoplasmic membrane-bound vacuoles containing granular material and laminated membranous profiles, present in alveolar septal cells (a, scale bar equals 3 microns) and endothelial cells of a capillary loop (b, scale bar equals 1 micron).
histopathology.7 In addition, the ‘‘atypical’’ lymphocytes originally reported in the peripheral smear were actually monocytes with foamy cytoplasmic inclusions. Storage material within many cell types in the lung demonstrated the systemic nature of the disease and suggests the reason for the significant pulmonary dysfunction observed in our patient. Furthermore, although the definitive mechanisms by which lysosome dysfunction and sialyloligosaccharide accumulation leads to severe fetal hydrops is not established, the vacuolization of pulmonary endothelium on electron microscopy supports theories that suggest an insult to vascular integrity and alteration in Starling forces as the basis for this disorder.8,9 Sialidosis results from a defect in a-neuraminidase, which is part of a multienzyme complex within lysosomes that also includes Journal of Perinatology 2005; 25:491–494
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protective protein/cathepsin A (PPCA), and the glycosidases b-galactosidase and N-acetylgalactosamine-6-sulfate sulfatase.10,11 Association of a-neuraminidase with PPCA, and formation of the multienzyme complex assures the efficient intracellular transport and lysosomal activation of a-neuraminidase.12,13 Sialidosis is typically classified according to clinical presentation, although this classification scheme does not correlate with specific mutations or the degree of enzyme activity. Type I sialidosis is characterized by juvenile or adult onset of myoclonus, dementia, ataxia and tremors.14 Type II sialidosis has an earlier and more dramatic presentation with dysmorphic features, hepatosplenomegaly, dysostosis multiplex, nephrotic syndrome, mental retardation, and progressive neurological deterioration.15 Within the ‘‘Type II’’ classification is a congenital form that presents as hydrops fetalis.16 In vitro studies suggest that the severity of disease appears to be a function of not only the residual activity of the enzyme, but also its intracellular localization. Thus, mutations that result in retention of a-neuraminidase in the endoplasmic reticulum (ER) or Golgi are associated with the severe congenital forms of sialidosis, including those with hydrops fetalis. In contrast, mutant protein that reaches the lysosome and is catalytically inactive results in juvenile Type II sialidosis, whereas mutant protein that is lysosomal and partially active is responsible for the milder Type I disease.17 Interpreting genotype–phenotype correlation is further complicated by phenotypic heterogeneity even within family members sharing the same mutation. Of the many mutations in NEU1 that are responsible for sialidosis, only a small number have been described that cause hydrops fetalis. The mutations identified in our patient would appear to support this mechanistic scheme: the 45G>A mutation, which introduces a premature stop codon [W15X] in the signal peptide, is likely to be a null mutation while the 1022G>A mutation predicts insertion of 32 additional amino acids that would likely result in a misfolded peptide with ER retention and/or degradation.18 In the case described here, establishing the biochemical diagnosis permitted a deliberate and informed decision regarding prognosis and direction of care. Furthermore, with the molecular etiology established, accurate prenatal diagnosis will be available for the family in future pregnancies. It has become increasingly clear that many cases of ‘‘idiopathic’’ nonimmune hydrops fetalis are due to lysosomal storage diseases; thus it is imperative that these conditions be considered early in the evaluation when the more common causes have been excluded. Detailed examination of readily available specimens can provide early clues that a storage disease may be present. When combined with sophisticated, but available, biochemical and molecular diagnostic testing, the ability to arrive at a specific diagnosis in more patients with nonimmune hydrops fetalis should decrease the number of cases deemed ‘‘idiopathic’’. Journal of Perinatology 2005; 25:491–494
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Acknowledgements We thank Kim Detjen MSW at St. Louis Children’s Hospital NICU for providing support to the family and care team. Supported by NIH DK52025, GM60950, Assisi Foundation of Memphis, the Cancer Center Support Grant CA21765, and the American Lebanese Syrian Associated Charities (ALSAC) (Ad’A & YC); NIH HL-65174 (AH). Ad’A holds an Endowed Chair in Genetics and Gene Therapy from the Jeweler Charity Fund.
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16. Beck M, Bender SW, Reiter HL, et al. Neuraminidase deficiency presenting as nonimmune hydrops fetalis. Eur J Pediatr 1984;143:135–9. 17. Bonten EJ, Arts WF, Beck M, et al. Novel mutations in lysosomal neuraminidase identify functional domains and determine clinical severity in sialidosis. Hum Mol Genet 2000;9:2715–25.
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18. Pattison S, Pankarican M, Rupar CA, Graham FL, Igdoura SA. Five novel mutations in the lysosomal sialidase gene (neu1) in type II sialidosis patients and assessment of their impact on enzyme activity and intracellular targeting using adenovirus-mediated expression. Hum Mutat 2004; 23:32–9.
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