Difficult airways: a 3D printing study with virtual fibreoptic endoscopy.

Head and neck cancer patients present unique airway challenges, and oropharyngeal, laryngeal, and hypopharyngeal tumours considerably distort and narrow the anatomy of the airway. We describe the use of 3D augmented reality software combined with 3D printed models to assess the anatomy of difficult airways and to assist in the formulation of the most optimal airway management strategy in such patients. The reported patients had computed tomograms (CT) of the neck prior to their anaesthetic and surgical management. DICOM files of the respective scans were imported to 3D rendering software (OsiriX, Pixmeo). We constructed volume rendered models for initial assessment of the airway then generated serial surface rendered models to create a virtual endoscopic path of the airway to simulate the fibreoptic approach. To further facilitate the study of difficult airways we have subsequently printed 3D models of those that were most difficult using rapid prototyping. Head and neck tumours significantly distort the airway. Thorough study of the relevant anatomy prior to airway management for operating reasons enhances communication between the surgeon and anaesthetist, and aids selection of the most appropriate intubation approach. In conclusion, this paper highlights a useful and novel pre-assessment strategy that allows a virtual, visual, 3-dimensional assessment of the airway anatomy combined with 3D modelling and 3D printing. This enables the airway specialist, anaesthetist, and head and neck surgeon to anticipate any critical steps and adjust the plan accordingly.

Marked elevation in plasma trimethylamine-N-oxide (TMAO) in patients with mitochondrial disorders treated with oral l-carnitine.

Oral supplementation with l-carnitine is a common therapeutic modality for mitochondrial disorders despite limited evidence of efficacy. Recently, a number of studies have demonstrated that a gut microbiota-dependent metabolite of l-carnitine, trimethylamine oxide (TMAO), is an independent and dose-dependent risk factor for cardiovascular disease (CVD). Given the limited data demonstrating efficacy with oral l-carnitine therapy and the newly raised questions of potential harm, we assessed plasma TMAO levels in patients with mitochondrial disease with and without oral l-carnitine supplementation. Nine subjects were recruited and completed the study. Eight out of 9 subjects at baseline had plasma TMAO concentrations <97.5th percentile (<15.5 µM). One subject with stage 3 renal disease, had marked elevation in plasma TMAO (pre 33.98 µm versus post 101.6 µm). Following at least 3 months of l-carnitine supplementation (1000 mg per day), plasma TMAO levels were markedly increased in 7out of 9 subjects; overall, plasma TMAO significantly increased 11.8-fold (p < 0.001) from a baseline median level of 3.54 µm (interquartile range (IQR) 2.55-8.72) to 43.26 (IQR 23.99-56.04) post supplementation. The results of this study demonstrate that chronic oral l-carnitine supplementation markedly increases plasma TMAO levels in subjects with mitochondrial disorders. Further studies to evaluate both the efficacy and long term safety of oral l-carnitine supplementation for the treatment of mitochondrial disorders are warranted.

Clinical and molecular features of mitochondrial DNA depletion due to mutations in deoxyguanosine kinase.

Published mutations in deoxyguanosine kinase (DGUOK) cause mitochondrial DNA depletion and a clinical phenotype that consists of neonatal liver failure, nystagmus and hypotonia. In this series, we have identified 15 different mutations in the DGUOK gene from 9 kindreds. Among them, 12 have not previously been reported. Nonsense, splice site, or frame-shift mutations that produce truncated proteins predominate over missense mutations. All patients who harbor null mutations had early onset liver failure and significant neurological disease. These patients have all died before 2-years of age. Conversely, two patients carrying missense mutations had isolated liver disease and are alive in their 4th year of life without liver transplant. Five subjects were detected by newborn screening, with elevated tyrosine or phenylalanine. Consequently, this disease should be considered if elevated tyrosine is identified by newborn screening. Mitochondrial DNA content was below 10% of controls in liver in all but one case and modestly reduced in blood cells. With this paper a total of 39 different mutations in DGUOK have been identified. The most frequent mutation, c.763_c.766dupGATT, occurs in 8 unrelated kindreds. 70% of mutations occur in only one kindred, suggesting full sequencing of this gene is required for diagnosis. The presentation of one case with apparent viral hepatitis, without neurological disease, suggests that this disease should be considered in patients with infantile liver failure regardless of the presence of neurological features or apparent infectious etiology.

Carnitine transporter defect due to a novel mutation in the SLC22A5 gene presenting with peripheral neuropathy.

The carnitine transporter defect (McKusick 212140) is an autosomal recessive disorder caused by mutations in the SLC22A5 gene, which encodes the high-affinity carnitine transporter OCTN2 (Wang et al 2001). Diagnosis is suspected when plasma carnitine levels are extremely low and secondary causes of carnitine loss are excluded. The disease can present with recurrent Reye-like episodes of hypoketotic hypoglycaemia or with cardiomyopathy associated with myopathy (Stanley et al 1991). Here we report novel clinical findings in a 3-year-old with primary carnitine deficiency.

Pseudo-lysosomal storage disease caused by EMLA cream.

Prilocaine-lidocaine emulsion (EMLA cream) is a topical anaesthetic commonly used prior to diagnostic and therapeutic procedures. While undergoing clinical investigation for the suspicion of a metabolic disorder, a series of children underwent skin biopsy with EMLA cream pretreatment. In each case, the pathologist identified ultrastructural features consistent with a lysosomal storage disorder, yet the clinical features were not consistent with the pathological findings. Ultrastructural artefact was suspected, resulting from the use of the EMLA cream. All patients underwent repeat skin biopsy without EMLA cream. Biopsies were reviewed by two pathologists blinded to the previous biopsy findings. Electron microscopy repeated without the use of EMLA cream was normal. It is concluded that the use of EMLA cream causes ultrastructural artefact and should be avoided prior to skin biopsy for electron microscopy.

A case of sporadic infantile histiocytoid cardiomyopathy caused by the A8344G (MERRF) mitochondrial DNA mutation.

The A8344G mitochondrial DNA (mtDNA) mutation is best known for the MERRF phenotype (myoclonic epilepsy, myopathy, and ragged red fibers). We describe a sporadic case of an infant with the A8344G mtDNA mutation who presented with failure to thrive and sudden unexpected death at 11 months of age. The autopsy revealed a histiocytoid cardiomyopathy, diffuse steatosis of the liver, and bilateral retinal hypoplasia. Electron micrographs of cardiac myocytes showed striking mitochondrial hyperplasia, dispersing the sarcomeres. Special stains of frozen heart muscle showed an absence of complex IV (cytochrome c oxidase) in many of the myocytes. Both complexes I and IV of the respiratory chain were reduced in cardiac muscle. The A8344G mtDNA mutation was detected in both liver and cardiac muscle tissue. To our knowledge, this is the first description of the A8344G mtDNA mutation presenting as a sporadic case of fatal infantile cardiomyopathy and the first occurrence of this mutation associated with histiocytoid cardiomyopathy.

Novel point mutation (W184R) in neonatal type 2 Gaucher disease.

Gaucher disease is the most prevalent inherited sphingolipidosis and results from deficient glucocerebrosidase activity. Three clinical forms of Gaucher disease have been described: type 1, or non-neuronopathic; type 2, or acute neuronopathic; and type 3, or subacute neuronopathic. We have identified a novel mutation in a patient of Russian-British descent who died of type 2 Gaucher disease a few hours after birth. A heterozygous T --> C transition mutation in exon 6, cDNA nucleotide position 667, results in the substitution of tryptophan by arginine at amino acid residue 184 (W184R) of glucocerebrosidase. This mutation creates a new cleavage site for the restriction endonuclease Hinf1. We developed a method that utilizes Hinf1 restriction endonuclease analysis to confirm the presence of the mutation and test family members. The second mutation identified in the other glucocerebrosidase allele of the patient is mutation L444P, a severe mutation frequent in type 2 and 3 Gaucher disease. Since the patient died very shortly after birth, we postulate that the W184R/L444P genotype may result in little or no detectable glucocerebrosidase activity and thus a poor prognosis.

Identification of 6 new mutations in the iduronate sulfatase gene. Mutation in brief no. 233. Online.

Mucopolysaccharidosis type II (Hunter syndrome) is an X-linked lysosomal storage disorder caused by a deficiency of the enzyme iduronate-2-sulfatase. We sequenced genomic DNA and RT-PCR products in the iduronate sulfatase (IDS) gene in 6 unrelated patients with Hunter syndrome to assess genotype/phenotype relationships and offer carrier testing where required. Six novel mutations were identified: four missense mutations, one four-base pair deletion (596-599delAACA) and a cryptic splice site mutation. Three of the missense mutations were significant amino acid substitutions (S143F, S491F, E341K) of which the latter two involve amino acids conserved amongst sulfatase enzymes. The patients identified with these mutations all had a severe clinical phenotype. One missense mutation with a minimal amino acid substitution (H342Y), in a non-conserved region of the gene, was associated with a mild clinical phenotype. We identified a novel cryptic splice site (IVS5+934G>A) with some normal (wild type) mRNA processing. We predict that the normal mRNA product confered some residual functional enzyme, resulting in a mild phenotype associated with the absence of overt central nervous system disease.

D-2-Hydroxyglutaric aciduria: biochemical marker or clinical disease entity?

D-2-Hydroxyglutaric aciduria has been observed in patients with extremely variable clinical symptoms, creating doubt about the existence of a disease entity related to the biochemical finding. An international survey of patients with D-2-hydroxyglutaric aciduria was initiated to solve this issue. The clinical history, neuroimaging, and biochemical findings of 17 patients were studied. Ten of the patients had a severe early-infantile-onset encephalopathy characterized by epilepsy, hypotonia, cerebral visual failure, and little development. Five of these patients had a cardiomyopathy. In neuroimaging, all patients had a mild ventriculomegaly, often enlarged frontal subarachnoid spaces and subdural effusions, and always signs of delayed cerebral maturation. In all patients who underwent neuroimaging before 6 months, subependymal cysts over the head or corpus of the caudate nucleus were noted. Seven patients had a much milder and variable clinical picture, most often characterized by mental retardation, hypotonia, and macrocephaly, but sometimes no related clinical problems. Neuroimaging findings in 3 patients variably showed delayed cerebral maturation, ventriculomegaly, or subependymal cysts. Biochemical findings included elevations of D-2-hydroxyglutaric acid in urine, plasma, and cerebrospinal fluid in both groups. Cerebrospinal fluid gamma-aminobutyric acid was elevated in almost all patients investigated. Urinary citric acid cycle intermediates were variably elevated. The conclusion of the study is that D-2-hydroxyglutaric aciduria is a distinct neurometabolic disorder with at least two phenotypes.

DNA-based diagnosis of arylsulfatase A deficiencies as a supplement to enzyme assay: a case in point.

OBJECTIVE: To identify the molecular basis of arylsulfatase A deficiency in a family at risk for metachromatic leukodystrophy (MLD) and determine the genetic risk in the offspring. METHODS: Mutations in the arylsulfatase A gene were identified by PCR amplification and restriction enzyme digestion. Individuals had previously been tested for arylsulfatase A activity. RESULTS: Assays of arylsulfatase A activity had resulted in ambiguous results for MLD carrier identification. DNA analysis clearly identified two MLD mutations in the family, and an unsuspected arylsulfatase A pseudodeficiency. The DNA information immediately clarified the MLD risk for the family and confirmed that a newborn with low arylsulfatase A activity was unaffected. CONCLUSIONS: The overlap between activities for various combinations of MLD and pseudodeficiency alleles and the variability inherent in the assay of arylsulfatase A complicate the interpretation of activity levels in families at risk for MLD. Use of simple molecular biological tests for pseudodeficiency and the common MLD mutations in combination with the enzyme data can facilitate carrier identification and prenatal diagnosis.

An improved method for quantification of very long chain fatty acids in plasma.

Most peroxisomal disorders can be detected via analysis of very long chain fatty acids (VLCFA) and phytanic acid in plasma or serum. Previous methods utilizing gas-liquid chromatography (GLC) alone are time consuming and are hampered by interference from cholesterol derivatives. We describe here a GLC-mass spectrometry method for the simultaneous quantification of VLCFAs and phytanic acid. The method employs single ion monitoring with deuterated internal standards. We studied 38 normal controls and 12 patients with peroxisomal diseases and found complete discrimination between the two groups. Comparison with other methodology is discussed. We believe this to be a practical and accurate method for the quantification of both VLCFAs and phytanic acid in serum or plasma. It should be useful for laboratories involved in the diagnosis of biochemical disorders.