Birgit Uhlenberg

Mutations in antiquitin in individuals with pyridoxine-dependent seizures

We show here that children with pyridoxine-dependent seizures (PDS) have mutations in the ALDH7A1 gene, which encodes antiquitin; these mutations abolish the activity of antiquitin as a Δ1-piperideine-6-carboxylate (P6C)–α-aminoadipic semialdehyde (α-AASA) dehydrogenase. The accumulating P6C inactivates pyridoxal 5′-phosphate (PLP) by forming a Knoevenagel condensation product. Measurement of urinary α-AASA provides a simple way of confirming the diagnosis of PDS and ALDH7A1 gene analysis provides a means for prenatal diagnosis.

Mutations in the Gene Encoding Gap Junction Protein α12 (Connexin 46.6) Cause Pelizaeus-Merzbacher–Like Disease

The hypomyelinating leukodystrophies X-linked Pelizaeus-Merzbacher disease (PMD) and Pelizaeus-Merzbacher-like disease (PMLD) are characterized by nystagmus, progressive spasticity, and ataxia. In a consanguineous family with PMLD, we performed a genomewide linkage scan using the GeneChip Mapping EA 10K Array (Affymetrix) and detected a single gene locus on chromosome 1q41-q42. This region harbors the GJA12 gene, which encodes gap junction protein alpha 12 (or connexin 46.6). Gap junction proteins assemble into intercellular channels through which signaling ions and small molecules are exchanged. GJA12 is highly expressed in oligodendrocytes, and, therefore, it serves as an excellent candidate for hypomyelination in PMLD. In three of six families with PMLD, we detected five different GJA12 mutations, including missense, nonsense, and frameshift mutations. We thereby confirm previous assumptions that PMLD is genetically heterogeneous. Although the murine Gja12 ortholog is not expressed in sciatic nerve, we did detect GJA12 transcripts in human sciatic and sural nerve tissue by reverse-transcriptase polymerase chain reaction. These results are in accordance with the electrophysiological finding of reduced motor and sensory nerve conduction velocities in patients with PMLD, which argues for a demyelinating neuropathy. In this study, we demonstrate that GJA12 plays a key role in central myelination and is involved in peripheral myelination in humans.

Polyoma virus‐associated progressive multifocal leukoencephalopathy after renal transplantation: Regression following withdrawal of mycophenolate mofetil

Weber SC, Uhlenberg B, Raile K, Querfeld U, Müller D. Polyoma virus‐associated progressive multifocal leukoencephalopathy after renal transplantation: Regression following withdrawal of mycophenolate mofetil. Pediatr Transplantation 2011: 15:E19–E24.

Analysis of human alternative first exons and copy number variation of the GJA12 gene in patients with Pelizaeus–Merzbacher-like disease†

Pelizaeus–Merzbacher‐like disease (PMLD) is a heterogeneous disease with primary hypomyelination of the central nervous system. Only the minority of patients have mutations in the coding region of the GJA12 gene encoding gap junction protein alpha 12, a subunit of intercellular channels highly expressed by oligodendrocytes, the myelin forming cells of the central nervous system. No other gene has been found so far to be mutated in PMLD besides GJA12. We therefore extended the mutational screening in the GJA12 gene, searched for alternative first exons—as described in mice—determined the human 5′‐end of the gene, screened therein for mutations and analyzed for copy number variations of the GJA12 gene in 14 patients with PMLD. Unlike in mice we did not find alternative first exons but detected a unique 79 bp first exon in human adolescent brain and spinal cord. No mutation in this non‐coding region was found in our cohort. Copy number variation of the GJA12 gene was assessed by real‐time PCR TaqMan® gene expression technology, but neither patient showed an aberrant copy number. These data confirm that GJA12 alterations are a rare cause of PMLD—even after extending the screening for copy number variation and for mutations in the non‐coding region of GJA12. Full genome scans in informative families and further screenings of candidate genes are feasible approaches to elucidate the genetic background of the majority of patients with PMLD.

Atypical tetanus in a completely immunized 14-year-old boy.

We report the uncommon clinical course of tetanus in a completely immunized 14-year-old boy. His initial symptoms, which included a flaccid paralysis, supported a diagnosis of botulism. Preliminary mouse-test results with combined botulinum antitoxins A, B, and E, obtained from tetanus-immunized horses, backed this diagnosis. The change in his clinical course from paralysis to rigor and the negative, more specific, botulinum mouse test with isolated botulinum antitoxins A, B, and E, obtained from nonvaccinated rabbits, disproved the diagnosis of botulism. Tetanus was suspected despite complete vaccination. The final results of a positive mouse test performed with isolated tetanus antitoxin confirmed the diagnosis. Adequate treatment was begun, and the boy recovered completely.

Genome-wide linkage analysis is a powerful prenatal diagnostic tool in families with unknown genetic defects

Genome-wide linkage analysis is an established tool to map inherited diseases. To our knowledge it has not been used in prenatal diagnostics of any genetic disorder. We present a family with a severe recessive mental retardation syndrome, where the mother wished pregnancy termination to avoid delivering another affected child. By genome-wide scanning using the Affymetrix (Santa Clara, CA, USA) 10k chip we were able to establish the disease haplotype. Without knowing the exact genetic defect, we excluded the condition in the fetus. The woman finally gave birth to a healthy baby. We suggest that genome-wide linkage analysis – based on either SNP mapping or full-genome sequencing – is a very useful tool in prenatal diagnostics of diseases.

Epithelioid hemangioendothelioma of the lung presenting with pneumonia and heart rhythm disturbances in a teenage girl.

The epithelioid hemangioendothelioma (EHE) is a rare low-grade tumor of vascular origin that may arise at any site. However, lung and liver represent the 2 main locations. Symptoms of the pulmonary EHE are usually nonspecific and mild. Distant metastases of PEHE are frequent. However, heart metastases have only been reported in connection with primary EHE of the liver. We describe the case of a 15-year-old girl presenting with an abscess forming pneumonia and severe rhythm disturbances associated with an EHE of the lung. The untypical fulminant clinical course, the surgical interventions, and the involvement of the heart as a life threatening complication, eventually on the basis of cardiac metastases of PEHE, are emphasized.

Oligodendroglial transcription factor (OLIG1 and OLIG2) mutations are not associated with pelizaeus-merzbacher-like leukodystrophy

The human phenotype with primarily impaired myelination is represented by hypomyelinating leukodystrophies. The most frequent form is Pelizaeus–Merzbacher disease, which is due to alterations in the PLP1 gene encoding the major myelin protein. Another form, Pelizaeus–Merzbacher‐like disease, is partly associated with mutations in the GJA12 gene encoding gap junction protein alpha 12, but seems to be heterogeneous. Olig1 and Olig2 are transcription factors in oligodendrocyte development. We postulated that disturbed oligodendroglial maturation could be associated with primary hypomyelination in humans and analyzed the coding sequence of OLIG1 and OLIG2 in 13 patients from 12 unrelated families which were thoroughly characterized with regard to phenotype and magnetic resonance imaging results. From our findings we conclude that mutations in OLIG1 and OLIG2 are not likely to be associated with this subgroup of hypomyelinating disorders.

Atypical manifestation of childhood primary cerebral lymphoma restricted to the leptomeninges.

We present the atypical manifestation of a primary cerebral lymphoma in childhood due to exclusive affection of the leptomeninges. Retrospectively, adequate treatment was delayed by early administration of glucocorticoids before the histological diagnosis was confirmed. The patient was an otherwise healthy 8-year-old boy who complained of recurrent headaches. On account of a left facial paresis and detection of Borrelia IgM antibodies in serum (CSF was not analysed at this time) he was treated on an outpatient basis with oral penicillin and cortisone for 2 months and full recovery of his facial paresis was achieved. One year later he again reported headaches, a cerebral CT scan was normal and because of an elevated Borrelia IgM level in CSF (18 cells/ll, exclusively lymphoblasts, protein 78 mg/dl, glucose 30 mg/dl) he was treated with cephalosporins for recurrent neuroborreliosis. One month later he displayed right facial paresis, a cerebral MRI scan was normal and a mildly elevated cell count was found in CSF (55 cells/ll, protein 143 mg/dl, glucose 44 mg/dl). Because of his reduced general condition, MRI with contrast medium was done again but found to be uninformative. CSF displayed 60 cells/ll (again exclusively lymphocytes, no lymphoblasts), CSF glucose had fallen to 19 mg/dl and protein was 160 mg/dl. Despite a negative tuberculosis polymerase-chain-reaction in CSF but because of progressively low glucose in CSF, tuberculostatic treatment was started, including rifampicin, pyracinamide, ethambutol, isoniazid and dexamethasone. Two weeks later he developed generalised seizures, became tetraparetic and mechanical ventilation was started. Intravenous amphotericin B was added, spinal MRI and bone marrow histology were normal. Tetraparesis was interpreted as a meningeal compression close to the anterior horn cells and lead to a biopsy of the leptomeninges and underlying cortex yielding the definite diagnosis of primary Burkitt-like lymphoma. The family decided against chemotherapy and the boy died 20 months after initial symptoms. We present the rare case of a primary cerebral B-cell lymphoma in which the appropriate diagnosis was delayed by its atypical presentation due to mere leptomeningeal affection. Sole leptomeningeal affection and dissemination were clearly shown by histological investigations (Fig. 1). To our knowledge, histological proof of mere leptomeningeal affection has not been published so far. Clinical symptoms of cerebral lymphoma usually comply with a fast growing tumour and signs of elevated cranial pressure [5]. However, atypical presentations have rarely been reported and include acute blindness as the solely presenting sign [4]. Additionally, early diagnosis was delayed by administration of glucocorticoids known to lead to a transient amelioration of neurological deficits, marked shrinkage of the tumour as well as B. Uhlenberg (&) Æ A. von Moers Department of Neuropaediatrics, Charité University Medical School, Augustenburger Platz 1, 13353 Berlin, Germany E-mail: birgit.uhlenberg@charite.de Tel.: +49-30-450566112 Fax: +49-30-450566920

Mutations in RARS cause a hypomyelination disorder akin to Pelizaeus–Merzbacher disease

Pelizaeus-Merzbacher disease (PMD) is a rare Mendelian disorder characterised by central nervous system hypomyelination. PMD typically manifests in infancy or early childhood and is caused by mutations in proteolipid protein-1 (PLP1). However, variants in several other genes including gap junction protein gamma 2 (GJC2) can also cause a similar phenotype and are referred to PMD-like disease (PMLD). Whole-exome sequencing in two siblings presenting with clinical symptoms of PMD revealed a homozygous variant in the arginyl-tRNA synthetase (RARS) gene: NM_002887.3: c.[5A>G] p.(Asp2Gly). Subsequent screening of a PMD cohort without a genetic diagnosis identified an unrelated individual with novel compound heterozygous variants including a missense variant c.[1367C>T] p.(Ser456Leu) and a de novo deletion c.[1846_1847delTA] p.(Tyr616Leufs*6). Protein levels of RARS and the multi-tRNA synthetase complex into which it assembles were found to be significantly reduced by 80 and 90% by western blotting and Blue native-PAGE respectively using patient fibroblast extracts. As RARS is involved in protein synthesis whereby it attaches arginine to its cognate tRNA, patient cells were studied to determine their ability to proliferate with limiting amounts of this essential amino acid. Patient fibroblasts cultured in medium with limited arginine at 30 °C and 40 °C, showed a significant decrease in fibroblast proliferation (P<0.001) compared to control cells, suggestive of inefficiency of protein synthesis in the patient cells. Our functional studies provide further evidence that RARS is a PMD-causing gene.