ABSTRACT
Recurrent copy number variants (CNVs) are found in a significant proportion of patients with congenital heart disease (CHD) and some of these CNVs are associated with other developmental defects. In some syndromic patients, CHD may be the first presenting symptom, thus screening of patients with CHD for CNVs in specific genomic regions may lead to early diagnosis and awareness of extracardiac symptoms. We designed a multiplex ligation-dependent probe amplification (MLPA) assay specifically for screening of CHD patients. The MLPA assay allows for simultaneous analysis of CNVs in 25 genomic regions previously associated with CHD. We screened blood samples from 402 CHD patients and identified 14 rare CNVs in 13 (3.2%) patients. Five CNVs were de novo and six where inherited from a healthy parent. The MLPA screen led to early syndrome diagnosis in two of these patients. We conclude that the MLPA assay detects clinically relevant CNVs and suggest that it could be used within pediatric cardiology as a first tier screen to detect clinically relevant CNVs and identify syndromic patients at an early stage.
Subject(s)
DNA Copy Number Variations/genetics , Gene Dosage/genetics , Heart Defects, Congenital/genetics , Adolescent , Aged , Child , Child, Preschool , Chromosome Aberrations , Female , Heart Diseases/genetics , Humans , Infant , Infant, Newborn , Male , Multiplex Polymerase Chain Reaction/methods , Nucleic Acid Amplification TechniquesABSTRACT
Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is an autosomal recessive disease characterized by early infantile macrocephaly and delayed motor and cognitive deterioration. Magnetic resonance imaging (MRI) shows diffusely abnormal and swollen cerebral white matter and subcortical cysts. On follow-up, atrophy ensues. Approximately 80% of MLC patients have mutations in MLC1. We report 16 MLC patients without MLC1 mutations. Eight retained the classical clinical and MRI phenotype. The other 8 showed major MRI improvement. They lacked motor decline. Five had normal intelligence; 3 displayed cognitive deficiency. In conclusion, 2 phenotypes can be distinguished among the non-MLC1 mutated MLC patients: a classical and a benign phenotype.
Subject(s)
Cysts/genetics , Leukoencephalopathies/genetics , Leukoencephalopathies/pathology , Membrane Proteins/genetics , Mutation/genetics , Adolescent , Adult , Brain Diseases/complications , Brain Diseases/pathology , Child , Cysts/complications , DNA Mutational Analysis , Female , Follow-Up Studies , Humans , Leukoencephalopathies/complications , Magnetic Resonance Imaging/methods , Male , Retrospective Studies , Young AdultABSTRACT
Mutations in the gene MLC1 are found in approximately 80% of the patients with the inherited childhood white matter disorder megalencephalic leukoencephalopathy with subcortical cysts (MLC). Genetic linkage studies have not led to the identification of another disease gene. We questioned whether mutations in CLCN2, coding for the chloride channel protein 2 (ClC-2), are involved in MLC. Mice lacking this protein develop white matter abnormalities, which are characterized by vacuole formation in the myelin sheaths, strikingly similar to the intramyelinic vacuoles in MLC. Sequence analysis of CLCN2 at genomic DNA and cDNA levels in 18 MLC patients without MLC1 mutations revealed some nucleotide changes, but they were predicted to be nonpathogenic. Further, in electrophysiological experiments, one of the observed amino acid changes was shown to have no effect on the ClC-2-mediated currents. In conclusion, we found no evidence suggesting that the CLCN2 gene is involved in MLC.
Subject(s)
Chloride Channels/genetics , Alternative Splicing , Animals , CLC-2 Chloride Channels , Chloride Channels/chemistry , Chloride Channels/metabolism , DNA, Complementary/genetics , Dementia, Vascular/genetics , Female , Genetic Predisposition to Disease , Humans , In Vitro Techniques , Membrane Proteins/genetics , Mice , Mice, Knockout , Mutant Proteins/chemistry , Mutant Proteins/genetics , Mutant Proteins/metabolism , Mutation , Oocytes/metabolism , RNA, Messenger/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , XenopusABSTRACT
The presence of chromosome-specific low-copy repeats (LCRs) predisposes chromosome 22 to deletions and duplications. The current diagnostic procedure for detecting aberrations at 22q11.2 is chromosomal analysis coupled with fluorescence in situ hybridization (FISH) or PCR-based multiplex ligation dependent probe amplification (MLPA). However, there are copy number variations (CNVs) in 22q11.2 that are only detected by high-resolution platforms such as array comparative genomic hybridization (aCGH). We report on development of a high-definition MLPA (MLPA-HD) 22q11 kit that detects copy number changes at 37 loci on the long arm of chromosome 22. These include the 3-Mb region commonly deleted in DiGeorge/velocardiofacial syndrome (DGS/VCFS), the cat eye syndrome (CES) region, and more distal regions in 22q11 that have recently been shown to be deleted. We have used this MLPA-HD probe set to analyze 363 previously well-characterized samples with a variety of different rearrangements at 22q11 and demonstrate that it can detect copy number alterations with high sensitivity and specificity. In addition to detection of the common recurrent deletions associated with DGS/VCFS, variant and novel chromosome 22 aberrations have been detected. These include duplications within as well as deletions distal to this region. Further, the MLPA-HD detects deletion endpoint differences between patients with the common 3-Mb deletion. The MLPA-HD kit is proposed as a cost effective alternative to the currently available detection methods for individuals with features of the 22q11 aberrations. In patients with the relevant phenotypic characteristics, this MLPA-HD probe set could replace FISH for the clinical diagnosis of 22q11.2 deletions and duplications.