224 research outputs found
Release and uptake of lysosomal enzymes : studied in cultured cells
The purpose of the experimental work described in this thesiswas to investigate some aspects of the release and uptake
of lysosomal enzymes. The experiments involved the use of
normal human and animal fibroblasts and some other cell
types such as hepatocytes and hepatoma cells as sources of
hydrolytic enzymes, and fibroblasts from patients with lysosomal
storage diseases associated with a single lysosomal
enzyme deficiency and with "1-cell" disease as recipient
cells. In a number of studies co-cultivation of normal and
mutant cells was applied. as a means to study the release
and uptake of lysosomal enzymes at physiological concentrations.
We have also studied some characteristics of released
lysosomal enzyme activities in· normal and mutant cell cultures
in comparison with the corresponding intracellular
activities. Finally, we have used somatic cell hybridization
to investigate the genetic background of the metabolic abnormalities
in some of the human mutant cells. Our studies
have provided information on the normal transport of lysosomal enzymes within the cell and between cells. Furthermore,
we hope that more knowledge about recognition and
uptake of lysosomal enzymes by deficient cells from patients
with lysosomal storage diseases will have future implications
for enzyme therapy
The cystic fibrosis defect approached from different angles - New perspectives on the gene, the chloride channel, diagnosis and therapy
Abstract
The search for the basic defect in cystic fibrosis (CF) has reached a decisive stage since the recent identification of the responsible gene. Electrophysiological and biochemical research had defined the CF defect as a dysregulation of epithelial chloride channels. The putative protein product of the now identified gene shares properties with other known transport proteins, but it is not necessarily itself a chloride channel protein. Elucidation of the primary cellular defect will certainly have important aetiological and hopefully therapeutic implications. The identification of the major gene mutation already has significant consequences for genetic counselling and prenatal diagnosis. Heterozygote detection at the population level awaits identification of the probably heterogenous mutations on about 30% of the CF chromosomes. At present, about 50% of CF patients are homozygous for the recently identified major CF mutation
Characterization of the cytosolic tuberin-hamartin complex. Tuberin is a cytosolic chaperone for hamartin
Tuberous sclerosis (TSC) is an autosomal dominant disorder characterized
by a broad phenotypic spectrum that includes seizures, mental retardation,
renal dysfunction and dermatological abnormalities. Mutations to either
the TSC1 or TSC2 gene are responsible for the disease. The TSC1 gene
encodes hamartin, a 130-kDa protein without significant homology to other
known mammalian proteins. Analysis of the amino acid sequence of tuberin,
the 200-kDa product of the TSC2 gene, identified a region with limited
homology to GTPase-activating proteins. Previously, we demonstrated direct
binding between tuberin and hamartin. Here we investigate this interaction
in more detail. We show that the complex is predominantly cytosolic and
may contain additional, as yet uncharacterized components alongside
tuberin and hamartin. Furthermore, because oligomerization of the hamartin
carboxyl-terminal coiled coil domain was inhibited by the presence of
tuberin, we propose that tuberin acts as a chaperone, preventing hamartin
self-aggregation
Combined cardiological and neurological abnormalities due to filamin A gene mutation
Background: Cardiac defects can be the presenting symptom in patients with mutations in the X-linked gene FLNA. Dysfunction of this gene is associated with cardiac abnormalities, especially in the left ventricular outflow tract, but can also cause a congenital malformation of the cerebral cortex. We noticed that some patients diagnosed at the neurogenetics clinic had first presented to a cardiologist, suggesting that earlier recognition may be possible if the diagnosis is suspected. Methods and results: From the Erasmus MC cerebral malformations database 24 patients were identified with cerebral bilateral periventricular nodular heterotopia (PNH) without other cerebral cortical malformations. In six of these patients, a pathogenic mutation in FLNA was present. In five a cardiac defect was also found in the outflow tract. Four had presented to a cardiologist before the cerebral abnormalities were diagnosed. Conclusions: The cardiological phenotype typically consists of aortic or mitral regurgitation, coarctation of the aorta or other left-sided cardiac malformations. Most patients in this category will not have a FLNA mutation, but the presence of neurological complaints, hyperlaxity of the skin or joints and/or a family history with similar cardiac or neurological problems in a possibly X-linked pattern may alert the clinician to the possibility of a FLNA mutation
Array-Based FMR1 Sequencing and Deletion Analysis in Patients with a Fragile X Syndrome–Like Phenotype
Background: Fragile X syndrome (FXS) is caused by loss of function mutations in the FMR1 gene. Trinucleotide CGG-repeat expansions, resulting in FMR1 gene silencing, are the most common mutations observed at this locus. Even though the repeat expansion mutation is a functional null mutation, few conventional mutations have been identified at this locus, largely due to the clinical laboratory focus on the repeat tract. Methodology/Principal Findings: To more thoroughly evaluate the frequency of conventional mutations in FXS-like patients, we used an array-based method to sequence FMR1 in 51 unrelated males exhibiting several features characteristic of FXS but with normal CGG-repeat tracts of FMR1. One patient was identified with a deletion in FMR1, but none of the patients were found to have other conventional mutations. Conclusions/Significance: These data suggest that missense mutations in FMR1 are not a common cause of the FXS phenotype in patients who have normal-length CGG-repeat tracts. However, screening for small deletions of FMR1 may be of clinically utility
Angelman syndrome in an inbred family
Angelman syndrome (AS) is characterized by severe mental retardation, absent speech, puppet-like movements, inappropriate laughter, epilepsy, and abnormal electroencephalogram. The majority of AS patients (≃ 65%) have a maternal deficiency within chromosomal region 15q11-q13, caused by maternal deletion or paternal uniparental disomy (UPD). Approximately 35% of AS patients exhibit neither detectable deletion nor UPD, but a subset of these patients have abnormal methylation at several loci in the 15q11-q13 interval. We describe here three patients with Angelman syndrome belonging to an extended inbred family. High resolution chromosome analysis combined with DNA analysis using 14 marker loci from the 15q11-q13 region failed to detect a deletion in any of the three patients. Paternal UPD of chromosome 15 was detected in one case, while the other two patients have abnormal methylation at D15S9, D15S63, and SNRPN. Although the three patients are distantly related, the chromosome 15q11-q13 haplotypes are different, suggesting that independent mutations gave rise to AS in this family
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