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Biomedical subjects

H Galjaard

Publications and source records attributed to H Galjaard.

At least 127 records · Page 7Linked to original sources
PubMed1980

Prenatal detection of a probable heterozygote for ADA deficiency and severe combined immunodeficiency disease using a microradioassay.

A pregnancy at risk for adenosine deaminase deficiency and severe combined immunodeficiency disease has been investigated by assay of adenosine deaminase activity in cultured amniotic fluid cells using a microradioassay. A low-normal level of activity consistent with heterozygote status in the foetus was found and confirmed after birth by assay of red cell and fibroblast adenosine deaminase activities. It is suggested that the radioassay method offers significant advantages in sensitivity and specificity over the standard spectrophotometric procedure.

Adenosine Deaminase↗
PubMed1980

Correction of I-cell defect by hybridization with lysosomal enzyme deficient human fibroblasts.

I-cell fibroblasts with a multiple intracellular lysosomal enzyme deficiency were hybridized with cells from patients with different types of single lysosomal enzyme defects. Fusion with G(M2) gangliosidosis, type 2, (Sandhoff disease) fibroblasts resulted in a restoration of the hexosaminidase activity, in a normalization of the electrophoretic mobility of the isoenzymes, and in a decreased activity in the medium. Fusion of I-cells with fibroblasts from G(M1) gangliosidosis, type 1, led to enhancement of beta-galactosidase (beta-gal) activity. This complementation must be the result of the presence of normal polypeptide chains in I-cells, whereas the other cell types provide a factor that causes the intracellular retention of the enzymes. Restoration of beta-gal was also observed in heterokaryons after fusion of I-cells with beta-galactosidase/neuraminidase-deficient (beta-gal(-)/neur(-)) variants, indicating that the neuraminidase(s) and the posttranslational modification of beta-gal are affected in a different way in I-cell disease and in beta-gal(-)/neur(-) variants. Fusion of I-cells with mannosidosis fibroblasts resulted in a restoration of the acidic form of alpha-mannosidase and in a decrease of the extracellular activity of both this enzyme and the hexosaminidase enzyme, indicating that fusion of I-cells with different types of fibroblasts with a single lysosomal enzyme deficiency not only leads to complementation for one particular enzyme but also to a correction of the basic defect in I-cells.

Cell Fusion↗
PubMed1979

Quantitative cytochemical analysis of (single) cultured cells.

Human fibroblasts or amniotic fluid cells can be cultivated in special dishes with a bottom of thin transparent plastic foil. After quick freezing and freeze-drying small groups of cultured cells or single cells can be isolated by dissecting small pieces of plastic foil under the stereomicroscope. These can be incubated in (sub)microlitre volumes of substrate and the absorbance, fluorescence or radioactivity can be measured and expressed per cell. When absorbance is measured with a microscope spectrophotometer in 1-10 microliter, microcuvettes, the minimum amount of coloured product that can be determined is of the order of 10(-11) moles. Incubation under paraffin oil in 0.05 microliter of substrate, followed by fluorescence measurements in 1-5 microliter with a microscope fluorometer, allows down to 10(-14) mol of methylumbelliferone, which is the end-product in many analyses of lysosomal enzymes, to be detected. (Re)cycling procedures have an even greater sensitivity (10(-15)-10(-19)mol) but they are more complicated and limited to NAD(P)(H)-dependent reactions. Several radiometric procedures have also been adapted for the analysis of small numbers of cultured cells but most of them still require 10(4)-10(5) human fibroblasts. These microtechniques allow rapid prenatal diagnosis of at least 20 different genetic metabolic diseases to be made within 7-14 days after amniocentesis at the 14th-16th week of pregnancy. Enzyme assays on single cultured cells have also enabled complementation studies to be done on heterokaryons after hybridization of different types of mutant fibroblasts, as well as investigations of the intercellular exchange of lysosomal enzymes between normal and mutant cells.

Amniotic Fluid↗
PubMed1979

Adult type GMl-gangliosidosis: a complementation study on somatic cell hybrids.

Two adult siblings with progressive pyramidal and extrapyramidal lesions, and generalized muscle atrophy had a profound deficiency of beta-galactosidase in all the cells and body fluids examined. Neuraminidase activity was normal in fibroblasts. The fused fibroblasts of infantile GMl-gangliosidosis and each of these adult patients had beta-galactosidase activity as expected for the average value in a mixture of equal numbers of parental cells. However, there was a remarkable increase in the activity of beta-galactosidase when the cells from each of these cases were fused with those from the beta-galactosidase-deficient adult with cherry-red spots, cerebellar ataxia, myoclonus and neuraminidase deficiency in fibroblasts. It was concluded that the two siblings represent a new genetic variant (adult type) of GMl-gangliosidosis.

Adult↗
PubMed1978

Lysosomal enzyme activities in different types of amniotic fluid cells measured by microchemical methods, combined with interference microscopy.

In primary amniotic fluid cultures, four distinct types of cells were characterized as epithelioid (E I and E II), fibroblast-like (F), And large cells, Small numbers (1-200) of freeze-dried cells were isolated from colonies of each cell type and analyzed for the activity of three lysosomal enzymes: beta-N-acetylglucosaminidase, beta-galactosidase, and alpha-glucosidase. When expressed per cell, the activities for each of the enzymes were not significantly different among the small types of cells (EI, EII, and F). However, 5 to 10-fold higher enzyme activities were found in the large cells. The dry mass of individual large cells, as measured by microinterferometry, was also 5 to 10 times higher than that of the smaller cell types. When expressed per unit of dry mass, the enzyme activities tested, appeared to be independent of the type of amniotic fluid cell. The significance of this observation for the rapid prenatal diagnosis of metabolic diseases is discussed.

Acetylglucosaminidase↗
PubMed1978

Biochemical, immunological, and cell genetic studies in glycogenosis type II.

Fibroblasts from patients with the adult, juvenile, and infantile form of glycogenosis type II (Pompe disease) were cultured under standardized conditions, and the activity of acid alpha-glucosidase (E.C.3.2.1.20) towards glycogen, maltose, and 4-methylumbelliferyl-alpha-D-glucopyranoside was measured. Glycogen levels in muscle biopsies and in cultured fibroblasts from patients were determined. Residual enzyme activities varying from 7%-22% were detected in fibroblasts from patients with the adult form but not from patients with the infantile form of glycogenosis II. An inverse correlation was found between the severity of the clinical manifestation and the degree of residual enzyme activity in the fibroblasts. The kinetic and electrophoretic properties of acid alpha-glucosidase in fibroblasts from the adult patients and from control individuals were similar. Immunological studies suggested that the decrease of acid alpha-glucosidase activity is caused by a mutation that affects the production or degradation of the enzyme rather than its catalytic activity. Complementation studies were carried out by fusing fibroblasts from patients with the adult, juvenile, and infantile form of glycogenosis II, but neither conventional assays on multikaryons nor enzyme assays on single binuclear heterokaryons gave any evidence for genetic heterogeneity among these forms.

Adolescent↗
PubMed1977

Characterization of residual hexosaminidase activity in Sandhoff's disease using man-Chinese hamster cell hybrids.

To obtain information about the nature of the residual hexosaminidase activity in Sandhoff's disease, hybrid cell lines between fibroblasts from a patient with Sandhoff's disease and Chinese hamster cells were isolated. In these hybrid cell lines, a heteropolymeric isoenzyme was detected that is composed of human alpha- and Chinese hamster hexosaminidase subunits. Due to the electrophoretic and immunological behavior of the heteropolymeric molecules in interspecies hybrids with normal fibroblasts and with cells from a patient with Sandhoff's disease, we conclude that Sandhoff cells contain an alpha-subunit of hexosaminidase with normal characteristics.

Animals↗
PubMed1977

An unexpected high frequency of trisomic fetuses in 229 pregnancies monitored for advanced maternal age.

In 229 pregnancies monitored because of advance maternal age, 16 (7%) abnormal fetal karyotypes were detected. We found 13 cases of trisomy 21, twice a trisomy 18, and once an additional marker chromosome. The frequency of abnormal fetal karyotypes in different maternal age groups was found to increase from 1:20 at 38--40 years, to 1:16 at 41--43 years, and finally to 1:4.5 in women of 44--46 years. The overall incidence of chromosomal aberrations and specifically of trisomy 21 is considerably higher than that described in retrospective studies.

Adult↗
PubMed1977

The effect of increased crypt cell proliferation on the activity and subcellular localization of esterases and alkaline phosphatase in the rat small intestine.

The activity and ultrastructural localization of alkaline phosphatase and esterase has been studied in normal rat intestine and after the increased crypt cell proliferation that occurs during recovery after 400 rad X-irradiation. Alkaline phosphatase activity is not present in crypt cells of normal intestine, but becomes apparent after the cell has migrated on to the villus. The enzyme is localized in the microvilli, along the lateral cell membranes and in dense bodies. Its activity increases 10 to 15-fold from the base to the tip of the villus. Morphometric analysis of the cell structures where this enzyme is localized reveals no marked changes in their relative proportions during crypt cell development. The expansion of the proliferative cell compartment along the whole length of the crypt which occurs during recovery after irradiation (72 hr after 400 rad X-irradiation) results in a marked reduction of alkaline phosphatase activity in the lower 10-15 cell positions at the base of the villus. During subsequent migration of these cells, the activity increases with cell age but normal values are not attained. From a morphometric analysis it was found that the ultrastructural development is similar to that in controls. These results suggest that during cell maturation, normal values for alkaline phosphatase activity are only attained after 10-12 hr period of maturation in a non-proliferative state and only after the cell has migrated on to the functional villus compartment. In normal intestine, esterase activity shows a 3-fold increase from the bottom to the tip of the crypt and 3 to 4-fold increase during migration up to the middle of the villus. Enzyme activity is localized in the endoplasmic reticulum, the dense bodies and the perinuclear space. Morphometric analyses reveal a 2 to 3-fold increase in the absolute size of these subcellular compartments during crypt cell differentiation and a 2-fold increase at the crypt-villus junction. The relative sizes increase 1-5-fold during crypt cell differentiation and at the time of transition of the cells on to the villus. Increased crypt cell proliferation after irradiation leads to a marked decrease in esterase activity both in crypts and villi. Morphometric analyses of electron micrographs indicate that these changes in activity are not related to any changes in the subcellular structures in which the enzyme is localized. It appears that the normal development of esterase activity depends both on the functional state of the cell and its localization in the crypt or villus.

Alkaline Phosphatase↗