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Biomedical subjects

J van Blerkom

Publications and source records attributed to J van Blerkom.

7 recordsLinked to original sources
PubMed1996

The oxidizing agent tertiary butyl hydroperoxide induces disturbances in spindle organization, c-meiosis, and aneuploidy in mouse oocytes.

It has been recently proposed that a concomitant generation of oxidative stress of oocytes with increasing maternal age may be a major factor responsible for the age-related increase in aneuploid conceptions. As a preliminary step in the testing of this hypothesis, we need to confirm that oxidative stress in itself can induce errors in chromosome segregation. In order to achieve this goal, germinal vesicle (GV)-stage mouse oocytes from unstimulated ICR and (C57BL x CBA) F1 hybrid female mice were matured in vitro for 9 h for metaphase I (MI) oocytes or 16 h for metaphase II (MII) oocytes in the presence of varying concentrations of the oxidizing agent tertiary-butyl hydroperoxide (tBH). MII oocytes from (C57BL x CBA) F1 hybrid mice were fixed and C-banded for karyotyping analysis. MI and MII oocytes from ICR mice were fixed and stained with the DNA-fluorescent probe 4',6-diamidino-2-phenylindole (DAPI) to detect abnormalities in chromosomal distribution. Meiosis I and meiosis II spindles from ICR mice were visualized by confocal immunofluorescence microscopy. Data from these experiments demonstrate that in-vitro exposure of mouse oocytes to tBH during meiosis I reduces the length (pole-to-pole distance) and width (diameter at the equator of the spindle) of meiosis I and meiosis II spindles. This reduction is associated with an increase in the percentage of oocytes showing chromosome scattering and clumping on the MII plate, and of aneuploidy (hyperhaploidy) in MII oocytes. However, tBH at the concentrations used in the present study has only a minimal negative effect on the frequency of meiotic maturation. These results suggest that oxidative stress during meiotic maturation in vitro may induce chromosomal errors that are undetectable in the living oocyte and whose developmental consequences may become manifest after fertilization.

Aneuploidy↗
PubMed1983

Computer-assisted analysis demonstrates that polypeptides induced by natural and recombinant human interferon-alpha are the same and that some have related primary structures.

The biological effects on diploid and trisomy 21 human fibroblasts of pure human interferon IFLrA, a single IFN-alpha species produced from cloned DNA, were compared with those of partially purified natural IFN-alpha. Twelve interferon-induced polypeptides were visualized by two-dimensional gel electrophoresis and autoradiography. Seven of these were shown to have related primary structures and are therefore products of related genes or are related through post-translational modification. Qualitative visual comparisons and computer-aided quantitation of autoradiograms revealed no differences in the patterns of polypeptide induction following treatment with the two types of IFN-alpha, and the two interferons also induced (2'-5') oligoisoadenylate synthetase equally. By these criteria, the activities of the two interferons are qualitatively and quantitatively indistinguishable. In addition, the effects of trisomy 21 on IFLrA-induced polypeptide synthesis and on antiviral response were similar to those previously demonstrated with natural IFN-alpha.

Computers↗
PubMed1982

Implications of cytoskeletal interactions for cellular architecture and behavior.

The three major filamentous components of the cytoskeleton (microfilaments, microtubules and intermediate filaments) do not just coexist in the cell, but interact with each other in various ways. This paper discusses some examples of structural interactions visualized in critical-point-dried cells by stereoscopic high-voltage electron microscopy. The relative contribution of two classes of interactions to the consolidation of different cytoskeletal domains is considered. One class is represented by T-junctions (end-to-side contacts) of actin filaments with other filaments, and the other by 3 nm links. Attention is then turned to what may be called the behavioural consequences of cytoskeletal interactions. As an example of a coordinated interplay between events at the cell membrane and the cytoskeleton, we discuss changes in cytoskeletal organization of polymorphonuclear leucocytes upon stimulation with a chemotactic factor. These changes culminate in some cells in centriole separation and the establishment of two microtubule asters, each centred around a single centriole.

Actins↗
PubMed1981

Stabilization and the cytoplasmic ground substance in detergent-opened cells and a structural and biochemical analysis of its composition.

Treatment of epithelial BSC-1 cells with low concentrations of the detergent Brij 58 results in partial or complete removal of the plasmalemma and partial extraction of internal membrane-bound organelles without causing massive release of "cytosolic" proteins from the cytoplasmic ground substance. Stereoscopic high-voltage electron microscopy of such extracted and fixed cells demonstrates a system of slender (4-20 nm) strands in a three-dimensional "microtrabecular" arrangement similar to that observed in unextracted whole-mount preparations. Extraction of Brij-extracted cells with Triton X-100 dissolves many of the microtrabecular strands, leaving, as a more stable structure, a characteristic cytoskeletal network composed of various filaments and microtubules. Two-dimensional polyacrylamide gel electrophoresis of 35S-labeled polypeptides performed concurrently with the morphological studies demonstrates that Triton extraction of Brij-extracted cells releases a large number of polypeptides. This release parallels the loss of structural components observed by electron microscopy. Labeling of Brij-extracted cells with heavy meromyosin subfragment 1 decorates actin filaments with characteristic arrowhead complexes which are readily visualized only after subsequent Triton extraction. These observations support the concept that many cytoplasmic proteins are structure-bound and, in addition to the components comprising the cytoskeleton, are structure-forming. We conclude that a metastable association of various proteins of the cytoplasmic ground substance exists whose morphological integrity is maintained, at lest temporarily, after removal of the plasmalemma in solutions containing Brij 58.

Animals↗
PubMed1981

Structural interaction of cytoskeletal components.

Three-dimensional cytoskeletal organization of detergent-treated epithelial African green monkey kidney cells (BSC-1) and chick embryo fibroblasts was studied in whole-mount preparations visualized in a high voltage electron microscope. Stereo images are generated at both low and high magnification to reveal both overall cytoskeletal morphology and details of the structural continuity of different filament types. By the use of an improved extraction procedure in combination with heavy meromyosin subfragment 1 decoration of actin filaments, several new features of filament organization are revealed that suggest that the cytoskeleton is a highly interconnected structural unit. In addition to actin filaments, intermediate filaments, and microtubules, a new class of filaments of 2- to 3-nm diameter and 30- to 300-nm length that do not bind heavy merymyosin is demonstrated. They form end-to-side contacts with other cytoskeletal filaments, thereby acting as linkers between various fibers, both like (e.g., actin- actin) and unlike (e.g., actin-intermediate filament, intermediate filament-microtubule). Their nature is unknown. In addition to 2- to 3-nm filaments, actin filaments are demonstrated to form end-to-side contacts with other filaments. Y-shaped actin filament "branches" are observed both in the cell periphery close to ruffles and in more central cell areas also populated by abundant intermediate filaments and microtubules. Arrowhead complexes formed by subfragment 1 decoration of actin filaments point towards the contact site. Actin filaments also form end-to-side contacts with microtubules and intermediate filaments. Careful inspection of numerous actin-microtubule contacts shows that microtubules frequently change their course at sites of contact. A variety of experimentally induced modifications of the frequency of actin-microtubule contacts can be shown to influence the course of microtubules. We conclude that bends in microtubules are imposed by structural interactions with other cytoskeletal elements. A structural and biochemical comparison of whole cells and cytoskeletons demonstrates that the former show a more inticate three-dimensional network and a more complex biochemical composition than the latter. An analysis of the time course of detergent extraction strongly suggests that the cytoskeleton forms a structural backbone with which a large number of proteins of the cytoplasmic ground substance associate in an ordered fashion to form the characteristic image of the "microtrabecular network" (J.J. Wolosewick and K.R. Porter. 1979. J. Cell Biol. 82: 114-139).

Actins↗