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P Drochmans

Publications and source records attributed to P Drochmans.

At least 19 recordsLinked to original sources
PubMed1978

Structure and biochemical composition of desmosomes and tonofilaments isolated from calf muzzle epidermis.

Complexes of plasma membrane segments with desmosomes and attached tonofilaments were separated from the stratum spinosum cells of calf muzzle by means of moderately alkaline buffers of low ionic strength and mechanical homogenization. These structures were further fractionated by the use of various treatments including sonication, sucrose gradient centrifugation, and extraction with buffers containing high concentrations of salt, urea, citric acid, or detergents. Subfractions enriched in desmosome-tonofilament-complexes and tonofilament fragments were studied in detail. The desmosome structures such as the midline, the trilaminar membrane profile, and the desmosomal plaque appeared well preserved and were notably resistant to the various treatments employed. Fractions containing desmosome-tonofilament complexes were invariably dominated by the nonmembranous proteins of the tonofilaments which appeared as five major polypeptide bands (apparent molecular weights: 48,000; 51,000; 58,000; 60,000; 68,000) present in molar ratios of approx. 2:1:1:2:2. Four of these polypeptide bands showed electrophoretic mobilities similar to those of prekeratin polypeptides from bovine hoof. However, the largest polypeptide (68,000 mol wt) migrated significantly less in polyacrylamide gels than the largest component of the hoof prekeratin (approximately 63,000 mol wt). In addition, a series of minor bands, including carbohydrate-containing proteins, were identified and concluded to represent constituents of the desmosomal membrane. The analysis of protein-bound carbohydrates (total 270 microgram/mg phospholipid in desmosome-enriched subfractions) showed the presence of relatively high amounts of glucosamine, mannose, galactose, and sialic acids. These data as well as the lipid composition (e.g., high ratio of cholesterol to phospholipids, relatively high contents of sphingomyelin and gangliosides, and fatty acid pattern) indicate that the desmosomal membrane is complex in protein and lipid composition and has a typical plasma membrane character. The similarity of the desmosome-associated tonofilaments to prekeratin filaments and other forms of intermediate-sized filaments is discussed.

Animals↗
PubMed1977

Ultrastructural and metabolic studies of isolated and cultured hepatocytes.

Cells isolated from adult rat liver form distinct cell populations: the parenchymal cells or hepatocytes and the non-parenchymal cells, mainly endothelial and Kupffer cells. These two groups are easy to separate by centrifugation methods, including centrifugal elutriation. The single-cell suspension of hepatocytes seems to be constituted of subfractions which are located roughly in the centrilobular and perilobular regions of the liver lobule and are differentiated so that they form cell lines with distinct metabolic activities. The basic means of characterizing the isolated hepatocytes consists of determining their size distribution, their sedimentation and elutriation properties, and their metabolic activities. Cultures of hepatocytes offer the possibility of prolonging their survival and of studying the differentiation of new bile canaliculi and the reconstitution of biliary polarity in the cytoplasm. The combination of Kupffer cells with hepatocytes improves the culture condition, as a result of elimination of cell debris by phagocytosis by the non-parenchymal cells. The application of the isolation and partition methods to pre-neoplastic liver allows us to classify the cells according to their degree of hyperplasia and opens up a new field of investigation.

Animals↗
PubMed1977

Adult rat hepatocytes in primary monolayer culture. Ultrastructural characteristics of intercellular contacts and cell membrane differentiations.

Primary monolayer cultures were obtained in 60-mm petri dishes by incubating 3 X 10(6) isolated hepatocytes at 37 degrees C in Dulbecco's medium supplemented with 17% fetal calf serum. The ultrastructure of monolayer cells was examined after various incubation periods. Within 4 h of plating, the isolated spherical cells adhere to the plastic surface, establish their first contacts by numerous intertwined microvilli, and form new hemidesmosomes. After 12 h of culture, wide branched trabeculae of flattened polyhedral cells extend in all directions. Finally, after 24 h of culture, bile canaliculi are reconstituted, and a biliary polarity is recovered: the Golgi elements, which are scattered throughout the cytoplasm in the isolated cells, are reassembled in front of the newly formed bile canalculi, symmetrically in the adjacent cells; lysosomes are concentrated in that region, and microtubules reappear. Concomitantly, plasma membrane differentiations, namely desmosomes and tight junctions, develop. Tight junctions sealing the bile ducts constitute a barrier to the passage of ruthenium red and horseradish peroxidase. De novo formation of these junctions was studied by the freeze-etching technique: 10-nm particles compose a network of anastomosed linear arrays in the vicinity of the bile canaliculi; in the next step of differentiation, the particles fuse, form short ridge segments and finally continuous branched smooth strands, characteristic of the mature tight junction.

Cell Adhesion↗
PubMed1977

Effect of insulin on ultrastructure and glycogenesis in primary cultures of adult rat hepatocytes.

Insulin in the presence of high concentrations of glucose has a beneficial trophic effect on the development of primary cultures of hepatocytes. Compared to the situation observed in hormone-free control cultures, the flattening of the reaggregated hepatocytes is enhanced, and the reconstituted cell trabeculae are enlarged and tend to form a confluent monolayer after 3 days; the survival time is prolonged from 3 to 5 or 6 days. Ultrastructural modifications are also initiated by insulin; numerous glycogen particles appear after 24 h, in between the cisternae of the proliferated smooth endoplasmic reticulum. After 48 h, large amounts of glycogen are stored, and numerous polysomes are present. A small number of cells showed an increased synthesis of lipid droplets in the lumen of the smooth endoplasmic reticulum and form liposomes at the same time. After 72 h, cytolysomes filled with glycogen develop, simulating glycogenosis type II. Simultaneously, microtubules and microfilaments, closely related to numerous polysomes, appear in cytoplasmic extensions constituting undulating membranes. The biochemical data demonstrate that, in the absence of insulin, a high concentration of glucose stimulates glycogenesis and hinders glycogenolysis. This effect of glucose on polysaccharide synthesis is progressively lost. The addition of insulin to the culture induces after 48 and 72 h, a three- to fivefold increase of the glucose incorporation into glycogen, as compared to the controls. The presence of insulin is required to maintain the hepatocyte's capacity to store glycogen. Glycogen synthetase is converted into its active form under the influence of glucose. Insulin increases the rate of activation.

Cells, Cultured↗
PubMed1975

Ultrastructural aspects of muscle and nerve in Werdnig-Hoffmann disease.

The authors describe in biopsies from 6 cases of Werdnig-Hoffmann disease, including 2 of the more benign type, the ultrastructural typical aspects of denervation. They compare their findings with those of other workers. The striking points are the great variation in the diameter of the muscle fibres and the myofibrils, the disorganisation of the myofibrils, the sarcomeres and the filaments, with persistance of the relations between thick and thin filaments at various levels, the modifications of the Z-band and the triads in chains. The folds and the basement membrane are examined. Centrioles are present in a muscle fibre and in a satellite. Glycogen is very abundant. The nerves seem normal but some Schwann cells contain pi granules which are not observed usually at the age of the patient. The end plates and a muscle spindle are normal.

Basement Membrane↗
PubMed1975

Isolation and subfractionation on ficoll gradients of adult rat hepatocytes. Size, morphology, and biochemical characteristics of cell fractions.

The recirculating perfusion of adult rat liver with a Ca-++-free Hanks' solution produces a release of the adhesiveness of cells and a cleaving of the desmosomes. The addition of collagenase and hyaluronidase to the perfusion medium leads to complete dissociation of the liver tissue into a mixture of isolated cells and cell cords in which the hepatocytes remain connected with specific junctional differentiations, namely the gap and tight junctions. Individual cells are released by submitting the suspension of cell trabeculae to a gentle rolling. The gap junctions are ruptured at least in one of the two adjacent cells and remain generally attached to the other cell taking with them a small portion of cytoplasm. This technique of isolation of hepatocytes yields about 60-65% of the parenchymal cells contained in a liver; endothelial cells and other cells of the connective tissue are not recovered. The ultrastructural preservation of the isolated hepatocytes is excellent and the glucose-6-phosphatase activity, confined to the endoplasmic reticulum, appears unaltered in most cells. Protein, DNA and RNA recovery in the preparations of isolated hepatocytes is satisfactory, amounting to 70% of that found in liver homogenate; glycogen, the most labile component examined, is partly lost or degraded during the manipulations. Cell diameters measured by different methods confirm the preservation of the original volume of the in situ hepatocytes and the presence of more than one type of parenchymal cell. By submitting this heterogeneous cell population to an isopycnic density gradient centrifugation, two types of hepatocytes can be distinguished: the light hepatocytes, with a mean diameter of 20.5 mum and a mean density of 1.10, are characterized by an extended smooth-walled endoplasmic reticulum entrapping dispersed alpha-glycogen particles; the heavy hepatocytes, with a mean diameter of 19.0 mum and a mean density of 1.14, present a relatively reduced compartment of smooth endoplasmic reticulum, but large accumulations of glycogen. It is suggested that the cell fraction of low density is enriched in centrolobular cells and the high density fraction in perilobular hepatocytes.

Animals↗
PubMed1975

Isolation of centrolobular and perilobular hepatocytes after phenobarbital treatment.

Daily phenobarbital (PB) injections, on 3-7 consecutive days, induce an intense proliferation of smooth endoplasmic reticulum (ER) associated with a decrease of the glucose-6-phosphatase activity. This situation first affects the centrolobular hepatocytes, enhancing the degree of liver lobule heterogeneity. This experimental model was used for isolation and further subfractionation of hepatocytes on Ficoll density gradients, as described in the preceding paper. Profiles of protein, DNA, RNA, glycogen, phosphorylase, and glucose-6-phosphatase were determined all along the gradient. Two liver cell populations were distinguished: (a) light hepatocytes (mean density 1.10) present the same morphological characteristics as centrolobular cells, i.e., an abundant smooth ER composed of tubular elements, numerous small mitochondria, and few glycogen particles; (b) heavy hepatocytes (mean density 1.14) are characterized by large and compact glycogen areas and prominent rough endoplasmic cisternae, as are the perilobular cells. After incubation in the Wachstein-Meisel medium, Centrolobular hepatocytes exhibit dispersed reaction sites of glucose-6-phosphatase activity, whereas perilobular cells present a continuous and intense reaction. Morphometric determinations were carried out for both cell populations. Centrolobular PB hepatocytes are considerably enlarged (mean diameter: 23.7 mum); perilobular hepatocytes have a significantly smaller mean diameter of 19.2 mum, which is close to the values of control liver cells.

Animals↗