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Biomedical subjects

M R Bernfield

Publications and source records attributed to M R Bernfield.

At least 19 recordsLinked to original sources
PubMed1979

Collagen reduces glycosaminoglycan degradation by cultured mammary epithelial cells: possible mechanism for basal lamina formation.

Collagenous substrates are reported to promote the accumulation of extracellular matrix materials by epithelia in culture. Glycosaminoglycan (GAG) metabolism is compared in secondary cultures of mouse mammary epithelial cells maintained on plastic or type I collagen gel substrates. The incorporation of 35SO42- into GAG during brief labeling indicates no difference between substrates in the rate of GAG synthesis. During prolonged labeling, however, accumulation of [35S]GAG in cultures on colllagen exceeds that of cultures on plastic. This increased accumulation is due to a markedly reduced rate of GAG degradation. GAG degradation does not occur in the medium, indicating that degradation is localized to the cells. The cultures on collagen contain a slowly degrading cell-associated [35S]GAG pool and a ruthenium red-stained basal lamina, neither of which is present in cultures on plastic. The cell-associated [35S]GAG in cultures on collagen is, in part, localized to the site of the ultrastructurally identified basal lamina. Formation of the basal lamina, therefore, may result from collagen-mediated reduction in the degradation of GAG-containing molecules.

Animals↗
PubMed1978

Acquisition of synchronous beating between embryonic heart cell aggregates and layers.

Synchronous beating between chick embryonic heart cell aggregates and heart cell layers was used to study the relationship between intercellular adhesion and ionic coupling. Adhesion was measured by counting the proportion of aggregates which were not to be removed from cell layers by gentle washing after a 30 min incubation. Synchrony between bound aggregates and contiguous layers was assessed by phase microscopy. The first evidence of synchrony was seen 1.5 h after addition of aggregates to layers, following which there was an increase in the percentage of aggregates beating synchronously, reaching over 50% at 7 h and slowly increasing to a maximum of 65% by 24 h. Scanning electron microscopy and autoradiography of thymidine-labeled cells suggest that synchrony does not depend on cell movement at the interface between aggregate and layer. Acquisition of synchrony can be prevented completely by inhibiting protein synthesis, although pulsation of aggregates and layers continues in proportions unchanged from controls. After reversal of protein synthesis inhibition, synchrony is acquired at a rate and to an extent closely resembling that of newly adherent controls. These data indicate that ionic coupling is neither an inevitable nor an immediate consequence of adhesion. Since ionic coupling has been shown to correlate with the presence of gap junctions, the findings suggest that gap junctions are not involved in the initial events responsible for intercellular adhesion in vitro and that their formation following adhesion in this system may depend upon protein synthesis.

Animals↗
PubMed1978

Morphological and functional correlates of synchronous beating between embryonic heart cell aggregates and layers.

We have examined correlations between morphological and functional evidence of cell coupling between aggregates of beating embryonic heart cells and underlying layers. Synchronously beating aggregate-layer pairs were compared with asynchronous pairs. Intracellular microelectrode studies demonstrated that asynchronously beating aggregate-layers could not be induced to beat synchronously by electrical stimulation of the aggregate, whereas 86% of synchronous instances showed propagation of stimulating current pulses from aggregate to layer. By freeze fracture we have found significant differences both in the number and in the total area of gap junctions between the aggregate-layer interfaces of synchronous and asynchronous preparations. The data suggest that synchronous beating is a reliable functional indication of effective ionic coupling, and requires a certain area and number of gap junction/cell.

Action Potentials↗
PubMed1977

Basal lamina of embryonic salivary epithelia. Production by the epithelium and role in maintaining lobular morphology.

The role of the basal lamina in maintaining the normal morphology of mouse embryo submandibular epithelia was assessed by examining its production as well as the cellular and organ culture changes associated with its removal and replacement. The lamina was removed from epithelia isolated free of mesenchyme by brief treatment with testicular hyaluronidase in the absence of calcium. The treatment causes rounding-up of the cells, loss of cellular cohesion, appearance of microvilli, and changes in the organization of cytoskeletal structures. The lamina is not removed and the cellular alterations do not occur in the absence of hyaluronidase in calcium-free medium or when both enzyme and calcium are present, possibly because digestion of chondroitin sulfate, a component of the lamina, is inhibited by calcium. Within 2 h after treatment, in the absence of mesenchyme or biological substrata, the epithelia deposits a new lamina, which is identical by several criteria to the preexisting lamina, and reverses the cellular alterations. Epithelia treated with hyaluronidase lose lobular morphology during culture with mesenchyme. Delaying culture with mesenchyme, to allow restoration of the lamina and of normal cellular architecture, prevents the loss of lobular morphology. The results indicate that the basal lamina imposes morphologic stability on the epithelium, while the mesenchyme apparently affects processes involved in changes in morphology, possibly by selective degradation of the basal lamina.

Animals↗
PubMed1977

Basal lamina of embryonic salivary epithelia. Nature of glycosaminoglycan and organization of extracellular materials.

The ultrastructural organization and the composition of newly synthesized glycosaminoglycan (GAG) in the epithelial basal lamina of mouse embryo submandibular glands were assessed. The labeled GAG accumulating in the lamina is distinct from that in its tissue of origin, the epithelium, or from that in the surrounding mesenchyme. In the lamina, hyaluronic acid accounts for approximately 50% of the labeled GAG, chondroitin-4-sulfate is twice the chondroitin-6-sulfate, and there is a low proportion of chondroitin. This composition is constant regardless of whether the lamina is labeled by whole glands or, in the absence of mesenchyme, by isolated epithelia retaining a lamina and by isolated epithelia generating a lamina de novo. The results andicate that the labeled GAG are bona fide components of the lamina, and suggest that laminar GAG is deposited in units of constant composition. Ultrastructural observations following ruthenium red staining or tannic acid fixation extablish that the lamina is a highly ordered specialization of the basal cell surface. Discrete structures in macroperiodic arrays apparently attached to the plasmalemma are visualized. This organization is seen in intact glands and in the laminae produced by epithelia in the absence of mesenchyme or biological substrate. The data are interpreted as indicating that the basal lamina contains supramolecular complexes of hyaluronic acid and proteoglycan which are organized into an extracellular scaffolding which imposes structural form on the epithelium.

Animals↗
PubMed1976

Relationship of transformation, cell density, and growth control to the cellular distribution of newly synthesized glycosaminoglycan.

Mouse 3T3 cells and their Simian Virus 40-transformed derivatives (3T3SV) were used to assess the relationship of transfromation, cell density, and growth control to the cellular distribution of newly synthesized glycosaminoglycan (GAG). Glucosamine- and galactosamine-containing GAG were labeled equivalently by [3H=A1-glucose regardless of culture type, allowing incorporation into the various GAG to be compared under all conditions studied. Three components of each culture type were examined: the cells, which contain the bulk of newly synthesized GAG and are enriched in chondroitin sulfate and heparan sulfate; cell surface materials released by trypsin, which contain predominantly hyaluronic acid; and the media , which contain predominantly hyaluronic acid and undersulfated chondroitin sulfate. Increased cell density and viral transformation reduce incorporation into GAG relative to the incorporation into other polysaccharides. Transformation, however, does not substantially alter the type or distribution of newly synthesized GAG; the relative amounts and cellular distributions were very similar in 3T3 and 3T3SV cultures growing at similar rates at low densities. On the other hand, increased cell density as well as density-dependent growth inhibition modified the type and distribution of newly synthesized GAG. At high cell densities both cell types showed reduced incorporation into hyaluronate and an increase in cellular GAG due to enhanced labeling of chondroitin sulfate and heparan sulfate. These changes were more marked in confluent 3T3 cultures which also differed in showing substantially more GAG label in the medium and in chondroitin-6-sulfate and heparan sulfate at the cell surface. Since cell density and possibly density-dependent inhibition of growth but not viral transformation are major factors controlling the cellular distribution and type of newly synthesized GAG, differences due to GAG's in the culture behavior of normal and transformed cells may occur only at high cell density. The density-induced GAG alterations most likely involved are increased condroitin-6-sulfate and heparan sulfate and decreased hyaluronic acid at the cell surface.

Cell Count↗
PubMed1975

Subcellular redistribution of seryl-transfer RNA during estrogen-induced phosvitin synthesis and specificity of the estrogen effect.

The estrogen-induced hepatic synthesis of the serine-rich protein, phosvitin, in chickens is accompanied by an increase in the serine acceptance of total hepatic tRNA, which is limited to two sering isoacceptors. To investigate the role of the tRNA alterations in the synthesis of phosvitin, the relative amounts of various seryl-tRNA species in isolated nuclei and in free and membrane-bound ribosomes were determined. Estrogen treatment causes a shift in the subcellular distribution of hepatic seryl-tRNA. Of the four serine isoacceptors, the amount of tRNASerAGU,AGC was specifically increased in nuclei and in membrane-bound ribosomes. Changes in total hepatic tRNA occurring during physiologic estrogenization were compared with those occurring by varying steroid hormones, times after estrogen administration, estrogen doses, animal ages, and tissue types. The changes observed demonstrate that the seryl-tRNA alterations are closely correlated with the synthesis of phosvitin. The coincident change in seryl-tRNA levels and in phosvitin synthesis, together with the specific change in cellular localization, suggests that the amount and subcellular distribution of each tRNA species are separately controlled in a manner dependent upon its frequency of use in translation.

Animals↗
PubMed1972

Cytochalasin B: lack of effect on mucopolysaccharide synthesis and selective alterations in precursor uptake.

Synthesis and secretion of mucopolysaccharide in mouse 3T3 fibroblasts and in embryonic submandibular glands are unaffected by amounts of cytochalasin B that alter the morphology of these cells and tissues. The drug markedly and reversibly inhibits incorporation of [(3)H]glucosamine into mucopolysaccharide by preventing cellular uptake of the precursor, but does not affect incorporation of radiosulfate. Cytochalasin does not alter DNA, RNA, or protein synthesis, but stimulates the uptake of orotic acid and markedly inhibits the uptake of glucose. These selective effects on the transport of small molecules suggest that the primary action of the drug may be on cell membranes. Since processes unrelated to microfilament disruption may be altered by cytochalasin, great caution must be exercised in interpreting studies with the drug.

Animals↗
PubMed1972

Acid mucopolysaccharide (glycosaminoglycan) at the epithelial-mesenchymal interface of mouse embryo salivary glands.

Acid mucopolysaccharide (glycosaminoglycan) has been demostrated at the epithelial-mesenchymal interface of mouse embryo submandibular glands by (a) specific staining for polymeric sulfate with Alcian blue 8 GX at various magnesium concentrations, (b) specific staining for polymeric uronic acid by selective oxidation of these residues to Schiff-reactive compounds, (c) electron microscope localization of ruthenium red staining, (d) radioautographic localization of glucosamine-(3)H and (35)SO(4), and (e) by susceptibility of the glucosamine radioactivity at the interface to digestion with protease-free hyaluronidase. Moreover, material labeled with glucosamine-(3)H and (35)SO(4) and with chemical characteristics identical with those of acid mucopolysaccharide were isolated from the glands. Acid mucopolysaccharide is distributed over the entire epithelial surface. The amount of acid mucopolysaccharide, as revealed by the staining procedures, is nearly equivalent at all sites. In contrast, the rate of accumulation of glucosamine-labeled mucopolysaccharide is greater at the surface of the distal ends of the growing and branching lobules. This distribution of newly synthesized acid mucopolysaccharide at the sites of incipient cleft formation suggests that surface-associated acid mucopolysaccharide is involved in the morphogenetic process. A mechanism of branching morphogenesis is proposed which accounts for the distribution of collagen fibers and total and newly synthesized acid mucopolysaccharide at the epithelial surface.

Ammonia↗