bluuebunny/crossref_metadata_2025 huggingface dataset:
Human serotonectin: A blood glycoprotein that binds serotonin and is associated with platelets and white blood cells
Authors: Tamir H. , Payette R. F. , Huang Y. L. , Liu K. P. , Gershon M. D.
Published: April 2021

A glycoprotein that circulates in blood, binds to the surface of platelets, and also binds serotonin with high avidity and specificity, has previously been found in rats. This glycoprotein has been called serotonectin. We now report the purification and characterization of a similar circulating glycoprotein in human blood that we propose be called human serotonectin, to distinguish it from the rat protein. Human serotonectin binds serotonin with high affinity (Kd1=36nM; Kd2= 1·1 μM). Monospecific antisera were raised in rabbits to purified human serotonectin. These antibodies were used to locate human serotonectin immunocytochemically, for quantitative estimation of the glycoprotein, and for rapid preparation of material purified by affinity chromatography. Evidence was obtained that indicated that human serotonectin circulates in plasma and also binds to the surfaces of white blood cells and platelets but not to red blood cells. In bone marrow it is found on megakaryocytes and on developing white cells of the eosinophil line. The protein can be completely removed by washing with isotonic sucrose or salt solutions from the surfaces of white cells but similar treatment only partially (63% sucrose wash/73% salt wash) removes human serotonectin from platelets. Antibodies to human serotonectin antagonize the uptake of serotonin by platelets but do not inhibit platelet aggregation. These data show that humans, like rats, have a circulating serotonin-binding glycoprotein that is also present as a peripheral membrane protein on platelets. The human also differs from the rat serotonectin in binding to white cells. The material may function in platelet serotonin uptake in both humans and rats; however, its function, if any, with respect to white cells is obscure.
Skin fibroblasts obtained from cancer patients display foetal-like migratory behaviour on collagen gels
Authors: Schor S. L. , Schor A. M. , Durning P. , Rushton G.
Published: April 2021

When plated on the surface of collagen gel substrata, all types of fibroblasts rapidly begin to migrate down into the three-dimensional collagen matrix. We have previously demonstrated that normal (adult and foreskin), foetal and transformed fibroblasts may be distinguished from each other by virtue of their differential migratory response to changes in cell density. The effects of cell density on fibroblast migration into the gel may be expressed by a single numerical value, the ‘cell density migration index’ (CDMI). We now present evidence that ostensibly normal skin fibroblasts obtained from the majority of patients we examined with carcinoma of the breast, malignant melanoma, familial polyposis coli, retinoblastoma and Wilms’ tumours display aberrant CDMI values falling within the foetal range. Skin fibroblasts obtained from the majority of patients examined with genetic or chronic diseases (e.g. rheumatoid arthritis, Duchenne muscular dystrophy) displayed CDMI values falling within the normal range.
Isolation and characterization of macronuclei of Paramecium caudatum infected with the macronucleus-specific bacterum Holospora obtusa
Authors: Freiburg Manfred
Published: April 2021

Macronuclei from Paramecium caudatum infected with Holospora obtusa may be isolated on sucrose step gradients. Macronuclei containing primarily infectious forms can be separated from those bearing predominantly reproductive forms. RNA polymerase activity in infected macronuclei is greater by a factor of 5 than that in uninfected macronuclei. Proteinase activity is also significantly higher.
Sperm-oocyte interaction in the sea-urchin
Authors: Dale Brian , Santella Luigia
Published: April 2021

A study was made of the electrical and morphological changes in the sea-urchin oocyte during interaction with spermatozoa. The first event, a small step depolarization accompanied by a 20—40% decay in input resistance occurs within seconds of attachment. The evidence suggests that this electrical event is the result of sperm—oocyte fusion, and that the ion channels that lower the resistance across the oocyte-sperm complex are located in the sperm plasma membrane. This primary electrical event does not necessarily lead to sperm incorporation. A second, determinative, event occurs at 50 s, which leads to sperm entry and the formation of a cytoplasmic protrusion at the site of sperm entry. This second event probably results from the transfer of a soluble component from the spermatozoon into the oocyte cytoplasm, which leads to sperm incorporation and formation of the protrusion. The changes in the oocyte following insemination are compared with the events of egg activation.
Apoptosis as the mode of cell death in antibody-dependent lymphocytotoxicity
Authors: Stacey N. H. , Bishop C. J. , Halliday J. W. , Halliday W. J. , Cooksley W. G. E. , Powell L. W. , Kerr J. F. R.
Published: April 2021

A light and electron microscopic study of antibody-dependent lymphocytotoxicity was carried out with the object of elucidating the mechanisms responsible for the cell killing, the basis for the research being the relationship that has recently been shown to exist between the morphology of cell death and its pathogenesis. Chang liver cells coated with a rabbit anti-human antibody were used as targets and normal human peripheral-blood lymphocytes as effector cells.

Cytotoxicity assays using release of 51Cr demonstrated extensive K-cell killing, thus validating the suitability of the model for morphological studies. Cell death displaying the features of apoptosis correlated with K-cell activity. A small amount of cell death by classical necrosis was observed, but its extent appeared to be unrelated to the presence of lymphocytes, to pre-treatment of the target cells with antibody, or to the magnitude of 51Cr release.

The results support evidence indicating that lymphocytotoxicity depends on activation of a selfdestruct program within the target cell. They do not favour a mechanism involving the production of plasma membrane lesions analogous to those responsible for complement-mediated immune cytolysis.
Production and functional properties of eosinophils from bone marrow cultures
Authors: Strath Malcolm , Sanderson Colin J.
Published: April 2021

Bone marrow cultures have been established from mice infected with Mesocestoides corti and undergoing parasitic eosinophilia. In the absence of added conditioned medium, eosinophil differentiation ceases, and eosinophils are undetectable by 7 days, whereas neutrophil production continues over several weeks as with normal bone marrow. Eosinophil production can be induced by adding pokeweed mitogen-stimulated spleen supernatants (MSSS) or specific antigen-stimulated spleen supernatants (ASSS) produced from spleen cells of M. corti-infected mice. In contrast to the continuous production of neutrophils, eosinophil production is transient, suggesting that there is no continued production of eosinophil progenitor cells in these cultures. More eosinophils are produced when MSSS is added at the initiation of cultures, compared to after a delay of 2 weeks, and establishing the cultures at 33 °C does not appear to enhance eosinophil production.

The eosinophils produced are shown to express the eosinophil differentiation antigen defined by monoclonal antibody NIMP-R13, they produce eosinophil peroxidase in similar amounts to eosinophils taken from mice. They show normal phagocytic activity of antibody-coated erythrocytes and lyse red cells coated with antibodies of IgGl, IgG2a, IgG2b, but not IgM isotypes.
Correlations between characteristics of some free-living Chlorella sp. and their ability to form stable symbioses with Hydra viridis
Authors: Rahat M. , Reich V.
Published: April 2021

Aposymbiotic polyps of Hydra viridis were infected with 17 strains of in vitro cultured Chlorella sp. Larvae of Artemia fed with the chlorellae were used as an infecting vector.

Of the 17 strains, seven formed stable symbioses and one formed a transient infection that disappeared within several weeks. Chlorellae of the nine other strains were cleared out of the infected hydra within 2–3 days.

There was a distinct correlation between the ability of the chlorellae to form stable symbioses and their ability to adapt and grow in media enriched with 0·5% proteose peptone. Only strains that grew in the latter medium formed symbioses with the hydra.

The symbioses formed with the different strains of chlorellae differed from one another. Hydra infected with some strains greened completely while those infected with other strains greened only partially. The degree of infection varied also within each population, and there were differences in the distribution of the various chlorellae along the stalk and inside the digestive cells of the hydra. Growth rates of the infected hydra were all less that those of aposymbiotic hydra or of hydra hosting native zoochlorellae.

We conclude that adaptability to a nutrient-rich environment inside the perialgal vacuole of the digestive cell and a sufficient growth rate therein are crucial to the ability of chlorellae to form stable symbioses with H. viridis. In time, co-adaptation of hydra and chlorellae would restore the normal growth rate of the former and bring about regularity to the form and extent of infection by the latter.
Morphological changes in rat tracheal cells during the adaptive and early growth phase in primary cell culture
Authors: Chang Ling-Yi , Wu Reen , Nettesheim Paul
Published: April 2021

The purpose of our studies was to determine the fate of different cell types present in early primary cultures of tracheal epithelial cells and, if possible, to elucidate the role they play in the establishment of the cultures. Epithelial cells were isolated from rat trecheas with 0’5% Pronase and were cultured on collagen-coated dishes as described previously. Light and transmission electron microscopic studies showed that the cell population harvested from rat trachea was composed of approximately 30% ciliated cells, 50% granule-containing cells and 20% undifferentiated cells (presumably basal cells). Upon seeding the tracheal cell suspensions into culture, ∼ 40% of the cells attached. Cell attachment was virtually complete after 16 h. Roughly 60% of the cells attaching during the first 12 h were neither ciliated nor granulated, suggesting that undifferentiated cells played a major role in establishment of the early cultures. Between 20 and 35% of the cells attaching during this time were identified as granulated cells (mucous cells). Ciliated cells did not start to attach in significant numbers until 8 h after seeding. They never amounted to more than 8–12% of the attached cell population.

After 12 h of culture, the cell population underwent a progressive loss of differentiation. The number of poorly differentiated cells (i.e. those showing neither cilia nor mucous granules) increased correspondingly. This loss of differentiation preceded the onset of DNA synthesis and cell growth which began at about 24 and 40 h, respectively. Continuous [3H]thymidine-labelling studies showed that at 48 h after the start of culture about 90% of all attached cells had entered DNA synthesis at least once. This finding is consistent with the interpretation that the ciliated cells are terminally differentiated cells and are probably the only part of the tracheal cell inoculum not participating in the growth of the cultures.

At 72 h, the cultures (now in mid-log growth phase) were composed of uniformly undifferentiated cells lacking cilia and mucous granules. The cells nevertheless showed unequivocal epithelial characteristics such as tight junctions and desmosomes.

The studies suggest that both basal and mucous cells are responsible for the establishment and growth of the rat tracheal epithelial cell cultures.
Ultrastructural changes in gap junctions associated with co2 uncoupling in frog atrial fibres
Authors: Mazet F. , Dunia I. , Vassort G. , Mazet J. L.
Published: April 2021

The correlation between gap junction morphology and the state of electrical coupling was investigated in the frog auricle, which presents an atypical gap-junction organization. Electrical uncoupling of the tissue was achieved by perfusion with CO2 -saturated Ringer medium. The tissue was fixed with glutaraldehyde and freeze-fractured before and during the application of CO2 -saturated Ringer medium and after returning to the initial medium. The electrical tissue coupling was assayed by microelectrode recording just before fixation. At least 97% uncoupling was induced by CO2 -saturated Ringer medium, as estimated from double sucrose gap experiments on several single atrial trabeculae.

Several modifications were induced by CO2 -saturated Ringer medium. A decrease in the number of particles per junctional assembly and dispersion of these assemblies in the plane of the membrane, indicate a decrease in the organization of the gap junctions. In parallel, loose clusters of particles became evident on the P-fracture face of the membrane. The size of the particles in these clusters was larger than the size of the background particles, and of the gap junction particles. They never corresponded with complementary pits on the E-fracture face. On return to the initial Ringer perfusion medium the cell coupling was reversed and the gap junction dispersion was also reversed. However, the gap junctions remained small. The relationship between these morphological modifications and the conducting state of the tissue is discussed.
A subclass of luminal epithelial cells in the human mammary gland, defined by antibodies to cytokeratins
Authors: Bartek J. , Durban E. M. , Hallowes R. C. , Taylor-Papadimitriou J.
Published: April 2021

Two monoclonal antibodies, BA16 and BA17, have been developed using a detergent-insoluble extract of human mammary epithelial organoids as immunogen. Indirect immunofluorescent staining of cultured cells showed that the component reacting with the antibodies was filamentous and the intensity of staining was stronger in mitotic cells. Immunoblotting of cell extracts showed that both antibodies react with only one band of 40× 103 molecular weight, which was present in keratin-enriched extracts of cells or organoids. Furthermore, the tissue distribution of the component reacting with the antibodies was that predicted for human keratin 19. The antibodies showed differences in the intensity of staining of cells or tissue sections fixed and prepared in different ways indicating that they reacted with different epitopes. The pattern of expression of the 40×103Mr keratin by normal mammary epithelial cells was investigated by immunoperoxidase staining of tissue sections, cultured milk cells, and organoids of different sizes cultured in collagen gels. It was found that basal or myoepithelial cells did not express this keratin. Some heterogeneity of expression of this component was seen in luminal epithelial cells, found almost exclusively in the smaller structures. These cells did, however, express other keratins characteristic of luminal cells. The distribution in the mammary tree of the luminal cells that did not express the 40 × 103Mτ keratin appears to be similar to that expected for cells with the proliferative potential to produce new terminal ductal lobular units or an increase in branching of existing terminal ductal lobular units. It is shown that these cells have considerable proliferative potential by the fact that they form large colonies in milk cell cultures.
Capsella embryogenesis: The chalazal proliferating tissue
Authors: Schulz Patricia , Jensen W.A.
Published: April 2021

Electron-microscope and histochemical procedures were used to study the development and breakdown of the chalazal proliferating tissue in Capsella. This tissue is formed by the enlargement of several layers of nucellar cells at the chalazal end of the embryo sac. When the embryo reaches the early globular stage these enlarged cells start to disintegrate, beginning with those immediately bordering the embryo sac and continuing until all have broken down. Characteristic ultrastructural changes accompany the development and breakdown of the chalazal proliferating cells. The mature cells form increased numbers of dictyosomes and large amounts of endoplasmic reticulum (ER). This is accompanied by a thickening of the cell wall. As the cells begin to break down, electron density increases, plastids become polarized in the cells, dictyosomes disappear and the ER is dispersed and fragmented. Plastids, some mitochondria, and pieces of ER appear to be digested in autophagic vacuoles. Cell disorganization is accompanied by an increased number of microbodies and multivesicular bodies per cell. Finally, the nucleus breaks down and the plasmalemma disappears. The end wall ruptures and releases intact mitochondria, ribosomes, and portions of degenerated cytoplasm into the endosperm. Histochemical changes accompany these events. Also discussed are the antipodals and the destruction of the proximal part of the chalazal nucellus by the expanding megagametophyte prior to the development of the chalazal proliferating tissue.
The analysis of malignancy by cell fusion: I. hybrids between tumour cells and l cell derivatives
Authors: Klein G. , Bregula U. , Wiener F. , Harris H.
Published: April 2021

A wide range of different kinds of malignant cell were fused with certain derivatives of the L cell line and the ability of the resulting hybrid cells to grow progressively in vivo was examined. In all cases the highly malignant character of the tumour cells was suppressed by fusion with the L cell derivatives, whether or not these had metabolic defects that facilitated selection of the hybrid cells. So long as the hybrid cells retained the complete chromosome complements of the two parent cells, their ability to grow progressively in vivo was very limited, for tumours composed of such unreduced hybrids were not found. However, when they lost certain specific, but as yet unidentified, chromosomes, the hybrid cells regained the ability to grow progressively in vivo and gave rise to a tumour. These findings thus indicated that the L cell derivatives contributed something to the hybrid that suppressed the malignancy of the tumour cell, and that this contribution was lost when certain specific chromosomes were eliminated.
The quantitative glucose and mineral nutrient requirements of mouse LS (suspension) cells in chemically defined medium
Authors: Birch J. R. , Pirt S. J.
Published: April 2021

The influences of glucose, phosphate, magnesium and potassium concentration on growth rate and maximum population density were studied. From the data obtained the growth yields for glucose and phosphorus were found to be 2·4× 106 cells (i.e. 0·76 mg dry weight)/mg glucose and 2·2 × 106 (i.e. 69 μg dry weight) μg P. The growth response to magnesium and potassium was complex since there were threshold concentrations of these metals below which no growth occurred. The growth yield for potassium (2·4 × 105 cells (i.e. 75μg dry weight) μg K) was obtained by measuring potassium uptake during growth. Omission of calcium ions from the medium resulted in poor reproducibility of growth.
The extracellular matrix ofVolvox: A comparative study and proposed system of nomenclature
Authors: Kirk David L. , Birchem Regina , King Nancy
Published: April 2021

The structure of the extracellular matrix (ECM) of representatives of all four sections of the genus Volvox was examined by a combination of light- and electron-microscopic methods. On the basis of these observations, plus published descriptions of aspects of ECM organization in other members of the order Volvocales, a system of nomenclature is proposed, to facilitate discussion of comparative morphology and phylogeny of the ECM in the order. In this system the ECM is divided into four main zones: the flagellar zone (FZ), which consists of attachments to and specializations of the ECM around the flagella; the boundary zone (BZ), which consists of portions of the ECM that (except in periflagellar regions) are continuous over the surface of the organism and are not structurally continuous with deeper layers; the cellular zone (CZ), which consists of specializations, other than those of the FZ, around individual cells; and the deep zone (DZ), which consists of components that fill the central region of the organism, internal to CZ. An empirically based set of hierarchical subdivisions of these zones is then proposed that permits specific identification of most morphologically distinct ECM components. The fact that not all zones and subzones are present in all members of the order means that this system permits identification of those ECM structures that have been gained or lost during Volvocalean evolution.

Species-specific differences in the structure of virtually all aspects of the ECM were seen among the Volvox species examined in this study. However, the fact that such differences cannot always be used as diagnostic characters for the four divisions of the genus was demonstrated by the observation that in certain ECM features two members of the samedivision (V. carteri f. nagariensis and V. carteri f. weismannia) differ markedly in structure from one another, with one member of the pair resembling a member of another division. Thus many details of ECM organization appear to be under separate control, and capable of independent evolution.
Actin cytoskeleton of spread fibroblasts appears to assemble at the cell edges
Authors: Svitkina Tatjana M. , Neyfakh Alexander A. , Bershadsky Alexander D.
Published: April 2021

The action of metabolic inhibitors on actin cytoskeleton of cultured quail embryo fibroblasts has been studied using electron microscopy of platinum replicas and immunofluorescence microscopy. Sodium azide as well as other inhibitors (oligomycin and dinitrophenol) caused the disassembly of all types of actin structures: actin meshwork at the cell active edges, microfilament sheath underlying the cell surface, and microfilament bundles. Studying the time- and dose-dependence of the destruction process we have found that the active edge meshwork and microfilament sheath are much more labile than microfilament bundles. After the removal of metabolic inhibitors actin cytoskeleton restoration begins at the cell edges. The first sign of this process is the formation of actin meshwork along the whole cell perimeter (l-10min of recovery). Sometimes fragments of this meshwork bend upwards forming ruffles. Later (10-20 min of recovery) the microfilament sheath appears at the cell periphery as a narrow band. The sheath seems to be formed from the edge meshwork, since ruffles in the process of transformation to sheath could be seen. During the following restoration the microfilament sheath gradually expands towards the cell centre. The last step of actin cytoskeleton restoration (60-120 min of recovery) is the formation of bundles.

We suggest that the actin in spread fibroblasts polymerizes predominantly at the cell edges and then moves centripetally, forming the microfilament sheath and bundles.
Three-dimensional electron microscopy of the internal nucleolus-associated chromatin and of the nucleolar vacuoles during early germination of Sinapis alba
Authors: Deltour R. , Mosen H. , Bronchart R.
Published: April 2021

Spatial relationships between the internal nucleolus-associated chromatin (NAC) and the numerous nucleolar vacuoles that appear during early germination have been studied in nucleoli of quiescent (non-germinated) and early germinating embryos of Sinapis using serial sections. In quiescent non-vacuolated nucleoli, the transcriptionally inactive internal NAC is a short strand about 900 nm thick that in cross-section appears as heterogeneous fibrillar centres (FCs). At 4 and 6 h after germination one or several large networks of interconnected nucleolar vacuoles develop around the dispersing internal NAC. Clumps of dense chromatin are still present within the nucleolar vacuoles and are probably unfolding into deoxyribonucleoprotein (DNP) fibres (about llOnm thick), which rapidly intrude within the nucleolar body and form thin chromatin threads. At 24 h after germination the internal NAC is more dispersed and forms, for its greatest part, a long thread (about 240 nm in diameter) wrapped up with a few dense fibrillar component, the whole forming the first outline of a nucleolonema. In cross-section most of the internal NAC appears as homogeneous FCs but short portions remain more condensed and appear as heterogeneous FCs always associated with a nucleolar vacuole. From 48 h the internal NAC is a longer thinner strand (about 160 nm in diameter), probably continuous and surrounded entirely by a homogeneous muff of dense fibrillar component, the whole forming a typical nucleolonema (about 950 nm thick) meandering throughout the nucleolus. Small amounts of the internal NAC still remain undispersed in the form of heterogeneous FCs associated with a nucleolar vacuole. The repeated association of nucleolar vacuoles and dispersing internal NAC suggests that they could play a role in chromatin dispersion and, or, activation by creating a favourable microenvironment.
Control of cell proliferation in the human embryonic cornea: An autoradiographic analysis of the effect of growth factors on dna synthesis in endothelial and stromal cells in organ culture and after explantation In Vitro
Authors: Hyldahl Louise
Published: April 2021

A novel technique for studying the growth properties of human embryonic corneal endothelial and stromal cells in organ culture was devised. Human embryonic eye globes were microdissected so that a passage was opened between the outer environment and the anterior chamber, which rendered free access of tissue culture medium to the endothelial cell monolayer. The dissected eye globes were maintained in organ culture for 24 h in the continuous presence of tritiated thymidine. Cross-sections were cut through the whole eye globes and subjected to autoradiographic analysis in order to estimate the mitogenic response of each corneal cell type to externally supplied growth factors and hormones.

It was found that the corneal endothelial cells could be stimulated to initiate DNA synthesis by exposure to epidermal growth factor (EGF). The stimulatory effect of this growth factor could be enhanced if either insulin-like growth factor I (IGF I) or a combination of insulin, transferrin and high density lipoprotein (HDL) was simultaneously added. Similar results were obtained by adding growth factors and hormones to primary cell cultures from human embryonic corneas. It was also found that the stromal cell could be stimulated to initiate DNA synthesis by the addition of EGF and IGF I or a combination of insulin, transferrin and HDL.

Taken together, these results suggest that the proliferation of corneal endothelial cells and stromal cells is dependent on EGF-like factors as well as on some insulin-like substance during embryogenesis.
Characterization of early compartments in fluid phase pinocytosis: a cell fractionation study
Authors: Casey Kimberly A. , Maurey Karen M. , Storrie Brian
Published: April 2021

Flotation through a 5·6% Percoll gradient of pinosomes from Chinese hamster ovary (CHO) cells labelled during a 10min internalization period with horseradish peroxidase (HRP), a solute, revealed two pinosomal populations, the expected low-buoyancy population and an unexpected buoyant population. The buoyant pinosomes that sedimented similarly to plasma membrane were not an artifact of HRP trapping during homogenization or of cell surface-adherent HRP. No trapping or cell surface adherence of HRP could be detected by biochemical or cytochemical assays, even after internalization periods as short as 15 s to 1 min. With short uptake times, the buoyant pinosome population was the major HRP positive vesicle population, suggesting a precursor-to-product relationship between the two populations. In pulse-chase experiments, the buoyant pinosome population was shown to be highly exocytic and the precursor to later pinosomes. By electron-microscope cytochemistry, rapidly labelled, HRP positive pinosomes (15 s to 1 min uptake) were typically smooth vesicles with a median diameter of ≈0·30 μm and a size range from ≈0·10 μm to greater than 1·0 μm in diameter. We suggest that these rapidly labelled structures are a very early stage in the intracellular processing of pinocytic vesicles.
Differential redistribution of lectin receptor classes on clonal rat myotubes and myoblasts
Authors: Sawyer James T. , Akeson Richard A.
Published: April 2021

To evaluate the relative mobilities of cell surface glycoconjugates during myogenesis we have studied the redistribution of fluorescein-conjugated plant lectins on L6 rat myogenic cells. Previous experiments had demonstrated that the receptors for the lectins soybean agglutinin (SBA), wheat germ agglutinin, concanavalin A and Lens culinaris agglutinin all were relatively uniformly distributed on both myoblasts and myotubes, and that SBA receptors were capable of rapid redistribution on myotubes but not myoblasts at 4°C (Sawyer & Akeson, 1983). Here we show that when SBA-labelled myoblasts are incubated at 37°C, or for extended times at 4°C, the lectin aggregates as on myotubes. So it appears that SBA-binding components show a quantitative rather than qualitative change in their mobility during L6 differentiation. In addition, the redistribution of the three other lectins on myoblasts and myotubes was either less prominent (i.e. showing fewer apparent surface clusters) or occurred less rapidly than with SBA. None of these three lectins showed striking differences in mobility between myoblasts and myotubes. Thus, it appears that SBA binds to a subset of surface glycoconjugates that is relatively highly mobile, and that this mobility is specifically enhanced with differentiation.
Cellulose microfibril orientation in Oocystis solitaria: Proof that microtubules control the alignment of the terminal complexes
Authors: Quader H.
Published: April 2021

In the green alga Oocystis solitaria microtubules control the regular deposition of cellulose microfibrils. Although it has frequently been suggested that the influence of the cortical microtubules is mediated through the alignment of structures in the plasma membrane, e.g. the cellulose-synthesizing enzymes, experimental proof is lacking. In Oocystis the putative cellulose-synthesizing units, the so-called terminal complexes, can be visualized following freeze-fracture. With respect to the synthesis of a given layer of microfibrils two distinct situations are observable: terminal complex doublets occur before the start of cellulose formation, but are subsequently separated into single terminal complexes by pressure exerted by the crystallizing microfibrils. In order to investigate the effect of anti-microtubular substances on the orientation of the terminal complexes, the state of cellulose deposition at the time of drug application was marked by short (15–30 min) treatment with Congo Red, which causes a morphological change in the terminal complexes. The characteristic alignment of the terminal complexes, both doublets and fragmented single ones, is severely disturbed in cells treated with the herbicide amiprophosmethyl, which is known to interfere with the action of microtubules. The results provide strong evidence that microtubules control the alignment of the putative cellulose-forming units in Oocystis. The observed pattern of interference indicates that the microtubules most probably achieve their control by imposing fluidity channels on the membrane and not via direct links with the terminal complexes.
A gradient in the density of intramembrane particles is formed during capping induced by concana valin a
Authors: Bennett Holly , Condeelis John
Published: April 2021

During capping of concanavalin A (ConA) by amoebae of Dictyostelium discoideum, each cell becomes polarized, with the ConA at one end and newly extended pseudopodia at the opposite end of the cell. This new polarity is stable until the cap is shed or internalized.

Intramembrane particles (IMPs) are widely believed to represent large integral membrane proteins, many of which are ion pumps and channels. Since asymmetric ion currents have been implicated in the development of cell polarity, we have used morphological landmarks associated with the capped cell in freeze-fracture to make a morphometric analysis of the IMP distribution relative to the axis of polarization of the capped cell.

Untreated cells in suspension extend pseudopodia randomly from their surfaces. In these cells the numerical density of IMPs is random. However, capped cells demonstrate a density gradient of IMPs with the lowest density usually in the pseudopodia and the highest in the cap. The difference in density between the cap and other regions of the cell is two- to threefold for all IMPs, but can be as much as sevenfold for >12 nm IMPs.

This study is the first to document that the numerical density of IMPs is altered in response to ligand-induced capping and demonstrates that the distribution of IMPs in a capped cell is related to the axis of polarization of the cell. These results suggest that the development of cell polarity during capping in Dictyostelium amoebae may be due to the asymmetric distribution of IMPs, which may cause asymmetric ion currents across the cell.
Characterization and partial purification of a plasma membrane receptor for Bacillus thuringiensis var. kurstaki lepidopteran-specific δ-endotoxin
Authors: Knowles Barbara H. , Ellar David J.
Published: April 2021

The lepidopteran-specific Pl ô-endotoxin of Bacillus thuringiensis var. kurstaki HD-1 was activated in vitro using insect gut proteases and found to be highly specific for the lepidopteran cell line Choristoneura fumiferana CF1 among a wide range of lepidopteran and dipteran cell lines tested. The toxicity of Pl against CF1 cells is inhibited by.’V-acetylgalactosamine (GalNAc), and the lectins soybean agglutinin (SBA) and wheat-germ agglutinin. Protein blotting was used to identify a glycoprotein of 146×103Mr in the plasma membrane of CF1 cells, capable of binding both the toxin and SBA, which is specific for GalNAc. This glycoprotein was labelled using galactose oxidase and sodium boro-[3H] hydride and solubilized in Triton X-100 before partial purification by affinity chromatography on SBA-agarose. We propose that this glycoprotein is a good candidate for the cellular receptor of the lepidopteran-specific Pl δ-endotoxin of B. thuringiensis var. kurstaki HD-1.
Nuclear matrix generation during reactivation of avian erythrocyte nuclei: An analysis of the protein traffic in cybrids
Authors: Woodcock Christopher L. F. , Woodcock Hilary
Published: April 2021

It has previously been shown that an internal nuclear matrix is generated during the reactivation of the chick erythrocyte nucleus in mouse L-cell cytoplasts. This experimental system has now been used to identify the major polypeptides that migrate into the nucleus during the reactivation process. Mouse L-cells were prelabelled with [35S]methionine, enucleated using cytochalasin B, and fused with 14-to 17-day embryonic chick erythrocytes. Sixteen hours post-fusion, the redistribution of the labelled proteins was examined by electron microscopic autoradiography, and two-dimensional polyacrylamide gel fluorography of the isolated nuclei was used to identify the major imported species. After allowing for cytoplasmic contamination, 15 nucleus-associated polypeptides were identified, two of which also matched with counterparts in the L-cell nuclear preparation. Five of the nucleus-associated polypeptides were tentatively identified (on the basis of one-dimensional gel matches) as nuclear matrix proteins; these five included the two that had counterparts in L-cell nuclei. The autoradiographic results showed that 16 h post-fusion, the specific activity (silver grains/unit area) of the reactivated nucleus was about half that of the cytoplasm, with no evidence for an accumulation of labelled protein at the nuclear periphery. When well-reactivated nuclei were distinguished from poorly reactivated nuclei on the basis of the extent of chromatin decondensation, it was found that their specific activities were quite similar, but because of the difference in size, the well-reactivated nuclei contained about twice as much labelled protein. Estimates of the protein traffic based upon the autoradiographic data indicated that the nuclei had increased in mass by 10·20% during the 16 h reactivation period.
Effect of taxol on first and second meiotic spindle formation in oocytes of the surf clam, Spisula solidissima
Authors: Kuriyama Ryoko
Published: April 2021

The effect of taxol on first and second meiotic spindle formation was examined in oocytes of the surf clam, Spisula solidissima, by immunofluorescence staining with anti-tubulin antibody. The first meiotic spindle appeared to form as in untreated control cells. However, the spindle did not migrate toward the periphery of taxol-activated oocytes, resulting in blockage of the formation of the first polar body. In spite of inhibited microtubule depolymerization and failure of spindle disappearance, the pole separation in telophase that is typical of this material began at the same time as in untreated cells. Polymerization of the second spindle microtubules onto the spindle persisting from the first meiosis led to the formation of a triple form of spindle connected at the poles of each other. The subsequent emergence of ring-shaped microtubule-containing structures in mature activated eggs was not affected by taxol. The mechanism of meiotic spindle formation thus seemsto be different from that in mitosis, where taxol has been shown to block spindle formation completely.
Selection of kidney cell types in primary glomerular explant outgrowths by in vitro culture conditions
Authors: Oberley Terry D. , Yang An-Hang , Gould-Kostka Jeanine
Published: April 2021

Adult guinea pig glomeruli were grown in vitro either in serum or in a chemically defined medium. Glomeruli were plated either directly into plastic flasks or into plastic flasks that had been coated with the extracellular matrix produced by the PF-HR-9 mouse teratocarcinoma endodermal cell line. Both the composition of the medium and the nature of the culture substrate affected whole glomerular attachment and the type of cells produced in culture. Quantitative studies demonstrated selection of cell types by different culture conditions. Three colony types, each composed of distinctive cell types, could be identified by morphological features. The cells constituting two of these colony types were epithelial in nature, but they were identified as different epithelial types by both histochemical and ultrastructural criteria. Previous studies suggested that one epithelial cell type was derived from the glomerular visceral epithelial cell. This study demonstrates that this cell type could be selectively grown in defined medium on plastic. A second cell type showed several features of renal tubular epithelial cells, including histochemical staining for catalase, cell surface microvilli and cilia, and formation of hemicysts and structures that resembled tubules after prolonged periods in culture. To demonstrate that the ‘glomerulus-derived’ tubular cells were indeed tubular epithelium, we isolated purified renal cortical tubules (>99% pure) and cultured them on the HR-9 matrix in a serum-free chemically defined medium. The resultant outgrowths had morphological properties identical to those of the glomerulus-derived tubular cells. It seems likely that small tubular fragments attached to a minority of the glomeruli are the source of these glomerulus-derived tubular cells. Neither epithelial cell type could be subcultured on plastic, but both could be passaged on the HR-9 matrix. A third cell type, the spindle-shaped cell, was easily propagated on both plastic and the HR-9 matrix. The origin of this cell type is not clear. Our results demonstrate the important effect of culture conditions on the selection, growth and differentiation of kidney cell types in vitro.
Phorbol esters activate protein kinase c and glucose transport and can replace the requirement for growth factor in interleukin-3-dependent multipotent stem cells
Authors: Whetton A. D. , Heyworth C. M. , Dexter T. M.
Published: April 2021

Interleukin 3 (IL-3) promotes the survival, proliferation and development of progenitor cells from several distinct haemopoietic lineages and can also stimulate the self-renewal of stem cells. We have explored the mode of action of this growth factor in promoting survival and proliferation, using a multipotent haemopoietic stem cell line FDC-Mix 1. In the absence of IL-3 these cells died within 16–48 h. However, this requirement for IL-3 could be replaced by 12-O-tetradecanoyl-phorbol-13-acetate (TPA) plus Ca2+ ionophore, which promoted not only survival but also DNA synthesis with no concomitant loss of the multipotential nature of these cells. TPA and Ca2+ ionophore, respectively, could also interact synergistically with IL-3 to promote DNA synthesis. Both IL-3 and TPA stimulated the translocation of protein kinase C (PK-C) from the cytosol to a membrane-bound form in FDC-Mix 1 cells. Previously we suggested that IL-3 can activate the primary metabolism of IL-3-dependent cells so that increased glucose transport and glycolysis lead to maintenance of ATP levels and cellular survival. To investigate whether TPA and, or, Ca2+ ionophore could also influence cellular survival via an activation of glucose uptake we assessed the effects of these agents on hexose transport. TPA ± Ca2+ ionophore activated hexose transport to the same degree as does IL-3 but these agents cannot superstimulate FDC-Mix 1 hexose transport in cells that already exhibit an activated transport system from preincubation with IL-3. We conclude that IL-3 maintains FDC-Mix 1 cells via its ability to activate PK-C and increase cytosolic levels of Ca2+, and that an IL-3-mediated activation of PK-C may promote cellular survival via its ability to enhance hexose uptake by phosphorylating the glucose transport protein.
Growth and regrowth of actin bundles in Chara: bundle assembly by mechanisms differing in sensitivity to cytochalasin
Authors: Williamson Richard E. , Hurley Ursula A.
Published: April 2021

Cytochalasin is known to inhibit cytoplasmic streaming rapidly in characean cells without disassembling their actin bundles. Lower cytochalasin concentrations than those needed for streaming inhibition are now shown to disrupt bundle assembly and, over longer periods, assembled bundles. After local wounding, cytochalasin limited bundle regeneration to the production of polygons and straight, discontinuous bundles that rarely connected to bundles outside the wound. The regenerated bundles supported only scattered organelle movements, whereas long, oriented bundles of control cells were connected to those outside the wound and supported bulk endoplasmic streaming. Unwounded Chara plants cultured for up to 2 weeks in 1 μM-cytochalasin maintained normal bundle orientation and rapid cytoplasmic streaming, but the mean number of bundles per file of chloroplasts fell from 5·2 in controls to 2·0 in growing cells and 3·4 in nongrowing cells. These structural effects seem more likely than the streaming inhibition to reflect cytochalasin’s in vitro effect of blocking extension at the barbed but not the pointed end of F-actin. In particular, cytochalasin inhibited the extension into the wound of bundles in which only the barbed ends of filaments would be exposed. However, short lengths of isolated bundles grew within the wound and bundle growth in the intact cell continued, albeit in modified form. It is suggested that these examples of continuing bundle growth involve cytochalasin-resistant mechanisms that are not wholly dependent on barbed-end filament growth.
Role of cell adhesion in contact-dependent transfer of a lysosomal enzyme from lymphocytes to fibroblasts
Authors: Olsen I. , Oliver T. , Muir H. , Smith R. , Partridge T.
Published: April 2021

Normal lymphocytes were found to adhere strongly to monolayer cultures of fibroblasts deficient in the lysosomal enzyme, j3-glucuronidase. During this co-culture, the fibroblasts acquired from the lymphocytes substantial amounts of this enzyme, which often accumulated at sites of contact between the two types of cell. Enzyme transfer was prevented by addition to the co-cultures either of purified lymphocyte plasma membranes or of antibody raised against such plasma membranes, but it was not inhibited by the addition of antibody raised against lymphocyte-derived β-glucuronidase. An active role for the lymphocyte in this contact-dependent process was suggested by interference contrast, immunofluorescence and scanning electron-microscopic studies. These revealed extensive arrays of projections of the lymphocyte that ramified over the fibroblast cell surface. By transmission electron microscopy, conspicuous clusters of micropinocytotic vesicles were evident in the cytoplasm of the ‘recipient’ fibroblasts, subjacent to the surface in regions closely apposed to adherent lymphocytes. Such high frequencies of these vesicles were restricted to sites of lymphocyte-fibroblast contact, suggesting that they may play an important part in the transfer of enzyme between these two types of cell.
Sporidesmin and gliotoxin induce cell detachment and perturb microfilament structure in cultured liver cells
Authors: Jordan T. W. , Pedersen J. S.
Published: April 2021

Changes in cell morphology and cell adhesion occurred when cultured cells from the rat liver cell strain C3 were exposed to the fungal toxins, sporidesmin or gliotoxin. Both toxins caused loss of attachment of the cells to the plastic of tissue culture plates and this effect was preceded by loss of actin cables. Other changes included cytoplasmic vacuolation and blocked entry into S-phase of the cell cycle. Under these conditions [3H]thymidine incorporation into the cells was also diminished but changes were not detected in the amount of cellular actin, or in the accessibility of cell surface proteins to iodination carried out by the Bolton and Hunter method. The observations suggest that disruption of microfilaments is one of the earliest effects of these toxins on eukaryotic cells.
Evidence for the persistence of a decondensed chromosome scaffold in the interphase nucleus
Authors: Bekers AD G. M. , Pieck Anna C. M. , Rijken Antonia A. M. , Wanka Friedrich
Published: April 2021

Nuclei of in vitro cultured bovine liver cells, deprived of the membranes by Triton X-100, were treated with 2 M-NaCl and DNase. Changes in ultrastructure and protein composition were studied at successive steps during treatment. Electron micrographs of nuclei treated with 2 M-NaCl showed a peripheral lamina and an internal system of randomly coiled filaments embedded in a mass of DNA fibres. After partial removal of the DNA the filaments could be seen to serve as backbones for the DNA attachment. Artificial redistribution occurring during fixation with glutaraldehyde suggests that the salt-resistant filaments are not stably cross-bridged into a three-dimensional network. The existence of reversible cross-bridges in vivo cannot be excluded, however. From the available data it is inferred that the filaments represent a decondensed from of the chromosome scaffolds and play a basic role in the organization of the genome throughout the nuclear cycle.