Utilization of -glutamine in intercellular adhesion: Ascites tumor and embryonic cells

-Glutamine is required for the synthesis of complex carbohydrates required for the intercellular adhesion of mouse teratoma cells. It remained to be seen if these pathways were of general importance in the adhesion of other cell types. In this study, using an electronic particle counter assay to measure cell adhesion, Ehrlich ascites, Sarcoma 180 and Taper liver ascites tumor cells require exogenous -glutamine to aggregate. This effect is concentration dependent and the amino sugar, -glucosamine, replaces the glutamine requirement. Structural analogs of the active compounds are substantially less effective and metabolic inhibitors block the activity of the effective compounds. Two specific glutamine antagonists, DON (6-diazo-5-oxo- -norleucine) and azaserine (O-diazoacetyl-serine) decrease the action of -glutamine but not of -glucosamine. Trypsin dissociated six day old chick embryo neural retina cells do not require -glutamine to reaggregate, though the rate of aggregation is enhanced after preincubation with glutamine. Dissociation of small clumps of neural retina and inhibition of reaggregation of these cells are facilitated by preincubation with azaserine for 3–5 h. -Glutamine reduces the effect of azaserine on retina cells. These results are consistent with known metabolic pathways and suggest that -glutamine is involved in the synthesis of complex carbohydrates necessary for adhesion in a variety of cell types. The defective adhesion of the tumor cells examined may result from inability to produce glutamine synthetase, or effectively store cr transport -glutamine.     

CHO cell aggregation induced by fibronectin-coated beads. Differences between wild-type and adhesion-variant cells

ADvF11 (F11), a Chinese Hamster Ovary (CHO) cell variant, is defective in its ability to adhere to fibronectin (Fn)-coated substrata but will adhere to substrata coated with poly-L-lysine, conA or extracellular matrix (ECM) [1]. We have observed that both F11 and CHO wild-type (WT) cells were able to bind 3H-Fn beads in a similar manner; however, only WT cells and not F11 cells aggregate in the presence of Fn beads. Both cell types aggregated similarly in the presence of lectins. Fn-bead-mediated aggregation was blocked by low temperature and aggregation did not occur when formaldehyde-fixed WT cells were used. Colchicine, tetracaine and cytochalasin B were not effective in blocking aggregation induced by Fn beads. These results suggest that: 1. Both WT and F11 cells have surface membrane-binding sites for Fn. 2. The aggregation defect in F11 cells is distal to the initial interaction between the cell surface and Fn, but proximal to the cytoskeletal rearrangements required for cell adhesion.     

Calcium-dependent adhesion of Drosophila embryonic cells

Summary By using an in vitro functional assay, we have shown that Drosophila embryonic cells possess Ca²⁺-dependent adhesive sites, which resemble in many respects those described for vertebrate cells and tissues. The cells, obtained by mechanical disruption of gastrulastage embryos, form aggregates within 30 min when maintained under constant rolling. The aggregation is completely dependent on the presence of Ca²⁺ in the medium. In its absence, the cells remain dispersed but the process is reversible by readdition of Ca²⁺. In addition the aggregation is temperature-dependent. No aggregation occurs at 4° C but it can be restored by raising the temperature to 25° C. These properties are characteristic of these cells: established cell lines do not aggregate under the same conditions and mixing of cell lines and embryonic cells does not result in chimeric aggregates, thus pointing towards cell-type selectivity with respect to aggregability. Observations in electron microscopy have shown that the embryonic cells in the aggregates tightly adhere to one another and form, as early as after 30 min, maculae adherens junctions. Drosophila embryonic cells have adhesion sites that are protected from trypsin proteolysis in the presence of Ca²⁺ and sensitive in its absence. The cells' aggregation can be inhibited by a mouse antiserum directed against cell-surface components and a good correlation exists between neutralization of the inhibitory activity of the antiserum and the presence of trypsin-sensitive sites on the cells. These data are in favour of cell-cell adhesion mediated by specific adhesion proteins.     

The multicomponent nature of teratoma cell adhesion factor

The multicomponent nature of teratoma cell adhesion factor has been demonstrated. Fractionation of crude ascites fluid on a DEAE cellulose ion exchange column shows that two or more components are involved in teratoma adhesion factor (TAF) activity. Glycoproteins (or proteoglycans) in fractionated ascites fluid were localized in polyacrylamide gels. The possible role of these sugar-containing molecules in teratoma cell adhesion and current hypotheses on the mechanism of carbohydrate involvement in intercellular adhesion are discussed.     

Dual adhesion systems of chick myoblasts

Cultured chick myoblasts (Mb) were resuspended by incubation with 100 micrograms/ml trypsin/2.5 mM CaCl2 (to yield TC-Mb), or with 5 micrograms/ml trypsin/2.5 mM EDTA (to yield LTE-Mb). As measured in a particle counter, TC-Mb aggregation was Ca2+ dependent, whereas LTE-Mb aggregated equally well in the presence of CaCl2 or EDTA. Cells subjected to the same treatments in sequence, like cells dissociated directly with 100 micrograms/ml trypsin/2.5 mM EDTA, did not aggregate significantly in the presence or absence of Ca2+. Adhesive specificity was assessed by mixing unlabeled cells with cells labeled with a fluorescent dye and then analyzing the distribution of fluorescent and nonfluorescent cells in aggregates. No adhesive specificity was seen in controls (i.e., TC-Mb aggregated randomly with TC-Mb, or LTE-Mb with LTE-Mb), but TC-Mb and LTE-Mb did not cross-adhere. These results indicate the existence of two independent, noncomplementing, adhesion systems, and suggest that the differential treatments preserve or activate one system while destroying the other. Myoblasts dissociated with 2.5 mM EDTA in the absence of exogenous trypsin (E-Mb) have both adhesion systems active on their surfaces, as do Mb grown in Ca2+-free medium and then dissociated with 0.7 mM EDTA (Knudsen, K. A., and Horwitz, A. F., Dev. Biol. 58, 328-338, 1977). Although aggregation of E-Mb is largely Ca2+ independent and that of Knudsen/Horwitz-Mb is largely Ca2+ dependent, they adhere well to each other and to LTE-Mb while segregating from TC-Mb. Fibroblasts also have dual adhesion systems, one Ca2+ dependent and the other Ca2+ independent, but TC-Fb do not cross-adhere to TC-Mb (nor E-Fb to E-Mb). Cell type-specific adhesive selectivity may thus contribute to the selectivity of myocyte fusion.     

Cell density-dependent stimulation of glutamine synthetase activity in cultured mouse teratoma cells

A density dependent stimulation of glutamine synthetase (GS) activity has been observed in cultures of mouse teratoma cells. GS specific activity increased as cultures approached confluency to a level greater than 2-fold over the basal level found in sparse cultures. After confluency the GS specific activity returned to the basal level found in sparse cultures. The enzyme increase could not be attributed to age of cultures, medium or glutamine depletion, cell leakage of GS, or change in the amount of cellular protein. Dibutyryl cyclic AMP (db-cAMP) plus theophylline lowered GS specific activity both in cultured teratoma and in teratoma obtained from ascites grown tumors. The enzyme increase observed in cultured teratoma cells could be prevented by cycloheximide, and enhanced by hydrocortisone or actinomycin D.     

Selective adhesion of mast cells to tracheal epithelial cells in vitro

In allergic and nonallergic lung diseases, if intraluminal mast cells adhere to airway epithelium, inflammatory mediators released from activated mast cells may reach high local concentrations and thus greatly affect airway function. To determine whether mast cells adhere to airway epithelial cells, radiolabeled or unlabeled dog mastocytoma cells were incubated with cultured dog tracheal epithelial cells, with extracellular matrix substrates, and with cryostat-cut sections of dog trachea. Mast cells adhered well to cultured epithelial cells (35 +/- 13% adhesion, mean +/- 1 SD, n = 23) but adhered poorly to types I and IV collagen or to fibronectin (less than 7.5% mean adhesion in all cases). Similarly, in tracheal tissue sections, mast cells adhered preferentially to epithelial cells in surface epithelium or in submucosal glands but not to basal membrane or connective tissue. Adhesion to cultured epithelial cells was a characteristics of a subpopulation of mast cells, could persist for more than 48 h, did not require energy or the presence of divalent cations, and was not mediated by a known family of leukocyte-associated adhesion glycoproteins. Adhesion was completely abolished by pretreatment of mast cells with pronase E or proteinase K but not with trypsin (up to 10 micrograms/ml at 37 degrees C for 20 min each). In contrast, pretreatment of cultured epithelial cells with any of these proteinases had no effect on adhesion. It is concluded that dog mastocytoma mast cells adhere to dog tracheal epithelial cells and do so selectively. It is suggested that mast cell adhesion to airway epithelium may play a role in the effectiveness of mast cell-epithelial cell interactions, and thus, in certain lung diseases, airway function may be affected by intraluminal mast cells more than is currently appreciated.     

The anti-invasive flavonoid (+)-catechin binds to laminin and abrogates the effect of laminin on cell morphology and adhesion

To study the effect of the flavonoid (+)-catechin on cell-matrix interactions two cell types with a different morphology on and adhesion to laminin were used. MO4 virally transformed fetal mouse cells adhere and spread when cultured on top of laminin-coated coverslips or on human amnion basement membrane. M5076 mouse reticulum cell sarcoma cells poorly adhere to these substrates and remain round. Both cell types are invasive in confronting cultures with embryonic chick heart fragments. (+)-Catechin binds to laminin in a pH-dependent way. Pretreatment of laminin-coated coverslips or amnion basement membrane with 0.5 mM (+)-catechin abrogates the effect of laminin on cell morphology and adhesion. MO4 cells do not adhere to the pretreated substrates and remain round, while M5076 cells now adhere and spread. (+)-Catechin inhibits the invasion of MO4 cells but not of M5076 cells into embryonic chick heart in vitro. We speculate that the anti-invasive activity of the flavonoid to MO4 cells is the result of its interference with MO4 cell adhesion to laminin. Invasion of M5076 cells does not imply adhesion to and spreading on laminin.     

Mechanisms of adhesion among cells of the early chick blastoderm. Role of calcium ions in the adhesion of extraembryonic endoderm cells

Summary Cells from the extraembryonic endoderm of the gastrulating chick embryo adhere to one another in the absence of divalent cations. The addition of Mg²⁺ ions to the medium has no effect on the aggregation kinetics but the addition of Ca²⁺ ions increases the number of cells which aggregate and also stabilizes adhesion. Some aggregation also occurs when cells are suspended in saline devoid of Ca²⁺ and Mg²⁺ ions and supplemented with EGTA, a Ca²⁺ ion complexing agent, but adhesion is not stabilized. Shear sensitive and shear resistant bonds form in Ca-containing as well as in EGTA-containing saline. These results suggest that extraembryonic endoderm cells have Ca²⁺ indepedent and Ca²⁺ dependent mechanisms of adhesion.     

cAMP-induced phenotypic reversion of adhesion, aggregation, and endocytosis in adhesion-defective CHO cell variants

ADvF11 cells are a CHO adhesion variant which, unlike wild type (WT) cells, are not able to adhere to fibronectin (Fn) coated substrata or to be aggregated by Fn-beads. However, ADvF11 cells bind Fn-beads to the same extent as WT cells, thus suggesting that the defect(s) associated with ADvF11 cells are distal to the initial receptor-ligand binding event (Cheung and Juliano, Exp. Cell Res. 152:127, 1984). In this communication we report that cAMP analogs such as dibutyryl-cAMP (dbcAMP) and 8-bromo-cAMP are able to correct defect(s) associated with ADvF11 cells enabling them to adhere to Fn-coated dishes and to aggregate in the presence of Fn-beads. However, only approximately 40% of ADvF11 cells were found to be responsive to dbcAMP suggesting heterogeneity in the cell population with respect to dbcAMP sensitivity. Further analysis of this partial response led us to isolate a subclone of ADvF11 cells, F11CA11, which is highly responsive to dbcAMP treatment. Induction of Fn-mediated cell adhesion and aggregation in F11CA11 by dbcAMP is both time and dose dependent. Optimal responses were obtained after overnight incubation in alpha-MEM containing, 1% fetal calf serum, 4% bovine serum albumin, 0.5 mM dbcAMP and 0.2 mM methyl-isobutyl-xanthine (MIX), a phosphodiesterase inhibitor. Under these conditions, 70-80% of F11CA11 cells were found to be adherent, compared to 5-7% of untreated F11CA11 cells and 95-100% of WT cells. Aggregation of dbcAMP-MIX treated F11CA11 cells induced by Fn-beads also approached that of WT cells. In addition, treatment with dbcAMP-MIX markedly increased the ability of F11CA11 cells to internalize Fn-beads. The maintenance of the adherent phenotype required the constant presence of dbcAMP-MIX. Removal of dbcAMP-MIX from the incubation medium resulted in return to the original nonadhesive phenotype. Thus, elevation of cAMP levels can dramatically modify the behavior of F11CA11 cells with respect to fibronectin mediated adhesion, aggregation and endocytosis, in effect causing a phenotypic reversion of all three parameters to wild type status. This suggests that the mechanisms for adhesion, aggregation and endocytosis may each involve regulation by cyclic AMP-protein kinase systems.     

Cellular and metabolic specificity in the interaction of adhesion proteins with collagen and with cells

Fibronectin mediates the adhesion of fibroblasts to collagen substrates, binding first to the collagen and then to the cells. We report here that the interaction of the cells with the fibronectin-collagen complex is blocked by specific gangliosides, GD_{1 a} and GT₁, and that the sugar moieties of these gangliosides contain the inhibitory activity. The gangliosides act by binding to fibronectin, suggesting that they may be the cell surface receptor for fibronectin. Evidence is presented that other adhesion proteins or mechanisms of attachment exist for chondrocytes, epidermal cells, and transformed tumorigenic cells, since adhesion of these cells is not stimulated by fibronectin. Chondrocytes adhere via a serum factor that is more temperature-sensitive and less basic than fibronectin. Unlike that of fibroblasts chondrocyte adhesion is stimulated by low levels of gangliosides. Epidermal cells adhere preferentially to type IV (basement membrane) collagen but at a much slower rate than fibroblasts or chondrocytes. This suggests that these epidermal cells synthesize their own specific adhesion factor. Metastatic cells cultured from the T241 fibrosarcoma adhere rapidly to type IV collagen in the absence of fibronectin and do not synthesize significant amounts of collagen or fibronectin. Their growth, in contrast to that of normal fibroblasts, is unaffected by a specific inhibitor of collagen synthesis. These data indicate the importance of specific collagens and adhesion proteins in the adhesion of certain cells and suggest that a reduction in the synthesis of collagen and of fibronectin is related to some of the abnormalities observed in transformed cells.     

The mucin epiglycanin on TA3/Ha carcinoma cells prevents alpha 6 beta 4- mediated adhesion to laminin and kalinin and E-cadherin-mediated cell- cell interaction

TA3/Ha murine mammary carcinoma cells grow in suspension, do not adhere to extracellular matrix molecules, but do adhere to hepatocytes and form liver metastases upon intraportal injection. Recently we showed that the integrin alpha 6 beta 4 on the TA3/Ha cells is involved in adhesion to hepatocytes. However, despite high cell surface levels of alpha 6 beta 4, TA3/Ha cells do not adhere to the alpha 6 beta 4 ligands laminin and kalinin. Here we show that this is due to the mucin epiglycanin that is highly expressed on TA3/Ha cells. Some monoclonal antibodies generated against epiglycanin induced capping of most of the epiglycanin molecules. TA3/Ha cells treated with these mAb did adhere to laminin and kalinin, and an epithelial monolayer was formed on kalinin, with alpha 6 beta 4 localized in HD1-containing hemidesmosome- like structures and E-cadherin at the cell-cell contact sites. Similar results were obtained after treatment of TA3/Ha cells with O- sialoglycoprotein endopeptidase which removes all epiglycanin. In addition, the enzyme induced E-cadherin-mediated cell-cell aggregation. Both treatments also enhanced the adhesion to hepatocytes, but given the potent antiadhesive effect of epiglycanin it is remarkable that nontreated TA3/Ha cells adhere to hepatocytes at all. We found that during this interaction, epiglycanin was redistributed. We conclude that epiglycanin can completely prevent both intercellular and matrix adhesion, but that this effect can be overcome in certain intercellular interactions because of the induced redistribution of the mucin.     

Adhesion site composition of murine fibroblasts cultured on gelatin-coated substrata

Fibroblasts in vivo adhere to a collagenous extracellular matrix. We present here a combined morphological and biochemical analysis of the adhesion sites of fibroblast-like cells cultured in vitro on gelatin-coated plastic, for comparison with earlier model studies using serum (plasma-fibronectin [pFn])-coated plastic. Scanning electron microscopy shows that cell adhesion to the gelatin is quite similar to that on plastic, but with some morphological differences reminiscent of those caused by higher concentrations of fibronectin adsorbed to the substratum. Measurement using 125I-radiolabeled pFn shows the level of substratum-bound pFn adsorbed from serum in the growth medium is, however, comparable on gelatin or plastic; thus, differences due to pFn must be attributed to the quality of the adsorbed protein; not its absolute quantity. Gel electrophoretic analysis of cellular adhesion sites formed on the two substrata shows their compositions to be qualitatively similar, suggesting again that the same fundamental adhesion processes are involved. However, three protein bands do change; notably, cellular fibronectin is increased on gelatin. These three proteins are also the most resistant to saline extraction, suggesting their intrinsic importance in the adhesion sites. The nature of the growth substratum thus appears to modulate a fundamentally unvarying morphology and adhesion site composition of the cells that adhere to it.     

Leukocyte-platelet aggregate adhesion and vascular permeability in intact microvessels: role of activated endothelial cells

Leukocyte-platelet aggregation and aggregate adhesion have been indicated as biomarkers of the severity of tissue injury during inflammation or ischemic reperfusion. The objective of this study is to investigate the mechanisms of the aggregate adhesion and quantitatively evaluate its relationship with microvessel permeability. A combined autologous blood perfusion with single microvessel perfusion technique was employed in rat mesenteric venular microvessels. The aggregate adhesion was induced by systemic application of TNF-alpha plus local application of platelet-activating factor (PAF). Changes in permeability were determined by measurements of hydraulic conductivity (Lp) before and after aggregate adhesion in the same individually perfused microvessels. The compositions of the adherent aggregates were identified with fluorescent labeling and confocal imaging. In contrast to leukocyte adhesion as single cells resulting in no increase in microvessel permeability, aggregate adhesion induced prolonged increases in microvessel Lp (6.1 +/- 0.9 times the control, n = 9) indicated by the initial Lp measurements after 3 h of blood perfusion, which is distinct from the transient Lp increase caused by PAF-induced endothelial activation in the absence of blood. Isoproteronol (Iso) attenuated aggregate adhesion-mediated Lp increases if applied after autologous blood perfusion and prevented the aggregate adhesion if the initial endothelial activation is inhibited by applying Iso before PAF administration but showed less effect on single leukocyte adhesion. This study demonstrated that leukocyte-platelet aggregate adhesion via a mechanism different from that of single leukocyte adhesion caused a prolonged increase in microvessel permeability. Our results also indicate that the initial activation of endothelial cells by PAF plays a crucial role in the initiation of leukocyte-platelet aggregate adhesion.     

The effect of cell surface components on adhesion ability of Lactobacillus rhamnosus

The aim of this study was to analyze the cell envelope components and surface properties of two phenotypes of Lactobacillus rhamnosus isolated from the human gastrointestinal tract. The ability of the bacteria to adhere to human intestinal cells and to aggregate with other bacteria was determined. L. rhamnosus strains E/N and PEN differed with regard to the presence of exopolysaccharides (EPS) and specific surface proteins. Transmission electron microscopy showed differences in the structure of the outer cell surface of the strains tested. Bacterial surface properties were analyzed by Fourier transform infrared spectroscopy, fatty acid methyl esters and hydrophobicity assays. Aggregation capacity and adhesion of the tested strains to the human colon adenocarcinoma cell line HT29 was determined. The results indicated a high adhesion and aggregation ability of L. rhamnosus PEN, which possessed specific surface proteins, had a unique fatty acid content, and did not synthesize EPS. Adherence of L. rhamnosus was dependent on specific interactions and was promoted by surface proteins (42–114 kDa) and specific fatty acids. Polysaccharides likely hindered bacterial adhesion and aggregation by masking protein receptors. This study provides information on the cell envelope constituents of lactobacilli that influence bacterial aggregation and adhesion to intestinal cells. This knowledge will help to understand better their specific contribution in commensal–host interactions and adaptation to this ecological niche.     

Plasma membrane lipid packing and leukocyte function-associated antigen-1-dependent aggregation of lymphocytes

A simple model system for study of adhesion mediated by leukocyte function-associated antigen-1 (LFA-1) is aggregation of lymphocytes stimulated in vitro. Although aggregation is blocked by monoclonal antibodies to LFA-1, not all lymphocytes expressing LFA-1 aggregate, indicating that LFA-1 is necessary but not sufficient for aggregation. To investigate whether the lipid bilayer plays a role in the functional activation of LFA-1, human peripheral blood lymphocytes and murine splenic lymphocytes were stimulated in culture, and measurements made of aggregation vs. packing of plasma membrane lipids. Progression of cells into aggregates was paralleled by a decrease in lipid packing of the population as a whole, as monitored by increased staining with the fluorescent probe merocyanine 540. Cells from aggregates stained more intensely than nonaggregated cells from the same population, indicating that aggregates are preferentially formed from cells in the population with the loosest packed membrane. In contrast, aggregated cells were found to express equivalent or even lower amounts of LFA-1 than nonaggregated cells. Looser lipid packing is therefore associated with the development of LFA-1-dependent aggregation, and might be involved in the functional activation of this cell adhesion molecule. © 1993 Wiley-Liss, Inc.     

Increase of β2-integrin on adhesion of THP-1 cells to collagen vitrigel membrane

When human monocyte-derived leukemia (THP-1) cells, which are floating cells, are stimulated with lipid peroxides, or Streptococcus suis, these cells adhere to a plastic plate or endothelial cells. However, it is unclear whether or not non-stimulated THP-1 cells adhere to collagen vitrigel membrane (CVM). In this study, firstly, we investigated the rate of adhesion of THP-1 cells to CVM. When THP-1 cells were not stimulated, the rate of adhesion to CVM was high. Then, to identify adhesion molecules involved in adhesion of THP-1 cells to CVM, expressions of various cell adhesion molecules on the surface of THP-1 cells adhering to CVM were measured. β-actin, β-catenin, and β1-integrin expressions did not change in non-stimulated THP-1 cells cultured on CVM compared with those in cells cultured in a flask, but β2-integrin expression markedly increased.     

Introduction to the Symposium and Studies on the Surfaces of Separated and Synchronized Tumor and Embryonic Cell Populations

Tumor and embryonic cell surfaces are examined in this symposiumwith respect to their roles in cell-cell interactions and inearly development and malignancy. Three sets of studies havebeen recently performed in my laboratory to help elucidate thenature of tumor and embryonic cell surfaces and the means bywhich these cells adhere to each other. We separated an in vivo129/J ascites mouse teratoma into specific subpopuladons ofcells by velocity sedimentation in shallow density gradients.The teratoma consistently separated into two major populations:"large" and "small" cells. Only the large cells displayed "malignant-like"surface characteristics in terms of their agglutinability withcarbohydrate binding lectins. The teratoma cells were also synchronizedin culture with thymidine plus colcemid. In these synchronizedcultures, cellular adhesiveness and glutamine synthetase specificactivity displayed oscillatory patterns with peaks of glutaminesynthetase specific activity occurring just prior to peaks ofadhesivenesss. Also, both glutamine synthetase specific activityand cellular adhesiveness were enhanced by two compounds: actinomycinD and hydrocortisone. Based upon previous work that implicatesL-glutamine in intercellular adhesion, it is not unreasonableto speculate that glutamine synthetase specific activity andcellular adhesiveness may be causally related. The problem ofaltered tumor cell adhesiveness is important because it seems,in part, to be responsible for tumor spread. Finally, the seaurchin embryo system was utilized to identify specific cellsurface carbohydrates that may be involved in intercellularadhesion. In 15 separate experiments with each sugar and with15 different saccharides, D-galactose and N-acetyl-D-galactosaminewere the best inhibitors of rotation-medicated reaggregationof 24-hr sea urchin embryo cells dissociated by removal of divalentcations. ß-galactosidase also inhibited reaggregationof these cells. These results implicate galactopyranosyl-likeresidues in the adhesion of 24-hr sea urchin embryo cells witheach other.     

Relationship of adhesiveness of cells in culture with specific enzyme activity.

l-Glutamine is required by mouse teratoma cells and other mouse ascites tumor cells in the synthesis of complex carbohydrates involved in intercellular adhesion. Since l-glutamine is synthesized by the enzyme glutamine synthetase (GS) (EC 6.3.1.2), these studies were undertaken to determine if a relationship exists between cellular adhesiveness and GS specific activity. Two types of experiment were performed to examine this relationship. Actinomycin D enhanced both teratoma cell GS specific activity and cellular adhesiveness over controls in batch cultures at confluency. Also, the relationship between cell adhesiveness and GS specific activity during the cell cycle was studied using cell populations synchronized with thymidine plus Colcemid. In these synchronized cultures, cellular adhesiveness displayed an oscillatory pattern with peaks of GS specific activity occurring just prior to peaks of adhesiveness. The levels of GS specific activity and intercellular adhesiveness were enhanced by the addition of hydrocortisone, a steroid known to induce GS specific activity in mouse teratoma cells. These results demonstrate a correlation between GS specific activity and cellular adhesiveness. Based upon previous work which implicates l-glutamine in intercellular adhesion, it is not unreasonable to speculate that GS specific activity and cellular adhesiveness may be causally related.     

Cells from Rana pipiens gastrulae and arrested hybrid gastrulae show differences in adhesion to fibronectin-sepharose beads

Experiments were performed to examine adhesion of Rana pipiens gastrula cells and arrested hybrid gastrula cells to fibronectin-Sepharose beads (FN-beads). Blastula cells from both normal and hybrid embryos show poor adhesion to FN-beads. Beginning at the early gastrula stage, however, normal cells show a progressively increasing tendency to adhere to beads. In two different arrested hybrid embryos, cells from all developmental stages lack the ability to adhere to beads. A third hybrid shows an increase and then a decrease in cell-bead adhesion. A fourth hybrid shows a late increase in cell-bead adhesion in animal-half cells and no increase at all in vegetal-half cells. Blastula-stage cells have the ability to adhere to con A-beads and two kinds of Cytodex beads but will not adhere to FN-beads. Similarly, some cells from arrested hybrid embryos lack the ability to adhere to FN-beads but will adhere to con A-beads and cytodex beads. Observations in the light and scanning electron microscope show that normal cells form lamellipodia on FN-beads and move about actively on them, much like they do in vivo on surfaces coated by fibrils containing fibronectin. For adherent hybrid cells attached to beads, one kind does so by small pseudopodia but does not move on them and another kind forms active lamellipodia at the tips of fusiform cells and moves on beads.