Cellular adhesiveness and extracellular substrata.
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Biomedical subjects
Publications and source records attributed to F Grinnell.
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Studies have been carried out to determine the effects of cold-insoluble globulin (CIG) on the attachment and spreading of baby hamster kidney cells on various collagen substrata. Cell attachment to native collagen substrata occurred in the absence of CIG just as fast as attachment to dried collagen or gelatin substrata occurred in the presence of CIG. On the other hand, cell attachment to dried collagen or gelatin was markedly reduced in the absence of CIG. Cell spreading also occurred on native collagen in the absence of CIG; however, CIG was absolutely required for cell spreading to occur on dried collagen or gelatin. Finally, anti-CIG antiserum or lactoperoxidase treatment inhibited cell spreading on CIG-coated substrata but not on native collagen substrata. The data are discussed in terms of the interaction of fibroblasts with collagen in situ.
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In the presence of serum-containing medium, BHK cells attached and spread during a 1-h period onto a 3-5 nm thick serum layer absorbed on the substratum surface. The closest approach of the plasma membrane to the serum layer was observed to be about 9nm, which was determined by tilting the sectioned cells in a goniometer holder. Bundles of microfilaments or other cytoplasmic specializations were not observed in association with the regions of close contact. However, in the space between the plasma membrane and the adsorbed serum layer, a diffusely stained material could be visualized after fixation/staining by the tannic acid-glutaraldehyde technique. This technique also permitted increased clarity of visualization of trilaminar appearance of the plasma membrane. The distribution and mobility of anionic sites on the surfaces of attached and spreading cells was determined by labeling with polycationic ferritin. We observed movement of polycationic ferritin into large clusters on the cell surface, collapse of cell surface microextensions, and endocytosis, all of which were similar to our previous findings utilizing cells in suspension. However, the absolute amount of ferritin bound to the upper cell surface was less than that previously observed when suspended cells were put under similar labeling conditions. Also, polycationic ferritin did not appear to penetrate between the lower cell surface and the substratum.
The evidence is reviewed that two types of cell attachment occur, depending upon the presence or absence of serum in the medium. In the absence of serum, attachment has many characteristics of a nonphysiological process. In the presence of serum, attachment occurs as a series of steps: adsorption of serum components onto the substratum, contact between the cell and substratum, initial attachment, and progressive attachments leading to cell spreading. Although there is a close interdependence of these events, they could be experimentally distinguished. Studies are reported indicating that cell spreading requires the adsorption of a specific serum glycoprotein onto the substratum surface. The relationship between cell adhesiveness and the altered behavior of malignant cells is discussed.
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The distribution and mobility of anionic sites on the surfaces of baby hamster kidney cells were studied by utilizing the multivalent ligand, polycationic ferritin, as a visual probe. Our observations revealed that anionic sites are distributed over the entire cell surface, with the highest density of sites being located on cell surface microextensions. Following the initial binding of polycationic ferritin to the surface of unfixed cells, the ligand-bound anionic sites redistributed by migrating from the surface of microextensions to the surface of the cell body. In 20 min, this migration resulted in a total clearing of anionic sites from the surface of microextensions concomitant with the formation of patches of anionic sites on the surface of the cell body. Polycationic ferritin-induced migration and patch formation of anionic sites was not prevented by 2,4-dinitrophenol, N-ethylmaleimide, colchicine, or cytochalasin B. However, the ligand-induced redistribution of cell surface anionic sites was prevented by prefixation of cells with glutaraldehyde.
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Normal and transformed baby hamster kidney (BHK) cells attach to Falcon polystyrene with the same first order rate constant. The longer the cells are attached to the bottles, the more difficult they are to remove. Sulfhydryl (-SH) binding reagents inhibit both the attachment of BHK cells and the increase in adhesive strength of attached cells. Attached BHK cells bind fewer molecules of [1-(14)C]N-ethylamleimide (an -SH binding reagent) than do suspended cells. Incubation of cells with high concentrations of trypsin results in a reversible loss of cell adhesiveness. The recovery of adhesiveness of trypsin-treated cells is inhibited by cycloheximide.
The strength of attachment of normal and transformed baby hamster kidney cells was markedly increased when attached cells were treated with concanavalin A (Con A). The cells became less sensitive to detachment by physical shear or by treatment with trypsin or EDTA; however, their morphology, as observed by phase contrast microscopy, did not change. The effects of Con A were prevented by the simultaneous addition of either D-glucose or alpha-methyl-D-glucoside with the Con A. Also addition of these reagents to the attached cells after Con A treatment partially reversed the effects caused by Con A. Pre-treatment of the culture flasks with Con A before cell attachment resulted in an increase in the strength of cell attachment to the culture flasks as compared to untreated controls.