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Biomedical subjects

L Glaser

Publications and source records attributed to L Glaser.

At least 73 records · Page 4Linked to original sources
PubMed1982

Mitogenicity of brain axolemma membranes and soluble factors for dorsal root ganglion Schwann cells.

Previous studies in this laboratory have shown that membranes derived from dorsal root ganglia (DRG) neurites are mitogenic for cultured Schwann cells derived from the same source [Salzer et al (1980): J Cell Biol 84:767-778]. Improved procedures are described for preparing Schwann cells derived from dorsal root ganglia that are highly responsive to various mitogens. Under these conditions, the cells respond not only to the neurite mitogen but also to pituitary extracts, dibutyryl cyclic AMP, and cholera toxin that have been shown previously to be good mitogens for Schwann cells derived from sciatic nerve [Raff et al (1978): Cell 15:813-822], thus reconciling discrepancies in the response of these different Schwann cell preparations to mitogens. Searching for a source of membranes more suitable for biochemical characterization of the neurite mitogen, we found that bovine brain axolemma, prepared by the method of DeVries et al [(1977):Brain Res 147:339-352] is highly mitogenic for Schwann cells. The mitotic index of Schwann cells was increased by the addition of axolemma from 0.5%-2% to 30%-50% during 24-h incubation with [3H]thymidine. Half maximal effect was obtained at about 0.4 microgram axolemma protein per microwell containing 2-4 X 10(3) cells. The axolemma mitogen appears to be an integral membrane protein that remains bound to the membrane under various ionic conditions but can be extracted in a partially active form with deoxycholate. Like the DRG neurite mitogen, the mitogenic activity of axolemma was abolished by trypsin treatment. Unlike the neurite preparation, however, the mitogenic activity of axolemma was only partially inactivated by heat treatment (60%-70% inactivation). A significant difference between the mitogenic activity of axolemma membranes and neurite membranes is the fact that axolemma membranes fail to stimulate Schwann cell proliferation in a defined, serum-free medium (N-2), whereas neurites show significant mitogenic activity in this medium. These findings indicate a possible difference between DRG neurites and brain axolemma either in the mitogen itself or surface components responsible for recognition between the membranes and the cells.

Animals↗
PubMed1982

Binding of isolated 3T3 surface membranes to growing 3T3 cells and their effect on cell growth.

We have quantitated by autoradiography the binding of [125I]labeled 3T3 plasma membrane fragments to 3T3 cells growing on the surface of plastic dishes; ie, the same conditions in which these membranes specifically arrest the growth of 3T3 cells early in the G1 phase of the cell cycle. We have been able to demonstrate that binding of membranes to cells is coincidental with the expression of the growth inhibitory activity of protein(s) present in the membrane fragments. Treatments that reduce binding (heat denaturation of the membranes or culture in the presence of high serum) also reduce growth inhibitory activity. [125I]labeled membranes bound to cells are located primarily on the cell surface (as determined by electron microscope autoradiography) and are exchangeable with unlabeled membranes. We conclude that binding of membranes to cells is necessary but may not be sufficient for the expression of the growth inhibitory activity of these membranes. This approach provides information not only on the average level of binding of membranes to cells, but also provides a quantitative assessment of the variation of the level of membrane to cell binding between different cells in the population.

Animals↗
PubMed1982

Serum and epidermal growth factor transiently depolarize quiescent BSC-1 epithelial cells.

Continuous intracellular recording of membrane potential with microelectrodes in BSC-1 epithelial cells has been used to study the early ionic effects of the interaction of mitogens with cell surface receptors. Initial results show that (i) there is no significant difference in membrane potential between growing and quiescent cells, (ii) addition of epidermal growth factor or serum to quiescent BSC-1 cells induces a brief and transient depolarization, (iii) the mitogenic response of BSC-1 cells to epidermal growth factor and the transient depolarization show similar concentration dependences, and (iv) serum addition to quiescent BSC-1 cells induces a sustained increase in Na+ influx that is electroneutral and amiloride sensitive. Intracellular pH changes may be a primary event triggering the response of quiescent cells to mitogenic polypeptides.

Amiloride↗
PubMed1982

Resistance to phosphatase of thiophosphorylated epidermal growth factor receptor in A431 membranes.

Epidermal growth factor (EGF) increases the phosphorylation of its receptor and other membrane proteins, and these proteins can be rapidly dephosphorylated by membrane-bound protein phosphatase [Carpenter, G., King L., Jr., & Cohen, S. (1979) J. Biol. Chem. 254, 4884]. We report that [35S]-adenosine 5'-[gamma-thio]triphosphate is as effective as [gamma-32P]ATP as substrate for the EGF receptor-associated protein kinase in A431 membranes. Both the kinetics and the extent of the EGF-dependent thiophosphorylation at 0 degrees C are similar to those obtained with [gamma-32P]ATP, provided that ATP hydrolysis by the membrane preparation is inhibited by addition of adenosine 5'-[beta, gamma-imino]-triphosphate. The thiophosphorylation reaction requires Mn2+ but differs from the phosphorylation reaction in the inability of Mg2+ to serve as cofactor. Both EGF-dependent phosphorylated and thiophosphorylated membrane proteins yield the same two major bands of Mr 145,000-160,000 in autoradiograms of NaDodSO4/polyacrylamide gel electrophorograms. The rate of dephosphorylation of membrane proteins that have been thiophosphorylated in the presence of EGF is dramatically slower (factors of 1/20 to 1/40) than that of the phosphorylated proteins at both 0 degrees C and 32 degrees C. This increased metabolic stability of the thiophosphorylated proteins will be useful for investigation of the role of phosphorylation in the biological effects of EGF.

Adenosine Triphosphate↗
PubMed1982

Multiple controls for the synthesis of muscle-specific proteins in BC3H1 cells.

The regulation of the synthesis of muscle-specific proteins has been examined in BC3H1 cells, a smooth muscle-like cell line isolated by Schubert et al. (J. Cell Biol., 1974, 61: 398-413.). The synthesis of both creatine kinase and the acetylcholine receptor appear to be under dual control, a positive control due to cell-cell contact which increases the rate of synthesis of this protein, and a negative signal, elicited by serum components, that decreases the rate of synthesis of these proteins. Induction of muscle-specific proteins in BC3H1 cells is a reversible process and can be arrested after partial induction has taken place by the addition of serum or high-molecular-weight protein fraction from serum to these cells. The high-molecular-weight protein fraction from serum is not by itself mitogenic for Bc3H1 cells and cannot be replaced by a variety of known hormones (mitogenic factors).

Animals↗
PubMed1981

Developmental changes in glycoproteins of the chick nervous system.

Temporal changes have been noted previously in retinal glycoproteins that bind to wheat germ agglutinin by a technique in which the denatured glycoproteins are first separated according to size by polyacrylamide gel electrophoresis, and are then localized on the gel using [125I]lectin. As reported here this technique will also detect differences between dorsal and ventral halves of the neural retina from 8-day chick embryos, and using other lectins will detect temporal changes in the glycoprotein pattern of the optic tectum. Some of the glycoproteins detected by wheat germ agglutinin in the neural retina appear to be represented on the surface of the retinal cells since: (a) the temporal changes in retinal glycoproteins can also be observed in a plasma membrane enriched fraction prepared from neural retina cells; and (b) antibodies prepared in mice against various size categories of wheat germ lectin binding glycoproteins bind to intact retinal cells.

Aging↗
PubMed1981

Growth inhibitory protein(s) in the 3T3 cell plasma membrane. Partial purification and dissociation of growth inhibitory events from inhibition of amino acid transport.

It has been reported previously that a plasma membrane-enriched fraction from 3T3 cells arrests the growth of sparse 3T3 cells early in the G1 phase of growth (Whittenberger et al., '78, '79). Addition of membranes to sparse 3T3 cells also results in a decrease of the rate of transport of alpha-aminoisobutyric acid (Lieberman et al., '79). We have partially purified the growth-inhibitory proteins from plasma membrane of 3T3 cells. These growth-inhibitory proteins behave as intrinsic membrane proteins and do not require membrane lipids for activity. The partially purified proteins arrest cell growth without affecting the rate of alpha-aminoisobutyric transport; thus, inhibition of both transport and cell growth are not obligatorily coupled events.

Aminoisobutyric Acids↗
PubMed1981

Density-dependent regulation of cell growth: an example of a cell-cell recognition phenomenon.

Cell-to-cell contact can result in a variety of changes in the cell's physiology. For different cell types, this may include both the initiation as well as the cessation of cell growth and changes in the state of differentiation. This review examines in detail one such phenomenon, density-dependent inhibition of growth, which is observed with many fibroblasts in culture. Data are summarized which demonstrate that the cessation of growth at high cell density is in part a consequence of cell-to-cell contact. An approach to the study of the molecular basis of this phenomenon is presented based on the demonstration that plasma membranes, when bound to sparse growing cells, mimic contact inhibition of growth. The present status of attempts to purify plasma membrane proteins responsible for this effect are summarized, and the properties of these membrane proteins are compared to those of previously described "soluble" proteins that inhibit cellular growth.

Animals↗
PubMed1980

Dissociation of cell density and cell cycle effects on the rate of transport of alpha-aminoisobutyric acid in 3T3 cells.

The rate of transport of alpha-aminoisobutyric acid (AIB) has been measured in Swiss 3T3 cells at different cell densities in the range 10(3) to 5 x 10(4) cells/cm2. There is a pronounced increase in the rate of Na+-dependent AIB uptake below 4 x 10(3) cells/cm2, which can be observed both in growing cells (10% serum) or in cells arrested early in the G1 (G0) portion of the cell cycle in medium containing plasma-derived serum. The rate of AIB transport of cells arrested in G0 can be inhibited by the addition of a plasma membrane-enriched fraction prepared from 3T3 cells. The results are interpreted in terms of two elements that control the rate of AIB transport, one dependent on the position in the cell cycle and the other on cell contact. Preliminary observations suggest that BALB 3T3 cells behave in a similar way.

Aminoisobutyric Acids↗
PubMed1980

Regulation of the synthesis of S-100 protein in rat glial cells.

The influence of serum on the synthesis of the nervous system specific S-100 protein has been investigated in the rat glial cell clone C61a. In sparse cells, where S-100 synthesis is low, removal of serum leads to a 2- to 3-fold increase of the rate of S-100 synthesis relative to total soluble cell protein. Measurements of S-100 content by radioimmunoassay showed that sparse cells are accumulating S-100 much more rapidly in the absence of serum. Our findings suggest that serum components inhibit the expression of S-100 synthesis in sparse cells by a concentration-dependent mechanism. This effect does not correlate directly with the reduced cellular growth rate in low serum or serum-free medium. Lipid-free medium, which has little growth-promoting activity, maintains the capacity to lower the rate of S-100 synthesis. A partially purified extract of platelets can substitute for serum in repressing the synthesis of the S-100 protein.

Animals↗
PubMed1980

Studies of Schwann cell proliferation. II. Characterization of the stimulation and specificity of the response to a neurite membrane fraction.

When prepared by methods utilized in our laboratory, pure populations of Schwann cells in culture do not divide, but, after recombination with peripheral sensory neurons or their processes, proliferate rapidly (Wood and Bunge, 1975, Nature (Lond.) 256:661--664). In this paper, we demonstrate that a membrane fraction prepared from sensory ganglion neurites is also mitogenic for Schwann cells and increases the labeling index (assessed by autoradiography after incubation of cells with tritiated thymidine) from less than 0.2 to 10% for primary cells, and from 0.4 to 18--19% for replated cells. The increased responsiveness of replated cells may reflect their greater access to the neurite membranes which is a consequence of the elimination of multiple cell layers after replating and the removal of the basal lamina. This stimulation was specific; addition of membrane preparations from other cell types (3T3, C1300, etc.) was not mitogenic. Ultrastructural analysis demonstrated apparent binding of neurite membranes to Schwann cells as well as significant phagocytosis of the membranes by the cells. The uptake of nonmitogenic membranes suggests that phagocytosis per se is not the stimulus of proliferation.

Axons↗
PubMed1980

Studies of Schwann cell proliferation. III. Evidence for the surface localization of the neurite mitogen.

In the preceding paper (Salzer et al., 1980, J. Cell Biol. 84:753--766), evidence was presented that a neurite membrane fraction could be used to stimulate Schwann cell proliferation in culture. In this study, we present evidence that the mitogenic signal by which intact neurites or neurite membranes stimulate Schwann cell proliferation is located at the neurite surface. This conclusion is based on the following observations: (a) stimulation of Schwann cell proliferation by neurons requires direct contact between neurites and Schwann cells, separation of the two cells by a permeable collagen diaphragm 6 microns thick prevents Schwann cell proliferation; (b) treatment of intact neurites with trypsin before preparation of neurite membranes abolishes the ability of these membranes to be mitogenic for Schwann cells; and (c) the mitogenic activity of neurite homogenates is exclusively localized in the particulate rather than the soluble fraction of the homogenate. The mitogenic component on the neurite surface is heat labile, and is inactivated by aldehyde fixation. Preliminary data suggest that the mitogenic effect of neurite on Schwann cells is not mediated by 3',5'-cyclic AMP.

1-Methyl-3-isobutylxanthine↗