Search PubMed⌕ Search

Biomedical subjects

E E Medrano

Publications and source records attributed to E E Medrano.

49 records · Page 3Linked to original sources
PubMed1987

Role of thrombin in the proliferative response of T-47D mammary tumor cells. Mitogenic action and pleiotropic modifications induced together with epidermal growth factor and insulin.

The growth of the human metastatic cell line (T-47D) in a chemically defined medium (DM) is shown to be dependent on the presence of three traditional growth factors: epidermal growth factor, insulin, and transferrin. The addition of thrombin further stimulates its growth. The mitogenic action on a human mammary tumor cell line from epithelial origin is a novel action of thrombin. Cells in the DM show striking morphological changes which are dramatically enhanced by the addition of thrombin. These observations are part of a pleiotropic response to the growth factors: the protein content of the cells increases in the defined medium; the 2DG gels of the 35S- and 32P-labeled proteins show important changes in spots, several of which are probably of cytoskeletal origin. It is also shown that cells in a semisolid growth factor-supplemented medium have growth advantages over their counterparts grown with serum. All the phenotypic changes mentioned above reveal the important role of growth factors in the growth and behavior of this mammary cell line. The results obtained with thrombin indicate a new site of action of this enzyme which may be important in the metastatic spread of human mammary tumor cells.

Breast Neoplasms↗
PubMed1987

Subpopulations of MCF7 cells separated by Percoll gradient centrifugation: a model to analyze the heterogeneity of human breast cancer.

Exponentially growing MCF7 human breast cancer cells were separated in Percoll gradients into six different fractions of increasing density (A to F). These fractions could be subcultured and were found to contain different cellular subpopulations as defined by the following criteria: ability to generate other cellular subpopulations; growth rate; DNA synthesis; and expression of estrogen receptors, ras oncogene-encoded protein p21, and carcinoembryonic antigen. One of the minor fractions (E), which contained about 5% of the total cell number, appeared to contain the stem cells, on the basis of the following criteria: (i) its ability to reproduce the other cellular subpopulations, (ii) its high rate of growth and DNA synthesis, and (iii) the inability of the other subpopulations to generate it. The most differentiated subpopulation appeared to be the densest one (F), since it was the slowest growing and appeared to be the end point of the other subpopulations.

Breast Neoplasms↗
PubMed1986

A novel mechanism of resistance to alpha-difluoromethylornithine induced by cycloheximide. Growth with abnormally low levels of putrescine and spermidine.

Treatment of the chemically transformed fibroblasts BP-A31 and other cell lines with low concentrations of cycloheximide (CHM) for 72 h followed by the removal of the protein synthesis inhibitor leads to the proliferation of alpha-difluoromethylornithine (DFMO)-resistant phenotypes. These drug-resistant cells contain almost no ornithine decarboxylase (ODC) activity and concomitantly very low levels of putrescine and spermidine. Southern blot analysis and measurements of ODC activity and intracellular polyamine levels showed that the described mechanism of inducing resistance to DFMO triggered by CHM does not involve ODC gene amplification, altered transport of the drug or reduced affinity of the enzyme for DFMO.

Animals↗
PubMed1985

Dolichol-sugar derivative synthesis in human breast cancer cell line (T47D). Effects of estrogens and antiestrogens.

In the present paper we report evidence about the formation of polyprenyl-phosphate monosaccharides, their elongation products and the assembly of dolichyl-diphosphate-oligosaccharide to endogenous T47D clone 11 proteins upon incubation with [14C]glucose. The influence of estradiol and two nonesteroidal antiestrogens -nafoxidine and tamoxifen- was examined on the dolichol pathway in T47D cell cultures. Estradiol (1 nM) does not change the rate of synthesis of dolichyl-phosphate-sugar derivatives in contrast to nafoxidine and tamoxifen both a micromolar concentration, which induce a remarkable decrease in the formation of dolichol-sugar derivatives. In addition, T47D cells were pretreated with nafoxidine or tamoxifen during one hour, fresh medium supplemented with estradiol was added to the cells simultaneously with [14C]glucose. Results indicated that estradiol after nafoxidine induces a slight increase in the polyprenyl-sugar derivatives formation, however, estradiol after tamoxifen decreases the synthesis of these substances.

Breast Neoplasms↗
PubMed1985

Transcription of ribosomal RNA is differentially controlled in resting and growing BALB/c 3T3 cells.

Shortly after serum-deprived BALB/c 3T3 fibroblasts are stimulated to grow in medium containing 10% calf serum, the RNA polymerase I activity in permeabilized cells shows a two-fold increase over the values observed in either serum-deprived or density-inhibited resting cells. Inhibition of protein synthesis by pactamycin or cycloheximide specifically reduces the enhanced RNA polymerase I activity in serum-stimulated cultures without affecting the values in resting cells. On the other hand, inhibition of rRNA processing by the nucleoside analogs 5-fluoruridine and toyocamycin decreases the rate of 45S rRNA transcription in serum-stimulated cells but has no effect on the values found in resting cultures. These data suggest that the regulation of rRNA transcription occurs by two different mechanisms, depending on the growth state of the cell. One mechanism, in serum-stimulated cells, is dependent on a continuous protein synthesis and a correct 45S rRNA processing; the other, in resting cells, is independent of these two parameters.

Adenosine↗
PubMed1983

Inhibitory effect of 1-beta-D-arabinofuranosylcytosine on the synthesis of phosphatidyl-dCMP.

The synthesis of phosphatidyl-dCMP in mouse thymocytes is inhibited by the antineoplastic agent 1-beta-D-arabinofuranosyl cytosine (AraCyt) 50% inhibition (ID50) being reached at an AraCyt concentration of 0.18 mM. In the same cells, ID50 for DNA synthesis is 0.03 mM. This inhibition is probably mediated by the phosphorylated derivative of AraCyt (aCTP) since the synthesis of phosphatidyl-dCMP from dCTP using permeabilized thymocytes is inhibited by aCTP (ID50 = 0.11 mM). The incorporation of [3H]AraCyt into the organic phase could also be detected, suggesting that this drug may act as a substrate for the enzyme that catalyzes the transfer of dCTP into phosphatidic acid.

Animals↗
PubMed1983

Differential effect of alpha-difluoromethyl-ornithine on the proliferation of Balb 3T3 and chemically transformed 3T3 cells.

In order to study the effect of polyamine depletion on growth and proliferation of untransformed and chemically transformed cells, alpha-difluoromethyl-ornithine (DFMO) was added to cultures of 3T3 cells and their benzo[a]pyrene derivative BP-3T3. Both types of cells stopped their proliferation after 72 hr of treatment with the inhibitor. When DFMO was removed and cells were cultivated afterwards in fresh medium without the drug, untransformed cells resumed growth after a lag period, whereas transformed cells were unable to proliferate unless exogenous polyamine was added. These alterations showed a strict correlation with intracellular polyamine pools, since after removal of DFMO from the culture medium, polyamine concentrations increased to almost normal values in 3T3 cells, but remained at low levels or decreased even more in the transformed cells BP-3T3. The analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of labeled proteins of both cell extracts has indicated that the described control of cell proliferation by intracellular polyamine levels might be related to the synthesis of at least two proteins with molecular weights of about 36,000 and 55,000 daltons.

Animals↗
PubMed1982

Restriction point control of cell growth by a labile protein: evidence for increased stability in transformed cells.

It has been proposed that animal cells must accumulate a labile protein(s) before they can pass the restriction (R) point in the G1 phase of the cell cycle [Rossow, P. W., Riddle, V. G. H. & Pardee, A. B. (1979) Proc. Natl. Acad. Sci. USA 76, 4446--4450]. Here, we present evidence that this R protein acquires increased stability in transformed 3T3 cells, thereby allowing these cells to continue growth under conditions that arrest untransformed cells. Low doses of cycloheximide or histidinol drastically reduced the rate at which normal 3T3 (A31) fibroblasts in early G1 could enter DNA synthesis. These drugs had less effect on entry of two tumorigenic A31 derivatives, BPA31 and SVA31, in S, although measurement of [3H]leucine incorporation showed that the inhibitors were equally effective in the three cell lines. The hypothesis is that the transformed lines are less sensitive because moderate inhibition of their R protein synthesis is compensated by lower rates of protein degradation. To test this idea, we completely inhibited cytoplasmic protein synthesis for several hours shortly before A31 and BPA31 cells had reached the R point. After removal of inhibitor, A31 cells showed delays in the onset of S that were in excess of the inhibitor pulse, consistent with decay of labile protein during the pulse. BPA31 cells showed no excess delays, suggesting a much more stable R protein. The half-life of the R protein was estimated as 2.5 hr in A31 cells, indicating that, in these cells, R protein synthesis starts at the beginning of G1. In the BPA31 cells the R protein showed no signs of decay for at least 8 hr.

Animals↗
PubMed1981

On deciding which factors regulate cell growth.

Many conditions affect the growth of animal cells in culture. Some, such as essential nutrients, are necessary for growth as precursors of macromolecules. Other substances appear to have regulatory functions. Their presence or absence determines whether the cell will continue to grow or move into a resting, quiescent state. In particular certain factors (usually provided by serum) are thought to have regulatory roles. To decide which factors are regulatory is a difficult problem. We propose here that no single criterion is sufficient to permit a decision and that the best course at present is to apply several criteria. Three such criteria are proposed. To the degree that the tests are satisfied we can tentatively decide whether or not a given factor is regulatory.

Animals↗
PubMed1980

Prevalent deficiency in tumor cells of cycloheximide-induced cycle arrest.

Mammalian cell growth is regulated by a process that is completed at a restriction point in the late G1 part of the cycle. This process is highly sensitive to serum concentration and to moderate inhibition of protein synthesis by cycloheximide (CHM) or other agents. We have proposed that a cell must accumulate a labile protein in a critical amount before events related to its DNA synthesis can start. The accumulation of this protein requires conditions suitable for growth, including sufficient amounts of serum-derived factors. An important criterion for attributing a major role to such a regulatory mechanism is that cells whose growth control is modified--e.g., by mutation--should be defective in this process. Cells of this kind are produced by tumorigenic transformation. We show here that mouse 3T3 cells, human fibroblasts, and Chinese hamster CHEF/18 cells have stringent G1 growth control by CHM. In contrast, tumorigenic lines obtained from these cells by transformation with varius agents (DNA tumor virus, RNA tumor virus, chemical carcinogens) or spontaneously all showed relaxed growth control under the influence of CHM. In these lines, growth control was relaxed to different degrees; some lines were held in G1 by a combination of low serum concentration and CHM, but others were not. Serum concentration showed a synergistic effect with CHM. Low serum concentrations did not limit growth only by affecting the rate of protein synthesis. The labile-protein mechanism is likely to be basic to growth control by serum factors. Transformed tumorigenic cells in general may have relaxed this mechanism.

Animals↗
PubMedDate not supplied

Characterization of IIB-MEL-J: a new and highly heterogenous human melanoma cell line.

A highly heterogeneous cell line, IIB-MEL-J, was established from a human metastatic melanoma. This cell line contains small cells, dendritic cells, and megacells with multiple nuclei. IIB-MEL-J expresses S 100, cytokeratin intermediate filaments and the gangliosides GD2 and GD3. It requires growth factors (insulin, EGF, and transferrin) and antioxidants for optimal growth. When plated under optimal conditions, IIB-MEL-J grows with a doubling time of 70-80 hours. The cells may be fractionated by Percoll gradient centrifugation into several subpopulations (A, B, and C) with different characteristics. Subpopulation A is the slowest growing, and most of the DNA-synthesizing cells are concentrated in fractions B and C. Every subpopulation expresses S 100 and cytokeratin intermediate filaments, whereas only subpopulation B and C express GD2 and GD3. Pigmented cells are concentrated mainly in subpopulation C. Cytogenetic analysis of IIB-MEL-J revealed extensive chromosomal alterations, including a highly heterogeneous chromosome number and chromosomal rearrangements, gains, losses, isochromosomes, and double minutes. This highly heterogeneous cell line may be helpful to study cellular differentiation and interaction between different subpopulations in human melanoma.

Cell Transformation, Neoplastic↗