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  • 1980-1984  (655)
  • 1935-1939
  • 1980  (655)
  • Cell & Developmental Biology  (654)
  • Nuclear reactions
  • 101
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980), S. 199-207 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: It has been previously shown that newly synthesized nuclear low molecular weight RNA species C and D are first detected in the cytoplasm for a few minutes before they are finally found in the nucleus. The following are some of the observations made in the present study, regarding the formation of C and D RNA: (1) The 5′ end cap ribose methylation of the C RNA precursor is complete in its cytoplasmic stage; the internal ribose methylation of the precursor seems to be completed about the time of its apparent transition from cytoplasm to nucleus. (2) The few nucleotides lost from the D RNA precursor during maturation seem to be excised sometime near its apparent cytoplasmic → nuclear transition. Newly synthesized C RNA also appears to lose some of its non-conserved nucleotides about the time of that transition, while the other extra nucleotides are lost later, in the nucleus. (3) The maturation of C and D RNA is inhibited early during suppression of protein synthesis by cycloheximide, while their synthesis is not. (4) The cytoplasmic precursors of C and D RNA are not associated with ribonucleoprotein particles as large as those reported for mature C and D RNA, although they do not appear to be free in the cytoplasm. (5) When the cellular UTP pool is depleted by exposure of the cells to amino sugars, and the synthesis of C, D, and other RNA species decreases, the level of[3H]uridine labeling of C and D RNA increases, while that of 4 S and 5 S RNA does not. These data are compatible with the existence of more than one nuclear UTP pool.
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  • 102
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980), S. 175-181 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A procedure has been investigated for sorting viable cells according to their DNA content. Cells are stained with the U.V. activated fluorochromes 4′6-diamidino-2-pheylindole (DAPI), Hoechst 33258 or Hoechst 33342, and sorted with a Fluorescence Activated Cell Sorter. Hoechst 33342 is a suitable vital stain for a varietyof cell types. Hoechst 33258 and DAPI, however, are quantitative vital stains for CHO cells only. Cloning efficiency is unaffected by the sorting procedure, and these stains are not mutagenic at concentrations suitable for vital staining. Potential applications of this procedure to cell biology are discussed.
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  • 103
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980), S. 193-197 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Membrane potential was measured in perfused rat liver and was shown to increase from -33 ± 1.0 mV in livers from normal rats to -50 ± 1.1 mV in livers from rats 12 hr after partial hepatectomy. The hyperpolarization of the membrane in regenerating liver was no longer evident after perfusion with 1 mM ouabain for 5 min. Ouabain had a small (4 mV) depolarizing effect on membrane potential in normal liver. The potential measured in normal and regenerating liver decreased as a function of the external potassium concentration above 5 mM; however, the potential was more electronegative in regenerating liver compared to normal liver at all values of external potassium concentration, and the differences in potential between the two kinds of cells did not decrease at higher concentrations of external potassium. Thus, a plot of membrane potential vs external potassium concentration resulted in approximately parallel curves for the two different cell types. We conclude that hyperpolarization of the liver cell membrane is an early event during rat liver regeneration and results from an electrogenic Na-K pump.
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  • 104
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980), S. 209-216 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cell cycle studies, using PLM analysis, were carried out on a mouse-Chinese hamster cell hybrid and its derivatives which stably retained all parental chromosomes during the year of study. Parameter estimates were obtained from the PLM curves, using conjugate gradient curve fitting procedures. The hybrid initially grew very slowly, and all phases (especially G1) were longer than those of either parent. During propagation, mean generation time decreased progressively, and the phase times approached those of the mouse parent (which had the longer G1 and S). DNA replication could be scored separately in mouse and hamster chromosome sets; initially termination was highly asynchronous, but during growth asynchrony was progressively reduced as DNA synthesis in the hamster set was prolonged.We conclude that cell hybrids may undergo progressive modifications of the cell cycle, even in the absence of significant chromosome segregation, and suggest that such changes may at least partly account for the great variety of relationships between the growth rates and phase times of parent and hybrid cells which have been reported. Because of the complexity of these changes in the cycles of interspecific cell hybrids, we believe that somatic cell genetic analysis of the regulation of the cell cycle would be more usefully applied to intraspecific hybrids whose parents differ in only one specific cycle characteristic.
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  • 105
    Electronic Resource
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980), S. 233-243 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The progenitor cells of neutrophil granulocytes and macrophages which are able to proliferate and differentiate in vitro (CFU-c) form a heterogeneous population. By the use of specific colony stimulating activities and cell separation by equilibrium density centrifugation, three subpopulations of CFU-c can be detected. These three CFU-c are characterized by buoyant densities of 1.070, 1.075 and 1.080 g.cm-3 and by their proliferative response to 18 h postendotoxin serum, colony stimulating factor from extracts of mouse embryos and uteri (CSF-pmue) and erythrocyte lysate, respectively. The three CFU-c are compared with respect to their differentiation potential, the maturation rate of their progeny cells and their proliferation capacity. It is shown that with increasing density of the CFU-c the maturation rate increases (sequential maturation of colonies derived from CFU-c with densities of 1.080, 1.075, 1.070 g.cm-3) and the proliferation capacity decreases (colony size decreases in the sequence of CFU-c with densities 1.070, 1.075, 1.080 g.cm-3). Concerning the differentiation potential it is shown that all three CFU-c detected have the capacity to form granulocytes as well as macrophages. On the basis of these results it is concluded that the CFU-c with densities of 1.070, 1.075 and 1.080 g.cm-3 represent a maturation sequence.
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  • 106
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 102 (1980) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 107
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 313-322 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mitochondrial and cytoplasmic composition were determined with electron probe analysis in freeze-dried guinea pig aortic smooth muscle cells cultured on stainless steel grids. The mitochondrial calcium content in normal cells was low: not significantly different from that detected in the cytoplasm. Mitochondrial calcium granules were found in less than 3% of the cells, and in these the cytoplasmic K/Na ratio was reduced, indicating that they were damaged. There were no major differences between the cytoplasmic concentrations of K, Cl, Ca, Mg, and S of cultured cells and those previously found in adult vascular smooth muscle (Somlyo et al. '79). There was no evidence of nuclear Na or Ca sequestration in cultured cells, and the transmitochondrial Na, Cl, and K gradients were small.Attempts to selectively remove adhering, extracellular ions with a 2-second wash with isotonic ammonium acetate were unsuccessful because they were accompanied by loss of cell K.
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  • 108
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 289-298 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cytoplasmic calcium levels are believed to be important in blood platelet activation. Upon activation, the discrete marginal microtubule band, which maintains the discoid shape of non-activated platelets, becomes disrupted. Present studies demonstrate that the extent of assembly of the marginal microtubule band is related to cytoplasmic calcium levels. The divalent cationophore, A23187, causes platelet aggregation, secretion, and contraction by promoting calcium transport from intraplatelet storage sites into the cytoplasm. A23187 caused disassembly of platelet microtubules. Quantitation of electron micrographs revealed that numbers of microtubules were reduced by approximately 80% after A23187 treatment. Secondly, assembled microtubules in homogenates of platelets, in which microtubules were stabilized prior to homogenization, were decreased in favor of free tubulin in A23187-treated platelets. Thirdly, A23187 increased 14C-colchicine binding by intact platelets; this also indicated a shift in the microtubule subunit equilibrium to favor free, colchicine-binding tubulin subunits. In control experiments, A23187 did not affect the stability of platelet tubulin, the colchicine binding reaction, or the total tubulin content of platelets. Stimulation of colchicine binding depended on A23187 concentration (0.05-0.5 μM) and did not require extracellular calcium. A23187-stimulation of colchicine binding was blocked by dibutyryl cyclic AMP (0.80 mM) and/or 3-isobutyl-1-methylxanthine (50 μM) and by indomethacin (10 μM). Cyclic AMP or indomethacin also interferes with A23187-induced platelet activation, but indomethacin is not likely to completely inhibit the perturbation of intraplatelet calcium gradients by A23187. It is suggested that A23187-induced microtubule disassembly may be an indirect effect of calcium on microtubules.
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  • 109
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 279-287 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cloudman S91 mouse melanoma cells lose their ability to demonstrate an MSH-induced increase in tyrosinase activity as cell density increases. This loss in hormone responsiveness occurs before confluency is reached and cannot be reversed by exposure of cells to increasing concentrations of MSH. The failure of high-density cultures to respond to MSH is apparently not the result of an inability of MSH to stimulate cAMP production, since either low- or high-density cultures exposed to MSH demonstrate equivalent increases in intracellular levels of cAMP. Further, neither theophylline (1mM), dibutyryl cyclic AMP (10-4M), or prostaglandin E1 (10-6M) is effective in stimulating tyrosinase activity in melanoma cells cultured at densities exceeding 6 ± 104 cells/cm2. This finding suggests that the decay of hormone responsiveness occurs at a cellular site distal to cAMP production. The decrease in tyrosinase stimulation by MSH as cell density increases is also apparently not the result of an increase in activity of any soluble inhibitor of the enzyme, for cytosol preparations from high-density cultures (105 cells/cm2) fail to inhibit tyrosinase activity in cell homogenates from low-density cultures treated with MSH.
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  • 110
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 349-353 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Adipocytes were isolated from the Wistar and Sprague-Dawley strains of rats of different ages. An impermeable reagent, mercury-[3H] dextran, was used to quantitate the sulfhydryl groups on the surface of these cells. The application of this reagent after the pre-reduction of cells with 2-mercaptoethanol yielded a measure of the total sulfhydryl and disulfide groups located there. Aging was found to significantly decrease the ratio of the number of exofacial sulfhydryl groups to the total number of exofacial sulfhydryl and disulfide groups.
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  • 111
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Bovine vascular endothelial cells continuously maintained and grown in the presence of FGF adopt at confluence the configuration of a cell monolayer composed of contact-inhibited cells which do not overgrow each other and which are highly flattened and closely apposed. Such cultures exhibit structural and morphological characteristics similar to those observed with their in vivo counterparts. These include the production of an extracellular matrix consisting mostly of basement membrane collagen and fibronectin localized exclusively beneath the cell monolayer, but not on top of it, as well as a nonthrombogenic, blood-compatible apical cell surface. Removal of fibroblast growth factor (FGF) from adult bovine aortic endothelial cell (ABAE) cultures results within three passages in the loss by the cells of their characteristic contact-inhibited morphology. The cells, which during their logarithmic growth phase divide with a greatly increased doubling time, become larger and more elongated. Confluent cultures, instead of adopting the morphology of a contact inhibited cell monolayer, are now composed of overgrowing cells. Parallel with the morphological alterations taking place within the culture, the cells also lose the polarity of cell surfaces characteristics of the vascular endothelium. Formation of an extracellular matrix composed primarily of fibronectin and collagen types I, III, and IV is observed on both the apical and basal cell surfaces. Platelets which previously did not bind to the apical cell surface now become capable of binding to it. CSP-60, a major cell surface protein present in highly confluent and contact-inhibited vascular endothelial cell cultures, can no longer be detected. Exposure of confluent endothelial cell cultures, maintained in the absence of FGF to medium conditioned by cells which had been grown in the presence of FGF, but maintained in its absence upon reaching confluence led, within four to eight days, to a reversion of the altered phenotype. This medium has little or no mitogenic activity and retains a full activity in the absence of serum or after depletion of its fibronectin content by affinity chromatography on a gelatin-Sepharose column. Cultures which were previously composed of cells growing in multiple layers reorganized into a single cell monolayer composed of closely apposed and highly flattened cells. The cultures thereby regained the contact-inhibited morphology characteristic of the vascular endothelium. Concomitant with this cellular reorganization, the extracellular matrix disappeared from the apical cell surface, the cells regained their nonthrombogenic properties, and CSP-60 reappeared as one of the major cell surface proteins. These results suggest that vascular endothelial cells secrete a soluble factor(s) which can restore the normal morphology and function lost following removal of FGF from the medium. Such a factor(s) may be involved in maintaining the differentiated state of the vascular endothelium.
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  • 112
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 355-362 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The numbers of CFU-S which developed in spleen colonies were measured 11 days after injection of irradiated mice with marrow from normal mice or mice which had been treated in one of a variety of ways. The broad spread of CFU-S numbers, seen by other authors, in colonies derived from normal marrow was confirmed. However, the range and distribution of CFU-S per colony was generally different in colonies derived from the marrow of mice which were recovering or had recovered from some form of depopulation. From the data obtained, the mean CFU-S/colony, M1, and the probability of self-renewal, p, of the CFU-S were calculated. These values are used to calculate the number of cell cycles undergone during development of the colony and, by making certain assumptions, the cell cycle time of the CFU-S. The plot of p against log M for the various samples measured should be linear if all CFU-S proliferate at the same rate in a growing colony. It is not linear, however, so that CFU-S obtained under different experimental conditions do not all undergo the same number of cycles. In general, treatments given to the mice result in a lowering of the capacity for self-renewal of their CFU-S and also to a shortening of their cell cycle time. Some of the possible implications of these findings are discussed.
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  • 113
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 114
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 385-392 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effect of culture conditions on calf dorsal aorta endothelial cells was studied. Population doubling time varied as a function of the cell seeding density, growth medium, serum supplement, and concentration of fibroblast growth factor (FGF). The shortest population doubling time was found for cells (population doubling level 0-30) grown in Eagle's Minimal Essential Medium (MEM) supplemented with 10% fetal bovine serum (FBS) and 100 ng/ml FGF. The stimulatory effect of FGF on bovine endothelial cell proliferation was dependent on cell inoculation density. FGF significantly increased cell division rate at cell inocula less than 1 ± 104 cells/cm2 but not at higher densities. The population doubling time and cell size increased as the mass culture population doubling level increased. The replicative lifespans of bovine endothelial cells grown in medium supplemented with 20% FBS were 10-15% greater than parallel cultures supplemented with 10% FBS. Cultures grown in medium supplemented with 10% FBS and 50 ng/ml FGF showed a 50% increase in replicative lifespan compared to cultures grown in medium supplemented with 10% FBS alone. When FGF was used the increase in the number of doublings was a function of the length of time the cells were grown in the presence of FGF. This report extends comparable observations on the in vitro aging of human diploid fibroblasts to bovine endothelial cells.
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  • 115
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Phenylalanine in high concentrations inhibits the growth of mouse A9 cells. Protein synthesis is inhibited earlier and more severely than RNA or DNA synthesis. Phenylalanine inhibits the uptake and decreases the intracellular pool of several amino acids. Certain amino acids added in excess reverse the phenylalanine inhibition. The strongest reversing amino acids appear to function by excluding phenylalanine. The phenylalanine inhibition does not appear to be due to a deficiency of any amino acid, but to the high intracellular phenylalanine concentration and/or an amino acid imbalance resulting from the large ratio of phenylalanine to other amino acids.
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  • 116
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 117
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 103 (1980), S. 1-9 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The specific activity of lactate dehydrogenase (LDH; EC 1.1.1.27) is induced two-fold by l-norepinephrine (NE) in C6TK- rat glioma cells, but not in NA mouse neuroblastoma cells or various other nonglial cells. Previous reports have shown that the induction is mediated by cyclic AMP (cAMP) and possibly protein phosphorylation, and that it requires RNA and protein synthesis. To study the block to LDH induction in nonglial cells, we hybridized C6TK- cells with NA cells and isolated a hybrid clone in which LDH is inducible by NE. Mouse and rat LDH from hybrid cells were separated by electrophoresis and quantitated by two independent methods, and it was found that mouse and rat LDH were induced equally when cells were exposed to NE. The results suggest that inducibility of LDH is not determined by a cis-acting control at the gene level, but rather by the presence or absence of an earlier component in the cAMP-mediated induction system, and that the induction system acts indiscriminately on all active LDH gene copies in the cell.
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  • 118
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In this study, the intracellular concentrations of six elements (mmole/kg dry weight) were directly measured in the muscle fibers of pectoralis major muscles of eight week old, genetically dystrophic and normal chickens by the X-ray microanalysis technique. The extent of muscle degeneration was evaluated by morphometric measurements of muscle fiber diameter and other histological changes. A significant increase in the concentration of intracellular sodium and chlorine was evident in dystrophic muscles. The concentration of intracellular sodium was 127.0 ± 35.0 in the muscle fibers of dystrophic chicks compared to 65.7 ± 16.5 in normal controls. The concentration of chlorine was 90.5 ± 27.5 and 54.1 ± 5.5 in the muscle fibers of dystrophic and normal chicks respectively. The intracellular concentrations of potassium, magnesium, phosphorous, and sulfur remained unchanged in the dystrophic condition. Morphometric studies revealed that the dystrophic pectoralis muscles contain fewer but thicker fibers per unit area compared to normal pectoralis muscles. The importance of these findings are discussed in relation to the results of earlier investigations.
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  • 119
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 104 (1980), S. 163-170 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cellular fibronectin is expressed only at very low levels on the surface of the spontaneously transformed Chinese Hamster Ovary (CHO) cell line, as detected by immunofluorescence studies and confirmed by resistance of CHO cells to complement-mediated lysis in the presence of anti-fibronectin antibody. Treatment of CHO cells with sodium butyrate results in a marked susceptibility to anti-fibronectin complement-mediated killing. Using selected complement-containing sera from rabbits, it is possible to demonstrate that killing of butyrate-treated CHO cells is absolutely and specifically dependent on the presence of antibody to fibronectin. The increased susceptibility of butyrate-treated cells to complement-mediated killing correlates with an increase in cell surface fibronectin detected by immunofluorescence studies. This increased antigen expression should allow the isolation of mutants with altered regulation of fibronectin expression in a cell line of proven value for somatic cell genetic studies.
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  • 120
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 104 (1980), S. 153-162 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: BALB/c or DBA/2 mice were infected with Abelson murine leukemia virus (A-MuLV), pseudotype Molony murine leukemia virus (M-MuLV). Infection of these mice with 104 focus-forming units of A-MuLV (M-MuLV) induced overt leukemia, detectable grossly or microscopically in 90% of the mice at 20-38 days. However, these methods did not detect leukemia at 17 days or before. Bone marrow cells from A-MuLV-infected leukemic or preleukemic mice were placed in tissue culture in a soft agarose gel. Cells from leukemic or preleukemic BALB/c mice grew to form colonies of 103 cells or more, composed of lymphoblasts, whereas marrow cells from normal uninfected mice did not. Cells from these colonies grew to form ascitic tumors after intraperitoneal inoculation into pristane-primed BALB/c recipient. Colony-forming leukemia cells could be detected in the marrow of A-MuLV-infected mice as early as 8 days after virus incoluation. The number of colony-forming leukemia cells increased as a function of time after virus inoculation.Colony-forming leukemia cells require other cells in order to replicate in tissue culture. Normal bone marrow cells, untreated or after treatment with mitomycin-C, provide this “helper” function. Only in the presence of untreated or mitomycin-C treated helper cells was the number of colonies approximately proportional to the number of leukemia cells plated.Marrow cells from leukemic BALB/c mice form more colonies than those from leukemic DBA/2 mice. The number of colonies formed per 103 microscopically identifiable leukemia cells plated was determined to be 2-3 for leukemic BALB/c mice and 0.3 for DBA/2 mice. Cocultivation of leukemic DBA/2 marrow cells with mitomycin-C treated normal BALB/c cells did not increase the number of colonies formed by the DBA/2 leukemic cells. Thus, the decreased ability of DBA/2 leukemia cells to form colonies appears to be a property of the leukemia cell population.
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  • 121
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The transport of selected neutral and cationic amino acids has been studied in Balb/c 3T3, SV3T3, and SV3T3 revertant cell lines. After properly timed preincubations to control the size of internal amino acid pools, the activity of systems A, ASC, L, and Ly+ has been discriminated by measurements of amino acid uptake (initial entry rate) in the presence and absence of sodium and of transportspecific model substrates. L-Proline, 2-aminoisobutyric acid, and glycine were primarily taken up by system A; L-alanine and L-serine by system ASC; L-phenylalanine by system L; and L-lysine by system Ly+ in SV3T3 cells. L-Proline and L-serine were also preferential substrates of systems A and ASC, respectively, in 3T3 and SV3T3 revertant cells. Transport activity of the Na+-dependent systems A and ASC decreased markedly with the increase of cell density, whereas the activity of the Na+-independent systems L and Ly+remained substantially unchanged. The density-dependent change in activity of system A occurred through a mechanism affecting transport maximum (Vmax) rather than substrate concentration for half-maximal velocity (Km). Transport activity of systems A and ASC was severalfold higher in transformed SV3T3 cells than in 3T3 parental cells at all the culture densities that could be compared. In SV3T3 revertant cells, transport activity by these systems remained substantially similar to that observed in transformed SV3T3 cells. The results presented here add cell density as a regulatory factor of the activity of systems A and ASC, and show that this control mechanism of amino acid transport is maintained in SV40 virus-transformed 3T3 cells that have lost density-dependent inhibition of growth, as well as in SV3T3 revertant cells that have resumed it.
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  • 122
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 105 (1980), S. 63-72 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: There have been very few studies on ribonucleotide reductase activity in human tissue. In this report we describe a rapid and convenient procedure for determining purine and pyrimidine ribonucleotide reduction in normal human diploid fibroblasts and use the method to examine some general properties of the activity in these cells. ADP and CDP reductase was characterized for its response to the positive effectors, ATP and dGTP, the negative effector dATP, and the reducing agent dithiothreitol. Apparent Km values for ADP and CDP were determined to be 0.1 mM and 0.04 mM respectively. The antitumor agent hydroxyurea inhibited both purine and pyrimidine reductase in a noncompetitive fashion, giving Ki values of 0.40 mM and 0.41 mM for ADP and CDP respectively. These Ki estimates are about four to five times higher than those reported for some permanent cell lines. An examination of the cytotoxic effects of hydroxyurea indicated a close correlation between the concentration of drug which inhibited enzyme activity and decreased colony-forming ability.Clearly the ability to investigate ribonucleotide reduction in low numbers of normal human diploid cells will be useful for genetic and biochemical studies.
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  • 123
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    Journal of Cellular Physiology 105 (1980), S. 93-103 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Viable interspecies cytoplasmic-nuclear hybrid cells were constructed by fusion of karyoplasts prepared from the highly tumorigenic A9 mouse fibroblast cell line and cytoplasts prepared from the Detroit 532 normal human diploid cell strain. The identity of the hybrid cells was ascertained using a variety of morphological, immunological, and genetic criteria, including: nuclear pattern of staining with the fluorochrome Hoechst 33258, appearance of the actin-myosin containing cytoskeleton, presence of fibronectin, and resistance to azaguanine and diphtheria toxin. About 90% of the hybrid cells were viable, that is, capable of division. Changes in the morphology of the hybrid cells, apparently nuclear directed, were observed before cell division occurred. Using the techniques described here, large numbers of interspecies hybrid cells suitable for many types of biochemical analyses can be routinely produced.
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  • 124
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    Journal of Cellular Physiology 105 (1980), S. 143-152 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The influence upon differnt cellular and humoral parameters of hemopoiesis of three structurally unrelated, highly purified bacterial cell-wall components (BCWC) was investigated. The spleens of C57BL/6 mice assayed 6 days after the injection of either lipid A or outer-membrane lipoportein, but not murein, showed a marked increase in granulocyte-macrophage, eosinophil, and megakaryocyte progenitor cell levels. The number of pluripotent hemopoietic stem cells (CFU-S) also increased in the spleens of mice treated with either lipid A or lipoprotein. Similar results were obtained following the injection of lipoprotein or lipid A into CBA or C57BL/6.nu mice. Genetically anemic Wf/Wf mice were found to have spontaneously elevated numbers of splenic progenitor cells, which increased further after the injection of lipid A. The proportions of the different splenic progenitor cell types were similar in both untreated and lipid A treated Wf/Wf mice, and in normal littermate controls.When tested in vitro, unfractionated or partially purified post-lipid A serum was found to stimulate the growth of granulocyte-macrophage progenitor cells (GM-CFC), but no detectable stimulation of eosinohphil, megakryocyte, or erythroid progenitor cells was observed. The data suggest that the rise in splenic levels of the different progenitor cells is not mediated by the corresponding types of CSF, but more likely by proliferation and differentiation of CFU-S.
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  • 125
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    Journal of Cellular Physiology 105 (1980), S. 181-184 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 126
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    Journal of Cellular Physiology 105 (1980) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 127
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    Journal of Cellular Physiology 105 (1980), S. 185-185 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 128
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    Journal of Cellular Physiology 105 (1980), S. 191-196 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Macrophages, when maintained in vitro, take up glucose from the medium and oxidize it to CO2. The rate of oxidation of glucose varies considerably, depending on the physical state of the cell preparation. Cells in suspension oxidize glucose at a level six-fold that of cells in monolayers. The differences cannot be attributed to change in the rates of transport of glucose. On the other hand, an increse in intracellular glycogen (about three-fold) and free glucose plus glucose-6-P (many-fold) was found in the cells prepared as monolayers. During subsequent incubation with glucose-14C, this could be the cause of an isotope dilution effect and could explain the lower production of 14CO2 by the adherent cells. Since oxidation of glucose-1-14C to 14CO2 is used by many investigators to indicate the functional state of macrophages, we suggest close attention be paid to the system used, i.e., monolayers vs. suspensions.
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  • 129
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    Journal of Cellular Physiology 105 (1980), S. 235-246 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A human osteosarcoma cell line, U-2 OS, cultured under serumfree conditions, was shown to produce a growth factor (osteosarcoma-derived growth factor, ODGF) for human-cultured glial cells, fibroblasts, and other cells. ODGF, collected from the spent medium of 2 OS cultures, was purified by a sequence involving heparin-Sepharose chromatography, hydrophobic chromatography, gel chromatography, and preparative gel electrophoresis in SDS. Purified ODGF, at a concentration of 3 ng/ml, elicited a mitogenic response in human glial cells equivalent to 50% of that afforded by human serum at a final concentration of 1%. The preparation was estimated to be 〉 50% pure.The biological activity of ODGF resided in a cationic, relatively heat-resistant, reduction-susceptible protein with a molecular weight of 30,000 (by gel chromatography and SDS-gel electrophoresis). The electrophoretic behaviour of radioiodinated ODGF suggested that the protein was composed of two different polypeptide chains (about 13,000-14,000 and 16,000-17,000 daltons, respectively) linked via disulphide bonds. The molecular makeup of ODGF was thus similar to that of platelet-derived growth factor. 125I-ODGF could be precipitated by an antibody to platelet-derived growth factor, indicating that the two factors were immunologically related. Resemblance with platelet-derived growth factor was also indicated by the finding that the latter (but not, e.g., fibroblast growth factor or epidermal growth factor) competed with 125I-ODGF for binding to human-cultured glial cells.
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  • 130
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    Journal of Cellular Physiology 105 (1980), S. 259-266 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have analyzed ongoing DNA replication in ts BN-2, a dna- mutant of BHK-21 cells (Nishimoto et al. '78). At the non-permissive temperature of 39.5°C, inhibition of 3H-thymidine into acid-precipitable material begins 1 to 2 h after the cells are released from a block at the start of the S-phase. The fraction of nuclei incorporating 3H-thymidine is similar to that of wild-type cells through the synchronized S-phase of 8 h. Alkaline sucrose gradient analysis shows tht pulse-labeled DNA from mutant cells is incorporated into high molecular weight material after 3 h at eithr the permissive or non-permissive temperature. DNA fiber autoradiograms reveal that, at 39.5°C, the rate of replication fork movement is about 30% increased in the mutant as compared to wild-type cells. In the mutant cells, however, the interval between adjacent initiation sites is increased and the relative frequency of initiation events is decreased at the restrictive temperature. The results indicate that there is a block to ongoing replication in ts BN-2 at the level of initiation of synthesis on individual replication units; elongation of nascent chains is not inhibited.
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  • 131
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    Journal of Cellular Physiology 105 (1980), S. 335-346 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Protein degradation has been measured in confluent monolayers of eleven lines of contact-inhibited cells and ten transformed lines as the rate of release of trichloroacetic acid-soluble radioactivity after prelabeling cell protein with [3H]leucine. Insulin, at concentrations from 10-12 M to 10-6 M, has been added at the beginning of the 4-hour degradation period to detect selective effects of this hormone as an inhibitor of the inducible proteolysis occurring in serumfree medium. In addition insulin binding measurements have been performed on selected cell lines in an attempt to relate receptor properties to insulin action. Substantial effects of insulin are found in most cells with a selective inhibition at low insulin concentrations noted in several of the transformed lines. The difference in insulin sensitivity is not entirely definitive because temperature-sensitive transformation mutants of NRK cells are not more sensitive to insulin at a temperature where they show the transformed phenotype. Although insulin receptors on different cell lines have similar binding properties, two of the hepatomas used, H35 and MH1C1, show inhibition of protein degradation at insulin concentrations where receptor occupancy is extremely low. Calvarial osteoblast-like cells have a high rate of protein degradation which can be reduced by growth factors but not by insulin. The lack of an insulin response is a consequence of poor insulin binding to the cells. Insulin binds to the osteogenic sarcoma cells in substantial amounts. However, its normal action to inhibit the induced proteolysis is restricted because with these cells no increase of proteolysis occurs in serum-free medium. Generally higher rates of protein degradation are observed in the contact-inhibited lines than the transformed cells. We suggest that this difference may provide a selective growth advantage to transformed cells.
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  • 132
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    Journal of Cellular Physiology 105 (1980), S. 313-325 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A key attribute of the stringent response of bacteria is the rapid inhibition of ribosomal RNA synthesis mediated by unusual nucleotides in response to uncharged tRNA. The question as to whether mammalian cells show a stringent response analogous to that of bacteria was critically tested by the effective rapid amino acid starvation of both normal and transformed cells. Rapid starvation giving a high proportion of uncharged tRNA for leucine was produced within 7 minutes of expression of a nonleaky ts leucyl tRNA synthetase mutation in transformed CHO cells (tsH1) and in its normal growth control revertant (L-73). To control for the effect of temperature alone, tsrevertants of tsH1 and L-73 were included in the study, and to control for effects due simply to the inhibition of protein synthesis, the translational elongation inhibitor cycloheximide was used. In addition, rapid starvation for histidine was effected by incubation of both the CHO cell lines and of freshly explanted normal Chinese hamster embryo fibroblasts in histidine-free medium containing high concentrations of histidinol.The rate of preribosomal RNA synthesis and the extent of its maturation to mature rRNA was measured using (3H-methyl) methionine as a donor of methyl groups during synthesis and methylation of pre-rRNA. There was no effect on pre-rRNA synthesis of the rapid generation of uncharged tRNA for 45 minutes for any of the cell types tested. A nonspecific inhibition of maturation of 18S rRNA and late (3 hour) inhibition of pre-rRNA synthesis was observed, but could be mimicked by the inhibition of protein synthesis to comparable levels with cycloheximide. Less severe amino acid starvation resulting in a more physiological inhibition of protein synthesis to 30% also had no specific effect on pre-rRNA synthesis and maturation.Intracellular nucleotide pools were also examined for the appearance of unusual nucleotides such as guanosine tetraphosphate or pentaphosphate and for changes in the levels of normal nucleotides after severe amino acid starvation. No such changes could be detected.We conclude that although mammalian cells may have some biochemical reactions which respond to uncharged tRNA, they do not possess a macromolecular control system analogous to the stringent response of bacteria.
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  • 133
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    Journal of Cellular Physiology 105 (1980), S. 355-361 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The formation of erythroid colonies from bone marrow and spleen cells infected with the polycythemic strain of the Friend virus (FV-P) was characterized in an in vitro methyl cellulose colony-forming system in response to prostaglandin E1 and the beta-2 adrenergic agonist, albuterol. Both drugs markedly inhibited the formation of CFU-E colonies of FV-P-infected bone marrow and spleen in the absence or presence of erythropoietin. The albuterolmediated inhibition of CFU-E colonies (FV-P-infected) was selectively blocked by butoxamine, a beta-2 antagonist. Adenylate cyclase (AC) activity was also determined in FV-P spleen membrane preparations in response to albuterol and PGE1. Both agents stimulated enzyme activity, and butoxamine blocked the stimulation seen with albuterol. The ability of albuterol and PGE1 to stimulate AC activity in the FV-P-infected cells suggests that the effects of these agents on CFU-E formation may be mediated by specific beta-2 adrenergic and PG receptors through the adenylate cyclase-cyclic AMP system.
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  • 134
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    Journal of Cellular Physiology 105 (1980), S. 369-378 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Primary cultures of baby mouse kidney epithelial cells can grow without fibroblast overgrowth in a hormone-supplemented serum-free medium (Medium K-1) designed for an established kidney epithelial cell line, MDCK. The five supplements in Medium K-1 are insulin, transferrin, PGE1, T3, and hydrocortisone. Medium K-1 also supports the growth of kidney epithelial cell cultures from a number of animls, including man, withoug fibroblast overgrowth. Outgrowth of kidney epithelial cells from kidney explants was also observed with Medium K-1. Thus, the medium appears to be selective for epithelial cell growth.The physiological properties of primary cultures of baby mouse kidney epithelial cells were studied in detail. Baby mouse kidney epithelial cells grew at equal rates (0.5 doublings/day) in Medium K-1 and serum-supplemented medium. Medium K-1 also supported the formation of baby mouse kidney epithelial colonies at low cell densities. The dependence of baby mouse kidney epithelial colony formation upon the five factors in Medium K-1 was examined. These studies indicated that the formation of baby mouse kidney epithelial colonies in defined medium depended upon all the five supplements in Medium K-1, in a manner similar, although not identical, to MDCK colonies.Primary cultures of baby mouse kidney epithelial cells grown in Medium K-1 retained kidney cell-associated properties, including the ability to form multicellular domes, a phenomenon associated with transepithelial salt transport. Amiloride-sensitive Na+ uptake and the mucosal surface enzyme leucine aminopeptidase were also observed in baby mouse kidney cultures. Similar functions were observed in MDCK monolayers.
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  • 135
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    Journal of Cellular Physiology 105 (1980), S. 401-407 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have examined a number of events relating to ADP-ribose metabolism during serum-stimulated growth of BHK-21/C13 fibroblasts. Both the intracellular NAD+ content and the ADP-ribose polymerase activity were found to increase after serum stimulation of cells that were previously arrested by growth in low-serum medium. NAD+ content increased about two-fold, reaching a maximum of 4.2 nmol/μg of DNA 8 hr after serum step-up. The polymerase exhibited a sharp rise in activity, reaching a peak at about 5 hr after step-up; the activity declined below initial values by 10 hr, and then increased again to reach a plateau at 20 hr. We also report evidence which suggests a possible effect of ADP-ribosylation on the activity of DNA-dependent RNA polymerase I. The activity of this enzyme is diminished in isolated nuclei, and in a subsequent (NH4)2SO4 extract, when the nuclei are incubated with NAD+, the substrate for poly(ADP-ribose) polymerase. This inhibitory effect on the RNA polymerase is abolished when nuclei are incubated also with nicotinamide, a powerful inhibitor of the poly(ADP-ribose) polymerase.
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  • 136
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    Journal of Cellular Physiology 105 (1980), S. 423-429 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Studies were performed to determine whether decreases in transport of calcium and glucose might be among the earliest changes triggered by the antigen-antibody reactions occurring on the cell surface of murine leukemia L5178Y cells after treatment with rabbit antisera. After treatment with antisera, in the absence of complement, these cells exhibited a decreased uptake of 45Ca, 2-deoxy[3H]glucose, and 3-0-methyl[3H]glucose. These changes occurred rapidly, within 2 minutes after the addition of antiserum, in contrast to the previously reported inhibitory effects of antiserum on DNA, RNA, and protein synthesis, which became demonstrable only after 4 to 8 hours. The kinetics of uptake of the radioactive substrates was biphasic, with a very rapid initial uptake followed by less rapid linear uptake. The precise mechanism of cell growth inhibition remains to be elucidated, but one of the initial effects of antiserum treatment may be a perturbation at the cell membrane such that transport of specific nutrients is decreased, resulting in the observed effects on macromolecular synthesis.
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  • 137
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    Journal of Cellular Physiology 105 (1980), S. 461-468 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The intracellular pH (pHi) of sea urchin eggs was measured using 31P nuclear magnetic resonance (NMR) spectroscopy. The mean value of pHi of unfertilized eggs was about 6.2 (H. pulcherrimus and A. crassispina) to 6.3 (P. depressus). In contrast to results of other studies, pHi of sea urchin eggs was not changed after fertilization. During exposure of the eggs to NH4Cl or procaine-containing natural sea water (NSW), however, pHi rose about 0.4-0.8 pH units; the pHi fell to its initial value upon washing the eggs with weak basefree NSW. These changes of pHi by weak base treatment agreed well with the data obtained by other workers. In order to understand the discrepancy of pHi changes in fertilized eggs between NMR data and other measuring procedures, we measured acid production and O2 uptake, so that CO2 accumulation and proton release did not result in alkalinization. The invariance of the fertilized eggs under anaerobic conditions; otherwise NMR showed a different answer from other measuring procedures, because of its particular characteristics such as non-destructivity and compartmentation of pH.
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  • 138
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    Journal of Cellular Physiology 105 (1980), S. 489-502 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Rabbit reticulocytes were incubated with rabbit transferrin labelled with 59Fe and 125I in media in which the NaCl was replaced by other electrolytes or sucrose. Iron and transferrin uptake by the cells was affected by changes in the pH, ionic strength, ionic composition, and the osmolarity of the medium. Uptake was maximal at pH 7.4. A reduction in ionic strength produced by replacing NaCl with sucrose inhibited the uptake in a concentration-dependant manner, greatest inhibition occurring at lowest salt concentration. Similar results were obtained when KCl, LiCl, RbCl, Na2SO4, or K2SO4 were used instead of NaCl. Low ionic strength was found to inhibit the endocytotic uptake of transferrin labelled with colloidal gold, but had only a small effect on transferrin binding to cell membrane receptors. It was concluded that low ionic strength inhibits iron uptake primarily by blocking the endocytosis of transferrin. Three salts, NH4Cl, CaCl2, and MgCl2, produced different results from the above. NH4Cl inhibited iron uptake at all concentrations used. This action was due to an effect on the release of iron from transferrin, which appeared to be taken up by the cells in a normal manner. When the ionic strength of the sucrose medium was increased by adding low concentrations of CaCl2, iron uptake was greater than with equivalent concentrations of NaCl. However, with CaCl2 concentrations above 10 mM, iron uptake was inhibited, due to inhibition of transferrin uptake, possibly by blocking endocytosis. By contrast, MgCl2 stimulated iron uptake at all concentrations used. The results are discussed in terms of the possible effects of ionic strength, pH, and ionic composition of the extracellular fluid on the three main steps involved in iron uptake by immature erythroid cells: transferrin-receptor interaction, endocytosis, and iron release from transferrin.
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  • 139
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    Journal of Cellular Physiology 105 (1980), S. 513-517 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A single subcutaneous injection of the active androgen DHT (5α-androstan-17β-01-3-one or 5α-dihydrotestosterone; 5 mg/Kg body weight) into rats 7 days after orchidectomy stimulated DNA synthesis in the seminal vesicle, which began between 30 and 35 hr after injection and peaked at 48 hr. Mitotic activity began between 35 and 45 hr after DHT injection and peaked around 55 hr. These proliferative responses appeared to be confined mainly to the epithelial cells of the seminal vesicles. The DHT injection did not rapidly (between 5 min and 2 hr) increase the cyclic AMP level, but it did result in two small later surges of the cyclic nucleotide; the first of which peaked between 18 and 28 hr, but well before the onset of DNA synthesis, and the second of which peaked between 48 and 60 hr, when both the DNA-synthetic and mitotic activities were near or at their peak values.
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  • 140
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    Journal of Cellular Physiology 105 (1980), S. 527-532 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: TsAF8 is a temperature-sensitive (TS) mutant of BHK21 cells that arrests at nonpermissive temperatures in the mid-G1 phase of the cell cycle. TsAmaR-1 is a TS for growth mutant of CHO cells with a Ts- and α-amanitin-resistant (AmaR) RNA polymerase II activity. Hybrid TsAmaR-1 x TsAF8 cell lines were constructed at permissive temperatures. Such hybrid cells did not grow at nonpermissive temperatures; the two TS mutations did not complement. Two different AmaR derivatives of TsAF8 were isolated. Each contained only AmaR polymerase II activity, indicating that this RNA polymerase II gene locus in TsAF8 is functionally hemizygous, as would be expected for a locus in which the recessive TsAF8 mutation had occurred. One of these AmaR isolates of TsAF8 had a partially reverted TS+ phenotype. Taken together these results suggest that the TS mutation in TsAF8 is in RNA polymerase II.
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  • 141
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    Journal of Cellular Physiology 105 (1980), S. 565-570 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: For malignant cells cultured from a human astrocytoma, electrophysiological characteristics of the plasma membrane included specific resistivity of 446.82 ± 279.5 ohm·cm2, specific capacitance of 0.758 ± 0.52 microfarads/cm2, time constant 0.318± 0.10 msec. The resting membrane potential averaged-14.07 ± 7.4 mV; the mean input resistance 8.1 ± 4.0 megohms. The average cell area was 1638 ± 585 ±2 for contactual and 1919 ± 989 ±2 for noncontactual cells. Changes in input resistance and resting membrane potential were observed with increasing time in culture, possibly reflecting cell cycling. There did not appear to be electrical coupling in this cell line.
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  • 142
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    Journal of Cellular Physiology 102 (1980), S. 37-43 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Exogenous ATP has been shown to cause a rapid and reversible increase in permeability in transformed 3T3 cells (3T6 and SV3T3) but not in untransformed 3T3 cells. The cells remain viable, but lose intracellular acid-soluble pools. Treatment of transformed cells with ATP greatly reduces incorporation of 14C-leucine into protein, which is restored by the incubation of the cells with Dulbecco's modified Eagle's medium or by the external additions of certain ions and energy sources. tRNA is not required for the restoration of protein synthesis. In the permeabilized cells the energy for protein synthesis can be provided by glycolysis, oxidative phosphorylation, or direct addition of ATP. These studies demonstrate the usefulness of this method for studying the control of metabolism and macromolecular synthesis in monolayer cultures of transformed mammalian cells.
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  • 143
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    Journal of Cellular Physiology 102 (1980) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 144
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    Journal of Cellular Physiology 102 (1980), S. 333-341 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Secretion of a granulocyte-macrophage colony-stimulating factor (GM-CSF) was accomplished by L-P3 cells in culture with a serum-free medium. Cell proliferation per se was not requisite for the production of GM-CSF; the cells continued secreting GM-CSF even after their growth had been suspended. The amount of GM-CSF accumulated in the conditioned medium was reasonably accounted by the daily rate of production, and the addition of a proteinase inhibitor such as leupeptin and pepstatin did not result in greater accumulation of GM-CSF in the culture. It is thus postulated that there is no significant proteolytic inactivation of the secreted GM-CSF in the culture. However, when partially purified GM-CSF preparation was chromatographed on a gel-filtration column in the presence of 0.1% Triton X-100, a derivative of the GM-CSF was yielded which had been diminished in the molecular weight and altered in the isoelectric point. On the other hand, when leupeptin was included in the solution during production and isolation of the factor, the yielded GM-CSF did not manifest such a detergent-induced transformation and maintained its isoelectric point at pH 3.5. It is thus assumed that, in the presence of the detergent, GM-CSF suffers deterioration by an endogenous proteinase and releases a sialoglycopeptide fragment without loosing its colony-stimulating activity.
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  • 145
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    Journal of Cellular Physiology 103 (1980), S. 179-191 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: UV damage to CHO cell DNA, measured by formation of thymine-containing dimers, increases from mitosis to early S phase. Computer simulation of UV absorption by the DNA of an idealized CHO cell at different stages in the cell cycle resembles the cycle dependence of UV damage.Incision at UV damage sites, measured by the accumulation of breaks in preexisting DNA during 30 minutes' post-irradiation incubation with the DNA synthesis inhibitors 1-β-D arabinofuranosylcytosine and hydroxyurea, increases from mitosis to interphase. Analysis of the dose dependence of DNA break accumulation indicates that both the affinity of the endonuclease for dimer sites and the maximum enzyme activity at saturating levels of dimers are significantly lower in mitosis than in interphase.The killing of CHO cells by UV is enhanced if repair is temporarily inhibited by ara C. The DNA gyrase inhibitor novobiocin prevents UV-induced incision.
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  • 146
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    Journal of Cellular Physiology 103 (1980), S. 173-178 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When primary cultures of hepatocytes are maintained for 2 weeks from the time of perfusion, the activity of the enzyme glucokinase decreases rapidly, so that the activity can no longer be detected after the fourth day in culture. Concomitantly, there occurs an increase in the activity of hexokinases, the low-KM isozymes, which predominate in fetal liver. We have made several modifications of the culture medium in an attempt to prevent the decrease in glucokinase activity. When the medium was supplemented with a mixture of insulin, thyroxine, glucagon, dexamethasone, testosterone, and estradiol, the activity of the enzyme in the hepatocytes was present at approximately 15% of in vivo levels after 2 weeks in culture. When this hormone mixture was present during the first 4 hrs of culture and when the hepatocytes were allowed to attach to the collagen support and were maintained thereafter in medium supplemented with fetal bovine serum, insulin, and dexamethasone, the activity of glucokinase increased after an initial decrease for 3 days and was maintained thereafter at levels comparable to those observed in vivo. This effect of the hormone mixture was found to be the result of the presence of glucagon in the mixture, since the presence of glucagon with no other hormones added, except insulin, during the attachment period produced the same pattern of increased glucokinase activity. Immunoprecipitation of glucokinase from the hepatocytes, using monospecific antibody, indicated that the increase in enzyme activity was the result of increased glucokinase enzyme protein and not an increased synthesis of the other hexokinase isozymes. These studies demonstrate the specific hormonal requirements for the maintenance of glucokinase levels in primary hepatocyte culture at those seen in vivo and lends support to the hypothesis that fetal gene expression in primary hepatocyte cultures is selectively regulated rather than being a general effect with a common regulatory mechanism.
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  • 147
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    Journal of Cellular Physiology 103 (1980), S. 201-208 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Treatment of murine bone marrow cultures with the cholinergic agonist carbamylcholine enhanced megakaryocytic colony growth by as much as 65%. In contrast, adrenergic agonists had no such effect. Addition to cultures of dibutyryl cyclic GMP (db-cGMP) also enhanced megakaryocytic colonies up to 50%, whereas dibutyryl cyclic AMP (db-cAMP) had no effect. Sodium nitroprus-side and sodium nitrite, putative guanyl cyclase activators, also enhanced colony numbers, as did imidazole, a postulated cGMP phosphodiesterase inhibitor. Preincubation of marrow for two hours with carbamylcholine resulted in both an increase in colony numbers (58%) and percent of progenitors in DNA synthesis (48%, compared to 14% for controls) as determined by tritiated thymidine suicide studies. Treatment of mice with the acetylcholinesterase inhibitor neostigmine resulted in an increase in CFU-M/humerus (62%) and percent in DNA synthesis (45%). These data indicate that (1) cholinergic, but not adrenergic, agonists modulate megakaryocytopoiesis in culture; (2) this effect may be mediated by cyclic GMP; and (3) only a brief period of exposure of marrow cells to agonist results in enhancement of megakaryocytic colonies.
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  • 148
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    Journal of Cellular Physiology 103 (1980), S. 209-216 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two predictions of the error/mutation hypothesis of cellular sensecence (Orgel, '73) namely, (a) exponential accumulation of somatic mutations during the replicative lifespan and (b) shortening of culture lifespan upon treatment with mutagens have been examined experimentally in a strain of cultured human diploid fibroblasts. Our studies show that as cells traverse the replicative lifespan (from 10 to 75 mean population doublings (MPD); total lifespan ⋍ 95 MPD), no rapid and exponential increase occurs in the accumulation of mutations measured by the frequencies of Thgr (thioguanine resistance) and Dipr (diphtheria toxin resistance) mutants. Furthermore, repeated cycles of treatment (from 1- to 14-times) of human fibroblasts with two mutagens, ethyl methane sulfonate (EMS) and N-methyl-N′nitro-nitrosoguanidine, which led to a marked increase in the mutation frequency for the Dipr marker (⋍ 100-fold), failed to shorten the lifespan of cultured fibroblasts. On the contrary, repeated mutagen treatment (12 times with EMS) prolonged the lifespan of one replicative culture (110 MPD versus 94-98 MPD). These results strongly indicate that mutations are unlikely to be the primary event in cellular senescence and suggest instead that senescence is probably controlled by one or more (specific) gene(s) whose expression can be modified by mutations.
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  • 149
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    Source: Wiley InterScience Backfile Collection 1832-2000
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    Notes: Molecular changes occur at the surface of hemopoietic cells during differentiation from progenitor cells to mature granulocytes and macrophages. The differential expression of surface carbohydrate residues has been probed using lectins and the results used to purify normal mouse granulocyte-macrophage progenitor cells. Ten different lectins were screened for selective interaction with mouse hemopoietic colony-forming cells (CFCs), using agglutination or a quantitative analysis of the number of fluoresceinated lectin molecules bound per cell using a fluorescence activated cell sorter (FACS). Pokeweed mitogen (PWM), Helix pomatia agglutinin (HPA), soybean agglutinin (SBA), and peanut agglutinin (PNA) preferentially bound to CFCs so that it was possible to enrich 4 to 10-fold for these progenitor cells by sorting for the highly fluorescent cells. Further analysis of the low and high angle light scattering characteristics of the CFCs indicated that these cells were polydisperse, but could be enriched ten-fold by selecting for cells with high intensity low angle (90°) scatter and low intensity high angle (90°) scatter. PWM gave the best enrichment (10 to 15-fold) for adult bone marrow CFCs, for CFCs from fetal sources (fetal liver, fetal blood), and for CFCs from the spleens of mice injected previously with outer membrane lipoprotein from E. coli. Three parameter sorting for CFC using the FACS (low angle scatter, high angle scatter, and PWM-fluorescence) resulted in large enrichment factors (16 to 50-fold) for CFCs from all the above sources. Over 7% of the cells sorted from bone marrow, 10% of the cells sorted from post-lipoprotein spleen, and 28% of the cells sorted from fetal peripheral blood were hemopoietic CFCs. Ninety percent of the cells in these fractions had the morphology of blast cells or myelocytes. Thus, it was possible to identify the morphological characteristics of the hemopoietic progenitor cells. Screening of other developmental systems using quantitation of fluorescence with lectins should prove of general value for the purification of selected differentiation states.
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  • 150
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    Journal of Cellular Physiology 103 (1980), S. 239-246 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Addition of prostaglandin E2 (PGE2) significantly altered the cellular composition of murine long-term bone marrow cultures. After 4-5 weeks of culture, increased cellularity in the suspension phase was observed in all cultures containing prostaglandin. These suspension cells contained markedly higher proportions of differentiated neutrophils than did cells cultured in the absence of PGE2. Granulocyte-macrophage progenitor cell levels in the suspension layer were increased 3-20 fold after five weeks in prostaglandin-containing cultures compared with control cultures. Fewer cells comprised the adherent layer in cultures containing prostaglandin. The number of macrophages in this layer was reduced 3-8 fold in these cultures compared with control cultures, while the number of granulocytes was increased 2-3 fold. The progenitor cells biased toward macrophage development were selectively inhibited in the cultures with PGE2. There was no significant effect of PGE2 on pluripotent stem cell levels or on the longevity of the cultures. It is concluded that excessive monopoiesis in bone marrow may be limited by PGE2 without influencing either stem cell maintenance or the development of other marrow-derived cell types.
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  • 151
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    Journal of Cellular Physiology 103 (1980), S. 271-278 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Adsorption of Sendai virus to HeLa cells induced in them an increased permeability to K+, Na+, Ca++, deoxyglucose, but not to fluorescein. The stimulation of uptake of 42K was temperature-dependent, did not occur below 15°C, and was not inhibited by ouabain.The virus-induced increase in the uptake and release of 42K and of 3H deoxyglucose could not be mimicked by treatment of cells with linoleic acid, a procedure which increased the fluidity of the cellular membranes. The stimulatory effect of 0.5 mM ATP on the release of deoxyglucose was enhanced several fold in the presence of Sendai virus. These results seem to indicate the possible involvement of membranal enzymes such as e.g. protein kinase in the permeability changes induced by Sendai virus.
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  • 152
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    Journal of Cellular Physiology 103 (1980), S. 263-269 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Phosphatidylinositol (PI) synthesis and its role in controlling the cell cycle has been investigated using fibroblasts and liver cells in culture. PI synthesis as measured by incorporation of [3H]-myo-inositol into trichloroacetic acid precipitable material during 0-60 min after serum or growth factor stimulation of serum-starved cells is increased in primary fetal rat liver cells, rat embryo fibroblasts, and 3T3 mouse cells. In contrast, growth stimulation of 3T3 cells and hepatocytes rendered quiescent in G1 by amino acid starvation is not accompanied by increased incorporation of [3H]-myo-inositol into trichloroacetic acid precipitable material. This suggests that those cells might be arrested at a different point in G1 than cells arrested by serum depletion. Inhibition of PI synthesis by δ-hexachlorocyclohexane (HCH), a steric analog of myo-inositol, during early times (e.g., 0-4 hr) after growth stimulation, reversibly blocks initiation of DNA synthesis in 3T3 cells. The results support the idea that increased PI synthesis in response to growth stimulation in the cell types studied here is a prerequisite for progression through G1 and subsequent entry into S phase.
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  • 153
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    Journal of Cellular Physiology 103 (1980), S. 299-303 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: To ascertain whether the fidelity of protein synthesis declines during cellular aging in vitro, we have developed a cell-free protein synthesizing system from cultured human fibroblasts which actively incorporates phenylalanine into acid-insoluble material upon addition of poly (U). The accuracy of poly(U)-directed protein synthesis was determined by comparing the ratio of leucine to phenylalanine incorporation in extracts of early- and late-passage fibroblasts derived from normal persons and from subjects with two genetic disorders of premature aging, progeria, and Werner syndrome. The results show no decline in translational fidelity at late passage or in prematurely aging cells, and thus fail to support the error catastrophe theory of cellular aging.
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  • 154
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    Journal of Cellular Physiology 102 (1980), S. 259-266 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cultured rat embryo fibroblasts, when placed in media with 10% serum containing 20 mM NH4Cl, show an inhibition of protein degradation and, concurrently, an accumulation of numerous, large vacuoles, partially filled with cellular debris. Cells placed in a serum-free media exhibit an enhanced degradation of cell protein, which is also inhibited by NH4Cl. When these cells are removed from media containing NH4Cl and placed in fresh media, the material accumulated in these vacuoles is rapidly and quantitatively released to the media in both an acid-soluble and acid-insoluble form. NH4Cl inhibits rapidly and specifically the lysosomal proteolytic mechanism, and is without effect on the basal turnover mechanism. The lysosomal proteolytic mechanism accounts for approximately 25% of protein turnover, and, at least in low density cultures, can be stimulated to levels which account for more than half of the protein turnover in the cell. The major pathway for the degradation of fast turnover proteins appears to be separate from lysosomal mechanism.
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  • 155
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    Journal of Cellular Physiology 102 (1980), S. 279-286 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of L-cell conditioned medium which contains granulocyte/macrophage colony stimulating factor (CSF); of highly purified L-cell CSF; and the antiserum directed against L-cell CSF, have been investigated in long-term murine bone marrow cultures. Treatment of cultures with CSF containing conditioned medium led to a rapid decline in haemopoiesis. However, this inhibition of in vitro haemopoiesis is probably caused by materials other than CSF, since the addition of highly purified L-cell CSF had no appreciable effect upon long-term haemopoietic cell proliferation or differentiation. Furthermore, the inhibitory activity of L-cell conditioned medium was not abrogated following neutralization of the CSF activity by CSF antiserum. The direct addition of CSF antiserum did not inhibit granulocyte or macrophage formation. These results suggest that long-term cultures of murine marrow cells may show extensive interactions with stromal cells which are not influenced by exogenous stimulatory or inhibitory factors.
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  • 156
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    Topics: Biology , Medicine
    Notes: BFU-E from the blood of 14 normal adults have been grown by the plasma clot technique. The hemoglobins synthesized in burst colonies were purified from other proteins by affinity chromatography on Sepharose-haptoglobin. The radioactivity incorporated in the globin chains was estimated by CM-cellulose chromatography in urea. The number of bursts scored at the 14th day of culture fluctuated between 50-130 (average 86, s: 29) for 106 mononuclear plated cells. A constant reactivation of fetal hemoglobin was found (from 1.4% to 11%, mean value 5.8%, s: 3.07), but was lower than previously described, mainly because of the highly selective purification of Hb. This reactivation of fetal hemoglobin was not dependent upon the concentration of erythropoietin (from 1 U/ml to 6 U/ml) nor on the purity of the erythropoietin preparations (from 6 U/mg of protein to 70 000U/mg of protein). In addition, the same subject exhibited a constant proportion of Hb F synthesized in culture over a period of time up to 6 months. A positive correlation exists between the proportion of Hb F in culture and that of F cells present in the blood, with the exception of two subjects. Such findings suggest that Hb F in culture is a characteristic of each individual and that this reactivation often represents an amplification of the Hb F synthesis in vivo.
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  • 157
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    Journal of Cellular Physiology 102 (1980), S. 317-321 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The growth of human mammary epithelial cells is stimulated by cholera toxin and analogues of cyclic AMP, while the growth of breast derived fibroblasts is inhibited. These compounds have little effect on DNA synthesis in the absence of other mitogens but show a synergistic effect with serum and/or EGF. The results suggest that high intracellular levels of cyclic AMP in human mammary epithelial cells increase the growth response of the cell to mitogens.
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  • 158
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    Journal of Cellular Physiology 102 (1980), S. 343-349 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Lectin-dependent neutrophil cytotoxicity against autologous human red cells was studied using an 111In(indium)-release assay. Human red cells were not readily killed by neutrophils in the presence of phytohemagglutinin (PHA). However, removal of red cell membrane sialic acids (desialylation) markedly enhanced their susceptibility to PHA-dependent neutrophil cytotoxicity. This neutrophil cytotoxicity was dependent on the energy supplied by anaerobic glycolysis, but it was independent of erythrophagocytosis. Catalase, superoxide dismutase, KCN, and Na azide did not inhibit PHA-dependent neutrophil cytotoxicity. Neutrophils from a patient with chronic granulomatous disease, in the presence of PHA, also killed desialylated red cells normally. On the other hand, desialylation of neutrophils had no effect on the expression of their cytotoxic effect. The results suggest that desialylated red cells are much more susceptible to lectin-dependent neutrophil cytotoxicity than normal red cells, and that lectin-dependent neutrophil cytotoxicity is independent of reactive oxygen species.
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  • 159
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    Journal of Cellular Physiology 102 (1980), S. 351-365 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Granulocytic and erythrocytic colonies developed on the chick embryo chorioallantoic membrane (CAM) following the inoculation of chick embryo spleen cells. Dose response and kinetic experiments showed that the colonies were derived from cell aggregates present in the inoculum. Dissociation and reaggregation studies of the CAM colony-inducing cells (CAM-CIC) indicated that these cells must be present as aggregates in order to form colonies. Results from the morphology and cell marker experiments suggested that the colony-inducing aggregates (CAM-CIA) attract and support the differentiation of primitive host hemopoietic cells. The physical characteristics of the CAM-CIC, which are different from those of the hemopoietic progenitor cells, indicated that they represent a stromal cell population of the chick embryo spleen. Further evidence supporting this notion was provided by the radiation studies which showed that the colony-inducing ability of the CAM-CIC is radioresistant. The above characteristics of the CAM-CIC strongly suggest that they represent the stromal cells of the chick embryo spleen which influence hemopoiesis.
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  • 160
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    Journal of Cellular Physiology 102 (1980), S. 385-393 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Rat peritoneal macrophages were studied for their ability to undergo metabolic cooperation. Macrophages were unable to cooperate with human fibroblasts. This was true for macrophages which had been activated in vivo as well as for macrophages treated with various agents in vitro. Macrophages were also unable to undergo metabolic cooperation with rat fibroblasts or with other macrophages. In contrast, rat reticular cells, mesothelial cells, and fibroblasts were able to cooperate with human fibroblasts.
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  • 161
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    Journal of Cellular Physiology 103 (1980), S. 371-383 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Erythroid colony formation in agar cultures of CBA bone marrow cells was stimulated by the addition of pokeweed mitogen-stimulated spleen conditioned medium (SCM). Optimal colony numbers were obtained when cultures contained 20% fetal calf serum and concentrated spleen conditioned medium. By 7 days of incubation, large burst or unicentric erythroid colonies occurred at a maximum frequency of 40-50 per 105 bone marrow cells. In CBA mice the cells forming erythroid colonies were also present in the spleen, peripheral blood, and within individual spleen colonies. A marked strain variation was noted with CBA mice having the highest levels of erythroid colony-forming cells.In CBA mice erythroid colony-forming cells were mainly non-cycling (12.5% reduction in colony numbers after incubation with hydroxyurea or 3H-thymidine). Erythroid colony-forming cells sedimented with a peak of 4.5 mm/hr, compared with CFU-S, which sedimented at 4.25 mm/hr. The addition of erythropoietin (up to 4 units) to cultures containing SCM did not alter the number or degree of hemoglobinisation of erythroid colonies.Analysis of the total number of erythroid colony-forming cells and CFU-S in 90 individual spleen colonies gave a correlation coefficient of r = 0.93 for these two cell types.In addition to benzidine-positive erythroid cells, up to 40% of the colonies contained, in addition, varying proportions of neutrophils, macrophages, eosinophils, and megakaryocytes. Taken together with the close correlation between the numbers of CFU-S in different adult hemopoietic tissues, including individual spleen colonies, the data indicate that the erythroid colony-forming cells expressing multiple hemopoietic differentiation are members of the hemopoietic multipotential stem cell compartment.
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  • 162
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    Journal of Cellular Physiology 103 (1980), S. 393-398 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Epithelial and fibroblast cells from adult rat liver were found to differ markedly in their metabolism of the purine hypoxanthine. Both cell types took up hypoxanthine and possessed hypoxanthine-guanine phosphoribosyl transferase for phosphoribosylating the purine. However, in the transferase assay, lysates from epithelial cells converted hypoxanthine predominantly to inosine monophosphate, with small amounts of the nucleoside inosine as product, whereas fibroblast cell lysates converted hypoxanthine predominantly to inosine. The inosine appeared not to be produced by direct ribosylation of the base, since fibroblast cell lysates had less purine nucleoside phosphorylase activity than epithelial cell lysates. Rather, the inosine produced by fibroblast lysates appeared to be derived from inosine monophosphate through catabolism of the mononucleotide by 5′ nucleotidase. An inhibitor of 5′ nucleotidase, thymidine triphosphate, reduced the amount of inosine formed.
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  • 163
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    Journal of Cellular Physiology 103 (1980), S. 407-416 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The water permeability of V-79 Chinese hamster lung fibroblasts was determined by measuring the rate of cell shrinkage in hypertonic medium using a cell sizer. The water permeability appears to follow Arrhenius kinetics as a function of temperature with a sharp discontinuity at 21°C. An activation energy of 7.0±1.6 kcal/mole was found below 21°C and 22.8±3.1 kcal/mole above 21°C. The correlation time of rotation of the spin label 2,2-dimethyl-5-dodecyl-5-methyloxazolidine-N-oxide was measured as a function of temperature in the cellular membranes, and shows a break at 20°C. A discontinuity was also found in the membrane to water partitioning of the spin label 2,2-dimethyl-5-pentyl-5-butyloxazolidine-N-oxide near 20°C. These breaks may correspond to a membrane lipid phase transition. Dimethylsulfoxide, in the concentration range of 0.2-0.5 M, decreases the water permeability by a factor of two.
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  • 164
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    Journal of Cellular Physiology 103 (1980), S. 447-453 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mouse myeloma cell lines synthesize large amounts of immunoglobulin on their membrane-associated polyribosomes. Variants which no longer synthesize immunoglobulins have been studied and shown to have the same number of tightly bound membrane-associated polyribosomes as the parental cell lines. These polyribosomes are still active in the synthesis of nonimmunoglobulin proteins.
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  • 165
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    Journal of Cellular Physiology 104 (1980) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 166
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    Journal of Cellular Physiology 103 (1980), S. 489-502 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Previous studies (J. Biol. Chem, 253: 99-105, 1978) showed that thyrotropin-releasing hormone (TRH) acutely stimulated uridine uptake in pituitary cell (GH4C1) cultures. Studies on the role of protein synthesis in this response to TRH led to the finding that an inhibitor of ribosomal translation, cycloheximide, also stimulated uridine uptake acutely. Studies reported here attempt to determine the mechanism of cycloheximide action and whether cycloheximide and hormone stimulation of uridine uptake occurred by similar pathways. The experiments presented indicate that: (1) seven inhibitors of ribosomal translation stimulated uridine uptake; (2) in contrast, inhibition of protein synthesis at tRNA aminoacylation resulted in reduced rates of uridine uptake; (3) inhibition of tRNA aminoacylation blocked cycloheximide but not TRH stimulation of uptake; (4) cycloheximide stimulation of uptake was restricted to amino acid-depleted cultures; (5) amino acid supplementation stimulated uridine uptake with a time-course identical to that of cycloheximide; (6) cycloheximide and amino acid supplementation promoted reacylation of cellular tRNAs in amino acid-depleted cultures; and (7) cycloheximide stimulation of uridine uptake resulted from enhanced nucleoside phosphorylation rather than increased uridine transport. We conclude that cycloheximide and amino acid stimulation of uridine phosphorylation may be mediated through a common pathway involving the extent of amino-acylation of cellular tRNAs. Furthermore, cycloheximide and TRH stimulate uridine phosphorylation by pathways that are distinguishable. It is apparent that not all cellular effects of cycloheximde can be attributed solely to inhibition of the synthesis of proteins.
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  • 167
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    Journal of Cellular Physiology 104 (1980), S. 11-19 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Spleen cells were collected from normal mice and cultured in a medium containing 20% calf serum. Addition of lipopolysaccharide (LPS) in the culture significantly increased the production of granulocyte-macrophage colony-stimulating factor (GM-CSF), and a maximum induction was attained in 5 days. Irradiation of the spleen cells with 300 to 3,000R X-rays also enhacned the production of GM-CSF, but there was a latent period of about 5 days before the factor appeared in the culture medium. The observed difference between LPS and X-rays in the timing of inducing GM-CSF production in the spleen cell culture was consistent with the difference in timing of the increase of spleen cell proliferation observed in animals after the administration of LPS or during recovery from damages by X-irradiation. It was observed furthermore that the X-ray-induced GM-CSF differed from the LPS-induced GM-CSF in its molecular properties; the X-ray-induced factor was represented by an acidic (pI=3.0) 70,000-dalton species, while the LPS-induced factor was much smaller in size (M.W. 20,000) and less acidic (pI=5.4). These results suggest that different mechanisms of GM-CSF production operate in the spleen in response to either LPS or X-rays.
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  • 168
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    Journal of Cellular Physiology 104 (1980), S. 41-46 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A mouse embryo epithelial cell line, MMC-E, was used to study the effects of purified murine sarcoma growth factors (SGFs) on the growth of epithelial cells. Murine SGF was a partially purified preparation from the serum-free culture media of mouse fibroblasts transformed by Moloney murine sarcoma virus. SGFs stimulated DNA-synthesis in resting cells and induced them to grow to higher densities than control cells. With continued exposure to SGFs, MMC-E cells lost their postconfluency inhibition of division and formed small expanding foci. When the SGFs were removed and the cells were subcultured, they regained their normal phenotype, showing that the effects of SGFs are reversible on these epithelial cells. SGFs could also stimulate the MMC-E epithelial cells to grow in soft agar, like the syngeneic fibroblasts (MMC-F) from the same mouse embryo, but slower than the control fibroblastic clone. Microgram quantitites of SGFs were needed to stimulate soft ager growth of MMC-E cells. The results indicate that SGF can bring about a phenotypic change in the growth pattern of epithelial cells as well as fibroblastic cells.
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  • 169
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    Journal of Cellular Physiology 104 (1980), S. 61-72 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Events that are essential for progression through the G1 period begin immediately or shortly after resting chick embryo cells are given fresh medium with serum. The following observations support the contention that the critical events include the production of non-ribosomal RNAs: (1) Addition to the “shift-up” medium of either of two inhibitors of RNA formation, comptothecin or 5, 6-dichloro-1-β-D-ribofuranosylbenzimidazole, delays the onset of DNA replication by about the length of time the cells are exposed to the drugs. (2) Although entry into the S phase is delayed by the inhibitors, the slopes of the DNA response curves are identical to that of control cultures. (3) Neither drug reduces significantly the rate of overall protein synthesis. Observations (2) and (3) are taken to mean that expansion of the G1 period is not due to cell damage. (4) A third inhibitor of RNA synthesis, cordycepin, also delays passage of stimulated cells throgh the G1 phase, but, in this case, the length of the delay period is greater than that of the exposure period. (5) A low dose of actinomycin D does not impede movement towards the S phase, even though the synthesis of preribosomal RNA is considerably reduced.The possibility is considered that the essential G1 molecules are mRNAs.
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  • 170
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    Journal of Cellular Physiology 104 (1980), S. 97-103 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Deprivation of extracellular Ca or serum inhibits the proliferation of WI38 human diploid fibroblasts. Under these conditions, the cells become quiescent at a point in the cell cycle typical of early G1 or Go phase-arrested cells. Exit of the cells from this point in the cycle appears to require both the presence of serum and Ca simulataneously. If quiescent cells are serum-stimulated in low Ca medium (0.01 mM), they do not progress through G1 to the S phase, which normally requires 14-18 hr. However, they remain competent to do so. Addition of Ca for up to 48 hr after serum stimulation results in an equal fraction of the cells progressing G1 phase as compared to the presence of Ca at the time of serum addition. In contrast, if quiescent cells are serum-stimulated in the presence of Ca, which is then removed, the cells can remain competent to enter S phase for only 10-12 hr. Re-addition of Ca beyond this time does not allow G1 progression on a normal schedule.These data suggest that Ca and serum are both required to trigger, in whole or in part, the pleiotypic response. Ca appears also to render the cells competent to enter S phase, but this competence is labile; an observation consistent with the PDGF-induced competence observed previously in the 3T3 cell. These observations are in contrast to previous data from other cell types which suggest that Ca is required only in late G1 phase for successful entrance to S phase.
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  • 171
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    Journal of Cellular Physiology 104 (1980), S. 127-136 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: An aging process has been detected in stationary 3T3 cell cultures, especially in the presence of plasma-derived serum (PDS) from adult bulls. It leads to irreversible conversion of an increasing percentage of initially responsive cells of a stationary population into cells unresponsive to growth stimulation by newborn calf serum (NBCS) or reseeding at low cell density in the presence of NBCS. These unresponsive cells are viable in the sense that, following trypsinization, they reattach and spread on a new culture plate and can be maintained for many days. The conversion process is accelerated by increasing PDS concentration. It is antagonized by NBCS. It is accompanied by enhancement of growth-inhibiting interactions exerted by stationary 3T3 cell populations on SV 40-3T3 cells.
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  • 172
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    Journal of Cellular Physiology 102 (1980), S. 19-26 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: An intraperitoneal injection of the β-adrenergic agonist dl-isoproterenol hydrochloride (100 mg/Kg body weight) into a rat caused an early, very large (400-fold) cyclic AMP surge (peaking at 10 minutes) in the parotid gland which was followed by a second, much smaller (two-fold) surge 12 to 16 hours later. DNA synthesis began about 16 to 20 hours after the isoproterenol injection and peaked between 24 and 28 hours. The maximum level of DNA-synthetic activity at 24 hours was correlated positively to the magnitude of the small cyclic AMP surge at 12 hours, but not to the size of the much larger cyclic AMP surge at 10 minutes. An intraperitoneal injection of dl-propranolol hydrochloride (59 mg/Kg body weight) at 8 hours after isoproterenol injection abolished the second cyclic AMP surge at 12 hours and markedly (65-75%) reduced the incorporation of [3H]-thymidine into DNA. Injection of dibutyryl cyclic AMP (6.3 mg/Kg body weight) and theophylline (25 mg/Kg body weight) at 8 hours prevented propranolol from inhibiting DNA synthesis. Propranolol appeared specifically to affect the cyclic AMP-dependent pre-DNA-synthetic step because it did not reduce [3H]-thymidine incorporation when injected after the second cyclic AMP surge had passed and DNA synthesis had just begun. Thus, the initial, large cyclic AMP surge following β-adrenergic stimulation may not be necessary for the initiation of prereplicative development, while the much smaller second surge may be needed for the initiation of DNA synthesis.
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  • 173
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    Journal of Cellular Physiology 103 (1980), S. 467-474 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The cell line B16/C3 will undergo melanogenesis at a specific time after plating. We have found that this time can be modulated by varying the pH of the culture medium. At high pH levels (8.2-8.6) the onset of melanogenesis occurs in 3 or 4 days, while at lower pH (6.7-7.2) it occurs in 7 or 8 days. Furthermore, the time of onset is also sensitive to the extracellular ionic strength. The addition of sodium lactate, sodium chloride, or any other salt tested delays or blocks completely the onset of melanogenesis. These effects are not simply consequence of growth inhibition, nor can they be correlated with patterns of lactate accumulation.These cells are sensitive to pH or ionic strength after entering the stationary phase just prior to the time of onset of melanogenesis. The existence of a specific pH-and ionic-strength-sensitive phase may provide an important clue to the events responsbile for differentiation in this system.
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  • 174
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    Journal of Cellular Physiology 103 (1980), S. 435-445 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Colony-stimulating factors (CSFs) stimulate granulocyte-macrophage production from single hemopoietic progenitor cells. Various preparations of purified CSFs of two different subclasses have been shown here to stimulate a plasminogen-dependent fibrinolytic (plasminogen activator) activity from resident and starch-induced mouse peritoneal macrophages. Lymphocyte supernatants also stimulate macrophage plasminogen activator (PA) activitty. Since they contain colony stimulating activity, it is possible that one or more sublcasses of CSF in these supernatants is responsible for this effect. Since both colony-stimulating and macrophage growth activities have been detected at inflammatory sites, these findings could reflect a role for CSF in inflammatory processes.
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  • 175
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Quiescent 3T3 cells can be stimulated to enter S by defined factors. When used in combination, three polypeptide hormones (EGF, vasopressin, and insulin), or a tumor promotor and insulin, are very effective in stimulating DNA synthesis. Like serum, the defined factors also stimulate deoxyglucose uptake and induce the synthesis of ornithine decarboxylase during G1. The second stage of deoxyglucose uptake and the induction of ornithine decarboxylase are protein synthesis-dependent events. When added with the growth factors, mouse interferon inhibits the synthesis of DNA and the induction of ornithine decarboxylase but has no effect on the uptake of deoxyglucose. Kinetic experiments comparing the effect of inhibitors of translation or transcription on induction of ornithine decarboxylase with the effect of interferon suggest that interferon may affect the synthesis of enzyme by inhibiting both transcription and translation of message. The findings provide further support for the proposition that interferon exerts a differential effect on mitogen-stimulated events which are dependent on continuous protein synthesis.
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  • 176
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    Journal of Cellular Physiology 103 (1980), S. 475-487 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: As constituents of both extracellular matrix and the cell surface, glycosaminoglycans are in a strategic position to influence several basic cell features. The localization and turnover of glycosaminoglycans was investigated in cultured normal human embryo fibroblasts of lung origin (IMR-90). Attention was directed particularly toward that compartment of the culture which could be released by gentle proteloysis (trypsin, 0.1 mg/ml, 15 min) and is considered to represent the cell surface. In the presence of Na2SO4, sulfated glycosaminoglycans (S-GAGs) of the cell surface were labeled rapidly, but within 30 min some 35S-GAG appeared in the extracellular medium. The intracellular pool of S-GAGs labeled during a 10-min period was lost during the first hr of chase with a half-life of 18 min, compared with 16 hr for S-GAGs labeled over a 48-hr period. Pulse-labeled S-GAGs of the surface turned over with an initial half-life of 60 min, compared with 7 hr for surface material labeled over a 48-hr period. These rapid movements of the early chase period were followed by similar movement at a much slower rate. The results are consistent with a model in which most of the S-GAGs synthesized in the cell move rapidly to the surface. The surface GAGs are then released immediately to the medium or accumulate at the membrane to be shed more slowly at a later time or to be degraded. The S-GAG which left the cell layer most rapidly during chase was dermatan sulfate, while heparan sulfate made up an increasing percentage of the cell layer as chase progressed. These cultures produce a fibrillar matrix of fibronectin, but the kinetics of this study suggest that the S-GAGs of the surface are membrane-bound, and an extracellular glycosaminoglycan matrix does not form.
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  • 177
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    Journal of Cellular Physiology 104 (1980), S. 21-26 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mitogenesis of human blood lymphocytes in culture is inhibited by concentrations of ouabain that are approximately one order of magnitude lower than those that block Na and K transport. For example, the 50% inhibition (ID50) of Na-K transport, 280 nM, is seven-fold greater than the ID50 for RNA synthesis, DNA synthesis, or blastogenesis, ˜40 nM. Yet, inhibition of transport and consequent reduction in cell K is considered responsible for the effects of ouabain on mitogenesis. Since synthetic processes are assessed at least 24 hours after lymphocyte stimulation, this discrepancy could be explained by either 1) a progressive increase in K leak, or 2) a progressive inhibition of Na-K transport by ouabain during 24 hours of PHA treatment. We found that the lymphocyte membrane leak rate of K increased immediately after PHA treatment but did not increase further from 4 to 24 hours. In contrast, the ouabain sensitivity of 42K uptake was markedly increased with time: ID50 for 42K uptake of 35 nM at 24 hours as compared to 280 nM at 30 minutes. Measurement of ouabain binding revealed a seven-fold increase in the lymphocyte-associated ouabain after 24 hours compared to binding at 1 hour. These data indicate that the dose response of ouabain inhibition of active K transport and lymphocyte proliferation are closely correlated if one considers the slow membrane binding of ouabain at low concentrations.
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  • 178
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    Journal of Cellular Physiology 104 (1980), S. 35-40 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effect of modifying culture pH on the adipose convesion of BALB/c 3T3 cells was studied. We have previously shown that the tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) stimulates cellular lactic acid production, lowers the pH of the culture medium, and inhibits differentiation in this system. We now report that addition or organic acids such as D- or L-lactic acid or acetic acid to the culture medium mimicked the effect of TPA by lowering pH, stimulating cellular lactic acid production, and inhibiting the adipose conversion. Lowering the pH by changing the NaHCO3 concentration also inhibited differentiation.A comparison of Dulbecco's modified minimal essential medium (DMEM) vs minimal essential medium (MEM) indicated that in the former media, the rate and extent of differentiation was greater than in the MEM, and TPA neither inhibited differentiation nor stimulated lactate production. Our standard MEM medium could be made similar to DMEM with respect to the cells' response to TPA simply by raising the NaHCO3 concentration, hence, the buffering capacity, of MEM to that present in DMEM, suggesting that TPA can only inhibit differentiation in this system if it can lower culture pH via lactate production.
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  • 179
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    Journal of Cellular Physiology 104 (1980), S. 53-60 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Transmembrane potential (Em) and alpha-aminoisobutyric acid (AIB) transport were measured in primary monolayer cultures of rat hepatocytes obtained from unoperated control rats and from rats 12 hr following partial hepatectomy. Measurements were performed 20-24 hr after plating the cells. The capacity of both kinds of cells to concentrate AIB depended upon extracellular sodium; however, the steady-state accumulation in regenerating cells was twice that of control cells. Transmembrane potentials, recorded with glass microelectrodes, were -13 ± 0.6 mV and -27 ± 1.6 mV in control and regenerating cells, respectively. Ouabain (1 mM) depolarlized regenerating cells to -18 ± 1.0 mV, but it had no effect on control cells. The initial rates of 1 mM AIB transport into control and regenerating cells were 1.2 ± 0.1 and 3.1 ± 0.1 nanomoles/mg protein × 4 min, respectively. Ouabain (1 mM) reduced the initial rate of AIB transport into regenerating cells to 2.7 ± 0.1 nanomoles/mg protein × 4 min, but it had no effect on AIB transport into control cells. Glucagon (10-7 M) added to control cells 12 hr before measurements hyperpolarized Em to -31 ± 1.3 mV and increased AIB transport rate to 3.1 nanomoles/mg protein × 4 min. The results suggest a relationship between increases in Em and increases in AIB transport in rat hepatocytes. An electrogenic Na-K pump may be involved in both of these events.
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  • 180
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    Journal of Cellular Physiology 104 (1980), S. 73-81 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Resting cultures of primary chick embryo cells have been labeled with 3H-leucine for the first 2 or 3 hours after feeding basal medium or basal medium supplemented with 10% dialyzed serum. The labeling patterns of the 3H-polypeptides of the soluble cell fraction have been compared by fluorography of two-dimensional gels. Large and consistent differences are seen in only three of the more than 900 spots that can visualized. This report concerns two of the three spots. The 3H contents of the two polypeptides (41,000 daltons, pI 7.1, designated 41-7.1, and 34,000 daltons, pI 6.2, designated 34-6.2) are increased by serum by about ten-fold. The highly selective effect of serum on the labeling of the two spots does not appear to be an artifact related to the extractability, solubility, or state of aggregation of the polypeptides. The radio-intensities of both polypeptides decrease markedly when the cells are labeled later than 3 hours after “shift-up”.Drugs that inhibit RNA synthesis and are known to stop the progression of the chick cells through the G1 period, camptothecin, cordycepin and 5,6-dichloro-β-D-ribofuranosylbenzimidazole, depress, with great specificity, the enhanced labeling of polypeptide 41-7.1 in the stimulated cells, and all but camptothecin have a similar action on polypeptide 34-6.2. A high level of actinomycin D (10 μg/ml), but neither a low level of the drug (0.02 μg/ml) nor 5-fluorouridine prevents the increased labeling of the two polypeptides in serum-fed cells. That 5-fluorouridine enters the chick cells and is converted to its active form is shown by the inhibition of the processing of pre-ribosomal RNA.The observations with the RNA inhibitors are at least consistent with the conclusions that the enhanced labeling of the two sports results from increased rates of synthesis of the polypeptïdes that depend upon mRNA production but not on the formation of ribosomal RNAs, and that the polypeptides play a role in the regulation of DNA replication in the chick cells.
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  • 181
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have investigated the effects of the tumor promoter 12-0-tetradecanoyl-phorbol-13-acetate (TPA) on plasminogen activator production, hexose transport and metabolism, and the incorporation of choline into the acid soluble pool and into phosphatidylcholine in suspension cultures of mouse L, mouse P388 leukemia, human HeLa, and Chinese hamster ovary cells, and in monolayer cultures of baby hamster kidney (BHK), mouse 3T3, mouse 3T6, and mouse P388D1 macrophage-like cells. BHK, 3T3, P388D1, and P388 cells produced plasminogen activator constitutively, but no significant production was observed in the other cell lines. Plasminogen activator production was induced or stimulated by TPA only in P388 cells (10- to 20-fold by 100 ng TPA/ml). On the other hand, phosphatidylcholine synthesis was stimulated by TPA only in HeLa cells, and hexose transport, as measured with 3-0-methyl-D-glucose, only in 3T3 and P388D1 cells, as well as in human lymphocytes. The stimulation of hexose transport occurred more rapidly than the induction of plasminogen activator production and seemed to be part of the mitogenic response of cells to TPA treatment. A stimulation of deoxyglucose uptake was similarly limited to 3T3 and P388D1 cells. A significant decarboxylation of carbon 1 of deoxyglucose occurred in P388 and P388D1 cells, but not in Novikoff cells, and any decarboxylation that occurred was not stimulated by TPA. The results indicate that the various investigated responses of cells to TPA are unrelated and occur independent of each other. The time courses of the biochemical responses also differ significantly.
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  • 182
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    Journal of Cellular Physiology 104 (1980), S. 137-152 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cell lines resistant to ethidium bromide have been developed from cultured mammalian BHK21/C13 cells and these same cells transformed by Rous sarcoma virus (C13/B4). Cells resistant to 2 μg ethidium bromide per milliliter have been cloned. One clone of the control and one of the virus-transformed cell lines has been employed for characterization. The resistant cells, in the presence of 2 μg ethidium bromide/ml, grow at approximately the same rate as the untreated parental cells. The control cells possess a “normal” karyotype (44 chromosomes), while the corresponding ethidium bromide mutant has a reduced chromsome number of 41 and a number of translocations. The mitochondria displayed morphological alterations compared to the parental lines during the transition phase prior to the isolation of the ethidium bromide-resistant cells. The mitochondria of the ethidium bromide-resistant mutants appear somewhat enlarged with a normal morphology.The effect of ethidium bromide on selected respiratory enzymes in normal and virus-transformed ethidium bromide-resistant baby hamster kidney cells was determined. Ethidium bromide-resistant cells exhibited a depressed level of cytochrome aa3. This depression could not be reversed by growth in ethidium bromide-free media. Ethidium bromide-resistant cells possessed the same cytochrome b, c, and c1 levels per cell as their corresponding parental lines.Purified mitochondria isolated from virus-transformed ethidium bromide-resistant cells exhibited a depression in cytochrome oxidase-specific activity, while the ethidium bromide-resistant control cells did not. All cell lines studied showed a depression in NADH-ferricyanide and NADH-cytochrome c reductase-specific activities relative to parental BHK21/C13 cells. An increase in succinate dehydrogenase and succinate cytochrome c reductase-specific activities was observed in ethidium bromide-resistant control cells relative to their parental BHK21/C13 cells. No increase was observed in virus-transformed ethidium bromide-resistant cells.Ethidium bromide-resistant control cells exhibited a two-fold increase in oligomycin-insensitive adenosine triphosphatase activity relative to their parental cells. All of the cell lines studied possessed equivalent oligomycin-sensitive adenosine triphosphatase-specific activity except for the virus-transformed, dyeresistant mutant, whose activity was increased.
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  • 183
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    Journal of Cellular Physiology 104 (1980), S. 171-175 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The tyrosinase (EC 1.14.18.1) activity of cultured B-16 mouse melanoma cells (C2M) in the stationary phase depends greatly on whether the culture medium contains glucose or galactose. The activity in medium cotaining galactose was about ten times that in medium containing glucose at pH 7.2. This difference in tyrosinase activity was concluded to be due to a shift of balance between synthesis and degradation of the enzyme. Experiments were conducted with stationary phase cultures in the presence and absence of cycloheximide.The melanoma cells did not synthesize tyrosinase in medium containing glucose in the stationary phase. But when they were cultured under identical conditions, except that glucose was replaced by galactose, they continued to synthesize tyrosinase.The rate of synthesis in medium containing galactose at pH 6.3 was one third of that in the same medium at about pH 7, in which the increase in specific activity of tyrosinase per day was about 30 nmoles/mg cell protein per hr. The rate of degradation of the enzyme was practically the same in medium containing glucose as in medium containing galactose, and largely dependent on the pH of the culture medium. At pH 6.3 the half-life was about one third of that at pH 7.2, where it was about 1.8 days.The degradation at acidic pH values was much reduced by ammonium salt and was strongly inhibited by the protease inhibitor, leupeptin.
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  • 184
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    Journal of Cellular Physiology 104 (1980), S. 187-197 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Bone marrow from barrier-sustained specific pathogen-free (SPF) CBA and C57BL/6 mice gave relatively low numbers of BFU-E colonies in methylcellulose culture, as compared to conventional mice. Addition of thymocytes to the marrow cultures increased the yield of BFU-E colonies more than fourfold in SPF mice but only 1.5-fold in conventional mice. Colony size was also increased.Increased yield of BFU-E colonies was also obtained by co-culture of bone marrow with lymph node cells or with bone marrow or spleen cells from 900R whole-body-irradiated mice. The effect appeared to be cellular rather than humoral. It was not reproduced by conditioned medium from thymus or pokeweed mitogen stimulated spleen cells. The helper effect of thymus cells was eliminated or reduced by freezing and thawing, or by 48 hours of incubation after irradiation. Treatment of bone marrow cells in vitro with anti-theta serum and complement did not decrease the number of BFU-E colonies. The putative helper cells appear not to be T cells, were non-adherent to the plastic culture dish, and were cortisone resistant and radioresistant. The low BFU-E colony yield from SPF mouse marrow is presumed to be largely the result of deficiency of these non-T helper cells in SPF bone marrow, rather than of BFU-E progenitor cells.
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  • 185
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    Journal of Cellular Physiology 104 (1980), S. 215-223 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When human diploid fibroblasts IMR-90 are cultured in routinely used medium (Eagle's basal medium supplemented with 10% fetal calf serum), sulfhydryl compounds appear in the medium. The major component of these sulfhydryl compounds is cysteine, and it is shown that a part of medium cystine is converted into cysteine by the cells. It is also shown that the sulfhydryl groups of serum albumin, which are masked and barely detectable before the culture, are restored. Probably cysteine formed by the cells reacts with serum albumin to give rise to the protein sulfhydryl groups via sulfhydryl-disulfide exchange reactions. Total sulfhydryl concentrations in the medium are maintained in a considerable level throughout the culture, and a possible physiological function of these sulfhydryl groups is discussed.
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  • 186
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    Journal of Cellular Physiology 104 (1980), S. 241-248 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A novel synergistic effect of the antitumor agent alanosine (2-amino-3-(hydroxynitrosoamino) propionic acid), which specifically inhibits the enzyme adenylosuccinate synthetase (ASS) and guanine on the growth of Chinese hamster ovary (CHO) cells and human diploid fibroblasts (HDF) has been observed. In the presence of subinhibitory concentrations of alanosine, both CHO cells and the HDF show excessive sensitivity to exogenous guanine - a phenotype which closely resembles that seen with some of the mutants containing reduced enzymatic activity of ASS. The growth inhibitory effects of alanosine, or alanosine and guanine, on CHO cells are completely reverted by the addition of adenine to the culture medium, and the synergistic effect of guanine is not observed in mutants which lack the enzyme hypoxanthine-guanine phosphoribosyl transferase. These results suggest that guanine nucleotides exert a regulatory effect on the activity of the enzyme adenylosuccinate synthetase. The ability to confer the guaninesensitive phenotype and its modulation by subinhibitory concentrations of alanosine in different cell types indicates that alanosine provides a useful probe for investigating the regulation of purine nucleotide metabolism in mammalian cells.
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  • 187
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    Journal of Cellular Physiology 104 (1980), S. 249-252 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The relative importance of viral tumor antigen expression and the cellular background in the maintenance of a transformation phenotype was examined in five SV40-transformed teratocarcinoma-derived cell lines. These cell lines show qualitative differences in growth characteristics associated with transformation, and vary in their state of differentiation. Viral T antigen expression was evaluated by two criteria: 1) the amount of immunoprecipitated antigen in growing cells, and 2) the amount and rate of antigen synthesis in density-inhibited cells. There was no direct correlation found between retention, or rate of synthesis, of the viral T antigen and the degree of transformation. These findings imply that the cellular environment has a more important influence on the growth properties of a stably transformed cell than the quantitative levels of viral T antigen expression.
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  • 188
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    Journal of Cellular Physiology 104 (1980), S. 261-268 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The major integral membrane protein of red blood cells, the mouse equivalent of human band 3, was purified and used to raise a specific antiserum. The murine protein resembles its human counterpart in several of its properties, including susceptibility to digestion by chymotrypsin added to intact cells and an ability to bind to concanavalin A. The synthesis of 35S-labeled band 3 was detected in Friend erythroleukemia cells treated with DMSO by immuneprecipitation followed by SDS gel electrophoresis and fluorography. Induction with DMSO led to a greater than tenfold increase in the synthesis of band 3 and maximal synthesis was reached 3 to 4 days after the beginning of induction.
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  • 189
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    Journal of Cellular Physiology 104 (1980), S. 269-281 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of fresh medium and serum on protein synthesis in suspension-cultured HeLa cells after growth to high cell density (〉5 × 105 cells/ml) were studied. Cells which were resuspended in fresh medium plus serum and grown for 24 hours (control) were compared with cells grown for 2 hours after resuspension (stimulated). The spectrum of proteins being synthesized by control and stimulated cells does not appear to be grossly different; that is, the weight and number average molecular weights of newly synthesized whole-cell protein are about the same in both cultures. Also, no significant differences were observed in the number of ribosomes per polysome or in the fraction of total ribosomes in polysomes. However, the transit times (combined elongation and termination times) were found to differ significantly; the average transit time for control cells was 2.24 minutes, while the average transit time for stimulated cells was 1.26 minutes. (An appendex evaluating the methodology involved in measuring the transit time is included.) In agreement with the difference in transit time, the absolute rate of protein synthesis in stimulated cells was approximately 1.8 times the rate measured in control cells. These data are taken as evidence that under certain conditions, the rate of elongtion and/or termination of polypeptide chains limits the overall rate of translation, and that cells can respond to growth conditions by changing the elongation and/or termination rate of protein synthesis.
    Additional Material: 7 Ill.
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  • 190
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    Journal of Cellular Physiology 104 (1980), S. 295-308 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mouse hepatoma cells that secrete large amounts of α-fetoprotein (AFP) and albumin have been crossed with rat hepatoma cells that secrete only albumin, and in relatively small amounts, to investigate the influence of each parental genome upon the expression of serum proteins. All of the ten independent hybrid clones examined produce mouse AFP and both mouse and rat albumin; none produces rat AFP. The absence of production of rat AFP by the hybrids suggests that different mechanisms are involved in the initiation and in the maintenance of expression of this function. The secretion of the three proteins by the hybrid cells is coordinate: Whatever the growth phase (exponential or stationary) and irrespective of the amounts produced over a wide range, the ratio secreted of mouse AFP to mouse albumin is near to one, and that of mouse albumin to rat albumin is near to five. In addition, even though the pattern of protein secretion during the growth cycle of hybrid cells is different from those of both parents, the products of both parental genomes conform to the new hybrid pattern. Finally, some hybrids secrete less of the proteins with increasing numbers of cell generations, yet all three continue to be secreted in coordinate fashion. Since the rates of secretion of serum proteins probably reflect their rates of synthesis, we conclude that coordinate secretion indicates coordinate synthesis, and may reflect coordinate transcription of the relevant genes.
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  • 191
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    Journal of Cellular Physiology 104 (1980), S. 309-319 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The mode of permeation of uracil, 5-fluorouracil, and orotic acid into cells has been investigated in four established cell lines (Novikoff rat hepatoma, P388 mouse leukemia, mouse L, and Chinese hamster ovary cells) in attempts to assess the rate-determining step(s) in their incorporation into the nucleotide pool and nucleic acids. Uracil and 5-fluorouracil shared a saturable transport system (Km = 5 to 15 mM) capable of rapid equilibration of these substrates across the cell membrane (t1/2 at 25 in first-order range of concentration = 25 to 58 sec). Thus it seems unlikely that transport is limiting the incorporation hypoxanthine. Only the non-ionized form of fluorouracil was a substrate for the transporter; exclusion of charged pyrimidines may explain why orotate was not a substrate at physiological pH. Orotate permeated the cell membrane much more slowly (t1/2 = 2890 to 6930 sec); its permeation was apparently non-mediated and rate-determining in the conversion of extracellular orotate to intracellular nucleotides.
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  • 192
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    Journal of Cellular Physiology 104 (1980), S. 335-348 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The attachment and detachment behavior of three mouse fibroblast cell lines adhering to plastic tissue culture substrata coated with the serum protein cold-insoluble globulin (CIg) resembles that seen on the usual serumcoated substrata. The transformed cell line SVT2 spreads more extensively on the CIg-coated than on the serum-coated substratum, while the nontransformed Balb/c 3T3 line and concanavalin A-selected “revertant” of SVT2 are equally well spread on both substrata. In all three cases, immunofluorescence microscopy using antibodies to CIg suggests that the cells are more tightly apposed to the CIg-coated substratum than to the serum-coated substratum.Substrate-attached material (SAM), which contains cell-substratum adhesion sites and which is left after EGTA-mediated detachment of cells, is enriched for cell surface fibronectin and glycosaminoglycans (GAG). When cells are seeded onto CIg-coated substrata rather than serum-coated substrata, there is an increased deposition of GAG but a comparable deposition of cellular proteins. The protein distribution of the two types of SAM are identical as analyzed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis, including fibronectin content. This indicates that substratum-bound CIg cannot functionally substitute for cell surface fibronectin in these adhesion sites. Analysis of the GAG deposited on CIg-coated substrata reveals that hyaluronate and the chondroitins are increased to a much greater extent than heparan sulfate; however, the ratio of hyaluronate to the various chondroitin species is invariant. These data provide further evidence that hyaluronate and the chondroitins are deposited in adhesion sites in well-defined stoichiometric proportions, possibly as supramolecular complexes, and that CIg may mediate adhesion of cells in the serum layer by binding to GAG-containing proteoglycans.
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  • 193
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    Journal of Cellular Physiology 104 (1980), S. 349-357 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mouse myeloid leukemia cells (Ml) were induced to differentiate into mature macrophages and granulocytes by various inducers. The differentiated Ml cells synthesized and released prostaglandins, whereas untreated Ml cells did not. When the cells were prelabelled with [14C]arachidonate, the major prostaglandins released into the culture media were found to be prostaglandin E2, D2, and F2α in an early stage of differentiation, but the mature cells produced predominantly prostaglandin E2. The synthesis and release of prostaglandins were completely inhibited by indomethacin. Dexamethasone, a potent inducer of differentiation of Ml cells, did not induce production of prostaglandins in resistant Ml cells that could not differentiate even with a high concentration of dexamethasone. These results suggest that production of prostaglandins in Ml cells is closely associated with differentiation of the cells. Homogenates of dexamethasone-treated Ml cells converted arachidonate to prostaglandins, but this conversion was scarcely observed with homogenates of untreated Ml cells. Dexamethasone and the other inducers stimulated the release of arachidonate from phospholipids. Therefore, induction of prostaglandin synthesis during differentiation of Ml cells may result from induction of prostaglandin synthesis activity and stimulation of the release of arachidonate from cellular lipids. Lysozyme activity, which is a typical biochemical marker of macrophages, was induced in Ml cells by prostaglandin E2 or D2 alone, as well as by inducers of differentiation of the cells, but it was not induced by arachidonate or prostaglandin F2α. These results suggest that prostaglandin synthesis is important in differentiation of myeloid leukemia cells.
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  • 194
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    Journal of Cellular Physiology 104 (1980), S. 375-389 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cells of confluent cultures of the established pig renal epithelial line, LLC-PK1, accumulate α-methyl-D-glucoside against a concentration gradient. This transport system is strongly inhibited by phlorizin and 6-deoxy-D-glucose, moderately inhibited by phloretin, and only weakly inhibited by 3-0-methyl-D-glucose, paralleling the situation in mammalian kidney. The time course for the uptake of α-methyl-D-glucoside and for the carrier-mediated but passive uptake of 3-0-methyl-D-glucose are identical to those seen in mammalian kidney. Subconfluent cultures of LLC-PK1 cells are unable to accumulate α-methyl-D-glucoside, and their transport of this glucose analog is less sensitive to phlorizin inhibition than is the transport system in confluent cultures. Transmission electron micrographs show that cells from subconfluent cultures lack the microvillous surface seen in cells from confluent cultures. Cell density is thus a factor in the occurrence of structural and functional differentiated properties related to transport in these cells.
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  • 195
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Release of sulfated proteoglycans into the medium of fetal rat chondrocytes in monolayer culture was studied by contrasting the effects of 10% calf serum, long-acting cyclic nucleotides (8 Br-cAMP or DBcAMP), and lysine vasopressin (LVP). Eight hours after initiation of the experiment, the monolayer was pulsed for 2 hours with Na2[35SO=4], the radioactivity was chased, and the monolayer was reincubated for 6 hours with conditioned medium from replicate cultures. Immediately after labelling, the amount of newly synthesized sulfated proteoglycans was invariably higher in the insoluble matrix than in the medium compartment. Both additives selectively enhanced sulfate incorporation into chondroitin sulfate of the matrix when compared to serum controls, but only LVP stimulation caused increases in the medium. Remodeling (loss of cell layer and release into the medium at 6 hours) was suppressed by cAMP analogues and increased by LVP. This process was more active in cultures of lower cell density. Utilizing calibrated gel columns, no size difference of the glycosaminoglycans was found between the medium and cell layer compartments of the three treatment groups at the two time points. Because the cAMP analogues inhibit, while LVP stimulates cell division, our observations imply that the rate of degradation of the constraining matrix is increased when replication is favored, even when chondroitin sulfate synthesis is selectively stimulated.
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  • 196
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    Journal of Cellular Physiology 104 (1980), S. 425-431 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Viable cell samples containing 93% pure granulocytes were obtained from human blood using the techniques of dextran sedimentation followed by centrifugal elutriation. The resting transmembrane potential (Em) of human granulocytes was estimated using the fluorescent lipophilic cation, Di-S-C3(5), from the null point for potassium - i.e., the external K concentration at which there is no change in Em in response to valinomycin (a K ionophore). The Em of human granulocytes, as calculated from the Nernst potential for K at the null point, is approximately - 100 mV. Data indicate that this large transmembrane potential is due in part to the presence of an electrogenic Na-K pump in human granulocytes which is stimulated by external potassium and inhibited by ouabain.
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  • 197
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    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 105 (1980) 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 198
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    Journal of Cellular Physiology 104 (1980), S. 443-459 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Human lymphocytes contain a large, saturable fraction of K+ that exchanges slowly with K+ in the external medium, and a small non-saturable fraction that exchanges rapidly. We determined whether or not Na+ exchanges in a similar manner with external Na+. Cells were pre-equilibrated to ensure absence of net ion movements. Efflux was studied by loading with 22Na and transferring without washing to a non-labeled medium. Influx was studied by transferring to labeled medium and separating large samples of cells at 6,000g. There are fast, intermediate, and slow fractions of Na+ exchange, with half-times of 2, 14, and 120 minutes. At normal external K+, most cell Na+ exchanges rapidly, while at lower external K+ the Na+ that replaces cell K+ exchanges slowly. Parallel sources of fast and slow fractions, such as extracellular ones and subpopulations of cells, were ruled out by simultaneous 42K and 22Na fluxes and by a quantitative analysis of the combined K+ and Na+ content and flux data over a range of external K+ and Na+ levels. Five possible models of ion fluxes occurring in series were considered. Surface matrix, surface binding sites, and cytoplasmic channels with rapid nuclea exchange were eliminated as sources of the fast fractions. Therefore, the fast fractions of K+ and Na+ must reflect the permeability of the surface membrane. This left only two possible sources of the slow fractions. One, a subcellular compartment (e.g., nucleus), was eliminated by the combined content and flux data. We conclude that the slow fractions of ion flux are rate-limited by adsorption onto and desorption from cellular macromolecules. The data support the association-induction hypothesis and are understood by reference to two fundamental concepts: that of rapid solute exclusion from cell water existing in a polarized state; and that of solute accumulation limited by adsorption onto fixed anionic sites within the cell.
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  • 199
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    Journal of Cellular Physiology 104 (1980), S. 461-467 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: V79-8 cells lack a measurable G1 interval under normal growth conditions. We found that partial inhibition of protein synthesis using low levels of cycloheximide (0.05 μ/ml) could induce a measurable G1 in these cells without any significant effects on S, G2, or M. In view of these findings, recessive mutants selected from the V79-8 cell line, which each express G1, were analyzed for their rates of protein synthesis and degradation/loss. Three of the four mutants showed a decreased rate of protein synthesis sufficient to account for their G1 lengths. A fourth mutant, however, showed parental rates of both protein synthesis and degradation/loss. These results suggest not only that a G1 interval can be expressed as a result of a decreased rate of protein synthesis, but that other alterations (mutations) other than those simply affecting overall protein synthesis can result in the expression of a measureable G1 interval.
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  • 200
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    Journal of Cellular Physiology 105 (1980), S. 1-6 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Uptake of α-methyl-D-glucoside (AMG) by LLC-PK1 cells is inhibited by the uncoupler p-trifluoro-methoxyphenyl-hydrazone (FCCP) and by the absence of extracellular Na+, indicating that the transport system is energy- and Na+ -dependent. We have previously demonstrated that transport of AMG by LLC-PK1 cells proceeds against a concentration gradient and is phlorizin-sensitive (Mullin et al., '80). Uptake of AMG was also inhibited by ouabain (OUA) but not by ortho-vanadate (VAN). Rubidium uptake also was affected by OUA but not by VAN. VAN, however, caused collapse of the three-dimensional domes of confluent LLC-PK1 monolayers much more rapidly and thoroughly than OUA. Since domes are presumably dependent upon the Na+ pump, yet VAN is not acting on transport-related functions of the OUA-sensitive (Na+ + K+)-ATPase, we hypothesize a direct effect of VAN on the water permeability of these cells. We also suggest that OUA does not act on these cells until domes collapse in normal course, and access of the OUA to the extracellular surface of antiluminal membranes is then achieved.
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