bluuebunny/crossref_metadata_2025 huggingface dataset:
Circadian rhythms in the mitotic index of the basal epithelium and in the uptake rate of 3H‐thymidine by the tongue of the rat
Authors: Gasser Raymond F. , Scheving Lawrence E. , Pauly John E.
Published: February 2005

Circadian rhythms are demonstrated in the tongue of adult rats for both the mitotic index of the basal epithelium and the uptake rate of injected 3H‐thymidine by the tongue tip. The animals were entrained to a light‐dark cycle for four weeks prior to the experiments with the light phase extending from 0600 to 1800 hours (CST). The daily fluctuation is approximately 300% for the mitotic index and 185% for the uptake rate of 3H‐thymi‐dine. The highest mitotic index occurs at 1100, and the highest uptake of 3H‐thymidine occurs four hours earlier at 0700. The least activity for both parameters occurs during the first part of the dark span of the light‐dark cycle. Estimates of several other rhythmic parameters are determined by a computerized method.
Relationship between cytoagglutination and saturation density of cell growth
Authors: Weber Joseph
Published: February 2005

The agglutination with concanavalin A and wheat germ agglutinin of the established malignant cells, HEp 2, KB, HeLa, TDB‐3, HTC and RV 3T3, and of the putatively normal cells, BHK 21, 3T3 and Wi‐38 was examined as a function of their saturation densities in culture. A positive correlation between the saturation density of the cell lines and the capacity to agglutinate was demonstrated. Incubation for 15 minutes with 1.25 mg/ml of trypsin converted non‐agglutinating and poorly agglutinating cells into agglutinable ones, while leaving the highly agglutinating lines largely unchanged. The magnitude of change in agglutination after trypsin treatment correlated inversely with saturation density. Although the extent of agglutination varied with the saturation density, the agglutinability of a particular line remained relatively unchanged at different cell densities.
Selective hybrid cell formation by human lymphoid cells
Authors: Parkman Robertson , Hagemeijer Ann , Merler Ezio
Published: February 2005

Subpopulations of human lymphoid cells are capable of spontaneously fusing with fibroblasts of human or murine origin to form human‐human or human‐mouse hybrid cells. These cells were present in thymus, spleen, and bone marrow. After fractionation on discontinuous bovine serum albumin gradients, the cells were found in the less dense layers of the gradient. Cells of fetal origin, except for bone marrow, fused spontaneously at a higher rate than those of adult origin. The highest rate of fusion was found with adult bone marrow cells. These lymphoid cells appear to be “thymus‐derived” cells.
Nucleotide pools of novikoff rat hepatoma cells growing in suspension culture. IV. Nucleoside transport in cells depleted of nucleotides by treatment with KCN
Authors: Plagemann Peter G. W. , Erbe John
Published: February 2005

Incubation of Novikoff rat hepatoma cells in glucose‐free basal medium containing 2 mM KCN results in a rapid and almost complete loss of uracil and adenine nucleotides. By following the fate of radioactivity from 3H‐nucleoside pulse‐labeled cells during incubation with KCN it was shown that the nucleotides are degraded to nucleosides and bases which are released into the culture fluid. Depletion of the cells of nucleotides by incubation with KCN allows a direct analysis of the kinetics of uridine transport into the cell, since KCN‐treated cells fail to phosphorylate uridine. Uridine uptake follows normal Michaelis‐Menten kinetics with an apparent Kn of about 50 μm at 18°C. Uptake is by facilitated diffusion since it does not require energy and uridine is not transported against a concentration gradient. The effects of KCN are largely prevented by the presence of 10 mM glucose in the medium. They are also rapidly reversed by resuspending the cells in fresh medium without KCN. Upon removal of KCN, the cells rapidly regenerate their nucleotide pools and resume growth at the normal rate.
Cell separation by velocity sedimentation of postnatal mouse cerebellum
Authors: Barkley David S. , Rakic Ljiljana L. , Chaffee Jonathan K. , Wong Dona L.
Published: February 2005

Trypsinized cells of newborn mouse cerebellum have been separated by velocity sedimentation at unit gravity in shallow gradients of Ficoll. The two main technical difficulties were formation of gels around the dissociated cells and clumping of cells before and during the sedimentation procedure. These were solved by adding DNase to the dissociation medium and with holding serum, respectively. Proliferating cells of the external granular layer separated according to size differences in the cell generation cycle. Identification of Purkinje or other early‐forming neurons was made by labeling them with 3H‐thymidine on their birthdays. Many of the fractions contain viable cells capable of aggregating in culture.
Flow microfluorometric studies of lectin binding to mammalian cells. I. general features
Authors: Kraemer Paul M. , Tobey Robert A. , Van Dilla Marvin A.
Published: February 2005

Flow microfluorometry has been used to quantitate cell‐surface binding of fluorescein‐conjugated lectins. Frequency distributions of total surface binding of Concanavalin A per cell were prepared for a variety of cultured cell populations, including established cell lines, virus‐transformed lines and non‐transformed parental lines. In the case of growing Chinese hamster cells (line CHO), much of the variability of Con A binding per cell could be related to variability of cell size. Experiments with cells synchronized by mitotic selection indicated that the modal surface density of binding sites was almost constant throughout the cell cycle. However, as indicated by inhibition of binding with α‐methyl mannopyranoside and by the effect of trypsin, the sites on each cell were heterogeneous in chemical structure and/or exposure. Agglutinability of virus‐transformed cell lines or trypsin‐treated parental lines was demonstrated but could not be correlated closely with binding.
Colony formation by chicken hematopoietic cells and virus‐induced myeloblasts
Authors: Dodge W. H. , Moscovici C.
Published: February 2005

Leukemic myeloblasts and cells derived from normal chick hematopoietic tissue produced colonies in soft agar. Colonies produced by leukemic myeloblasts differed from normal chick tissue in their morphological characteristics, in the greater initial number of cells required for colony formation and in their decreased dependence on conditioned medium for development. The colony forming cells for both types were enriched when allowed to grow for several days in liquid growth medium.

In soft agar, myeloblasts differentiated into more mature granulocytic cells and macrophages. These differentiated cells accumulated between one and two weeks after seeding. When tested for release of avian myeloblastosis virus (AMV), 6 out of 18 colonies were releasing AMV at one week whereas 3 out of 39 were releasing AMV at two weeks. Five two week old colonies which were negative for AMV were producing myeloblastosis associated viruses (MAVs). Normal colony forming cells were present in leukemic buffy coat and although colonies made by these cells contained MAVs, no AMV could be detected.

The data obtained with normal avian tissues were similar to those obtained by others with mammalian hematopoietic tissue. Colony formation by normal hematopoietic tissues was strictly dependent on factors present in conditioned medium. Tissues producing colonies included bone marrow, yolk sac, spleen and peripheral leukocytes. Colonies were not obtained from thymus and bursa. Furthermore, the colony origin did not appear to be erythroid in nature.
Colony formation in agar by murine plasmacytoma cells: Potentiation by hemopoietic cells and serum
Authors: Metcalf D.
Published: February 2005

Clonal growth in semisolid agar medium was obtained using cells from 19 of 25 transplanted murine plasmacytomas when the medium was supplemented by whole mouse blood or washed red cells. With different tumors cloning efficiency ranged from 0.01% to 21.6%. With two exceptions, mouse blood did not potentiate colony formation in agar by cells from transplantable myelomonocytic, myeloid, and lymphoid leukemias, reticulum cell sarcomas and fibrosarcomas. The clonal growth of some plasmacytomas was also potentiated by syngeneic thymic, spleen or bone marrow cells. Plasmacytoma colony growth was not stimulated by normal mouse serum but serum from mice injected with endotoxin or polymerised flagellin stimulated colony growth by some plasmacytomas. The active serum factor was not the colony stimulating factor (CSF) and its appearance after antigenic stimulation was not T cell‐dependent. Preimmunised mice failed tq respond to antigenic stimulation. Whole body irradiation did not induce a rise in the capacity of serum to stimulate colony formation by plasmacytoma cells.
Isolation and characterization of revertant cell lines. II. Growth control of a polyploid revertant line derived from SV40‐transformed 3T3 mouse cells
Authors: Pollack Robert , Vogel Arthur
Published: February 2005

3T3 mouse fibroblasts in 10% calf serum cease to increase in number at confluence. SV40‐transformed 3T3 cells lose their sensitivity to this growth control and so are able to reach multilayered cell densities. We previously described a polyploid revertant line F1SV101, isolated from a SV40‐transformed cell, that has a low maximum cell density (Pollack et al., '68), despite the continued presence in this revertant of SV40‐specific DNA, RNA and Tantigen. We have examined the mechanism by which the revertant maintains a low saturation density, and find that at confluence the revertant and 3T3 are both reduced in mitotic index, fraction of cells synthesizing DNA, and rate of DNA synthesis. The transformed cell does not respond to confluence. None of the lines sheds intact cells into the medium at confluence.
The effects of inhibitors of RNA and DNA synthesis on protein synthesis and polysome levels in mouse L‐cells
Authors: Craig Nessly
Published: February 2005

Metabolic inhibitors of RNA synthesis, such as actinomycin D and MPB (2‐mercapto‐1‐(β‐4‐pyridethyl) benzimidazole) have often been used to test the possibility that transcription is required for some cellular process or to measure the „half‐life”︁ of messenger RNA (mRNA). The basic assumption of this work has been that the primary effect of these inhibitors is on transcription alone, and that any effect on translation is secondary to the inhibition of mRNA synthesis. This assumption has been tested by examining the effects of these inhibitors at different doses on various parameters of the transcriptional and translation processes in mouse L‐cells in culture, i.e., the rates of RNA, DNA, and protein synthesis; the level and size of cytoplasmic polysomes; the rates of polypeptide elongation and termination, and the role of peptide chain initiation in protein synthesis. The results indicate that the basic assumption is not correct since these inhibitors affect protein synthesis by inhibiting the rate of initiation and not the level of mRNA. Another implication of the experiments is that the average mRNA in L‐cells has a half‐life of at least one cell generation (16–18 hours), and that earlier values from experiments using actinomycin are underestimates. Cordycepin (3′‐deoxyadenosine) which inhibits the production of ribosomal and messenger RNA also affects protein synthesis at the level of initiation. In contrast, two inhibitors of DNA synthesis (hydroxyurea and cytosine arabinoside) inhibit protein synthesis and the level of polysomes by some other mechanism.
Calcium‐dependent stimulation by a phorbol ester (PMA) of thymic lymphoblast DNA synthesis and proliferation
Authors: Whitfield J. F. , Macmanus J. P. , Gillan D. J.
Published: February 2005

PMA (phorbol myristate acetate, i.e., 12‐0‐tetradecanoyl‐phorbol‐13‐acetate) a tumor‐promoting ester from croton oil, at its most effective concentration of 0.05 μg per milliliter, rapidly (within one hour) induces a large fraction of the lymphoblasts in suspended thymic lymphocyte populations to start making DNA, and these stimulated cells later progress into mitosis. This stimulatory PMA action is probably mediated by calcium because it disappears when calcium is omitted from the medium, and PMA strikingly increases the sensitivity of the lymphoblasts to calcium's stimulatory action.
Pyridine nucleotide metabolism in mammalian cells in culture
Authors: Hillyard David , Rechsteiner M. C. , Olivera Baldomero M.
Published: February 2005

The biosynthesis of pyridine nucleotides has been examined in a number of mammalian cell lines in culture. In all lines examined, nicotinamide is incorporated by a biochemical pathway distinct from the Preiss‐Handler pathway for nicotinic acid.

In at least the human cell line D98/AH2, there is no detectable endogenous synthesis of the pyridine ring from tryptophan. Although most cell lines examined (hamster BHK 21/13, mouse L929 and human D98/AH2) use either nicotinic acid or nicotinamide as a precursor for DPN and TPN, two mouse cell lines, 3T3‐4E and LM CIID, are unable to utilize nicotinic acid as a source of the pyridine ring.

If nicotinic acid is present in the medium, substantial amounts of intracellular desamido DPN accumulate suggesting that the last step (desamido DPN→DPN) is limiting in the Preiss‐Handler pathway. With nicotinamide, the only compound which accumulates in substantial amounts apart from DPN and TPN is nicotinamide ribose; there is no detectable NMN. The results of pulse‐labeling experiments suggest that nicotinamide ribose may be an intermediate in the nicotinamide pathway.

Following growth of D98/AH2 cells in high concentrations of niacin, biosynthesis of DPN from nicotinamide was completely inhibited for at least six hours. The converse experiment revealed no inhibition of niacin incorporation. This observation suggests that a niacin pathway intermediate, which present evidence indicates is desamido‐DPN. can inhibit nicotinamide utilization.

Newly synthesized DPN turns over with a half‐life of two hours in azaserine‐treated D98/AH2 cells. In the absence of azaserine, the nicotinamide moiety of newly synthesized DPN is lost from D98/AH2 cells to the medium with a half‐life of eight hours. About 80% of the nicotinamide is lost to medium as nicotinamide ribose.
Isolation and characterization of revertant cell lines. III. Isolation of density‐revertants of SV40‐transformed 3T3 cells using colchicine
Authors: Vogel A. , Risser R. , Pollack R.
Published: February 2005

Treatment of the SV40 transformed 3T3 cell line SV101 with colchicine permits the isolation of polyploid revertant sublines Which have lower saturation densities than SV101. These low saturation density lines have also reverted to a high serum requirement for growth, and are unable to form colonies in methocel. Normal SV40 has been recovered from these revertants.

3T3 cells are more resistant to colchicine than SV3T3 cells at all cell densities. Colchicine revertants do not display a 3T3‐like resistance to colchicine at low density, but do survive colchicine at confluent cell densities, presumably due to their increased contact inhibition.
On the kinetics of erythroid cell differentiation in fetal mice. III. DNA and hemoglobin measurements of individual erythroblasts during gestation
Authors: Steiner Rudolf
Published: February 2005

Microspectrophotometric absorption measurements were used to determine the hemoglobin content of erythroid cells derived from the yolk sac during gestation of fetal C3H mice, from day 9 to day 15. Using the DNA content as a marker for the mitotic state between 2C and 4C phase, five successive cell generations and their mean hemoglobin contents were distinguished: 12 pg (pg, picogram = 10−12 gm). 22.2 pg, 37 pg, 50 pg and 56 pg. In the final state, nucleated erythrocytes contained 98 ± 22 pg hemoglobin.

Erythroid cells derived from the liver were measured on day 15 of fetal gestation. The hemoglobin content of proerythroblasts was below 0.3 pg. The two cell generations in the basophilic state had 0.6 pg and 1.7 pg respectively. Polychromatic erythroblasts yielded a hemoglobin content of 5.1 pg in the first cell generation and 7.5 pg in the second one. Orthochromatic erythroblasts contained 8 pg, reticulocytes 12 pg and mature erythrocytes 28 ± 7 pg hemoglobin.

Calculations based on these data suggest that the rate of total hemoglobin synthesis is similar in both yolk sac and liver erythropoiesis. The difference between the final hemoglobin content in nucleated erythrocytes of yolk sac origin and that in hepatic erythrocytes can be explained by the different cell generation times.
Uridine uptake and its intracellular phosphorylation during the cell cycle
Authors: Stambrook Peter J. , Ebert James D. , Sisken Jesse E.
Published: February 2005

The rate of 5‐3H uridine uptake into Chinese hamster V79 cells and the rate of its incorporation into RNA increase tenfold during the cell cycle. Both reactions exhibit the same apparent Km(1.7 × 10−5 M). Chromatography of acid‐soluble material from cells incubated with 5‐3H uridine (0.25 μM) at different times of the cell cycle revealed that intracellular uridine was rapidly phosphorylated at all times, even though cells in late S and G2 take up roughly ten times as much uridine as cells in G1. Uridine kinase activity in synchronized cells increases about two and one‐half‐fold during the same time period, and in exponentially growing cells is not saturated until the external uridine concentration is raised above 200 μM. It is concluded that the change in uridine kinase activity during the cell cycle is not responsible for the tenfold increase in the rate of uridine transport, and that these two processes are independently regulated.
Tetrodotoxin‐sensitive and ‐insensitive action potentials, in myotubes
Authors: Land B. R. , Sastre A. , Podleski T. R.
Published: February 2005

The study of long‐term cultures of myogenic cells has proven that electrical excitability develops only after the development of electrical coupling between the cells. That is, neither surface contact in itself nor coupling in itself is sufficient to cause excitability to develop in these cells. Following the formation of multinucleated myotubes, several different types of electrical responses develop. Some of the action potentials are sodium‐dependent and are blocked by tetrodotoxin (TTX). Others are dependent upon sodium and possibly calcium and they are not blocked by TTX. Furthermore, these two types of responses may exist in a myotube at the same time. Under some circumstances the kinetics of the two systems are sufficiently different to result in action potentials that have two peaks. Under these conditions the first peak is always of shorter duration and it is always blocked by TTX.
Induction of lysosomal enzymes in contact inhibited 3T3 cells
Authors: Horvat A. , Ács G.
Published: February 2005

The pattern of changing activities of three lysosomal enzymes, N‐acetyl‐β‐D‐glucosaminidase, aryl sulfatase, and DNase II, and that of DNA polymerase was followed in homogenates of 3T3 cells during the logarithmic phase of growth and in stationary cultures. The change in activities of the polymerase and the lysosomal enzymes is antiparallel. DNA polymerase exhibits highest activity in growing cultures, and shows a three‐fold decline of the specific activity in stationary cultures. The lysosomal enzymes show a very marked increase in their specific activity after density saturation is reached, which can be prevented by the addition of cycloheximide. Colcemid added to logarithmically growing cultures also causes an increase in the specific activities of lysosomal enzymes.
The subcellular distribution of lysosomal hydrolases in the Harding‐Passey melanoma
Authors: Mufson R. Allan
Published: February 2005

Differential centrifugation of homogenates of Harding‐Passey melanoma demonstrated that aryl sulfatase A and β‐glucuronidase sediment with particles (i.e., lysosomes) distinct from those particles bearing tyrosinase (i.e., melanosomes). The sedimentation curves for the lysosomal enzymes and tyrosinase, however, demonstrated that an adequate separation of these particle types could not be obtained by differential centrifugation. Isopycnic density gradient centrifugation was used to obtain the necessary resolution. The results of the density gradient studies demonstrated that lysosomes and melanosomes could be separated by this technique, as judged by enzyme distribution among the fractions recovered from the gradients and from electron microscopic examination of the melanosome fractions. It was further evident that the purified and washed melanosomes contained significant amounts of both acid hydrolase activities. Indeed 24% to 27% of the total acid hydrolase activities recovered from the density gradients were associated with the melanosome fractions. The acid hydrolases associated with the melanosomes could not be solubilized by treatment with 0.1% (v/v) Triton X‐100, nor by exposure to hypo‐osmotic shock. The melanoma lysosomes, however, did release most of both their hydrolase activities into soluble form after treatment with the same percentage of detergent. The lysosomes were, however, very resistant to rupture by exposure to hypo‐osmotic conditions.
Glycolytic enzyme activity in developing red and white muscle
Authors: Dalrymple R. H. , Cassens R. G. , Kastenschmidt L. L.
Published: February 2005

The activities of the constant proportion enzymes of the Embden‐Meyerhof chain (triose phosphate isomerase (TIM), glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), phosphoglycerate mutase (PGM) and enolase (ENOL)), and the activity of lactic dehydrogenase (LDH) were studied in developing red (trapezius) and white (longissimus) muscles of the pig from a fetal stage to 24 weeks postnatal. Both muscles were differentiated by two weeks postnatal in the sense that they had reached the adult level of enzyme activity. Enzyme activities were two‐ to three‐fold greater in the longissimus than in the trapezius. Enzyme activity ratios based on GAPDH were not consistent in the fetal and day 1 samples but were consistent during later stages of growth. Ratios of enzyme activity based on activity at 105 days gestation revealed that TIM, PGK and PGM are grouped and follow the same pattern, but GAPDH and ENOL are quite different from each other and from the pattern shown by TIM, PGK and PGM. The constant proportion concept in developing muscle is therefore questioned.
Effect of tolbutamide on the metabolism of Tetrahymena
Authors: Blum J. J. , Connett R. J. , Porter Pamela
Published: February 2005

Tolbutamide partially inhibited the growth but increased the glycogen content of Tetrahymena pyriformis in logarithmically growing cultures. Tolbutamide slightly increased 14CO2 production from [1‐14C] and [6‐14HC] glucose and [2‐14C] pyruvate, but had little effect on the oxidation of [1‐14C] acetate when any of these substrates were added to the proteose‐peptone medium in which the cells had been grown. Measurement of 14CO2 production from [1‐14C] and [2‐I4C]‐glyoxylate showed that this substrate was primarily oxidized via the glyoxylate cycle, with little if any oxidation occurring via the peroxisomal glyoxylate oxidase. Addition of tolbutamide inhibited the glyoxylate cycle as indicated by a marked reduction in label appearing in CO2 and in glycogen from labeled acetate. In control cells, addition of acetate strongly inhibited the oxidation of [2‐14C]‐pyruvate whereas addition of pyruvate had little effect on the oxidation of [1‐14C]‐acetate. Acetate was more effective than pyruvate in preventing the growth inhibitory and glycogen‐increasing effects of tolbutamide. The data suggest that one effect of tolbutamide may be to interfere with the transfer of isocitrate and acetyl CoA across mitochondrial membranes.
Properties of protein polymers as substratum for cell growth in vitro
Authors: Macieira‐Coelho A. , Berumen L. , Avrameas S.
Published: February 2005

The behavior of two established cell lines was found to vary when subcultivated on protein polymers covered with either negatively or positively charged substances. Results indicate that the influence on cell behavior is conditioned by the charge rather than by structural differences or degree of attachment of the substances to the polymer. Furthermore, the substratum seems to have just a physical effect at the cell membrane without any direct influence on cell metabolism.

Cells were also seeded on a calf serum polymer and it was found that serum loses its properties on cultivated cells when used as substratum.
Potassium content of single human red cells measured with an electron probe
Authors: Gary Kirk R. , Crenshaw Miles A. , Tosteson D. C.
Published: February 2005

The potassium content of single human red cells was measured with an electron probe. Cells were placed on beryllium discs and coated with a thin layer of dibutyl pthalate to prevent loss of cellular contents. Samples were stable under the electron beam during analysis for more than 15 minutes and could be stored for long periods of time. Primary standards were prepared by loading red cells with varying known amounts of potassium in order to circumvent the corrections for absorption. The X‐ray intensity was found to be directly proportional to the potassium content of the cells.
Effects of lymphocytes from the thymus and lymph nodes on differentiation of hemopoietic spleen colonies in irradiated mice
Authors: Basford Nancy L. , Goodman Joan W.
Published: February 2005

Hemopoietic colonies were counted macroscopically and microscopically in spleens of hybrid mice seven or eight days after they had been irradiated and given parental bone marrow in donor‐host combinations exhibiting poor growth. Colonies counted microscopically were classified as to differentiation pathway. Lymphocytes from the thymus or lymph nodes were injected into some recipients at several different dosages and lymphocyte: bone marrow (L:B) ratios. In confirmation of earlier work it was found that thymocytes increased the number and size of colonies in recipients of marrow. A shift of differentiation toward granulopoiesis was also seen when thymocytes were given, although erythropoietic colonies were still the most frequently seen type except at very high L:B ratios. Lymph node lymphocytes shifted the pattern more markedly toward granulopoiesis, even at low L:B ratios. When lymphocytes from either source were given without marrow, only a few colonies could be found in recipients, and if differentiated they were almost exclusively granulopoietic. Irradiation (900 R) of lymphocyte donors reversed the shift so that a normal pattern of differentiation, like that resulting from marrow alone, was seen; irradiated lymphocytes were nonetheless capable of augmenting the size and total number of hemopoietic colonies.
A mammalian somatic “cell cycle” mutant defective in G1
Authors: Michael Liskay R.
Published: February 2005

Variants or “mutants” temperature‐sensitive (ts) for growth have been isolated by selection from a near‐diploid mouse cell line. Thus far. 10 ts mutants which grow normally at 33° C, but not at 39° C, have been isolated. These ts mutants were then studied to determine if any manifested their defect at a unique point or stage in the cell cycle. This type of ts mutant is termed a “cell cycle” mutant.

The first screen involves observing individual cells of an asynchronous culture for residual division after a shift from 33° C (permissive temperature) to 39° (nonpermissive temperature). A cell cycle mutant should show some fraction of the cells dividing only once at a normal rate after the shift. The ts variant B54 met this first criterion for a cell cycle mutant (i.e., 50% residual division) and was further analyzed.

The second screening technique monitors (1) the rate of entry into S, (2) the length of G2, and (3) the rate and duration of cells entering mitosis after a shift of an asynchronous culture to 39°. This experiment with B54 revealed that cells in G1 at the time of the shift to 39° failed to enter S while cells already into S completed the cycle at 39°. These results suggest that B54 is defective in a G1 function which is required for entry into S, but which is no longer needed once cells have entered S. Other results are presented which also support this hypothesis.

In addition the ts function of B54 is apparently required for recovery from a “high density” G1 arrest.
The negatively charged protein extracted from Tetrahymena pyriformis as an attractant in Amoeba proteus chemotaxis
Authors: Nohmi Mitsuo , Tawada Katsuhisa
Published: February 2005

We extracted a pure protein from Tetrahymena pyriformis as the chemotactic substance in Amoeba proteus. The protein was heat‐stable, negatively charged at neutral pH (since its isoelectric point was near pH 4.5) and composed of three subunits. Its molecular weight was 3 × 8,000.
Flow microfluorometric studies of lectin binding to mammalian cells II. Estimation of the surface density of receptor sites by multiparameter analysis
Authors: Steinkamp John A. , Kraemer Paul M.
Published: February 2005

A flow‐system multiparameter cell analyzer that simultaneously measures and processes fluorescence and cell volume signals from single cells was used to study the binding of fluorescein‐conjugated Concanavalin A (Con A–F) to the cell surface. Cells reacted with Con A–F were passed through a flow chamber where sensors measured both cell volume and fluorescence of each individual cell. Sensor signals were electronically processed by first converting the cell volume signals to two‐thirds power (proportional to surface area) and then forming the fluorescence‐to‐surface area ratio. These ratios, which were considered as estimates of the surface density of binding sites, were displayed as frequency distribution histograms using a multichannel pulse‐height analyzer for various cell populations differing in cell size. Comparisons between cell lines showed characteristic differences in binding site density. Cell cycle dependent changes were not found for CHO cells synchronized by mitotic selection. An important benefit of this analysis method was the ability to quantitate very weak cell surface fluorescence.
Myonemal contraction of Spirostomum I. Kinetics of contraction and relaxation
Authors: Hawkes R. B. , Holberton D. V.
Published: February 2005

A microphotometric method is introduced that allows measurement of the contraction‐relaxation kinetics of Spirostomum in response to electrical stimulation. The time course of contraction includes a rapidly contracting phase of some 4–5 mS during which cells shorten at a rate in excess of 100 cell lengths sec−1. While a stimulus strength‐duration curve determines the threshold of the response, the response to above threshold stimuli of different strengths and to trains of stimuli suggest that contraction of Spirostomum may not be an all‐or‐none event. The kinetics of relaxation following high stimulating voltages and repetitive after contractions also induced by high voltages are explained by excitation‐contraction coupling through a stimulus‐dependent intermediate effector, possibly the release of calcium ions. Changes in resting membrane potential detected by intracellular recording do not influence the initiation of contraction, while microinjection of calcium buffers above 10−5 M Ca2+ invariably induces contraction.
Role of pH in fibroblast proliferation
Authors: Froehlich Jeffrey E. , Anastassiades Tassos P.
Published: February 2005

Secondary cultures of human diploid fibroblasts were used to study the effect of pH on cellular proliferation. In nonconfluent cultures, the growth rate at pH 7.1 was similar to that at pH 7.7 regardless of serum concentration. However, the saturation density achieved at pH 7.7 at any serum concentration was always 2–4 times that achieved at pH 7.1, although the greatest differences in saturation density were observed at the higher serum levels. The results suggest that the effect of pH on saturation density is due to two factors. One, cells at pH 7.1 seem to have a greater ability to undergo contact‐inhibition than at pH 7.7, independent of any serum functions; and, two, confluent cells in medium at pH 7.1 are somewhat less sensitive to growth stimulation by increasing serum concentration than are confluent cells raised in medium at pH 7.7.
A temperature‐sensitive cell cycle mutant of the BHK cell line
Authors: Burstin Stuart J. , Meiss Harriet K. , Basilico Claudio
Published: February 2005

A temperature‐sensitive growth mutant derived from the BHK 21 cell Line, ts AF8, was found to have greatly reduced DNA synthesis at the nonpermissive temperature. This reduction is mainly due to a decrease in the frequency of cells synthesizing DNA. Upon shift up, ts AF8 becomes blocked in the G1 phase of the cell cycle. The cells acquire elevated cAMP levels and a unimodal distribution of DNA content, equivalent to that of G1 cells at the permissive temperature, Ts AF8 cells blocked at the G1/S boundary with hydroxyurea will enter S when shifted to the nonpermissive temperature. On the other hand, ts AF8 cells arrested m G1 by serum deprivation and shifted to the nonpermissive temperature at the moment of serum addition do not enter S, while those synchronized by isoleucine deprivation and shifted at the time of isoleucine addition will enter S. These data suggest that the cycle arrest point of the ts AF8 mutation is located in G1 between the blocks induced by serum starvation and isoleucine deprivation.

The reduction in DNA synthesis caused by the ts AF8 mutation is not reversed by infection or transformation with Polyoma virus. Mitochondrial DNA continues to be synthesized at wild‐type levels at the nonpermissive temperature.
The distribution of pyridine nucleotides between nucleus and cytoplasm
Authors: Rechsteiner Martin
Published: February 2005

Following enucleation of a portion of human culture cells containing 3H‐pyridine nucleotides, autoradiography revealed no difference in the grain density over enucleated and whole cells. These results provide evidence that the concentration of pydine nucleotides does not differ by more than threefold between nucleus and cytoplasm and probably does not vary at all.
Synthesis of glycolipids and phospholipids in hamster cells: Dependence on cell density and the cell cycle
Authors: Hirschberg Carlos B. , Wolf Barbara Anne , Robbins P. W.
Published: February 2005

Sakiyama et al. ('72) reported the isolation of a line of hamster cells (NIL 1c1) which contains only three glycolipids, hematoside, ceramide monohexoside and ceramide dihexoside. The incorporation of radiolabeled palmitate into hematoside during 24 hours was three fold higher in normal confluent, non growing cells than sparse, growing ones. Polyoma transformed cells did not exhibit this effect.

We have continued studies with the untransformed cell line and have found that the higher incorporation of radiolabeled palmitate into hematoside by normal confluent cells is not due to a higher rate of turnover of hematoside at confluence but represents a true chemical increase. We have also found that this increase is not a gradual process during cell growth but instead occurs only when the cells become confluent and stop growing. The increase of hematoside at confluence is not due to a higher rate of synthesis of hematoside during G1, relative to the other phases of the cell cycle. We found the rate of synthesis of hematoside to be constant throughout the cell cycle.

The rate of synthesis of phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine, phosphatidyl inositol and sphingomyelin was also studied as a function of the cell cycle. We found no large differences in the synthetic rate of any given phospholipid species throughout the cell cycle although the rate of synthesis of the glycerophospholipids was somewhat higher during late G1 and S. We did, however, find major differences in the rates of synthesis of the different phospholipid species.
Synthesis of glycolytic and peroxisomal enzymes in Tetrahymena following a change in culture conditions
Authors: Levy Michael R.
Published: February 2005

The specific activity of a peroxisomal enzyme, lactate oxidase, and of pyruvate kinase and lactate dehydrogenase, which are not peroxisomal, increased rapidly when shaken cultures of Tetrahymena were transferred to conditions of oxygen restriction and supplemented with glucose. Two other peroxisomal enzymes, catalase and TPN‐linked isocitrate dehydrogenase, did not increase substantially, nor did succinate dehydrogenase. The increases were reduced if glucose was not added at the time of transfer, and were prevented by actinomycin D or cycloheximide, but not by chloramphenicol. The results suggest an involvement of peroxisomes in the metabolism of glycolytic endproducts when the availability of oxygen to the cell is limiting.
Slow phase hemolysis in hypotonic electrolyte solutions
Authors: Chan T. K. , Lacelle P. L. , Weed R. I.
Published: February 2005

When a population of erythrocytes is partially hemolyzed the time course of hemolysis can be divided into a fast phase and a slow phase. The slow phase occurs with both rapid and gradual addition of the hypotonic medium (rapid and gradual hemolysis). There is no difference in the osmotic fragility of erythrocytes remaining at 60 minutes after rapid or gradual hemolysis.

Erythrocytes near their critical hemolytic volume have an equimolar ouabain‐insensitive sodium‐potassium exchange. Critical non‐hemolytic swelling with resulting stress on the membrane appears requisite to slow phase hemolysis since more non‐penetrant sucrose is required to prevent slow phase lysis rather than that which would be predicted from the intracellular colloid osmotic pressure due to hemoglobin. Sucrose protection from slow phase hemolysis thus depends not only on counter‐balancing the colloid osmotic pressure, but also removal of sufficient intracellular water to prevent critical membrane strain. This model is consistent with that proposed by Katchalsky.

Irreversible membrane changes associated with hypotonic stress manifested by persistent stomatocytic shape change and membrane wrinkling on return of cells to isotonicity appear to be due to critical changes in membrane components. Such cells, having normal indices and specific gravity are less deformable than control cells in 2.8 μm pore size polycarbonate filters.
Effect of chloramphenicol on replication of mitochondria in Tetrahymena
Authors: Gleason Florence K. , Ooka Mary P. , Cunningham William P. , Hooper Alan B.
Published: February 2005

Tetrahymena pyriformis ST (3 × 104 cells/ml) was treated with 0.1 mg/ml chloramphenicol (CAP). Cell division ceased after 1.5 divisions with no decreased viability. Total mitochondrial volume and succinic dehydrogenase (SDH) activity/liter increased 1.7‐fold and 3‐fold, respectively. SDH activity/cell decreased whereas malate dehydrogenase activity/cell and respiratory control ratios and P: O ratios of isolated mitochondria were unchanged in treated cells. During 12 hours of growth in CAP the total surface area of mitochondrial inner and outer membrane was essentially unchanged or increased 4‐fold, respectively. Mitochondria from cells treated with chloramphenicol had decreased size, buoyant density and protein: lipid ratio in the membranes. The membrane ubiquinone: protein ratio was unchanged.

Tetrahymena cells contained 3.6 × 10−12 g of mitochondrial DNA and 6,800 mitochondria in a volume of 41,000 μ3. A 4‐hour treatment with CAP caused a 4‐fold increase in the number of mitochondria/cell and a 10‐fold increase in mitochondria/liter in contrast to a 4‐fold increase in number of mitochondria/liter in control cells. Thus CAP stimulated division of mitochondria. Individual mitochondria of treated cells had one‐tenth the volume of control mitochondria. The rate of increase of mitochondrial DNA/liter was the same in control and CAP‐treated cultures. The amount of DNA/mitochondrion decreased 75% in CAP‐treated cells due to the rapid division of mitochondria. The cell volume, cell protein content and mitochondrial DNA content/cell decreased with growth of control cultures.
Influence of mono‐ and multivalent cations on the electrokinetic properties of Rana pipiens kidney cells
Authors: Kiremidjian L. , Stotzky G.
Published: February 2005

Changes in the cation composition of the ambient cell environment, at constant ionic strengths, altered the electrokinetic properties of the surface of normal adult Rana pipiens kidney cells in a manner which reflected more the physiochemical properties and binding affinities of the cation than its valence. The effect was expressed in the changes of the magnitude of the net surface charge, at any pH, and in the shift of the isoelectric point of the cell surface.
Variant HeLa cells selected for their resistance to ouabain
Authors: Rosenberg Howard M.
Published: February 2005

The cardiac glycoside, ouabain, normally kills HeLa cells at concentrations of about 10−7 m or greater. By treating a population of HeLa cells with increasingly higher concentrations of the drug, a variant population was obtained of HeLa cells capable of growing in medium containing 10−4 M ouabain.

Inhibition of volume regulation of cells subjected to hypotonic shock was used as a measure of inhibition of active transport of Na across the plasma membrane. In that way dose‐response curves for the rapid effects of ouabain and other inhibitors of active Na transport were obtained with both the original, ouabain‐sensitive (OS) and the variant, ouabain‐resistant (OR) cells. Three other cardiac glycosides (digoxin, digitoxin and hellebrin) and two aglycones (digitoxigenin and strophanthidjn) were found to be equally as effective as ouabain in inhibiting volume regulation of the OS cells; the concentration which produced half‐maximum inhibition, I(max/2), was about 6 × 10−7 M in each case.

Similar inhibition of the OR population by ouabain was observed only when the concentration exceeded 10−4 m [I(max/2)∼2.5 × 10−4 m], and the other steroid compounds had no effect on the variant cells at the highest concentrations tested (∼2 × 10−5 m). OR and OS cells differed also in their sensitivities to the cardioactive erythrophleum alkaloid, coumingine; I(max/2) for OS and OR cells was 5 × 10−8 m and 6 × 10−7 M, respectively.

These results, in addition to results of ouabain binding experiments and measurements of the rates of reversal of inhibition of volume regulation, suggest that a major reason for the differential sensitivities of the two phenotypes to these drugs is different affinities of their sodium pumps for inhibitors of active transport.
Characterization of the ATP‐phosphohydrolase activity of bovine spermatozoa flagellar extracts
Authors: Young Leona G. , Smithwick Eleanor B.
Published: February 2005

The ATP‐phosphohydrolase activity of extracts prepared from bovine spermatozoa flagella (BSFE), was characterized with respect to enzyme, substrate, activator ion and salt concentration, temperature dependence and time stability. BSFE required the presence of a divalent cation for activity: Mg++ or Ca++ could function as activator; Mn++, Zn++ and Cd++ could not. EDTA, but not EGTA, was inhibitory to enzymatic activity. Ca++ inhibited the Mg++ stimulated activity. ATP was dephosphorylated more rapidly than GTP>CTP>ITP, and ADP was dephosphorylated at 40% of the rate of ATP. The magnesium activated ATPase was stimulated by potassium and inhibited by sodium ions. Activation of BSFE ATP‐phosphohydrolase was maximal in the presence of Mg++ and ATP in equimolar concentrations and K+ (0.05–0.3 m) at 30°C. Although the enzymatic activity of the extract was found to decrease rapidly with time, it could be maintained for up to three days by the addition of 2‐β‐mercaptoethanol to the bovine spermatozoa flagellar extracts.
Exposure of an arginine‐rich protein at surface of cells in S, G2, and M phases of the cell cycle
Authors: Stein Sylvia M. , Berestecky John M.
Published: February 2005

Phenylglyoxal (PG) is shown to be a cell surface probe specific for arginine moieties in protein: (1) It does not enter the cell as evidenced by lack of PG in the cytoplasm. (2) It does not cause excessive cell leakage as measured by release of 51Cr. (3) It reacts with positively‐charged groups in proteins at the cell surface but not with those of phospholipids at the surface; since pronase removes PG from the surface, but phospholipase C does not. (4) Under the conditions used in these experiments, it reacts virtually exclusively with arginine moieties in protein (Freedman et al., '68; Takahashi, '68; Werber and Sokolovsky, '72).

Synchronized cells were exposed to radioactive PG to assess quantity of arginine moieties in protein at the surface. There is a sharp decrease in arginine at the cell surface at entry into G1 phase from M and a 24‐fold increase upon entry into S phase. There is a slight drop in exposed arginine in late S phase followed by an increase to 26 times the G1 level immediately prior to mitosis. Lactoperoxidase‐catalyzed iodination of tyrosine moieties in protein at the surface of synchronized cells shows a very gradual increase in protein as the cells move through the cycle and increase in size. Since the increase in arginine moieties in protein at the surface does hot reflect a similar increase in total protein at the surface, an arginine‐rich protein appears to be exposed at the cell surface during the division‐related phases of the cell cycle.
Requirement of bromodeoxyuridine for the maintenance of “transformed” characteristics in bromodeoxyuridine dependent cells
Authors: Horn Diane , Davidson Richard L.
Published: February 2005

A bromodeoxyuridine (BUdr) dependent cell line, called B4, which requires BUdr not only for optimal growth but also for the maintenance of the non‐contact inhibited state was described previously. We have now shown that contact inhibition in the B4 cells in the absence of BUdr is associated with a marked decrease in the percent of cells synthesizing DNA. The transition to the contact inhibited state in the absence of BUdr does not seem to be due to changes in cyclic AMP levels. It has also been shown that several but not all of the characteristics which distinguish transformed from untransformed cells also distinguish B4 cells grown in the presence of BUdr from B4 cells grown in the absence of BUdr. In addition to being contact inhibited, B4 cells grown in the absence of BUdr have a higher serum requirement, grow less well in soft agar, and are less agglutinable by wheat germ agglutinin than B4 cells grown in the presence of BUdr. Agglutinability by concanavalin A, however, is the same for B4 cells grown in the presence and absence of BUdr. Dependent cells maintained in the presence of BUdr do not form tumors and it is not yet clear how the transformed characteristics of the dependent cells are related to malignancy.
The pattern of density dependent growth inhibition in murine fibroblasts
Authors: Canagaratna Mary C. P. , Riley P. A.
Published: February 2005

Observations on the pattern of nuclear incorporation of 3H‐TdR in long term (8‐day) and short term (3‐day) 3T3 cultures with local cell densities between 0.2 × 104 and 6.2 × 104 cells/cm2 are reported. Contrary to a number of previous studies our observations indicate that density dependent inhibition is exhibited in relatively sparse cultures, commencing at 0.5 × 104 cells/cm2. Various possible mechanisms which could have caused the observed pattern of density‐dependent regression in labelling index are discussed.
Glutamine synthetase activity in WI‐38 cells
Authors: Viceps Dace , Cristofalo V. J.
Published: February 2005

In the presence of complete growth media (Eagle's MEM), human diploid WI‐38 cells have a low level of glutamine synthetase activity. The activity could be increased by depriving the cells of exogenous glutamine; addition of hydrocortisone to either glutamine‐deficient or complete medium had no effect on the activity of the enzyme. Cell growth ceased under conditions that enhanced glutamine synthetase activity, and hydrocortisone could not reverse this inhibition.