bluuebunny/crossref_metadata_2025_split huggingface dataset:
Residual radiation injury exhibited in long‐term bone marrow cultures
Authors: Reincke Ursula , Hannon Eileen C. , Hellman Samuel 
Published: February 2005

AbstractResidual radiation injury was demonstrated in long‐term primary cultures of mouse bone marrow. Control cultures underwent three phases of hematopoietic activity as distinguished by initial establishment, steady high (plateau) production of granulocytes, and gradual decline. Irradiation with 50, 300, or 550 rads, given at the end of the initial phase, did not prevent any culture flasks from entering the plateau phase. However, actual production levels and the time they were maintained varied inversely with the radiation dose so that the accumulated postradiation cell production corresponded to an exponential dose‐response relationship at any time after treatment. The accumulated cell productions were found to be similar in all groups when expressed by the number of stem cell doublings necessary to produce them. The findings cannot be explained by reproductive cell death and are consistent with the notion of a limited division capacity in hematopoietic stem cells.
Variation of the carbohydrates of glycoproteins of cells growing on different surfaces
Authors: Warren L. , Blithe D. L. , Cossu G. 
Published: February 2005

AbstractThe carbohydrate components of some glycoproteins of hamster cells differ as a function of their growth on various substrates; glass, plastic, or plastic coated with collagen. This observation is interpreted as an effect of the environment on cellular structure at the molecular level. The basis of the change and its possible significance are discussed.
The role of calcium in the initiation of superoxide release from alveolar macrophages
Authors: Holian Andrij , Daniele Ronald P. 
Published: February 2005

AbstractThe role of calcium in the release of superoxide anion (O) was examined in alveolar macrophages after stimulation with the Soluble stimuli: concanavalin A (Con A), N‐formyl methionyl phenylalanine (FMP), and the calcium ionophore, A23187. The release of O by Con A was unaffected over a wide range of extracellular calcium concentrations (20 μM to 3 mM), whereas increasing the extracellular calcium above 2 mM inhibited FMP‐stimulated O release. In contrast, A23187 did not stimulate O release in calcium‐free medium (≤ 30 μM). The addition of EGTA (50 μM) to calcium‐free medium had no effect on Con A stimulation of O release or FMP‐stimulated O release. These results suggest that, for the three soluble stimuli, there are different roles for Ca+2 in the activation and transmission of stimulatory signals across the cell membrane.Con A‐ or FMP‐stimulated calcium efflux from calcium‐loaded cells in either calcium‐free medium or 0.5 mM calcium‐containing medium. In calcium‐free medium, FMP transiently retarded 45Ca+2 uptake, while in 0.5 mM calcium‐containing medium, FMP transiently stimulated 45Ca+2 uptake. For either Con A or FMP, calcium efflux preceded O release by 30–45 sec. Quinine, an agent that blocks membrane hyperpolarization in macrophages, completely blocked O release by concanavalin A or FMP and inhibited 45Ca+2 efflux by 50% or more for both agents. These results support the hypothesis that redistribution of cellular Ca+2 is one of the initial steps leading to the release of O.
Control of melanin synthesis and secretion by B16/C3 melanoma cells
Authors: Laskin Jeffrey D. , Piccinini Linda , Engelhardt D. L. , Weinstein I. Bernard 
Published: February 2005

AbstractIn culture, B16/C3 murine melanoma cells grown in the presence of serum undergo melanogenesis at a specific time after plating. At this time, melanin is synthesized intracellularly and then secreted into the extracellular culture fluid. We have found that melanin secretion is dependent on the presence of serum in the growth medium. When confluent cultures are deprived of serum, that is, refed with serum‐free medium, cells remain viable but do not undergo melanogenesis. Addition of serum‐free medium supplemented with either melanocyte‐stimulating hormone (MSH) or dibutyryl cAMP induced melanogenesis in these cells but did not result in melanin secretion. Furthermore, when B16/C3 cells are grown in serum‐free, hormone‐supplemented medium, they also undergo melanogenesis but fail to release melanin. The addition of serum, however, to B16/C3 cells induced to undergo melanogenesis with MSH, dibutyryl cAMP, or hormone‐supplemented medium promotes melanin secretion. Fractionation studies hence revealed that serum contains specific factors capable of inducing melanin secretion. These results demonstrate that factors that regulate melanin synthesis are distinct from those that induce cells to release melanin into their extracellular environment. Furthermore, the ability to induce melanogenesis with single factors will permit us to study the precise sequence of events leading to differentiation in B16/C3 cells under chemically defined conditions.
Cell cycle perturbations in normal and transformed fibroblasts caused by detachment from the substratum
Authors: Campisi Judith , Medrano Estela E. 
Published: February 2005

AbstractExponentially growing, anchorage‐dependent fibroblasts were impeded in their progress through the cell cycle as a result of brief trypsinization from the substratum followed by replating. Untransformed mouse (3T3, clone A31), hamster (CHEF/18‐1) and human (FS2) fibroblasts were partially inhibited from entering the DNA synthetic (S) phase of the cell cycle for 8 or 12 hours after detachment, even though the cells reattached within an hour of replating and attained a spread morphology 5 or 8 hours later. The decline in the proportion of cells in S phase was accompanied accumulation of cells in G1 as measured by autoradiography and flow microfluorimetry. Cells removed from the substratum by EDTA alone showed identical disturbances of exponential growth. These cell cycle perturbations could be a result of the detachment per se, as opposed to the rounded morphology. Synchronized A31 cells, exposed to colcemid or cytochalasin B for two hours, were not delayed in their entry into S, whereas trypsinization delayed S phase entry by 4 to 5 hours. These drugs disrupt the cytoskeleton without causing detachment. Isotope incorporation experiments revealed no decreases in the rates of protein or RNA synthesis following replating. However, exponentially growing A31 cells, treated for 2 hours with an inhibitor of protein synthesis behaved similarly to those briefly detached from their substratum: 7 hours after treatment, there were fewer cells in S and more cells in G1 relative to untreated cells. Brief treatment with an inhibitor of hn‐RNA synthesis did not alter the cell cycle distribution of these fibroblasts. Three tumorogenic A31 derivatives were less affected by brief detachment from the substratum than were the untransformed cells. The derivative exhibiting the least in vitro growth control (an SV‐40 transformant) showed the least sensitivity to trypsinization, while that derivative having the most in vitro growth control (a Moloney sarcoma virus transformant) was most sensitive. A chemically [benzo(a)pyrene] transformed derivative gave intermediate results with respect to both growth control and sensitivity to detachment from the substratum. The results suggest that as yet unidentified protein(s) necessary for the normal transit through G1 may be quite sensitive to the presence of an anchoring substratum.
Mitogenic proteins of the 3T3 plasma membrane
Authors: Lieberman Michael A. 
Published: February 2005

AbstractA mitogenic factor from 3T3 plasma membranes has been identified and partially characterized. The factor appears to be a peripheral membrane protein that can be released by mild trypsin, chymotrypsin, or plasmin treatment. This component is sensitive to heat and acid, and has a molecular weight in the range of 150,000–200,000 daltons as determined by gel filtration. A similar mitogenic activity has also been found on the membranes of both SV‐40‐transformed 3T3 cells and human fibroblasts. The factor appears to be distinct from all previously described mitogenic components.
Synthesis of cathepsin B by cells derived from the HL60 promyelocytic leukaemia cell line
Authors: Burnett David , Crocker John , Vaughan Andrew T. M. 
Published: February 2005

AbstractCells of the human promyelocytic HL60 line were induced to differentiate into granulocyte‐like cells with dimethylsulphoxide (DMSO) or macrophage‐like cells with 12‐0‐tetradecanoylphorbol‐13‐acetate (TPA). The synthesis of Cathepsin B by these cells was studied by immunoperoxidase staining and assay of cell lysates using the fluorimetric substrate benzoyloxycarbonyl‐phenylanalyl‐arginine‐4‐methyl‐7‐coumarylamide. Only 2–5% of the uninduced HL60 cells and DMSO‐induced cells were immunohistochemically positive for Cathepsin B, compared with over 80% of the TPA‐induced cells. Cathepsin B activity was lowest in the lysates of uninduced HL60s. DMSO‐induced cells contained 1.5–2‐fold the enzyme activity of HL60s and TPA‐induced cell lysates demonstrated 5–14‐fold the activity of uninduced HL60s. Induction of Cathepsin B synthesis was therefore associated with differentiation of the promyelocytes into cells of the monocyte/macrophage type, but not granulocyte‐like cells. Cathepsin B was located immunohistochemically in human palatine tonsils. The enzyme was only demonstrated within macrophages in these tissues. Cathepsin B may therefore be a useful immunohistochemical marker for malignant and nonmalignant cells of the monocyte/macrophage lineage.
Synergistic killing of Hela cells by hydroxyurea and caffeine
Authors: Beetham Karen L. , Busse Paul M. , Tolmach L. J. 
Published: February 2005

AbstractSynchronous populations of HeLa S3 cells suffer synergistic killing during S phase in the presence of 0.5–5 mM hydroxyurea together with 5–10 mM caffeine. Both the rate and the extent of killing are greater than expected for independent action of the two drugs. Only simultaneous treatment is effective. The dependence of the synergistic killing on cell age resembles the age dependence for killing by hydroxyurea alone (>3 mM), but not that by high concentrations of caffeine. In addition, rapid killing occurs if caffeine is added to cultures that have been incubated in the presence of hydroxyurea from early G1 and are blocked at the beginning of S, although such cells are killed only slowly on continued incubation in ⩾ 10 mM hydroxyurea alone. Furthermore, cells that are incubated with the two drugs from early G1 begin to undergo synergistic killing at about 12 h after mitotic collection, but they do not commence DNA replication for another 2–3 h if the drugs are removed. It is concluded that cells that have reached a point in the cycle identical with or close to the end of G1 are sensitive to the combination whether or not they are able to synthesize DNA, and whether or not they are sensitive to hydroxyurea alone. A tentative model is proposed: hydroxyurea is postulated to kill cells by interacting with sites of replication in DNA, and the synergism is attributed to the extra replication points that caffeine is known to induce.
The study of human myeloid differentiation using bromodeoxyuridine (BrdU)
Authors: Keoffler H. Phillip , Yen James , Carlson John 
Published: February 2005

AbstractLittle is known concerning the mechanism of myeloid differentiation. A human promyelocytic cell line (HL‐60) differentiates to granulocytes or macrophage‐like cells when cultured with a variety of agents. How these agents trigger myeloid differentiation is not understood. This study shows that 1.0–10.0 μg/ml bromodeoxyuridine (BrdU) induced myeloid differentiation of HL‐60 in liquid culture. After 7 days, BrdU (3.0 μg/ml) produced only moderate inhibition of HL‐60 growth, but induced myeloid maturation with 40% of the cells becoming morphologically more mature; 41% developed the ability to reduce nitroblue tetrazolium (NBT); 19% phagocytized Candida albicans; and 18% developed Fc receptors. The action of BrdU was mimicked by 5‐iododeoxyuridine. Thymidine (Td) (1‐ to 10‐fold excess) competitively inhibited incorporation of [3H]BrdU into DNA of HL‐60 and inhibited the triggering of HL‐60 differentiation by BrdU. The BrdU‐induced maturation of HL‐60 correlated with the incorporation of BrdU into DNA of HL‐60. DNA buoyant density studies showed that about 46% of the Td was replaced by BrdU in each DNA strand of HL‐60 as the cells differentiated in culture containing 3 μg/ml BrdU for 7 days. We established 20 thymidine kinase (TK)‐deficient HL‐60 clones. The HL‐60 TK‐deficient cells were unable to phosphorylate Td, to incorporate either [3H]Td or [3H]BrdU or differentiate in the presence of BrdU (1‐1000 μg/ml). The HL‐60 TK‐deficient cells retained the ability to differentiate in the presence of other HL‐60 inducers. Taken together, the studies suggest myeloid differentiation of HL‐60 is triggered because of incorporation of BrdU into DNA of the cells.
Insulin‐like growth factor I/somatomedin‐C (IGF‐I/SM‐C) and glucocorticoids synergistically regulate mitosis in competent human fibroblasts
Authors: Conover Cheryl A. , Dollar Laura A. , Hintz Raymond L. , Rosenfeld Ron G. 
Published: February 2005

AbstractIn serum‐free medium, insulin‐like growth factor‐I/somatomedin‐C (IGF‐I/SM‐C) was weakly mitogenic for adult human fibroblasts in culture. However, in the presence of 0.5% human hypopituitary serum (HHS), which by itself had little effect, there was a marked dose‐dependent response to IGF‐I/SM‐C with a 10‐ to 20‐fold increase in [3H]thymidine incorporation at 25 ng/ml IFG‐I/SM‐C. With the further addition of dexamethasone or hydrocortisone to the combination of IGF‐I/SM‐C + 0.5% HHS, there was a dramatic synergistic effect resulting in a 60‐ to 70‐fold increase in [3H]thymidine incorporation. This stimulation was two times greater than that seen with 20% FCS. In contrast, glucocorticoids had no effect in serum‐free medium or with HHS alone. These [3H]thymidine incorporation results were clearly supported by cell replication studies. Dose‐response curves for 125I IGF‐I/SM‐C binding and IGF‐I/SM‐C stimulation of [3H]thymidine incorporation were similar with 1/2 maximal effects for both at 5 ng/ml. However, the striking synergism seen with glucocorticoids occurred in the absence of any glucocorticoid‐induced change in IGF‐I/SM‐C binding, indicating that the interaction of IGF‐I/SM‐C and glucocorticoids occurs at a postreceptor level. These data demonstrate that in the presence of a low concentration of HHS, IGF‐I/SM‐C and glucocorticoids stimulate complete cell cycle traverse and replication of human fibroblasts.
Calcium uptake by isolated rat intestinal cells
Authors: Bronner Felix , Pansu Danielle , Bosshard Arlette , Lipton Jeffrey H. 
Published: February 2005

AbstractIntestinal cells were isolated by a combination of mechanical and enzymatic means, and their calcium uptake was assayed by a rapid filtration procedure. Calcium uptake was a time‐ and concentration‐dependent process that was markedly elevated at 25 and 37°C, as compared to 0°C. Cells isolated from rat duodenum exhibited higher uptakes than cells from jejunum, which in turn took up more calcium than cells from the ileurn. Duodenal cells from vitamin D‐deficient animals took up less calcium than cells from vitamin D‐replete cells. In vivo vitamin D repletion with 1,25‐dihydroxyvitamin D3 raised calcium uptake by duodenal cells from treated animals toward that of cells from replete rats. Furthermore, calcium uptake by duodenal cells from vitamin D‐deficient animals approximated that of ileal cells from replete rats. These findings with isolated cells parallel prior findings of tissue calcium transport and suggest that cellular calcium uptake may be related to the saturable component of intestinal calcium absorption. Isolated intestinal cells may therefore constitute one experimental model for the study of transcellular calcium transport.
Slow internalization of human chorionic gonadotropin by cultured granulosa cells
Authors: Robinson Margaret S. , Rhodes James A. , Albertini David F. 
Published: February 2005

AbstractKinetic studies were performed on two‐day cultures of rat ovarian granulosa cells to follow the fate of surface‐bound 125l‐labeled human chorionic gonadotropin (125l‐hCG). Low pH was used to release hCG from its surface receptor, allowing us to distinguish between surface‐bound and internalized hormone. Because our results indicated that hormone is lost from the cell surface by dissociation as well as internalization, equations were derived to determine independent rate constants for each process. We calculate that if hormone binding were irreversible, the t1/2 for internalization would be 8.5 hours. Morphometric studies on the uptake of horseradish peroxidase indicate that the t1/2 for internalization of bulk membrane in granulosa cells is 55 to 77 minutes. Thus, the rate of uptake of surface‐bound hCG appears to be seven to nine times slower than the rate of uptake of bulk plasma membrane, which suggests that the LH/hCG receptor may be selectively excluded from the endocytic vesicles of granulosa cells.
Biosynthesis of small nuclear RNAs in human cells
Authors: Chandrasekharappa Settara C. , Smith James H. , Eliceiri George L. 
Published: February 2005

AbstractWe have examined some aspects of the biosynthesis of human small nuclear RNAs (snRNAs). The sensitivity of U5 and U4 snRNA synthesis to α‐amanitin in whole cells suggests that RNA polymerase II is involved in the synthesis of these RNA species, in addition to that of U1, U2, and U3 snRNA. Two RNA bands were detected, whose properties are compatible with being U3 and U4 RNA precursors. The cytoplasmic U1 RNA precursor (pU1) was retained by an anti‐RNP antibody column, while the cytoplasmic precursors to U1 and U2 (pU2) RNA were immunoprecipitated by monoclonal anti‐Sm antibodies. Therefore, soon after their transcription, these cytoplasmic RNA precursors assemble with the polypeptides which bear the RNP (pU1) and Sm (pU1 and pU2) antigenic determinants. It has been shown before that, shortly after protein synthesis is interrupted, the apparent cytoplasmic→nuclear transition of newly made U2 RNA is inhibited, while U2 RNA transcription is not. The present data indicate that the trimming of the U2 RNA precursor to mature U2 RNA is not affected early after suppression of protein synthesis.
The uncoupling of macromolecular synthesis from cell division in SV3T3 cells by glucocorticoids: The imposition of a G2 block
Authors: Das Hasi R. , Lavin Mary , Sicuso Anthony , Young Delano V. 
Published: February 2005

AbstractThrough a receptor‐mediated process glucocorticosteroids block cell division by 20–45 hours in SV40‐transformed 3T3 (SV3T3) mouse fibroblasts growing in a low calf serum (0.30% v/v) medium containing biotin. However, the rate of DNA synthesis, determined at various times after dexamethasone addition by the incorporation of radioactive thymidine into acid‐insoluble material, is not inhibited by this steroid as late as 66 hours. A modest decrease is observable by 91 hours. There is also no reduction in the uptake of exogenous thymidine into acid‐soluble cellular pools. Similarly, RNA synthesis and the uptake of radioactive uridine are not affected by the glucocorticoid up to 69 hours. Measurements of the amounts of cellular DNA (by the fluorescent dye, 4′,6‐diamidino‐2‐phenylindole) and protein revealed that both macromolecules are present in elevated quantities in steroid‐treated cells. (The constancy of the protein content in the nonproliferative stage suggests that protein synthesis and degradation are occurring at equal rates.) If the steroid is removed and fresh 10% calf serum medium added, cell division commences (even if nearly 90% of protein synthesis is inhibited by cycloheximide) as early as 45 minutes later such that by 2 hours the viable cell count increases by as much as 70%. Since the growth curve after recovery resembles a step function, it appears that the cells are partially synchronized by the glucocorticoid. These results demonstrate that the glucocorticoid cytostatic effect in SV3T3 cells is the result of a block not in G1, as previously thought, but in G2.
Demonstration of a late amiloride‐sensitive event as a necessary step in initiation of DNA synthesis by thrombin
Authors: Stiernberg Janet , Labelle Edward P. , Carney Darrell H. 
Published: February 2005

AbstractAmiloride, a Na+ influx inhibitor, has been shown to inhibit initiation of DNA synthesis by thrombin in mouse embryo fibroblast‐like cells. Long exoosures (24 hr) to high concentrations of amiloride inhibited incorporation of thymidine into the DNA of both thrombin‐stimulated and nonstimulated cells, suggesting that this inhibition might not be specific for thrombin‐initiated DNA synthesis. Fluorescence microscopy and spectrofluorimetry showed that amiloride was internalized with an apparent mitochondrial association and that the internalized amiloride was readily released from the cells after removing amiloride from the medium. Based on this reversibility, cells were exposed to amiloride for short periods of time during thrombin treatment to determine the temporal relationship between any amiloride‐sensitive event(s) and initiation of DNA synthesis. The presence of amiloride (100 μM) during a 12‐hr exposure to thrombin did not block thrombin‐initiated DNA synthesis or cell division but did delay the onset of DNA synthesis and the peak of thymidine incorporation into DNA by approximately 3 hr, suggesting that early initiation events might proceed in the presence of amiloride. 86Rb+ transport studies demonstrated that in this system ouabain‐sensitive K+ uptake via the Na, K‐ATPase was stimulated by thrombin during both an early and a late period. This stimulation was amiloride‐sensitive under the same conditions used for growth experiments, suggesting that amiloride was inhibiting thrombin‐stimulated Na+ transport in this system. Additional experiments showed that exposing cells to amiloride only during the first 8 hr after thrombin addition did not inhibit initiation. The presence of amiloride from 8–12 hr after thrombin addition maximally inhibited thrombin‐stimulated DNA synthesis. Together these results demonstrate that amiloride inhibits thrombin‐initiated DNA synthesis not by inhibiting an early event occurring during the first 8 hr, but rather by inhibiting some later event 8–12 hr after thrombin addition.