bluuebunny/crossref_metadata_2025_split huggingface dataset:
Thy‐1 is a differentiation antigen that characterizes immature murine erythroid and myeloid hematopoietic progenitors
Authors: Miller Barbara A. , Lipton Jeffrey M. , Linch David C. , Burakoff Steven J. , Nathan David G. 
Published: February 2005

AbstractTo determine the role of Thy‐1 antigen in murine hematopoietic differentiation, bone marrow was treated with anti‐Thy‐1.2 antibody and complement or complement alone. Growth of immature hematopoietic progenitors, erythroid burst‐forming units (BFU‐E), and granulocyte/macrophage colony‐forming units (CFU‐GM) was greatly reduced following antibody and complement treatment and was not restored by mitogen‐stimulated spleen cell supernatants. In contrast, more mature erythroid and myeloid progenitors, the erythroid colony‐forming unit (CFU‐E) and the macrophage progenitor stimulated by L‐cell‐conditioned media (LCM), were spared by anti‐Thy‐1.2 antibody and complement treatment. Here, to separate the effects of anti‐Thy‐1.2 antibody treatment on accessory cells from those on progenitors, splenic T cells and thymocytes were added to treated marrow at ratios of up to 200%. Growth of BFU‐E and CFU‐GM was not restored. To more precisely replace required accessory cells, male complement‐treated marrow was cocultured with female anti‐Thy‐1.2 antibody and complement‐treated marrow. Even marrow cells failed to restore female BFU‐E and CFU‐GM growth. Fluorescent‐activated cell sorting (FACS) and immune sheep red cell rosetting with anti‐Thy‐1.2‐labeled marrow were then performed to determine if immature hematopoietic progenitors bear Thy‐1.2‐positive fraction, demonstrating the presence of Thy‐1.2 on early murine hematopoietic progenitors. CFU‐E and CFU‐M were present in the Thy‐1.2‐negative fraction following FACS separation. These data demonstrate that Thy‐1.2 is a differentiation antigen, present on at least some murine BFU‐E and CFU‐GM and lost as they mature to CFU‐E and CFU‐M.
Altered patterns of keratin synthesis in human epidermal keratinocytes transformed by SV40
Authors: Steinberg Mark L. , Defendi Vittorio 
Published: February 2005

AbstractTransformation of human epidermal keratinocytes by the oncogenic virus SV40 is a stage‐specific process in which normal patterns of differentiation are progressively altered over time following infection. Within the context of this scheme, we examined the keratins produced by the infected cells. Immunofluorescence studies indicated that viral infection led to the formation of variant cells visibly lacking the normal keratin cytoskeleton after about 10–15 serial passages (60–90 cell generations) post infection. Analyses of variant cell formation in clonal populations grown on palladium islands revealed that the variants were derived within 2–3 cell divisions from cells containing an apparently normal keratin cytoskeleton, but that variant formation depended upon cell density. Immunoprecipitation of 35S‐methionine labelled keratins from the infected keratinocytes revealed a gradual loss of the normal 46, 50, 56 and 58Kd keratin species over a period of many months after infection. The loss of the normal keratins was accompanied by the appearance of at least two species in the 48–52Kd size range not present in uninfected cells and the enhancement of a third, 40Kd, protein quite early after infection. Analysis of the altered keratin patterns on two‐dimensional acrylamide gels using either isoelectric focusing (IEF) or non‐equilibrium pH gradient electrophoresis (NEPHG) along the first dimension showed that the infected cells came more transformed with serial passage including at least five isoelectric forms not seen in uninfected cells. Translation of poly A+ RNAs from the infected cells indicated that the altered keratin synthesis probably reflects changes in the translatable mRNA pool.
Cationic amino acid transport by two renal epithelial cell lines: LLC‐PK1 and MDCK cells
Authors: Sepúlveda Francisco V. , Pearson Jeremy D. 
Published: February 2005

AbstractLLC‐PK1 and MDCK cells take up cationic amino acids (lysine and arginine) by a specific sodium independent transport system. Uptake is inhibited by ornithine in LLC‐PK1 and MDCK cells either in the presence or absence of sodium and by glutamine or homoserine in MDCK cells in the presence of sodium. Trans‐stimulation of uptake occurs in the presence of intracellular cationic amino acids. Experiments with valinomycin or with different extracellular potassium concentrations suggest that uptake is dependent on the membrane potential of these cells. These transport features are similar to those previously ascribed to a transport system denominated y+ in other cells. Further experiments suggested that this carrier system is localised to the basolateral membrane in each cell type.
Ultrastructural analysis of the organization and distribution of insulin receptors on the surface of 3T3‐L1 adipocytes: Rapid microaggregation and migration of occupied receptors
Authors: Smith Robert M. , Cobb Melanie H. , Rosen Ora M. , Jarett Leonard 
Published: February 2005

AbstractMonomeric ferritin‐insulin and high‐resolution electron microscopic analysis were used to study the organization, distribution, and movement of insulin receptors on differentiated 3T3‐L1 adipocytes. Analysis of the binding to prefixed cells showed that insulin initially occupied single and paired receptors preferentially located on microvilli. The majority of receptors (60%) were found as single molecules and 30% were pairs. In 1 min at 37%C, 50% of the receptors on nonfixed cells were found on the intervillous plasma membrane and more than 70% of the total receptors had microaggregated. By 30 min only 7% of the receptors were single or paired molecules on microvilli. The majority were on the intervillous membrane, with 95% of those receptors in groups. The receptor groups on the intervillous plasma membrane could be found in both noncoated invaginations and coated pits. The concentration of occupied receptors in the noncoated invaginations and the coated pits was similar; however, ten times more noncoated invaginations than coated pits contained occupied insulin receptors. The observations in this study contrast with those reported on rat adipocytes using identical techniques (Jarett and Smith, 1977). Insulin receptors on adipocytes were initially grouped and randomly distributed over the entire cell surface and did not microaggregate into larger groups. Insulin receptors on rat adipocytes were found in non‐coated invaginations but were excluded from the coated pits. The differences in the organization and behavior of the insulin receptor between rat and 3T3‐L1 adipocytes suggest that the mechanisms regulating the initial organization of insulin receptors and the aggregation of occupied receptors may be controlled by tissue‐specific processes. Since both of these cell types are equally insulin sensitive, the differences in the initial organization and distribution of the insulin receptors on the cell surface may not be related to the sensitivity or biological responsiveness of these cells to insulin but may affect other processes such as receptor regulation and internalization. On the other hand, the microaggregates of occupied receptors on both cell types may relate to biological responsiveness.
Butyrate stimulates fibronectin synthesis in cultured rabbit cornea
Authors: Tanaka Hitoshi , Nishida Teruo 
Published: February 2005

AbstractTo understand corneal wound healing, in which fibronectin plays an important role, we investigated the regulatory mechanism of fibronectin synthesis in cultured rabbit corneal blocks in situ. The amount of fibronectin was determined by enzyme‐linked immunosorbent assay. We found that butyrate stimulated fibronectin synthesis in a dose‐response fashion, and that butyrate had a greater stimulatory effect than did any of its derivatives examined. This suggests that butyrate stimulates fibronectin synthesis specifically; 8Br‐cAMP also stimulated fibronectin synthesis. Additivity of stimulation by butyrate and by 8Br‐cAMP was observed at the saturated concentration of each; the present result indicated an independent mechanism of action for these two compounds.
Phospholipid asymmetry in human erythrocyte ghosts
Authors: Williamson Patrick , Algarin Lourdes , Bateman Judy , Choe Hye‐Ryun , Schlegel Robert A. 
Published: February 2005

AbstractUsing phospholipase digestion and the fluorescent probe merocyanine 540 the maintenance of phospholipid asymmetry in the plasma membrane of human erythrocyte ghosts was investigated. Digestion with phospholipase A2 indicated that ghosts prepare in the presence of Mg++ as the only divalent cation retained the normal phospholipid asymmetry characteristic of intact erythrocytes. These ghosts, like normal erythrocytes, also failed to stain with merocyanine 540. However, the presence of as little as 5–10 μM Ca++ during ghost preparation resulted in ghosts in which lipid asymmetry had been abolished, as indicated by phospholipase digestion. Moreover, these ghosts stained with merocyanine 540. In contrast to ghosts, intact erythrocytes treated with ionophore required millimolar levels of Ca++ ions to disrupt membrane lipid asymmetry. To discover the reason for this difference in behavior between ghosts and intact cells, ghosts were prepared from preswollen cells using only small volumes of buffer for lysis. These experiments demonstrated that as the cellular contents of erythrocytes are diluted, the asymmetric arrangement of phospholipids becomes more sensitive to disruption by Ca++.
Induction of differentiation in human myeloid leukemic cells by proteolytic enzymes
Authors: Fibach Eitan , Treves Aliza , Kidron Miriam , Mayer Michael 
Published: February 2005

AbstractExogenous serine proteases were found to induce differentiation in human myeloid leukemic cells from either in vitro established long‐term cell lines or in primary cultures of cells derived directly from patients with acute myeloid leukemia. Exposure of the human promyelocytic cell line HL‐60 to trypsin, chymotrypsin, or elastase induced the appearance, within 3–6 days, of neutrophilic granulocytes defined by their morphology, their ability to reduce nitroblue tetrazolium, and their efficient phagocytosis of latex particles. Upon further incubation monocytelike cells appeared. While these cells developed into fully mature macrophages other types of cells disappeared and on day 12 the culture consisted of a pure macrophage population. The inducing effect could be observed when the enzyme was presented alone, whereas a synergistic effect was noted when the protease was added in the presence of subthreshold concentrations of chemicals known to induce differentiation in this cell line such as dimethylsulfoxide, retinoic acid, butyric acid, or hexamethylene bisacetamide. Optimal induction of differentiation by trypsin required a 48 hr continuous exposure to the enzyme. When the protease was removed earlier no appreciable differentiation was noticed. The protease‐induced differentiation involved a direct interaction with the cells and was not due to a proteolytic cleavage of a serum component because it could be obtained in serum‐free cultures. The enzymatic activity of the protease was needed for its effect on cell maturation: Addition of protease inhibitors such as soybean‐trypsin inhibitor or trasylol completely blocked differentiation induced by the proteases but had no effect on differentiation induced by the other inducers. It is still to be determined whether a proteolytic process is a general molecular event in cell differentiation or induction by chemicals involves a mechanism different from that initiated by exogenous proteases.
Serum‐free conditions for the growth of avian granulocyte and monocyte clones and primary leukemic cells induced by AMV, and the apparent conversion of granulocytic progenitors into monocytic cells by a factor in chicken serum
Authors: Dodge W. H. , Sharma S. 
Published: February 2005

AbstractThe supportive activity of chicken serum for the soft‐agar growth of chicken granulocyte and monocyte clones could be replaced with defined ingredients [bovine serum albumin (BSA), conalbumin, selenium, linoleic acid and for routine work, a liquid soy lecithin preparation (59% phospholipids, 39% linoleic acid, and <2% of inositol and choline)]. The lecithin preparation could be replaced with L‐α‐phosphatidylcholine. The source of colony‐stimulating factor was medium conditioned by fibroblasts cultured under protein‐free conditions. AMV‐induced leukemic cells could be cloned under identical conditions. In the presence of both chicken serum (10%) and the replacement ingredients, most of the proliferative clones produced were monocytic (84%). In the presence of serum alone, all of them were monocytic. Under serumfree conditions, all the clones produced were granulocytic when a day‐3 conditioned medium (CM) was used; monocytic ones were also present when a day‐6 CM was used. When the serum was serially diluted in the presence of the replacement ingredients, the number of proliferative monocytic clones progressively decreased while the number of proliferative granulocytic clones progressively increased and the kinetics of each were essentially the same, only opposite in direction. Moreover, the total number of proliferative clones did not change more than 33% at any dilution (or in the absence of serum). We postulate the existence in the chick system of a serum macrophage differentiation factor (M‐DF) which converts early granulocytic progenitors (or exclusively diverts a common progenitor) into monocytic cells.
Pattern of steroid interaction with inducers of aryl hydrocarbon hydroxylase activity in several cloned estradiol‐responsive stromal cell strains from mouse endometrium
Authors: Stols L. , Iannaccone P. M. 
Published: February 2005

AbstractSeveral steroid responsive stromal cell strains were isolated from normal mouse endometrium. Both estrogen responsive and progesterone responsive strains were obtained. The estrogen responsive cells were also responsive to diethylstilbestrol (DES). The progesterone responsive strains were inhbited by estrone and estriol. One of the estrogen responsive cell strains, E041, was cloned in the presence and absence of 17‐β‐estradiol (E2). Of 14 clones developed in the absence of E2 and screened for responsivity of growth, six were significantly responsive to E2. Six of 26 clones developed in the presence of E2 were responsive to E2. Microsomes isolated from these clones were shown to have aryl hydrocarbon hydroxylase (AHH) activity which was inducible with either benzanthracene (BA) or 2,3,7,8 tetrachlorodibenzo‐p‐dioxin (TCDD). The basal levels of AHH activity were significantly reduced by E2 in E2 responsive clones. The TCDD induced levels of AHH activity in the E2 responsive and E2 non‐responsive clones were significantly increased by E2 in one of the clones. These results show that mouse endometrial stroma consists of a number of populations of cells with different specific proliferative and metabolic responses to steroids.
The effect of biotin on cellular functions in HeLa cells
Authors: Bhullar R. P. , Dakshinamurti K. 
Published: February 2005

AbstractHeLa cells cultured in a biotin‐deficient medium show reduced rate of protein synthesis, DNA synthesis and growth. Addition of exogenous biotin to the cells cultured in biotin‐deficient medium results in enhanced protein synthesis, DNA synthesis and cell growth. Continuous protein synthesis is required for the increase in DNA synthesis observed upon the addition of exogenous biotin to the cells cultured in biotin‐deficient medium. These results suggest that cells cultured in biotin‐deficient medium are arrested in the G1 stage of cell cycle and this block is removed upon the addition of biotin to the deficient medium.
The effect of transport system A and N amino acids and of nerve and epidermal growth factors on the induction of ornithine decarboxylase activity
Authors: Rinehart Clifford A. , Viceps‐Madore Dace , Fong Wang‐Fun , Ortiz José G. , Canellakis Evangelo S. 
Published: February 2005

AbstractThe induction of ornithine decarboxylase (EC 4.1 1.17) (ODC) by amino acids and by the peptide hormones nerve growth factor (NGF) and epidermal growth factor (EGF) in salts‐glucose media has been studied. Only those neutral amino acids taken into the cell via one of the Na+ dependent transport systems stimulate ODC activity. Asparagine and the nonmetabolizable α‐amino‐isobutyric acid (AIB) were used as representatives of this class of inducing amino acids, and their intracellular concentrations were related to the levels of ODC induced. A threshold intracellular concentration of asparagine or AIB has to be attained before ODC can be induced. Further slight increases in intracellular concentrations of asparagine or AIB produce disproportionately large increases of ODC, resulting in a sigmoidal curve of ODC by valine is associated with a concurrent decrease in the intracellular level of the inducing amino acid, suggest that the intracellular amino acid level is causally related to the induction of ornithine decarboxylase. Glutamic acid, EGF, and NGF do not induce ODC except in the presence of an inducing amino acid. They act synergistically with the inducing amino acid and produce higher ODC levels at the same intracellular concentration of the inducing amino acid.
Effect of cell density on metabolism in isolated rat hepatocytes
Authors: Jurin Richard R. , McCune Sylvia A. 
Published: February 2005

AbstractFreshly isolated rat hepatocytes show many changes in metabolic activities as a function of cell density in the incubation flask. Fatty acid synthesis, cholesterol synthesis, general protein synthesis, and rates of accumulation of pyruvate, lactate, citrate, acetyl‐CoA, acetoacetate and β‐hydroxybutyrate decrease as the cell density increases between 1 mg/ml and 60 mg/ml. Glucose release only decreases between 1–5 mg/ml and the concentration of ATP does not vary at any density. There is a small increase in the lactate/pyruvate ratio and a dramatic decrease in the β‐hydroxybutyrate/acetoacetate ratio with increasing cell concentration. When cells at 8 or 28 mg/ml were incubated with added lactate and pyruvate, or alanine, a two fold increase in fatty acid synthesis and 50% decrease in cholesterol synthesis were observed as compared to rates with endogenous substrate. With added glucose the synthetic rates were similar to those obtained with endogenous substrate. However, regardless of the type of substrate used, the less dense cells gave rates up to three times greater than that of the more dense cells. When conditioned medium isolated after incubation of cells at high density was added to the less dense cells, a decrease in the rates of fatty acid synthesis and cholesterol synthesis was observed in the less dense cells; however, when medium from the less dense cells after incubation for the same period was added to the more dense cells, there was no significant change in fatty acid or cholesterol synthesis. These results suggest that a factor may be released into the medium of incubating hepatocytes that progressively inhibits certain metabolic processes as the cell density increases.
Single high affinity binding of interferon α2 to receptors on human lymphoblastoid cells: Internalization and inactivation of receptors
Authors: Faltynek Connie R. , McCandless Sarah , Baglioni Corrado 
Published: February 2005

AbstractThe interaction of human recombinant interferon (rIFN) α2 with its receptor on lymphoblastoid cells was studied using competitive displacement binding. The data were analysed with the LIGAND program, which tests their fit to one‐site or multiple binding site models. The binding at 4° and 37°C fits a one‐site model, with a similar KD for both IFN‐sensitive and resistant cells. Binding at 37°C to Daudi cells at high density fits artifactually a two‐site mode only when the receptor concentration is close to that of the KD. The binding of IFN to its receptor, therefore, follows a simple bimolecular interaction. Furthermore, IFN‐sensitive and resistant cells internalize IFN at similar rates. We have examined whether IFN receptors are also internalized and whether they subsequently recycle to the cell surface. By measuring cell surface and total receptors, we have observed that after 2 h treatment with IFN total receptors remain constant whereas cell surface receptors decrease. After prolonged treatment with IFN, however, there is a loss of total receptors. By inactivating cell surface receptors with proteinase K, we have shown that a fraction of cell surface receptors becomes resistant to inactivation and is apparently internalized. Moreover, experiments which measure IFN receptors either during incubation in the presence of IFN or after IFN has been removed from the medium, show that receptors do not recycle to the cell surface after internalization. The addition of monensin, a drug which has been shown to inhibit receptor recycling, has no effect on the loss of IFN receptors.
Modulation of transferrin receptor expression by inhibitors of nucleic acid synthesis
Authors: Hedley David , Rugg Catherine , Musgrove Elizabeth , Taylor Ian 
Published: February 2005

AbstractWe investigated the effects of the iron chelator desferrioxamine on the expression of transferrin receptors (TfR) by CCRF‐CEM human T‐cell leukaemia and B16 mouse melanoma cells growing in tissue culture. Desferrioxamine (DFOA) enhanced TfR expression when added in the dosse range of 10−5−10−4 to CCRF‐CEM cells, but was toxic to these cells, the lower concentrations producing a slowing of cell growth with a build up in S‐phase, while higher concentrations caused cell death with a block at the G1/S‐phase interface. These toxic effects are compatible with its previously reported inhibition of teh non‐haen iron containing (M2) subunit of ribonucleotide reductase. In marked contrast, DFOA caused the growth of B16 melanoma cells to arrest in G1, without loss of cloning efficiency, and resulted in a fall in TfR expression to approximately 50% of control values. These results suggested that the effects of DFOA on TfR expression were linked to DNA synthesis rather than to a more generalised inhibition of iron‐depdendent cellular processes. It was subsequently found that inhibition of the M2 subunit of ribonucloetide reductase in CCRF‐CEM cells with 5 x 10−5 M hydroxyurea, which is not an iron chelator, also enhanced TfR expression, as did thymidine and Cytosine arabinoside, which have different enzyme targets. By measuring cellular DNA and RNA content simultaneously it was shown that all of these agents caused unbalanced growth, i.e., inhibited DNA synthesis more than RNA synthesis. In contrast, 6‐thioguanine was more inhibitory to RNA synthesis, and treatment with this drug caused a fall in TfR expression. Thus, although CCRF‐CEM cells treated with DFOA show enhanced TfR expression, similar effects are also seen with other inhibitors of DNA synthesis, provided thatRNA synthesis is allowed to continue. These results provide further evidence that the regulation of TfR expression by proliferating cells is specifically linked to DNA synthesis rather than to the iron requirements of other cellular processes.