bluuebunny/crossref_metadata_2025_split huggingface dataset:
Modulation of smooth muscle cell behaviour by platelet‐derived factors and the extracellular matrix
Authors: Wren Fiona E. , Schor Ana M. , Schor Seth L. , Grant Michael E. 
Published: February 2005

AbstractWe have studied the combined effects of platelet‐derived soluble factors and three types of macromolecular substrata on the proliferation and migration of smooth muscle cells in vitro. Bovine aortic smooth muscle cells were plated onto three‐dimensional gels of type I collagen or onto cell‐free extracellular matrices deposited on such gels by either bovine aortic endothelial cells or smooth muscle cells. The cells were cultured in the presence of whole‐blood serum (WBS) or platelet‐poor plasma (PPP). Smooth muscle cell proliferation on type I collagen gels was dependent on the presence of platelet‐derived factors, i.e. the cells proliferated in the presence of WBS but not in PPP. In contrast, cell proliferation on the extracellular matrices occurred at the same rate in PPP and WBS. Smooth muscle cells plated onto collagen gels rapidly migrated down into the gel matrix; the percentage of cells migrating was inversely proportional to cell density. The presence of extracellular matrices did not alter the rate of cell migration into the underlying gel matrix. Irrespective of the substratum used, smooth muscle cell migration was independent of platelet‐derived or plasma factors and occurred in the absence of proliferation. These results indicate that possible chemotactic, chemokinetic, and/or mitogenic factors produced by the vascular cells and deposited within the extracellular matrix may play an important role in modulating smooth muscle cell behaviour in the vascular wall.
Effect of 1,3,6‐triaminohexane and 1,4,7‐triaminoheptane on growth and polyamine metabolism in SV‐3T3 cells treated with 2‐difluoromethylornithine
Authors: McGovern Kathy A. , Clark Richard S. , Pegg Anthony E. 
Published: February 2005

AbstractThe possibility that one or both of the synthetic triamines, 1,3,6‐triaminohexane and 1,4,7‐triaminoheptane, could substitute for the naturally occurring polyamines in the growth of SV‐3T3 cells was investigated. It was found that these triamines did lead to a restoration of growth in cells in which spermidine content had been depleted by exposure to the ornithine decarboxylase inhibitor 2‐difluoromethylornithine. This resumption of a normal growth rate occurred prior to the reduction in the content of cellular decarboxylated Sadenosylmethionine, suggesting that this nucleoside (which increases in concentration several hundred‐fold in cells treated with 2‐difluoromethylornithine) does not cause the reduction of cell growth. However, unlike the increase in cell growth brought about by spermidine, which continued indefinitely, the increase produced by 1,3,6‐triaminohexane or 1,4,7‐triaminoheptane was transient. Cell growth in the presence of 2‐difluoromethylornithine and these triamines stopped after about three or four population doublings. This corresponded to the time at which the intracellular spermine content of the cells was reduced to values less than 20% of normal. It is suggested that the increased growth rate of spermidine‐depleted cells in response to these triamines is due to their uptake into the cell and ability to displace spermine from intracellular sites, thus making spermine available to fulfill the polyamine function(s) essential for growth. These results indicate that the naturally occurring polyamines spermidine or spermine are essential for continued cell growth and cannot be replaced by analogues containing only primary amino groups.
Regulation of the expression of the SV40 T‐antigen coding gene under the control of an rDNA promoter
Authors: Surmacz Ewa , Rønning Øystein , Kaczmarek Leszek , Baserga Renato 
Published: February 2005

AbstractWe have constructed a hybrid gene in which the SV40 T‐antigen coding gene is driven by a mouse rDNA promoter and we have compared its expression to that of an SV40 T‐antigen coding gene under the control of its own promoter. The comparison has been carried out in microinjected cells, in transfected cells, and in stable cell lines carrying the respective T‐antigen coding genes in an integrated form. These cell lines were derived from ts AF8 cells, a mutant which is temperature sensitive for RNA polymerase II activity. The hybrid gene clearly expresses T‐antigen, albeit less efficiently than when the T antigen coding gene is under the control of the SV40‐promoter. We also show that the expression of T‐antigen by the hybrid gene is 50% inhibited by an antibody against RNA polymerase I. In tsAF8 cells carrying the hybrid gene, T‐antigen is still expressed at the restrictive temperature (where RNA polymerase II is inactive) at a level again about 50% of controls. However, our findings also confirm those of Smale and Tjian (Mol. Cell. Biol. 5:352, 1985) that such hybrid genes are in part transcribed by RNA polymerase II and generate abnormal transcripts.
Interleukin 3 and cell cycle progression
Authors: Kelvin David J. , Chance Susan , Shreeve Mona , Axelrad Arthur A. , Connolly Joe A. , McLeod David 
Published: February 2005

AbstractInterleukin 3 (IL‐3) is a regulatory glycoprotein required for the proliferation and differentiation of cells from many if not all hemopoietic lineages. With the emergence of the competence‐progression model of cell proliferation, which predicts that growth factors function at specific stages of the cell cycle, we examined the possibility that IL‐3 functions at a specific stage of the cell cycle. C‐63 cells were developed as a cell line from normal murine bone marrow. They have a mast cell phenotype and require pokeweed‐stimulated spleen cell‐conditioned medium (CM), a rich source of IL‐3, for their continued growth. Exponentially growing cells were transferred from growth medium, which contains CM, to medium lacking CM or IL‐3. After 24 hours, cell viability had decreased 40–50%. The remaining viable cells did not incorporate 3H‐thymidine, and displayed a single peak at G1 in a DNA histogram. Restimulation of these cells with CM or IL‐3 resulted in a dramatic rise in 3H‐thymidine uptake 20–24 hours after restimulation. DNA histograms of restimulated cultures indicated that the cells were progressing in a wave‐like fashion throughout the remainder of the cell cycle. The length of time necessary for cells to be in contact with CM or IL‐3 before they could progress into the remainder of the cell cycle was also examined. Cells incubated with CM or IL‐3 for less than 16 hours could not progress into S phase, whereas cells incubated for 16 hours or longer could progress into S phase and through the remainder of the cell cycle. These data suggest that IL‐3 exerts its function at a specific stage of the cell cycle.
Insulin‐like growth factor I regulation of transcription and replicating enzyme induction necessary for DNA synthesis
Authors: Yang Henry C. , Pardee Arthur B. 
Published: February 2005

AbstractInsulin‐like growth factor I (IGF‐I) regulation of the sequence of transcriptional, translational, and postranslational events during G1 that are necessary for DNA synthesis, and the induction of thymidine kinase and in vivo thymidylate synthase activities was studied in synchronized confluent BALB/c 3T3 mouse fibroblasts. IGF‐I was the only growth factor necessary from 6 to 2½ hours before S phase for onset of DNA synthesis. 5,6 dichlororibofuranosylbenzimidazole (DRB), an inhibitor of transcription, was ineffective in blocking DNA synthesis and increase of both enzyme activities during an interval of about 2½ hours before S phase. Cycloheximide, an inhibitor of translation, was ineffective in blocking DNA synthesis and the increase of in vivo thymidylate synthase activity during an interval of about 1½ hours before S phase but was effective in blocking the increase of thymidine kinase activity up to the G1/S boundary. The results demonstrate that IGF‐I is dispensable beyond the R‐point (2–3 hours before S phase), the time when serum factors are no longer necessary for the initiation of DNA synthesis, and that IGF‐I regulates transcriptional events necessary for both DNA synthesis and the induction of thymidine kinase and in vivo thymidylate synthase. The results also demonstrate a posttranslational interval for in vivo thymidylate synthase, suggesting posttranslational modification of this enzyme.
Interferon pretreatment lowers the threshold for maximal heat‐shock response in mouse cells
Authors: Morange M. , Dubois M. F. , Bensaude O. , Lebon P. 
Published: February 2005

AbstractInterferons (IFNs) are proteins which have antiviral and antiproliferative properties and are known to affect various immunological processes. Some of these activities have been shown to be potentiated by increased temperatures. When cells are subjected to a rise in temperature, the synthesis of the heat‐shock proteins (HSPs) is ‘switched on.’ In this report we demonstrate a synergistic effect of IFN and stress (arsenite treatment or elevated temperature) on the heat‐shock response. On the one hand, IFN pretreatment enhances the accumulation of HSP mRNAs and the corresponding protein synthesis after a mild stress and, on the other hand, it amplifies the decrease of the total protein synthesis after a severe stress. Thus in IFN pretreated cells the range of temperatures leading to the heat‐shock response is shifted towards common physiological values.
Substrate utilization for lactate and energy production by heat‐shocked L929 cells
Authors: Lanks Karl W. , Hitti Ibrahim F. , Chin Nena W. 
Published: February 2005

AbstractThe hypothesis that heat shock protein (HSP) induction depends on inhibition of respiration was tested by examining the effects of heat shock on tricarboxylic acid (TCA) cycle function. In control L929 cell cultures, glucose and exogenous pyruvate were converted primarily to lactate, and glutamine was extensively oxidized, accounting for more than one‐half of the calculated ATP production. During heat shock at 42°C, lactate production from all of the labeled substrates and total unlabeled lactate production increased significantly while oxygen consumption increased slightly. TCA cycle oxidation of pyruvate decreased during this period while that of glutamine increased. Uncoupling of oxidative phosphorylation caused large increases in oxygen consumption at both 37°C and 42°C, indicating that the capacity of the respiratory chain is not exceeded during heat shock. The net effect of these alterations in substrate utilization were decreased ATP generation and increased NADH utilization. Both 14CO2 and lactate production declined during the 24‐h period after cultures were returned to 37°C. On the basis of these data, we conclude that while inhibition of respiration plays no apparent role other metabolic consequences of heat shock related to energy metabolism may be involved in HSP induction.
Age dependency of the metabolic conversion of polyamines into amino acids in IMR‐90 human embryonic lung diploid fibroblasts
Authors: Chen Kuang Yu , Chang Zee‐Fen 
Published: February 2005

AbstractWhen radioactive polyamines (putrescine or spermidine) were incubated with mammalian cells in tissue culture, the radioactivity was incorporated into cellular proteins via two different metabolic pathways; one is metabolic labeling of an 18,000‐dalton protein via hypusine formation, and the other is general protein synthesis employing radioactive amino acids derived from biodegradation of polyamines via GABA shunt and Krebs cycle. Aminoguanidine, a potent inhibitor of diamine oxidase, blocked the metabolic conversion of polyamines to amino acids but had no effect on the metabolic labeling of the 18,000‐dalton protein. We have investigated these two polyamine‐associated biochemical events in IMR‐90 human diploid fibroblasts as a function of their population doubling level (PDL). We found that (1) the metabolic labeling of the 18,000‐dalton protein was about two‐fold greater in young cells (PDL = 22) than that in old cells (PDL = 48), and (2) the metabolic labeling of other cellular proteins, employing amino acids derived from putrescine via polyamine catabolic pathway, was more than six‐fold greater in the old cells (PDL = 48) than in the young cells (PDL = 22). Since the rate of protein synthesis was about 1.4‐fold higher in the young cells as compared to the old cells, our data indicated that the activity of catabolic conversion of putrescine (or spermidine) to amino acids in old IMR‐90 cells was about eight‐fold greater than that in young cells. This remarkable increase of polyamine catabolism and the slight decrease of metabolic labeling of the 18,000‐dalton protein were also observed in cell strains derived from patients with premature aging disease.
Collagen synthesis and deposition during mammary epithelial cell spreading on collagen gels
Authors: Marinkovich Matt P. , Rocha Victor 
Published: February 2005

AbstractMouse mammary epithelial cultivated on collagen gels demonstrate active spreading as the cells form monolayers. In this novel system, initiation of cell spreading is preceded by de novo synthesis of type IV collagen. The newly synthesized collagen is partitioned such that after 48 hr, approximately 24% is found in the culture medium, 35% is intracellular, and 41% is deposited in the extracellular matrix of the developing epithelium. Cultures deprived of serum failed to spread and to synthesize collagen. Proline analogues were shown to inhibit cell spreading and to suppress collagen synthesis in a dosedependent manner. Cytochalasin D inhibition of F‐actin elongation was shown to prevent cell spreading but not to suppress total collagen synthesis. During cytochalasin D treatment, inhibition of cell spreading was shown to result from failure to deposit or to maintain deposited collagen in the epithelium extracellular matrix. The data indicate that synthesis and extracellular deposition of a major basal lamina component (viz. type IV collagen) must precede and then accompany epithelial cell spreading in collagen gel culture. It is suggested that the microfilament apparatus, through some hypothetical integral membrane protein, can anchor extracellular type IV collagen, which then provides a necessary condition for cell spreading.
Thrombin‐induced increase in albumin permeability across the endothelium
Authors: Garcia Joe G. N. , Siflinger‐Birnboim Alma , Bizios Rena , Del Vecchio Peter J. , Fenton John W. , Malik Asrar B. 
Published: February 2005

AbstractWe studied the effect of thrombin on albumin permeability across the endothelial monolayer in vitro. Bovine pulmonary artery endothelial cells were grown on micropore membranes. Morphologic analysis confirmed the presence of a confluent monolayer with interendothelial junctions. Albumin permeability was measured by the clearance of 125l‐albumin across the endothelial monolayer. The control 125l‐albumin clearance was 0.273 ± 0.02 μl/min. The native enzyme, α‐thrombin (10−6 to 10−10 M), added to the luminal side of the endothelium produced concentration‐dependent increases in albumin clearance (maximum clearance of 0.586 ± 0.08 μl/min at 10−6 M). Gamma (γ) thrombin (10−6 M and 10−8 M), which lacks the fibrinogen recognition site, also produced a concentration‐dependent increase in albumin clearance similar to that observed with α‐thrombin. Moreover, the two proteolytically inactive forms of the native enzyme, i‐Pr2 P‐α‐thrombin and D‐Phe‐Pro‐Arg‐CH2‐α‐thrombin, increased the 125l‐albumin clearance (0.610 ± 0.09 μl/min and 0.609 ± 0.02 μl/min for iPr2 P‐α‐thrombin and D‐Phe‐Pro‐Arg‐CH2‐α‐thrombin at 10−6 M, respectively). Since the modified forms of thrombin lack the fibrinogen recognition and active serine protease sites, the results indicate that neither site is required for increased albumin permeability. The increase in albumin clearance with α‐thrombin was not secondary to endothelial cell lysis because lactate dehydrogenase concentration in the medium following thrombin was not significantly different from baseline values. There was also no morphological evidence of cell lysis. Moreover, the increase in 125l‐albumin clearance induced by α‐thrombin was reversible by washing thrombin from the endothelium. The basis for the increased albumin permeability following the addition of α‐thrombin appears to be a reversible change in endothelial cell shape with formation of intercellular gaps.
Receptor occupancy by a bovine sialoglycopeptide inhibitor correlates with inhibition of protein synthesis
Authors: Bascom Charles C. , Sharifi Behrooz G. , Johnson Terry C. 
Published: February 2005

AbstractWe have isolated from bovine cerebral cortex cells and purified to homogeneity an 18,000 dalton, pl 3.0 sialoglycopeptide that inhibits protein synthesis and DNA synthesis of nontransformed but not transformed cells without affecting uptake of radiolabeled precursors. In this paper, we examine the relationship between the binding of the sialoglycopeptide inhibitor to 3T3 cells and inhibition of protein synthesis. Binding of the sialoglycopeptide to 3T3 cells was rapid at 37°C and reached a maximum at 30 min; the binding at 37°C was shown to be saturable and specific. Scatchard analysis of the binding indicated that 3T3 cells contained about 2 × 104 receptors/cell with a dissociation constant of 1.0–1.5 nM. Several lines of evidence indicated that receptor occupancy on 3T3 cells correlated with the protein synthesis inhibitory activity of the sialoglycopeptide. A comparison of the kinetics of inhibitor binding with the kinetics of protein synthesis inhibition demonstrated that binding directly correlated with the inhibition of protein synthesis, concentration‐dependent inhibition of protein synthesis directly correlated with concentration‐dependent receptor occupancy, and a direct correlation was also observed between the kinetics of inhibitor dissociation from its specific cell surface receptor and the kinetics of recovery from protein synthesis inhibition.
ATP‐induced cell contraction in dermal fibroblasts: Effects of cAMP and myosin light‐chain kinase
Authors: Ehrlich H. P. , Rajaratnam J. B. M. , Griswold T. R. 
Published: February 2005

AbstractWhen 1mM ATP is added to human dermal fibroblasts (DF) in monolayer culture permeabilized by glycerol, they undergo a rapid reduction in length and their intracellular actin filaments aggregate. This process is referred to as cell contraction. Treating glycerol‐permeabilized DF with alkaline phosphatase before adding 1mM ATP should cause dephosphorylation. Dephosphorylated preparations do not undergo cell contraction initiated by ATP. When myosin light‐chain kinase (MLCK) isolated from turkey gizzard is added with cofactors to cells dephosphorylated by alkaline phosphatase treatment, contraction is restored. DF incubated for 24 h with db cAMP or cholera toxin show elevated intracellular concentrations of cAMP and little cell contraction. Contraction is reestablished when MLCK with cofactors is incubated with these preparations before ATP is added. Fibroblasts from Epidermolysis Bullosa dystrophica recessive patients produce excess cAMP. Those cells show minimal contraction, however; treating them with MLCK and cofactors renews contraction brought about by ATP. When DF are incubated with trifluoperazine to block calmodulin‐dependent enzyme reactions, cell contraction is inhibited. Adding cytochalasin B disrupts microfilaments and also inhibits contraction. This work supports the idea that myosin ATPase is critical to cell contraction. Myosin ATPase is dependent on the phosphorylation of the regulatory peptide, myosin light chain. Elevating intracellular concentrations of cAMP or treatment of permeabilized cell preparations with alkaline phosphatase may inhibit myosin ATPase activity. The restoration of phosphorylation by adding MLCK with cofactors served to reestablish cell contraction.
Sensitization of rat hepatocytes to hyperthermia by calcium
Authors: Malhotra Arun , Kruuv Jack , Lepock James R. 
Published: February 2005

AbstractThe viability of isolated rat hepatocytes, as assayed by trypan blue exclusion, decreases in a dose‐dependent fashion during exposure to hyperthermia (D0 [43°C] = 105 ± 10 min, D0 [45°C] = 24 ± 4 min). Hyperthermic sensitivity varies as a function of extracellular Ca2+ concentration in a biphasic manner; optimum survival occurs at 1–5 mM Ca2+, with sensitization in the absence of Ca+ and increasing sensitization at Ca2+ concentrations greater than 10 mM. Ca influx does not correlate well with loss of viability for hepatocytes in 4 mM extracellular Ca2+; influx does not occur until viability decreases to less than 1%. Under sensitizing conditions, Ca2+ influx preceeds loss of viability. Influx begins within 15 min at 45°C in 15 mM Ca2+, and the ionophore A23187 is a potent hyperthermic sensitizer in the presence of extracellular Ca2+. Thus, Ca2+ influx, whether caused by high extracellular Ca2+ or A23187, increases cellular damage caused by supraoptimal temperatures, although some Ca2+ is necessary for maximum resistance, probably because of stabilization of Ca2+ binding proteins against thermal denaturation or possibly to Ca2+‐induced decrease in lipid fluidity.
Involvement of both heparanase and plasminogen activator in lymphoma cell‐mediated degradation of heparan sulfate in the subendothelial extracellular matrix
Authors: Bar‐Ner Matia , Mayer Michael , Schirrmacher Volker , Vlodavsky Israel 
Published: February 2005

AbstractThe effect of plasminogen on the ability of highly metastatic ESb mouse lymphoma cells to degrade heparan sulfate (HS) in the subendothelial extracellular matrix (ECM) was studied. A metabolically sulfate‐labeled ECM was incubated with the lymphoma cells, and labeled degradation products were analyzed by gel filtration on Sepharose 6B. Heparanase‐mediated release of low‐Mr (0.5 < Kav < 0.85) HS cleavage products was stimulated fourfold in the presence of plasminogen. Incubation of plasminogen alone with the ECM resulted in its conversion into plasmin, which released high‐Mr (Kav < 0.33) labeled proteoglycans from the ECM. Heating the ECM (80°C, 1 hr) abolished its ability to convert plasminogen into plasmin, yet plasminogen stimulated, through its activation by the ESb plasminogen activator, heparanase‐mediated release of low‐Mr HS fragments. Heparin inhibited both the basal and plasminogen‐stimulated degradation of HS side chains but not the total amount of labeled material released from the ECM. In contrast, aprotinin inhibited the plasminogen‐stimulated release of high‐ as well as low‐Mr material. In the absence of plasminogen, degradation of heated ECM by ESb cells was completely inhibited by aprotinin, but there was only a partial inhibition of the degradation of native ECM and no effect on the degradation of soluble HS proteoglycan. These results demonstrate that proteolytic activity and heparanase participate synergistically in the sequential degradation of ECM HS and that the ESb proteolytic activity is crucial for this degradation when the ECM‐associated protease is inactivated. Plasminogen may serve as a source for the proteolytic activity that produces a more accessible substrate to the heparanse.