bluuebunny/crossref_metadata_2025_split huggingface dataset:
Reconstituted basement membrane influences prolactin, LH, and FSH secretion from adult and fetal adenohypophyseal cells in vitro
Authors: Horacek Mark J. , Dada M. Olubunmi , Terracio Louis 
Published: February 2005

AbstractThese investigations tested the hypothesis that secretion of prolactin (PRL), LH, and FSH in vitro is influenced by the substratum on which adult or fetal adenohypophyseal cells are cultured. Adenohypophyses were removed from adult male Golden Syrian hamsters and from fetal hamsters on day 16 of gestation. The glands were dissociated and cultured in a 1:1 mixture of Dulbecco's Modified Eagle's Medium (DMEM) and Ham's F‐12 medium containing 10% fetal bovine serum (FBS), 25 mM Hepes, and antibiotics. The cells were cultured on three substrata: glass, laminin, and the reconstituted basement membrane of the Engelbreth‐Holm‐Swarm (EHS) tumor (Matrigel). Medium was collected and replaced every 48 h for 14–22 days. Concentrations of PRL, LH, and FSH in medium were measured by RIA. The substratum influenced hormone secretion. PRL concentrations were elevated in cultures of adult cells on Matrigel in each of four experiments. Adenohypophyseal cells on Matrigel maintained a rounded shape longer than cells on glass or laminin. In studies using fetal adenohypophyseal cells, PRL concentrations were elevated significantly in medium from cultures on Matrigel at and after 2 days as were concentrations of LH and FSH after 6 days. Additional experiments showed that the higher PRL concentrations in medium surrounding adult cells plated on Matrigel were not due to the release of soluble factors from Matrigel, differential cell attachment on Matrigel, the differential presence of adenohypophyseal fibroblasts, nor differential rates of cell proliferation. The results show that Matrigel maintains the secretion of PRL from adult adenohypophyseal cells in vitro more effectively than glass or laminin substrata and support the hypothesis that cell‐matrix interactions mediate the observed differences. The results also show that in long‐term cultures (14–22 days), fetal adenohypophyseal cells secrete significantly more PRL, LH, and FSH on Matrigel than they secrete when cultured on glass or laminin. Thus, Matrigel influences the function and possibly the maturation of adenohypophyseal cells in vitro. Furthermore, although laminin is the most abundant component in Matrigel, the effects of Matrigel on lactotrophs and gonadotrophs in vitro are probably not attributable solely to its laminin content. © 1992 Wiley‐Liss, Inc.
Renewed DNA synthesis in senescent WI‐38 cells by expression of an inducible chimeric c‐fos construct
Authors: Phillips Paul D. , Pignolo Robert J. , Nishikura Kazuko , Cristofalo Vincent J. 
Published: February 2005

AbstractAs normal human cells approach the end of their proliferative lifespan in vitro they lose responsiveness to a variety of growth factors, to which they respond with DNA replication when they are young. Recently it has been reported that the protooncogene c‐fos is not expressed in senescent cells (Seshadri and Campisi, 1990). In this study we have found that both c‐jun and jun B, partners of c‐fos in heterodimeric transactivating complexes, are equivalently expressed in young and senescent cells at both early (1–6 hr) and late (12 or 16 hr) time points following serum stimulation of quiescent cells. We have also investigated the effect of the enforced expression of c‐fos in senescent WI‐38 cells using an inducible construct carrying the murine c‐fos gene under the control of the sheep metallothionein promoter. We have found that the transient transfection and subsequent activation of the conditional promoter with Zn++ stimulated DNA synthesis in a significant fraction of senescent cells which had completed 90%–95% of their proliferative lifespan. However, populations which had completed 100% of their proliferative life span and nondividing cultures which had been selected with BrdU did not respond to the expression of the c‐fos gene. These results demonstrate that one of the primary events associated with senescence in human cells is the suppression of c‐fos gene expression, but additional phenotypic changes must also occur in order to explain the ultimate loss of proliferative responsiveness of these cells. © 1992 Wiley‐Liss, Inc.
Control of alkaline phosphatase activity in C3H10T1/2 cells: Role of retinoic acid and cell density
Authors: Reese David H. , Larsen Rachel A. , Hornicek Francis J. 
Published: February 2005

AbstractThe enzyme alkaline phosphatase (AP) has been shown to be lost or inappropriately expressed during carcinogenesis in some tissues. Because retinoic acid (RA) appears to play a role in the normal regulation of the enzyme (RA up‐regulates AP in a variety of cell types) we have suggested that altered AP expression in some cancers may be caused by a defect in the ability of the cells to respond normally to retinoid. We have begun to use the chemically transformable mouse embryo fibroblast cell, C3H10T1/2, to investigate this possibility. In this initial study we characterized AP regulation in normal C3H10T1/2 cells and show that: (1) 10−7 M RA increases AP activity within 3–4 h in serum‐free medium; (2) serum inhibits short‐term induction (0–8 h) in a concentration‐dependent manner (10% serum causes complete inhibition); (3) during long‐term RA exposure (24 h and 48 h), induction can be detected in serum‐containing medium; (4) AP induction is dose related at RA concentrations from 10−10 M to 10−6 M in serum‐free medium; (5) 10−5 M RA is ineffective at inducing AP in serum‐free medium during 8 h but is the most effective concentration in serum‐containing medium during 24 h and 48 h exposures; (6) AP inducibility by RA requires near‐confluent cell densities; and (7) when cultures become confluent, cells become constitutive for AP and no longer require RA for enzyme expression. The effects of serum and cell density on AP inducibility by RA and implications of the RA up‐regulation of AP for teratogenesis are discussed. © 1992 Wiley‐Liss, Inc.
The inward rectifier potassium conductance in rat basophilic leukemia cells
Authors: Piguet Pascale , North R. A. 
Published: February 2005

AbstractWhole‐cell recordings were made from rat basophilic leukemia cells. The properties of the cells in the potential range negative to −20 mV could be accounted for by two potassium conductances, a leakage conductance (0.54 ± 0.01 nS, mean ± s.e.m., n − 6) and an inward rectifier conductance. The inward rectifier conductance activated at a potential close to Ek and had a maximal value of 2.7 ± 0.22 nS (n = 6 ). Cesium (2 mM, n = 6) decreased the inward current at every potential. Strontium (10 mM, n = 7) and rubidium (2 mM, n = 3) had a similar effect, but they also increased the slope conductance between −63 and − 103 mV. Barium (1–100 pM) blocked selectively the inward rectifying current without affecting the leakage current; the effect was use‐, voltage‐, and temperature‐dependent. TEA decreased the current; the concentration giving 50% inhibition was 37.5 mM. The current was unaffected by phorbol esters as well as several hormones and transmitters that werg tested. © 1992 Wiley‐Liss, Inc.
Relationship between thermal tolerance and protein degradation in temperature‐sensitive mouse cells
Authors: Li Jian Jian , Dewey William C. 
Published: February 2005

AbstractThe induction of thermotolerance was studied in a temperature sensitive mouse cell line, ts85, and results were compared with those for the wild‐type FM3A cells. At the nonpermissive temperature of 39°C, ts85 cells are defective in the degradation of short‐lived abnormal proteins, apparently because of loss of activity of a ubiquitin‐activating enzyme. The failure of the ts85 cells to develop thermotolerance to 41–43°C after incubation at the nonpermissive temperature of 39°C correlated with the failure of the cells to degrade short‐lived abnormal proteins at 39°C. However, the failure of the ts85 cells to develop thermotolerance to 43°C during incubation at 33°C after either arsenite treatment or heating at 45.5°C for 6 or 10 min did not correlate with protein degradation rates. Although the rate of degrading abnormal protein was reduced after heating at 45.5°C for 10 min, the rates were normal after arsenite treatment or heating at 45.5°C for 6 min. In addition, when protein synthesis was inhibited with cyclohexmide both during incubation at 33°C or 39°C and during heating at 41–43°C, resistance to heating was observed, but protein degradation rates at 39°C or 43°C were not altered by the cycloheximide treatment. Therefore, there is apparently no consistent relationship between rates of degrading abnormal proteins and the ability of cells to develop thermotolerance and resistance to heating in the presence of cycloheximide. © 1992 Wiley‐Liss, Inc.
Induction of differentiation and c‐jun expression in human leukemic cells by okadaic acid, an inhibitor of protein phosphatases
Authors: Adunyah Samuel E. , Unlap Tino M. , Franklin Christopher C. , Kraft Andrew S. 
Published: February 2005

AbstractOkadaic acid, a protein phosphatase inhibitor, is a strong tumor promoter which activates protein phosphorylation. Because another activator of protein phosphorylation, phorbol esters, stimulates hematopoietic differentiation, we sought to determine whether okadaic acid could also induce the differentiation of the human leukemic cell line U937. Differentiation was assessed by measuring changes in the following: mRNA levels, cell growth, morphology, cell surface markers, and the ability to induce superoxide. We found that okadaic acid treatment of U937 cells induces immediate increases in total cellular levels of both c‐junand c‐fos mRNAs. Nuclear run‐on experiments demonstrate that initial increases are secondary to increases in transcription, whereas latter changes may be secondary to mRNA stabilization. Like phorbol esters, okadaic acid treatment also activates AP‐1 enhancer activity and induces the phosphorylation of c‐Jun protein. Approximately 6–12 hours after treatment with okadaic acid, mRNA levels of c‐myc, p34cdc2, and p58GTA, two cell cycle regulated protein kinases, decrease. Okadaic acid inhibits the growth of U937 cells, induces changes in nuclear morphology, stimulates increases in Mac‐1 and Leu 11 surface antigens, and induces these cells to produce superoxide. These changes, taken together, suggest that U937 cells have been induced by okadaic acid to differentiate towards a more mature cell type. © 1992 Wiley‐Liss, Inc.
Stimulation of endothelial cell proliferation by vanadate is specific for microvascular endothelial cells
Authors: Maher Pamela A. 
Published: February 2005

AbstractMicromolar concentrations of sodium orthovanadate stimulated the proliferation of bovine capillary endothelial cells, but not bovine aortic endothelial cells. Vanadate was equally potent at inducing protein tyrosine phosphorylation and changes in morphology in both types of cells. However, vanadate treatment lead to an inhibition of protein tyrosine kinase activity in the aortic endothelial cells, but not the capillary endothelial cells. In capillary endothelial cells, the effect of vanadate was additive with basic FGF (bFGF) at low concentrations of bFGF. There was no interaction between bFGF and vanadate in aortic endothelial cells. TGF‐β, which inhibits the induction of endothelial cell proliferation by bFGF, appeared to shift the dose response curve to vanadate in capillary endothelial cells, increasing the proliferative effect of vanadate at low vanadate concentrations, but decreasing the proliferative effect at higher vanadate concentrations. © 1992 Wiley‐Liss, Inc.
Adenosine receptors, cyclic AMP accumulation, and DNA‐synthesis in aortic smooth muscle cell cultures of adult and neonatal rats
Authors: Querol‐Ferrer Valérie , Hultgardh‐Nilsson Anna , Ringertz Nils R. , Nilsson Jan , Jonzon Bror 
Published: February 2005

AbstractThe effects of two adenosine analogs on cyclic AMP (cAMP) accumulation and DNA synthesis were studied in cultured smooth muscle cells (SMCs) isolated from adult and neonatal rat arteries. N‐ethylcarboxamido adenosine (NECA) dosedependently increased intracellular cAMP levels and appeared to be more potent in adult than in neonatal SMCs. R‐phenylisopropyl adenosine (R‐PIA), in nanomolar concentrations, counteracted the increase in cAMP evoked by 10 μM forskolin in adult but not in neonatal SMCs, indicating that the enhanced “A2” response seen in adult SMCs was not due to a lack of “A1” receptors in these cultures. Binding experiments performed using the adenosine antagonist XAC did not reveal any differences in the number or affinity of the adenosine receptors between neonatal and adult SMCs. This indicates effects presumably on the G‐protein level. A high capacity to spontaneously synthesize DNA and a weak response to platelet‐derived growth factor (PDGF) were seen in the neonatal SMCs. Furthermore, NECA had no effect on PDGF‐induced DNA synthesis in these cells. In contrast, adelt SMCs presented a low rate of spontaneous DNA synthesis and a marked proliferative response to PDGF, which was inhibited by NECA. This inhibition paralleled the increase in cAMP elicited by NECA. Our findings suggest that neonatal and adult SMCs differ both in their response to growth factors and growth inhibitors. © 1992 Wiley‐Liss, Inc.
Quick and accurate method to convert BCECF fluorescence to pHi: Calibration in three different types of cell preparations
Authors: James‐Kracke Marilyn R. 
Published: February 2005

AbstractA rapid, easy, and accurate method for converting the fluorescence of BCECF to pH, as an alternative to the nigericin method, is described. The ratio of the fluorescence intensities for BCECF can be converted to pH between 4 and 9 by a formula similar to the one used to calculate [Ca2+]i from the fluorescence of fura2. The formula is inverted because H+ binding to BCECF causes a decrease in fluorescence, whereas Ca2+ binding to fura2 causes an increase in fluorescence. The ratio of the fluorescence intensities is a sigmoidal function of the [H+] between pH 4 and 9 with an essentially linear mid region from pH 6 to 8. This calibration procedure in cells is similar to the popular method for fura2 where ionomycin, Ca2+, and an alkaline EGTA solution are added in succession to change the intracellular pCa from 4 to 9. For BCECF in cells, a protonophore, FCCP or CCCP, is added and the cells are titrated with acid to an intracellular pH of 4 and then back to pH 9 with base by observing the gradual change in fluorescence as it asymptotically reaches its limiting minimum and maximum values. This method does not require changing the medium to one with high KCl to depolarize the membrane potential nor does the proton concentration need to be equilibrated across the plasma membrane. The technique can be used to calibrate BCECF in sheets of cells, as well as suspensions of cells over a wide range of pH sensitivities. © 1992 Wiley‐Liss, Inc.
Biochemical events accompanying macrophage activation and the inhibition of colony‐stimulating factor‐1‐induced macrophage proliferation by tumor necrosis factor‐α, interferon‐γ, and lipopolysaccharide
Authors: Vairo Gino , Royston A. Keith , Hamilton John A. 
Published: February 2005

AbstractAgents that can arrest cellular proliferation are now providing insights into mechanisms of growth factor action and how this action may be controlled. It is shown here that the macrophage activating agents tumor necrosis factor‐α (TNFα), interferon‐γ (IFNγ), and lipopolysaccharide (LPS) can maximally inhibit colony stimulating factor‐1 (CSF‐1)‐induced, murine bone marrow‐derived macroohage (BMM) DNA synthesis even when added 8–12 h after the growth factor, a period coinciding with the G1/S‐phase border of the BMM cell cycle. This inhibition was independent of autocrine PGE2 production or increased cAMP levels. In order to compare the mode of action of these agents, their effects on a number of other BMM responses in the absence or presence of CSF‐1 were examined. All three agents stimulated BMM protein synthesis; TNFα and LPS, but not IFNγ, stimulated BMM Na+/H+ exchange and Na+, K+‐ATPase activities, as well as c‐fos mRNA levels. IFNγ did not inhibit the CSF‐1‐induced Na+, K+‐ATPase activity. TNFα and LPS inhibited both CSF‐1‐stimulated urokinase‐type plasminogen activator (u‐PA) mRNA levels and u‐PA activity in BMM, whereas IFNγ lowered only the u‐PA activity. In contrast, LPS and IFNγ, but not TNFα, inhibited CSF‐1‐induced BMM c‐myc mRNA levels, the lack of effect of TNFα dissociating the inhibition of DNA synthesis and decreased c‐myc mRNA expression for this cytokine. These results indicate that certain biochemical responses are common to both growth factors and inhibitors of BMM DNA synthesis and that TNFα, IFNγ, and LPS, even though they all have a common action in suppressing DNA synthesis, activate multiple signaling pathways in BMM, only some of which overlap or converge. © 1992 Wiley‐Liss, Inc.
Demonstration of an insulin‐insensitive storage pool of glucose transporters in rat hepatocytes and HepG2 cells
Authors: Hah Jongsik , Jo Inho , Chakrabarti R. , Jung Chan Y. 
Published: February 2005

AbstractThe subcellular distribution of glucose transporters in rat hepatocytes and HepG2 cells was studied in the absence and in the presence of insulin. Glucose transporters were quantitated by measuring glucose‐sensitive cytochalasin B binding and by protein immunoblotting using isoform‐specific antibodies. Plasma membrane contamination into subcellular fractions was assessed by measuring distribution of 5′‐nucleotidase and cell surface carbohydrate label. In hepatocytes, GLUT‐2 occurred in a low‐density microsomal (LDM) fraction at a significant concentration, and as much as 15% of cellular GLUT‐2 was found intracellularly that cannot be accounted for by plasma membrane contamination. In HepG2 cells which express GLUT‐1 and GLUT‐2, the two isoforms showed distinct subcellular distribution patterns: GLUT‐2 was highly concentrated in LDM while very little GLUT‐1 was found in this fraction, indicating that a large portion of GLUT‐2 occurs in intracellular organelles. Insulin treatment did not change the subcellular distribution patterns of glucose transporters in both cell types. Our results suggest that rat hepatocytes and HepG2 cells possess an intracellular storage pool for GLUT‐2, but lack the insulin‐responsive glucose transporter translocation mechanism. © 1992 Wiley‐Liss, Inc.
Rapid inverse changes in α1B‐and β2‐adrenergic receptors and gene transcripts in acutely isolated rat liver cells
Authors: Ishac Edward J. N. , Lazar‐Wesley Eliane , Kunos George 
Published: February 2005

AbstractIn vitro incubation of hepatocytes acutely isolated from adult male rats leads to a rapid conversion of the adrenergic activation of glycogenolysis from an α1‐receptor (α1AR) to a β2‐receptor (β2AR) mediated response within 4 h. In order to understand the underlying mechanism, we examined time‐dependent changes in α1‐ and β2‐adrenergic activation of glycogenolysis and second messenger systems, the cellular density and affinity of α1AR and β2AR, and the steady state levels of α1BAR and β2AR mRNAs. Incubation of hepatocytes for 4 h resulted in a decrease in phosphorylase activation and inositol 1,4,5 trisphosphate accumulation in response to phenylephrine, a 40% decrease in α1AR density, and a 70% decrease in α1BAR mRNA levels. Incubation of hepatocytes for 4 h also resulted in the emergence of a phosphorylase response to isoproterenol, an increase in isoproterenol‐induced but not in glucagon‐ or forskolin‐induced cAMP accumulation, no significant change in β2AR density, and a twofold increase in β2AR mRNA levels. Exposure of cells to cycloheximide, 2 μM throughout the 4 h incubation, prevented the emergence of the phosphorylase response to isoproterenol and reduced β2AR densities, while the decrease in α1AR density was not affected and the decrease in phosphorylase activation by phenylephrine was attenuated. The results indicate that dissociation of rat liver cells triggers a rapidly developing decrease in α1BAR mRNA and increase in β2AR mRNA levels and corresponding inverse changes in the synthesis of α1BAR and β2AR which account, at least in part, for the rapid conversion from α1‐ to β2‐adrenergic glycogenolysis. © 1992 Wiley‐Liss, Inc.
Reduced mRNA levels for the multidrug‐resistance genes in cAMP‐dependent protein kinase mutant cell lines
Authors: Chin Khew‐Voon , Chauhan Shyam S. , Abraham Irene , Sampson Kathleen E. , Krolczyk Arkadiusz J. , Wong Margaret , Schimmer Bernard , Pastan Ira , Gottesman Michael M. 
Published: February 2005

AbstractWe have previously shown that in Chinese hamster ovary (CHO) cells, a mutant cell line with a defective regulatory subunit (RI) for the cAMP‐dependent protein kinase (Abraham et al: Mol. Cell. Biol., 7:3098–3106, 1987), and a transfectant cell line expressing the same mutant kinase, showed increased sensitivity to a number of drugs that are known to be substrates for the multidrug transporter (P‐glycoprotein). In the current study we have investigated the mechanism by which cAMP‐dependent protein kinase controls drug resistance. We report here that the sensitivity of the kinase defective CHO cell lines to multiple drugs results from decreased RNA levels for the multidrug‐resistance gene. Similar results were obtained with mouse Y1 adrenal cells. Wild‐type Y1 cells had high levels of P‐glycoprotein due to expression of both the mdr 1b and mdr2 genes, whereas the cAMP‐dependent protein kinase mutant Kin 8 cells had decreased RNA levels for these genes. A Kin 8 transfectant with restored cAMP‐dependent protein kinase activity recovered mdr expression, indicating a cause and effect relationship between the protein kinase mutations and mdr expression. No changes in nuclear run‐off assays could be detected, suggesting a non‐transcriptional mechanism of regulation. Wild‐type Y1 cells are more drug sensitive despite having higher levels of P‐glycoprotein than the mutant cells. This paradoxical result may be explained by the higher rate of synthesis of steroids by the wild‐type Y1 cells, which appear to be inhibitors of P‐glycoprotein transport activity. © 1992 Wiley‐Liss, Inc.