bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mechanisms of enhanced angiotensin II–stimulated signal transduction in vascular smooth muscle by aldosterone
Authors: Ullian Michael E. , Fine Jana J. 
Published: February 2005

AbstractWe tested the hypothesis that mineralocorticoids potentiate angiotensin II–stimulated phospholipase C activation through an increased number of angiotensin II receptors in cultured rat aortic vascular smooth muscle cells. Exposure of cells to aldosterone for 24 h resulted in concentration‐dependent increases in angiotensin II receptor binding. Via studies of angiotensin II displacement by non‐peptide receptor antagonists, both basal and upregulated angiotensin II receptors were found to be of the AT1, subtype. Incubation with 1 μM aldosterone resulted in 50%–100% enhancement of angiotensin II (100 nM)–stimulated diacylglycerol formation and intracellular calcium mobilization. Exposure to 100 nM 1,25‐(OH)2 VitD3, which did not upregulate angiotensin II receptors, did not potentiate stimulated inositol phosphate formation. Incubation with aldosterone resulted in potentiation of inositol phosphate formation upon receptor occupation (100 nM angiotensin II) but not upon post‐receptor stimulation (25 mM NaF/10 μM AlCl3). Aldosterone did not increase basal phospholipase C activity or content of the inositol trisphosphate precursor phosphatidylinositol‐4,5‐bisphosphate. These data are consistent with the hypothesis that aldosterone potentiates angiotensin II–stimulated, phospholipase C‐dependent intracellular signals solely by coupling to an increased number of angiotensin II receptors. This mechanism may contribute to the sensitized vascular responses to angiotensin II observed in states of mineralocorticoid excess. © 1994 Wiley‐Liss, Inc.
High resolution analysis of cell cycle‐correlated vimentin expression in asynchronously grown, TPA‐treated MPC‐11 cells by the novel flow cytometric multiparameter BrdU‐hoechst/PI and immunolabeling technique
Authors: Giese Günter , Kubbies Manfred , Traub Peter 
Published: February 2005

AbstractHigh resolution, multiparameter analysis using the flow cytometric BrdU/Hoechst quenching technique has been applied to study cell cycle kinetics and vimentin expression in individual cells of asynchronously grown MPC‐11 mouse plasmacytoma cell cultures treated with 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) to induce in vitro differentiation. BrdU treatment up to 16 h in the absence or presence of TPA did not affect either cell cycle progression or the kinetics or quantity of vimentin expression. TPA‐treated cells became arrested in G1 phase of the second cell cycle; however, this G1 phase arrest was transient only. In addition, G1 phase cells located prior to a putative transition point at the beginning of TPA treatment were completely blocked in cell cycle progression. There is also evidence that cells located in G1 or G2/M phase at the beginning of TPA treatment finally expressed low levels of vimentin. On the contrary, cells located in S phase at TPA exposure showed high vimentin levels after treatment. The results presented here show that, with the flow cytometric BrdU/Hoechst quenching technique, one can correlate time‐dependent protein expression at the single cell level in asynchronously grown cultures not only with the actual cell cycle state, but also with the history of cell replication. © 1994 Wiley‐Liss, Inc.
Redistribution of plasma membrane proteins by electroosmosis elicits cytosolic calcium response in tumor mast cells
Authors: Feder Toni J. , Webb Watt W. 
Published: February 2005

AbstractActivation of mast cells and basophils by binding of ligands that crosslink and micro‐aggregate cell surface receptors leads to a series of responses including a phosphoinositide cascade, elevation of intracellular free calcium ([Ca2+]i), morphological changes in the cell plasma membrane, and ultimately, exocytosis of granules containing histamine and other mediators of the allergic response. In rat basophilic leukemia (RBL) cells, a tumor mast cell line, stimulation by immunoglobulin E receptor crosslinking induces these responses. In order to determine whether redistribution or aggregation of cell surface proteins is sufficient to induce a response in these cells without extrinsic crosslinking, we have redistributed cell surface proteins by electroosmotic segregation and looked for second messenger [Ca2+]i responses. Video imaging of calcium ion activity using the fluorescent calcium sensitive dye fura‐2 revealed the effects of receptor motion and aggregation induced by application of small (10 V/cm) electric fields. A synchronous, monotonic rise in [Ca2+]i generally occurs within a few minutes after a steady field has been applied, while the redistribution of surface proteins is still in progress. The oscillations in [Ca2+]i characteristic of antigen‐stimulated cells are not seen, nor are any effects observed in weak alternating fields (0.02, 60 Hz). The observed rise in [Ca2+]i induced by static electric fields is attributed to perturbation of [Ca2+]i regulation by the large‐scale redistribution of membrane constituents induced by surface electroosmosis. © 1994 Wiley‐Liss, Inc.
Influence of nidogen complexed or not with laminin on attachment, spreading, and albumin and laminin B2 mRNA levels of rat hepatocytes
Authors: Levavasseur Françoise , Mayer Ulríke , Guillouzo André , Clement Bruno 
Published: February 2005

AbstractNidogen/entactin is a Mr = 150,000 glycoprotein which is present within basement membranes in a noncovalent stable complex with laminin. We have studied the effects of nidogen/entactin complexed or not with laminin on attachment, spreading, and functions of adult rat hepatocytes in primary culture. Freshly isolated hepatocytes attached on either recombinant or EHS‐derived nidogen, although to a lesser extent than on laminin/nidogen complex, laminin, and E8 and P1 fragments of laminin. Hepatocytes bound on a nidogen fragment bearing the N‐terminal and rod‐like domains but not on either the N‐terminal globules or the rod‐like domain which contains a RGD sequence. Attachment of hepatocytes on nidogen and laminin/nidogen complex was inhibited by anti‐β 1 integrin antibodies. Hepatocytes remained rounded on nidogen and laminin, whereas they rapidly spread on laminin/nidogen complex and collagen IV. Nidogen, laminin, and laminin/nidogen complex transiently maintained high steady‐state albumin mRNA levels in cultured hepatocytes, but a decrease in albumin mRNA content was observed after 24 h, independently of the substrates. Actinomycin D and cycloheximide treatment indicated that the transient effect of these substrates on albumin expression was related to post‐transcriptional mechanisms. Laminin B2 mRNAs were not detectable in freshly isolated hepatocytes but were expressed in 4 h hepatocyte cultures. After 24 h, a dramatic increase in the steady‐state level of laminin B2 mRNA was found in hepatocytes cultured on nidogen and laminin/nidogen complex. This effect was slightly prevented in hepatocytes plated on laminin. These results show that interactions of hepatocytes with nidogen/entactin in vitro result only in a transient modulation of hepatocyte functions. © 1994 Wiley‐Liss, Inc.
Human Co115 colon carcinoma cells potentiate the degradation of laminin mediated by tissue‐type plasminogen activator
Authors: Tran‐Thang C. , Vouillamoz D. , Kruithof E. K. O. , Sordat B. 
Published: February 2005

AbstractThe plasminogen activation (PA) system of human Co 115 colon carcinoma cells was investigated. Analysis at the levels of protein and mRNA of cultured cells and of histozymography of tumor xenografts in nude mice showed that Co 115 cells produce only tissue type PA (t‐PA) and no urokinase (u‐PA). Also, mRNA for the u‐PA receptor and for PA inhibitorr type 2 (PAI‐2), but not for PAI‐I, were detected. We developed a quantitative degradation assay using glutaraldehyde‐immobilized125I‐Iaminin to investigate the capacity of Co 115 cells to degrade laminin. Laminin degradation by Co 115 cells was completely inhibited by 100 μg/ml of polyclonal anti‐t‐PA IgG, by the plasmin inhibitors aprotinin (100μg/ml) or ε‐aminocaproic acid (EACA; at 0.3 M), but not by antibodies against u‐PA or u‐PAR nor by nonimmune IgG. Cycloheximide‐treated Co 115 cells were unable to degrade laminin but increased laminin degradation induced by conditioned medium of Co 115 cells or recombinant t‐PA. No potentiation was observed when Co 115 cells and laminin were kept separated by Transwell™ inserts. Our results suggest that Co 115 human colon carcinoma cells degrade laminin by potentiating t‐PA‐mediated plasminogen activaton at the cells surface which requires close contact between tumor cells and laminin substrate. © 1994 Wiley‐Liss, Inc.
Early cellular events couple covalent binding of reactive metabolites to cell killing by nephrotoxic cysteine conjugates
Authors: Chen Qin , Jones Thomas W. , Stevens James L. 
Published: February 2005

AbstractAddition of the nephrotoxic cysteine conjugate, S‐(1, 2‐dichlorovinyl)‐L‐cysteine (DCVC), to the LLC‐PK1 line of renal epithelial cells leads to covalent binding of reactive intermediates followed by thiol depletion, lipid peroxidation, and cell death (Chen et al., 1990, J. Biol. Chem., 265:21603‐21611.) The present study was designed to determine if increased intracellular free calcium might play a role in this pathway of DCVC‐induced toxicity by comparing the temporal relationships among increased intracellular free calcium, lipid peroxidation, and cytotoxicity. Intracellular free calcium increased 1 hr after DCVC treatment, long before LDH release occurred. The elevation of intracellular free calcium and cytotoxicity was prevented by inhibiting DCVC metabolism with AOA. The cell‐permeable chelators, Quin‐2AM and EGTA‐AM, prevented the toxicity. Pretreatment of cells with a nontoxic concentration of ionomycin increased intracellular free calcium and potentiated DCVC‐induced LDH release. However, the antioxidant, DPPD, which blocks lipid peroxidation and toxicity, did not affect the increase in intracellular free calcium, whereas buffering intracellular calcium with Quin‐2AM or EGTA‐AM blocked both lipid peroxidation and toxicity without preventing the depletion of nonprotein sulfhydryls by DCVC. Ruthenium red, an inhibitor of mitochondrial calcium uptake, also blocked cell death. We hypothesize that covalent binding of the reactive fragment from DCVC metabolism leads to deregulation of intracellular calcium homeostasis and elevation of intracellular free calcium. Increased intracellular free calcium may in turn be coupled to mitochondrial damage and the accumulation of endogenous oxidants which cause lipid peroxidation and cell death. © 1994 Wiley‐Liss, Inc.
Capping and receptor‐mediated endocytosis of cell‐bound verotoxin (shiga‐like toxin) 1: Chemical identification of an amino acid in the B subunit necessary for efficient receptor glycolipid binding and cellular internalization
Authors: Khine A. A. , Lingwood C. A. 
Published: February 2005

AbstractThe glycolipid globotriaosylceramide (Gb3) is the plasma membrane receptor that mediates the internalization of verotoxin (VT1) into susceptible cells by capping and receptor‐mediated endocytosis (RME). Internalization of fluorescein isothiocyanate‐conjugated holotoxin into Daudi lymphoma cells was found to be slower than the pentameric receptor binding B subunit alone, suggesting that the A subunit may interact with the membrane to compromise the lateral mobility of the receptor bound B subunit. 3‐D reconstruction of fluorescent images by confocal microscopy confirmed the complete internalization of holotoxin. VT1 internalization and cytotoxicity was inhibited by monodansyl cadavarine, which supports a role for clathrin coated pits in the RME of VT1. Biotinylation of the B subunit (in contrast to fluorescein labelling) was found to prevent toxin internalization. This effect correlated with reduced binding of Gb3 and reduced cytotoxicity in vitro. By cleavage of the B subunit at the single tryptophan residue, the reduced Gb3 binding and lack of cellular internalization was shown to be due to the biotinylation of lysine 53 in the VT1 B subunit. This residue was not labelled with fluorescein isothiocyanate in the native protein. This conclusion was confirmed by the finding that biotinylation of VT2c (which contains lys 53) prevented glycolipid receptor binding, whereas biotinylation of VT2e (in which lys 53 is substituted by ile) had no effect. © 1994 Wiley‐Liss, Inc.
Differential activation by platelet‐derived growth factor‐BB of mitogen activated protein kinases in starved or nonstarved AKR‐2B fibroblasts
Authors: Hoppe Jürgen , Hoppe Viviane , Karenberg Thomas‐Andreas , Fenn Anke , Simm Andreas , Sachinidis Agapios 
Published: February 2005

AbstractMore than 90% of serum‐deprived (starved) AKR‐2B mouse fibroblasts are stimulated to divide by the addition of platelet‐derived growth factor (PDGF)‐BB. In density‐arrested (nonstarved) cells, PDGF‐BB affords protection from cell death without stimulation of cell division. In both cultivation conditions the cells express similar amounts of PDGF β‐receptors and the receptor kinase activity was identical as judged by its autophosphorylation capacity. Three signaling pathways were studied in detail: (1) Phospholipase C‐γ (PLC‐γ) and [Ca2+]i increase, (2) activation of the phosphatidylinositol‐3 kinase (PI‐3 kinase), and (3) activation of mitogen activated kinases I and II (MAP kinases I and II). There was no difference in starved or nonstarved cells regarding PLC‐γ activation, increase of [Ca2+]i, and stimulation of PL‐3 kinase activity. But most remarkably the activation of MAP‐I was largely suppressed in nonstarved cells. The implications of these signaling pathways in cell protection or cell division are discussed. © 1994 Wiley‐Liss, Inc.
Second messenger signaling of c‐fos gene induction by parathyroid hormone (PTH) and PTH‐related peptide in osteoblastic osteosarcoma cells: Its role in osteoblast proliferation and osteoclast‐like cell formation
Authors: Kano Junichi , Sugimoto Toshitsugu , Kanatani Masanori , Kuroki Yasuo , Tsukamoto Tatsuo , Fukase Masaaki , Chihara Kazuo 
Published: February 2005

AbstractThe present study was performed to clarify second messenger signaling in parathyroid hormone (PTH)‐induced c‐fos gene expression, to characterize the participation of the c‐fos gene in the regulation of osteoblast proliferation and function as well as osteoclast‐like cell formation by PTH and to compare these effects of PTH with those of PTH‐related peptide (PTHrP). Both human (h) PTH‐(1‐34) and hPTHrP‐(1‐34) at 10‐8 M induced a transient c‐fos gene expression to a similar degree in osteoblastic osteosarcoma cells, UMR‐106. N6, O2' ‐dibutyryl adenosine 3′, 5′‐cyclic monophosphate (dbcAMP) as well as Sp‐diastereoisomer of adenosine cyclic 3′,5′‐phosphorothioate (Sp‐cAMPS), an activator of cAMP‐dependent protein kinase (PKA), induced a weak c‐fos gene expression. Although Rp‐diastereoisomer of adenosine cyclic 3′,5′‐phosphorothioate (Rp‐cAMPS), an inhibitor of PKA, almost completely antagonized dbcAMP‐and Sp‐cAMPS‐induced expression of c‐fos gene, it did not cause an obvious inhibition of PTH‐or PTHrP‐induced expression. Phorbol 12‐myristate 13‐acetate (PMA), an activator of protein kinase C (PKC), induced an intense expression of the c‐fos gene, while 4α‐phorbol 12, 13‐didecanoate (4αPDD), incapable of activating PKC, and calcium ionophores (A23187 and ionomycin) did not. Protein kinase C inhibitor (H‐7, 50 μM) completely blocked the expression of the c‐fos gene by PTH as well as by PTHrP. Antisense oligodeoxynucleotides (as‐ODN) complementary to c‐fos mRNA, which have been shown to inhibit its mRNA translation, at 1 μM significantly antagonized PTH‐and PTHrP‐induced inhibition of [3H] thymidine incorporation and stimulation of osteoclast‐like cell formation in the presence of osteoblasts, but not an increase in alkaline phosphatase activity, compared to control oligodeoxynucleotides with same nucleotides as as‐ODN but with a random sequence. The present study indicates the involvement of PKC system in c‐fos gene expression by PTH as well as PTHrP and also indicates the involvement of the c‐fos gene in the regulation of bone cell physiology by PTH and PTHrP. © 1994 Wiley‐Liss, Inc.
Adaptation of EGF receptor signal transduction to three‐dimensional culture conditions: Changes in surface receptor expression and protein tyrosine phosphorylation
Authors: Mansbridge J. N. , Ausserer W. A. , Knapp M. A. , Sutherland R. M. 
Published: February 2005

AbstractA431 cells grown as three‐dimensional spheroids show growth stimulation in response to nanomolar concentrations of EGF in contrast to monolayer cultures that show inhibition. In investigating the alterations in EGF signal transduction that underlie this modification of the proliferative response, we have compared the expression of EGF receptors on A431 cells under these conditions and related our findings to tyrosine phosphorylation and the growth response. EGF receptors were measured by 125I‐EGF binding to trypsin‐dispersed cells. Unexpectedly, dispersion of the monolayers caused an 80% decrease in surface EGF receptor, although, after dispersion, EGF receptor was digested by trypsin with a half‐life of 69 ± 32 min. No evidence for a comparable loss of cellular EGF receptor was seen on trypsin dispersion of spheroids. After allowing for this effect, we found that the receptor density on nondispersed monolayers (5 × 106 per cell) was twentyfold greater than that on spheroids (0.25 × 106 per cell). EGF‐induced tyrosine phosphorylation was confined to the outermost cells of the spheroid, although the presence of surface‐expressed EGF pinding sites could be demonstrated throughout the structure and the number of EGF receptors/cell on dispersed spheroid cells showed a single distribution peak by flow cytometry, with no evidence for more than one population. Using RCM‐lysozyme as a substrate, tyrosine phosphatase activity in spheroids lay within the range observed in monolayer cultures. Autophosphorylation of the EGF receptor following EGF stimulation in monolayer cultures of A431 cells rose rapidly in the first 10 seconds and then slowly increased for at least 3 h. In spheroids, it reached a maximum within 10 seconds and then declined over 3 h. Since the microenvironment within a tumor resembles that in a spheroid, a similar reduction in surface EGF receptor expression may be expected in tumors relative to monolayer cultures, together with corresponding growth stimulation in response to EGF. © 1994 Wiley‐Liss, Inc.
Effects of P‐glycoprotein expression on cyclic AMP and volume‐activated ion fluxes and conductances in HT‐29 colon adenocarcinoma cells
Authors: Kunzelmann K. , Slotki I. N. , Klein P. , Koslowsky T. , Ausiello D. A. , Greger R. , Cabantchik Z. I. 
Published: February 2005

AbstractThe tissue distribution of P‐glycoprotein (Pgp) and the structurally related cystic fibrosis transmembrane conductance regulator (CFTR) is apparently mutually exclusive, particularly in epithelia; where one protein is expressed the other is not. To study the possible function(s) of Pgp and its potential effects on CFTR expression in epithelia, HT‐29 colon adenocarcinoma cells, which constitutively express CFTR, were pharmacologically adapted to express the classical multidrug resistance (MDR) phenotype (Pgp+). Concomitant with the appearance of Pgp and MDR phenotype (drug resistance, reduced drug accumulation and increased drug efflux), CFTR levels and cAMP‐stimulated Cl conductances were markedly decreased compared to wild‐type HT‐29 (Pgp−) cells (as shown using the whole cell patch clamp technique). Removal of drug pressure led to the gradual decrease in Pgp levels and MDR phenotype, as evidenced by increased rhodamine 123 accumulation (Pgp‐Rev). Concomitantly, CFTR levels and cAMP‐stimulated Cl− conductances incresed. The cell responses of Pgp/Rev cells were heterogeneous with respect to both Pgp and CFTR functions. We also studied the possible contribution of Pgp to hypotonically activated (HCS) ion conductances. K+ and Cl− effluxes from Pgp− cells were markedly increased by HCS. This increase was twice as high as that induced by the cation ionophore gramicidin; it was blocked by the Cl− channel blocker DIDS (4,4′‐disothiocyano‐2,2′‐disulfonic stilbene) and required extracellular Ca2+. In Pgp+ cells, the HCS‐induced fluxes were not significantly different from those of Pgp− cells. Verapamil (10 μM), which caused 80% reversal of Pgp‐associated drug extrusion, failed to inhibit the HCS‐evoked Cl− efflux of Pgp+ cells. Similarly, HCS increased Cl− conductance to the same extent in Pgp−, Pgp+ and Pgp‐Rev cells. Verapamil (100 μM), but not 1,9‐dideoxyforskolin (50 and 100 μM), partially inhibited the HCS‐evoked whole cell current (WCC) in all three lines. Since the inhibition by verapamil was not detected in the presence of the K+ channel blocker Ba2+ (3 mM), it is suggested that verapamil affects K+ and not Cl− conductance. We conclude that hypotonically activated Cl− and K+ conductances are similar in HT‐29 cells irrespective of Pgp expression. Expression of high levels of Pgp in HT‐29 cells confers no physiologically significant capacity for cell volume regulation. © 1994 Wiley‐Liss, Inc.
Enhanced expression of granulosa cell low density lipoprotein receptor activity in response to in vitro culture conditions
Authors: Reaven Eve , Tsai Louisa , Spicher Marjorie , Shilo Lotan , Philip Mohan , Cooper Allen D. , Azhar Salman 
Published: February 2005

AbstractPrevious studies have shown that the B/E (low density lipoprotein [LDL]) receptor pathway plays a minor role in cholesterol uptake in the intact rat ovary, but when granulosa cells are isolated and maintained in culture, the cells develop a fully functional B/E receptor system. In the current study we examined the development of the B/E receptor over time (96 h) in culture and compared its physiological function, expression of mRNA and protein levels, and morphological events to the upregulation induced in 24 h by hormone (human chorionic gonadotropin [hCG] or Bt2cAMP). With both protocols, increased progestin production occurs and is associated with elevated binding, uptake, and degradation of LDL in the medium although the impact of Bt2cAMP stimulation on all these measurements is several times that observed with time alone. Only the hormone‐stimulated LDL receptor response was associated with an increase in receptor protein (Western blot) or mRNA levels (RNase protection assay). We conclude that unstimulated granulosa cells show posttranslational increase in B/E receptor activity with time in culture, but transcriptional changes in B/E receptor follow stimulation with trophic hormone or its second messenger, cAMP. © 1994 Wiley‐Liss, Inc.