bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differential regulation of cerebroside sulfotransferase and 2′,3′‐cyclic nucleotide 3′‐phosphodiesterase by basic fibroblast growth factor in relation to proliferation in rat oligodendrocyte cultures
Authors: Fressinaud Catherine , Sarliève Louis L. , Dalençon Dominique , Labourdette Gérard 
Published: February 2005

AbstractPrevious results (Fressinaud, C., Sarliève, L.L., and Labourdette, G. J. J. Cell. Physiol., 141:667–674, 1989b) have shown that cerebroside sulfotransferase (CST; EC 2.8.2.11) is enriched in pure rat oligodendrocyte (OL) cultures and that its activity is increased by factors mitogenic for OL precursors and galactocere‐broside (GC) expressing OL, such as basic fibroblast growth factor (bFGF), platelet‐derived growth factor, and high insulin concentrations. In contrast, transforming growth factor β or low insulin concentrations were found to be ineffective in this culture system. As bFGF mainly enhanced the proliferation of OL precursors (GC negative cells) rather than that of differentiated (GC +) cells, a relationship between OL precursor proliferation and CST increase was suggested. This hypothesis was first tested in 20‐day‐old OL cultures grown in chemically defined medium. The dose‐response curve of [125I] lododeoxyuridine ([125I]dUrd) incorporation toward bFGF was parallel to that of CST specific activity, and maximal stimulation was reached at 5 ng/ml bFGF for both. In contrast, 2′,3′‐cyclic nucleotide 3′‐phosphodiesterase (CNP; EC 3.1.4.37) specific activity decreased after bFGF treatment. To determine if CST increase was linked to the proliferation of OL precursors induced by bFGF, cell proliferation was blocked by cytosine arabinoside (ARA‐C). From 10−8 to 10−5M ARA‐C there was a dose‐dependent inhibition of cell proliferation and a decrease in CST specific activity, whereas CNP specific activity was enhanced. When the cells were treated with bFGF and 10−6M ARA‐C together, the proliferation was completely blocked and CST activity decreased by 72% below control values, whereas CNP activity was not significantly decreased. Immunocytochemical studies showed that the number of sulfatide‐expressing cells and the number of cycling cells were increased after bFGF treatment, but that there was no overlapping between these two populations. Taken together these results suggest that CST activity and sulfatide expression appear shortly after the arrest of OL precursor division.
Low intracellular pH induces redistribution of fodrin and instabilization of lateral walls in MDCK cells
Authors: Eskelinen Sinikka , Huotari Virva , Sormunen Raija , Palovuori Riitta , Kok Jan Willem , Lehto Veli‐Pekka 
Published: February 2005

AbstractWe have studied the effect of intracellular pH on the establishment and maintenance of the cellular polarity in MDCK cells by utilizing nigericin which causes lowering of the cytoplasmic pH. At pH below 6.5, MDCK cells lost their polarized morphology and became roundish, with an increased apical area and shortened and unstable lateral walls. The lateral wall marker proteins uvomorulin and Na, K‐ATPase remained segregated to the lateral walls in the acidified cells, as shown by immunofluorescence microscopy. Fodrin, on the other hand, was released from its normal basolateral residence and was found in the cytoplasm. Actin, which normally co‐localizes with fodrin along the basolateral walls, showed a dotty distribution in the cytoplasm of acidified cells, while stress fibers remained intact. Microtubular network appeared flattened, but the Golgi complex retained its apical position. The pH change–induced alterations were readily reversible, as the normal basal‐apical polarity (columnar shape, distinct apical and lateral domains with apposing and stiff lateral membranes) was reformed within 10 minutes after restoring the normal pH gradient across the cell membrane. This coincided with the translocation of fodrin from the cytoplasm to the lateral walls. The results show that lowering of intracellular pH leads to temporary segregation of fodrin from the other components of the membrane skeleton assembly, and that association of fodrin with the lateral walls seems to be a prerequisite for their close apposition and for the maintenance of normal basal‐axial polarity.
Degradation of collagen in the bone‐resorbing compartment underlying the osteoclast involves both cysteine‐proteinases and matrix metalloproteinases
Authors: Everts Vincent , Delaissé Jean‐Marie , Korper Wolf , Niehof Anneke , Vaes Gilbert , Beertsen Wouter 
Published: February 2005

AbstractThe site of action of cysteine‐proteinases (CPs) and matrix metalloproteinases (MMPs) in the degradation of bone collagen by osteoclasts was investigated by evaluating the effects of the CP‐inhibitor trans‐epoxy‐succinyl‐L‐leucylamido (4‐guanidino)‐butane (E‐64) and the MMP‐inhibitor N‐(3‐N‐benzyloxycarbonyl amino‐1‐R‐carboxypropyl)‐L‐leucyl‐O‐methyl‐L‐tyrosine N‐methylamide (Cl‐1) in an in vitro model system of PTH‐stimulated mouse calvaria. In the presence of each of the two inhibitors a large area of collagen free of mineral crystallites was seen adjacent to the ruffled border of the osteoclasts. Following a culture period of 24 h this area proved to be about 10 times larger in inhibitor‐treated explants than in controls. Moreover the percentage of osteoclasts in close contact with such demineralized bone areas appeared to be significantly higher in inhibitor‐treated explants than in control specimens (60% and 5%, respectively). These effects were not apparent when the osteoclastic activity was inhibited with calcitonin. No significant differences were found between the effects of the two inhibitors, E‐64 and Cl‐1. Our observations indicate that under the influence of inhibitors of MMPs and CPs demineralization of bone by osteoclasts proceeded up to a certain point whereas matrix degradation was strongly inhibited. It is concluded that within the osteoclastic resorption lacuna both CPs and MMPs participate in the degradation of the collagenous bone matrix.
Subtypes of betaglycan and of type I and type II transforming growth factor‐β (TGF‐β) receptors with different affinities for TGF‐β1 and TGF‐β2 are exhibited by human placental trophoblast cells
Authors: Mitchell E. Jane , Fitz‐Gibbon Linda , O'Connor‐McCourt Maureen D. 
Published: February 2005

AbstractTransforming growth factor‐β is likely to be an important factor controlling placental activities, including growth, differentiation, invasiveness, hormone production, and immunosuppression. We have used a chemical cross‐linking technique with either 125I‐TGF‐β1 or 125I‐TGF‐β2 and bis(sulfosuccinimidyl) suberate (BS3) to characterize TGF‐β binding components on human placental cells in primary culture. Trophoblast‐enriched primary cultures exhibited a predominant affinity‐labelled complex characteristic of membrane‐anchored betaglycan (formerly termed the Type III TGF‐β receptor) and relatively low levels of the Type I and Type II TGF‐β receptor complexes. The results from affinity labelling saturation and competition experiments with TGF‐β1 and TGF‐β2 suggest the existence of two distinct subtypes of betaglycan: one subtype has a lower capacity and higher affinity, binds both TGF‐β1 and TGF‐β2, yet has a preferential affinity for TGF‐β2; the second subtype has a higher capacity and lower affinity and binds TGF‐β1 exclusively. In contrast, mesenchymal cell‐enriched placental primary cultures possessed only one subtype of the betaglycan component that binds the two TGF‐β isoforms with similar affinities and capacities as observed on most cell lines. These experiments demonstrate that the betaglycan component which exhibits a higher affinity for TGF‐β2 than for TGF‐β1, that we had observed previously on term placental membranes, is actually present on trophoblast cells. In addition to the two distinctive betaglycan subtypes, subtypes of the Type I and II TGF‐β receptors were detected on the trophoblast‐enriched cultures. In competition experiments, when 125I‐TGF‐β1 was used as the radiotracer, the Type I and II TGF‐β receptors show a much higher affinity for TGF‐β1 than for TGF‐β2, as observed with other cell types. However, when 125I‐TGF‐β2 was used, low abundance subtypes of both the Type I and II receptors that show similar affinities for TGF‐β1 and TGF‐β2 were also revealed.
Serotonin produces a configurational change of cultured smooth muscle cells that is associated with elevation of intracellular cAMP
Authors: Lee Sheu‐Ling , Fanburg Barry L. 
Published: February 2005

AbstractEarly passaged bovine pulmonary artery smooth muscle cells (SMC) respond to serotonin (5‐HT) by developing a reversible change in configuration. (Lee et al. J. Cell. Physiol. 138:145, 1989). This configurational change does not occur in pulmonary artery endothelial cells (EC) subjected to 5‐HT and is adenosine triphosphate (ATP) dependent, lost with passage of SMC, and inhibited by various agents that block high‐affinity 5‐HT uptake. We now report a second configurational change (also dendritic formation) of SMC produced by 5‐HT only in the presence of isobutylmethylxanthine (IBMX), an inhibitor of phosphodiesterase. This configurational change was also ATP dependent, but unlike the first response, (Lee et al., 1989), it occurred in both first and later passaged SMC and was not inhibited by blockade of 5‐HT uptake. Also, unlike the response with 5‐HT alone that failed to elevate cAMP, this one was associated with a large elevation of cAMP (eight fold above control values), similar to the response to the β‐agonist isoproterenol, plus IBMX. The second response was not blocked by a variety of 5‐HT receptor antagonists but was reproduced by (±)‐8‐hydroxy‐DPAT HBr (8‐OH‐DPAT), a reputed 5‐HT1A agonist. The response was not dependent upon Ca2+ and was blocked by 1–2 mM n‐phenylanthranilic acid or anthracene‐9‐carboxylic acid, electrically conductive Cl− channel inhibitors. Hence, 5‐HT in the presence of IBMX causes a marked elevation of cAMP of SMC and this elevation in cAMP likely results in a cellular configurational change through a Cl− channel‐dependent mechanism similar to that we previously described for EC in the presence of β‐adrenergic agonist stimulation (Ueda et al. Circ. Res. 66:951, 1990). EC do not show a similar response to 5‐HT possibly because cAMP is not adequately elevated, even in the presence of IBMX, to enhance Cl− channel activity. We propose that our observations indicate the presence of two sites of action of 5‐HT on the smooth muscle cell, one intracellularly and another at a cell surface receptor.
Lipopolysaccharides decrease angiotensin converting enzyme activity expressed by cultured human endothelial cells
Authors: Watanabe Kentaro , Lam George , Keresztes Roger S. , Jaffe Eric A. 
Published: February 2005

AbstractAngiotensin converting enzyme (ACE) is present on endothelial cells and plays a role in regulating blood pressure in vivo by converting angiotensin I to angiotensin II and metabolizing bradykinin. Since ACE activity is decreased in vivo in sepsis, the ability of lipopolysaccharide (LPS) to suppress endothelial cell ACE activity was tested by culturing human umbilical vein endothelial cells (HUVEC) for 0–72 hr with or without LPS and then measuring ACE activity. ACE activity in intact HUVEC monolayers incubated with LPS (10 μg/ml) decreased markedly with time and was inhibited by 33%, 71%, and 76% after 24 hr, 48 hr, and 72 hr, respectively, when compared with control, untreated cells. The inhibitory effect of LPS was partially reversible upon removal of the LPS and further incubation in the absence of LPS. The LPS‐induced decrease in ACE activity was dependent on the concentrations of LPS (IC50 = 15 ng/ml at 24 hr) and was detectable at LPS concentrations as low as 1 ng/ml. That LPS decreased the Vmax of ACE in the absence of cytotoxicity and without a change in Km suggests that LPS decreased the amount of ACE present on the HUVEC cell membrane. While some LPS serotypes (Escherichia coli 0111 :B4 and 055 :B5, S. minnesota) were more potent inhibitors of ACE activity than others (E. coli 026:B6 and S. marcescens), all LPS serotypes tested were inhibitory. These finding suggest that LPS decreases endothelial ACE activity in septic patients; in turn, this decrease in ACE activity may decrease angiotensin II production and bradykinin catabolism and thus play a role in the pathogenesis of septic shock.
Decreased response to epidermal growth factor during cellular senescence in cultured human microvascular endothelial cells
Authors: Matsuda Takao , Okamura Kazuki , Sato Yasufumi , Morimoto Akio , Ono Mayumi , Kohno Kimitoshi , Kuwano Michihiko 
Published: February 2005

AbstractWe have previously demonstrated that epidermal growth factor (EGF) induces cell migration, tissue‐type plasminogen activator synthesis, as well as tubular formation in microvascular endothelial cells from human omental tissue. In this study, we compared the responsiveness to EGF of late passaged (senescent) human omental microvascular endothelial (HOME) cells with that of early passaged (young) HOME cells. We have employed HOME cells derived from surgically resected omental samples from 14 patients. EGF stimulated cell migration significantly more in the young cells than in the senescent cells during serial cultivation (aging) in vitro. Scatchard analysis demonstrated that the number for both high and low affinity receptors for EGF in HOME cells was decreased dramatically during serial cultivation. The expression of EGF receptor mRNA was also decreased in the senescent HOME cells. Treatment of HOME cells with EGF significantly increased cellular mRNA levels of tissue‐type plasminogen activator, and two protooncogenes, c‐fos and c‐myc, in young HOME cells, but not in senescent HOME cells. Thus HOME cells aged in vitro show a decreased responsiveness to EGF, resulting in decreased migration of human endothelial cells. The serial cultivation of human endothelial cells in vitro may downregulate EGF receptor and decrease responsiveness to exogenous EGF, a potent angiogenic factor.
Identification of estrogen receptor mRNA and the estrogen modulation of parathyroid hormone‐stimulated cyclic AMP accumulation in opossum kidney cells
Authors: Stock John L. , Coderre James A. , Burke Elizabeth M. , Danner David B. , Chipman Stewart D. , Shapiro Jay R. 
Published: February 2005

AbstractThe opossum kidney (OK) cell was used as a model to test the hypothesis that estrogen directly affects proximal renal tubular epithelial cells. To demonstrate the expression of estrogen receptor in OK cells, we developed an approach using reverse transcription and the polymerase chain reaction. Analysis of the DNA amplified with nested primers revealed the predicted size fragment and restriction enzyme digestion products. To demonstrate the functional effects of estrogen, OK cells at confluence were preincubated in serum‐free medium for 7–10 days with or without 17β‐estradiol. Bovine PTH(1–34) (bPTH(1–34)) then stimulated a dosedependent intracellular accumulation of cAMP that was maximal after 1 min and then gradually declined. Cyclic AMP in the medium slowly increased over 60 min. Preincubation with 17β‐estradiol did not affect cell proliferation as measured by total protein content but caused an inhibition of bPTH(1–34)‐stimulated intracellular cAMP accumulation that was maximal at 10−11M 17β‐estradiol (71 ± 3% control, p < .001). bPTH(1–34) also increased cAMP release into the medium, an effect maximal using 10−10M 17β‐estradiol (118 ± 3% control, p < .001). Preincubation with the inactive isomer 17α‐estradiol caused no changes in cAMP accumulation or release. Coincubation with the antiestrogen tamoxifen blocked the effects of 17β‐estradiol. Sodium‐dependent phosphate transport was: (1) inhibited by 2‐h incubations with 10−8 or 10−10 M bPTH(1–34) and not affected by preincubation with 17β‐estradiol, and (2) not inhibited by a 20‐min incubation with 10−8 M bPTH(1–34) unless cells were preincubated with 10−8 M 17β‐estradiol, suggesting that any possible effects of estrogen on phosphate transport are not directly mediated by changes in cAMP. These studies demonstrate the presence of estrogen receptor mRNA in OK cells as well as direct and specific effects of physiologic concentrations of estrogen on cAMP accumulation in these cells. This system may be a good model for further study of estrogen and PTH effects on the kidney.
Contrasting effects of the protein kinase C inhibitor staurosporine on the interleukin‐1 and phorbol ester activation pathways in the EL4–6.1 thymoma cell line
Authors: Dornand Jacques , Bouaboula Monsif , D'Angeac Arnaud Dupuy , Favero Jean , Shire David , Casellas Pierre 
Published: February 2005

AbstractEL 4–6. 1 cells, variants of the murine EL4 thymoma cell line, can be activated by interleukin 1 (IL‐1) or phorbol 12‐myristate‐13‐acetate (PMA), or PMA + IL‐1 to secrete interleukin 2 (IL‐2) and interleukin 4 (IL‐4) and to express the IL‐2 receptor (IL‐2R). To compare the different activation pathways, we examined the effects of staurosporine (STAR) and 1‐(5‐isoquinolinylsulfonyl)‐2‐methylpiperazine (H7), two protein kinase C (PKC) inhibitors, on the induction of interleukin secretion and IL‐2R expression in these cells. We report here that nanomolar concentrations of STAR strongly potentiated (20‐ to 30‐fold) the production of IL‐2 or IL4, when EL 4–6.1 cells were induced by IL‐1 α (or IL‐1 β) alone. By contrast, at identical concentrations, STAR dose‐dependently inhibited the production of IL‐2 and IL‐4 resulting from PMA or PMA + IL‐1 cell treatment. STAR also negatively affected the expression of IL‐2R, which was dependent on PMA‐sensitive PKC with either IL‐1, PMA, or PMA + IL‐1 stimulation. The changes in interleukin production and IL‐2R expression in EL 4–6.1 activated cells were correlated with changes at the mRNA level measured by quantitative polymerase chain reaction (PCR). This finding suggests a pretranslational effect of the drug. At micromolar concentrations, H7 showed the same effects as STAR, but only increased IL‐1‐triggered interleukin secretions twofold. We observed that the action of PKC inhibitors did not result from modification of IL‐1 receptor (IL‐1R) expression in EL 4–6.1 cells. Thus, our data show that PKC inhibitors clearly distinguish between IL‐1 and PMA stimulatory pathways. In addition, they suggest that the IL‐1 stimulatory pathway involves PKC(s) [or other undefined kinase(s)] which regulate this pathway and differ from PKC(s) activated by PMA. © 1992 Wiley‐Liss, Inc.
Importance of tyrosine phosphatases in the effects of cell‐cell contact and microenvironments on EGF‐stimulated tyrosine phosphorylation
Authors: Mansbridge J. N. , Knüchel R. , Knapp A. M. , Sutherland R. M. 
Published: February 2005

AbstractWe have compared the EGF responses of A431 cells when grown as monolayers at a variety of cell densities of as multicellular spheroids in order to investigate the effects of cell contact and 3‐dimensional structure on signal transduction. Proliferation of the A431 squamous carcinoma cell line grown in our laboratory was unaffected by EGF when grown in monolayer culture. As 3‐dimensional, multicellular spheroids, however, growth was stimulated by EGF. The maximum volume attainable in the presence of EGF was more than 30 times that in its absence. EGF‐dependent tyrosine phosphorylation was compared under these conditions by immunohistochemistry and Western blotting. In initial experiments using published procedures, tyrosine phosphorylation was density‐dependent in monolayers and undetectable in spheroids. However, the density‐dependence was abolished by the addition of high concentrations of protein tyrosine phosphatase inhibitors (1 mM Zn++ and VO43−). The density dependence of EGF‐stimulated tyrosine phosphorylation in monolayers was, therefore, largely the result of changes in phosphatase activity rather than kinase. Using high concentrations of phosphatase inhibitors, phosphotyrosine was clearly visible by immunohistochemistry in the outermost cells of spheroids, but it was still not visible in the spheroid center. The lack of response within the spheroid was not related to the presence of EGF receptor nor diffusion of EGF. In companion experiments, we showed that staining for EGF receptor was present homogeneously throughout the spheroid and that EGF penetrated to its center under the conditions of the experiment. Thus, although an increase in tyrosine phosphatase activity was a major factor affecting tyrosine phosphorylation in the outer cells, other factors were important in the inner cells. We concluded that an increase of tyrosine phosphatase activity was the most important component of the adaptation of the EGF signal transduction system to high cell density in monolayer cultures. In spheroids, tyrosine phosphatases are also enhanced, but other factors, such as autocrine synthesis of TGF‐α and possibly the cellular distribution of EGF receptors and cell shape, play a role. © 1992 Wiley‐Liss, Inc.
Endothelium‐derived relaxing factor contributes to the regulation of endothelial permeability
Authors: Oliver Juan A. 
Published: February 2005

AbstractTo determine whether endothelium‐derived relaxing factor (EDRF) contributes to the regulation of endothelial permeability, the transendothelial flux of 14C‐su‐crose, a marker for the paracellular pathway across endothelial monolayers (Oliver, J. Cell. Physiol. 145:536–548, 1990), was examined in monolayers of bovine aortic endothelial cells grown on collagen‐coated filters. The permeability coefficient of 14C‐sucrose was significantly decreased by 10−3 M 8‐Bromoguanosine 3′,5′‐cyclic monophosphate or by 5 × 10−6 M glyceryl trinitrate, an activator of soluble guanylate cyclase. Depletion of L‐arginine from endothelial monolayers increased 14C‐sucrose permeability from 3.21 ± 0.59 to 3.88 ± 0.50 × 10−5 cm · sec−1 (mean ± SEM; n = 6; P < 0.05). The acute administration of 5 × 10−4 M L‐arginine to monolayers depleted of this amino acid decreased 14C‐sucrose permeability from 2.91 ± 0.27 to 2.52 ± 0.26 × 10−5 cm · sec−1 (n = 11; P < 0.05). 14C‐sucrose permeability was increased by 10−7 M bradykinin and this effect was enhanced by the presence of each one of the following compounds: 10−5 M methylene blue, 4 × 10−6 M oxyhemoglobin, 5 × 10−4 M NG‐methyl‐L‐arginine or 5 × 10−4 M Nω‐nitro‐L‐arginine. These results suggest that EDRF contributes to the sealing of the endothelial monolayer and that EDRF released by bradykinin acts as a feedback inhibitor attenuating the increase in endothelial permeability induced by this peptide. Because endothelial cells have the ability to contract and relax and possess guanylate cyclase responsive to nitric oxide, our results suggest that EDRF decreases 14C‐sucrose permeability by relaxing endothelial cells, thereby narrowing the width of endothelial junctions. © 1992 Wiley‐Liss, Inc.