bluuebunny/crossref_metadata_2025_split huggingface dataset:
Involvement of the Golgi region in the intracellular trafficking of cholera toxin
Authors: Nambiar Madhusoodana P. , Oda Tatsuya , Chen Chaohua , Kuwazuru Yasuo , Wu Henry C. 
Published: February 2005

AbstractThe intracellular pathway following receptor‐mediated endocytosis of cholera toxin was studied using brefeldin A (BFA), which inhibited protein secretion and induced dramatic morphological changes in the Golgi region. In both mouse Y1 adrenal cells and CHO cells, BFA at 1 μg/ml caused a 80–90% inhibition of the cholera toxin (CT)‐elevation of intracellular cAMP. The inhibition of the cytotoxicity of CT by BFA was also observed in a rounding assay of Y1 adrenal cells. The inhibition of CT cytotoxicity by BFA was dose dependent, with the ID50 value similar to the LD50 of BFA in Y1 adrenal cells. Binding and internalization of [125I]‐cholera toxin in Y1 adrenal cells was not affected by BFA. Unlike the BFA‐sensitive cell lines such as Y1 adrenal and CHO cells, BFA at 1 μg/ml did not inhibit the cytotoxicity of CT in PtK1 cells, of which the Golgi structure was BFA‐resistant. These results strongly suggest that a BFA‐sensitive Golgi is required for the protection of CT cytotoxicity by BFA. In contrast, elevation of the intracellular cAMP by forskolin, which acts directly on the plasma membrane adenylate cyclase, was not affected by BFA. These observations indicate that the intoxication of target cells by CT requires an intact Golgi region for its intracellular trafficking and/or processing. In this respect, CT shares a common intracellular pathway with ricin, Pseudomonas toxin, and modeccin, even though their structures and modes of action are very different. © 1993 Wiley‐Liss, Inc.
Nuclear protein redistribution in heat‐shocked cells
Authors: Warters Raymond L. , Chu G. L. , Wong R. S. L. , Lyons B. W. , Dewey W. C. 
Published: February 2005

AbstractAn increase was observed in the total protein mass of nuclei isolated from Chinese hamster ovary cells heated at 45°C or 45.5°C. An increase in the fractional recovery of DNA polymerase α and β, and of DNA topoisomerase activity coincided with this increase in the protein mass of nuclei from heated cells. Nuclear protein mass which was soluble in 2.0 M NaCl decreased 0.5 fold, while DNA‐associated and nuclear matrix‐associated protein mass increased 2.2 and 3.4 fold, respectively. The results indicate that the increase in nuclear protein mass observed in nuclei from heated cells is due in part to an increased binding, or precipitation, of nuclear proteins onto the cell's DNA and nuclear matrix. © 1993 Wiley‐Liss, Inc.
Heat shock alters centrosome organization leading to mitotic dysfunction and cell death
Authors: Vidair C. A. , Doxsey S. J. , Dewey W. C. 
Published: February 2005

AbstractTo identify the cellular target(s) responsible for thermal killing in the G1 phase of the cell cycle, synchronous cultures of Chinese hamster ovary cells (CHO) were heat shocked and studied for one cell cycle by time‐lapse videomicroscopy and immunocytochemistry. At the first mitosis post‐heating, the fraction of cells giving rise to multinucleated progeny approximately equaled the nonclonogenic fraction. In addition, the cells yielding multincleated progeny were delayed in prophase‐metaphase relative to the cells yielding two uninucleated progeny (clonogenic cells). To study the basis for the delay in prophase‐metaphase and subsequent formation of multinucleated cells, cells in mitosis were examined by immunofluorescence for spindle abnormalities. Multipolar mitotic spindles and chromosome misalignment were induced by heat. All multiple spindle poles induced by heat stained for pericentriolar material (PCM), the microtubule nucleating material of centrosomes. Heated cells in mitosis also contained additional foci of PCM which were not associated with the spindle. Cells made thermotolerant by a nonlethal heat shock were resistant to both thermal killing and the induction of multiple foci of PCM. Quantitative analysis revealed a good correlation between the fraction of cells with multipolar spindles, the fraction with more than two foci of PCM, and the nonclonogenic fraction. These data indicate that heat‐induced alterations to the PCM of centrosomes resulted in multipolar mitotic spindles, delay in prophase‐metaphase, and formation of multinucleated cells which were nonclonogenic. These results identify the centrosome as a G1 target for cell killing. © 1993 Wiley‐Liss, Inc.
Regulation of neutrophil interleukin 8 gene expression and protein secretion by LPS, TNF‐α, and IL‐1β
Authors: Fujishima Seitaro , Hoffman Andrew R. , Vu Thanh , Kim K. Jin , Zheng Hui , Daniel David , Kim Youngmee , Wallace Ellen F. , Larrick James W. , Raffin Thomas A. 
Published: February 2005

AbstractNeutrophils are possibly involved in the pathogenesis of various lung diseases through the release of numerous mediators. In the present study, we studied the regulation of IL‐8 gene induction and protein secretion in human blood neutrophils. Northern blot analysis revealed that LPS increased IL‐8 mRNA levels in neutrophils, with a maximal fivefold increase by 2 h. IL‐8 mRNA levels returned to baseline value within 12 h. In contrast, LPS‐stimulated monocytes demonstrated a sustained increase of IL‐8 mRNA levels for more than 24 h. TNF‐α, IL‐1β, and phorbol myristate acetate also increased IL‐8 mRNA levels in neutrophils. Immunohistochemical analysis confirmed that IL‐8 was localized within stimulated neutrophils. IL‐8 secretion by neutrophils and monocytes was quantified using a specific ELISA for IL‐8. Resting neutrophils secreted minimal IL‐8 activity. However when cells were stimualted with LPS, TNF‐α, or IL‐1bT, neutrophils secreted IL‐8. IL‐8 secretion was most marked during the first 2 h after stimulation and decreased thereafter. In contrast, monocytes maintained a high rate of IL‐8 secretion over 12 h. Although a single monocyte secreted 70‐fold more IL‐8 than did a single neutrophil after 4 h of incubation, the high abundance of neutrophils in peripheral blood made the neutrophil‐secreted IL‐8 more significant. During the first 2 h, neutrophils secreted ∼40% of the IL‐8 released by monocytes in the same volume of blood. This ratio decreased to 9% after 12 h. Neutrophil‐secreted IL‐8 may play an autocrine or paracrine role during the initial stage of inflammation. © 1993 Wiley‐Liss, Inc.
Modulation of expression of insulin and IGF‐I receptor by Epstein‐Barr virus and its gene products LMP and EBNA‐2 in lymphocyte cell lines
Authors: Kriauciunas Kristina M. , Goldstein Barry J. , Lipes Myra A. , Kahn C. Ronald 
Published: February 2005

AbstractThe receptors for insulin and insulin‐like growth factor I (IGF‐I) are two closely related integral membrane glycoproteins involved in signalling of cell growth and metabolism. We have used the unique paradigm of pairs of Burkitt lymphoma cell lines (BLO2, BL30, BL41) with or without Epstein‐Barr Virus (EBV) infection and cells transfected with EBV‐related genes to examine effects of EBV on expression of these receptors at the gene and protein functional level. In BL30 and BL41 cells, EBV infection increased surface insulin binding and total receptor number by 2‐and 18‐fold, respectively. By contrast, EBV infection decreased total IGF‐I receptors by 29 to 87% in all three cell lines. In general, there was a correlation between total receptor concentration and the level of insulin or IGF‐I receptor mRNAs, although in one cell line insulin binding increased while receptor mRNA levels decreased slightly, suggesting posttranslational effects. BL41 cells transfected with a vector expressing the EBV latent membrane protein (LMP) exhibited a 2.6‐ to 3.2‐fold increase in insulin receptor expression, whereas cells transfected with EBNA‐2 (one of the EBV nuclear antigens) alone exhibited no effect. However, EBNA‐2 appears to be required for the EBV effect on insulin receptor expression since cells infected with a mutant virus, P3JHRI, which lacks the EBNA‐2 gene failed to show an increase in insulin receptor number. These data indicate that EBV infection of lymphocytes increases expression of insulin receptors while simultaneously decreasing expression of IGF‐I receptors. The magnitude and sometimes even the direction of change, depends on host cell factors. A maximal increase in insulin receptors appears to require the coordinate action of several of the EBV proteins including LMP and EBNA‐2. © 1993 Wiley‐Liss, Inc.
Kinetic characterization of Na+/H+ antiport of Plasmodium falciparum membrane
Authors: Bosia A. , Ghigo D. , Turrini F. , Nissani E. , Pescarmona G. P. , Ginsburg H. 
Published: February 2005

AbstractIntraerythrocytic malaria parasites produce vast amounts of lactic acid through glycolysis. While the egress of lactate is very rapid, the mode of extrusion of H+ is not known. The possible involvement of a Na+/H+ antiport in the extrusion of protons across the plasma membrane of Plasmodium falciparum has been investigated by using the fluorescent pH probe 6‐carboxyfluorescein. The resting cytosolic pH was 7.27 ± 0.1 in ring stage parasites and 7.31 ± 0.12 in trophozoites. Spontaneous acidification of parasite cytosol was observed in Na+‐medium and realkalinization occurred upon addition of Na+ to the medium in a concentration‐dependent manner, with no apparent saturation. The rate of H+‐at the ring stage was higher than that at the trophozoite stage due to the larger surface/volume ratio of the young parasite stage. Na+‐H+‐was: 1) inhibited by the Na+/H+ inhibitors amiloride and 5‐(N‐ethyl‐isopropyl) amiloride (EIPA), though at relatively high concentrations; 2) augmented with rising pH6 (pHi = 6.2 [Na+]o = 30 mM); and 3) decreased with increasing pHi (pHo = 7.4; [Na+]o = 30 mM). The pHi and the pHo dependencies of H+‐were almost identical at all parasite stages. Only at pHi > 7.6 efflux was totally obliterated. The target of this inhibitory effect is probably other than the antiport. Results indicate that H+‐is mediated by a Na+/H+ antiport which is regulated by host and parasite pH and by the host cytosol sodium concentration. The proton transport capacity of the antiport can easily cope with all the protons of lactic acid produced by parasite's glycolysis. © 1993 Wiley‐Liss, Inc.
Occurrence and possible consequences of multipolar mitoses in primary cultures of adult rat hepatocytes
Authors: Eckl P. M. 
Published: February 2005

AbstractProliferating primary cultures of adult rat hepatocytes are characterized by the occurrence of multipolar mitoses, and chromosome loss resulting in the formation of micronuclei at telophase. The percentage of multipolar mitotic figures was determined to be 12.76 ± 7.9%, 80% of which were tripolar. Multipolar mitotic stages showed a high incidence of chromosome loss, increasing from meta‐ (61.7 ± 16.6%) to telophase (72.1 ± 19.3%). Regular bipolar mitotic figures on the other hand also showed chromosome loss, however, to a lesser degree and decreasing from meta‐ (49.5 ± 10.4%) to telophase (34.9 ± 7.9%). The incidence of chromosome loss even in regular mitotic figures is very high compared to other cells and appears to depend on another special feature of hepatocytes: they remain flat and well attached during mitosis, so that shearing forces could be responsible for the separation of chromosomes from the mitotic spindle. Additionally this morphology creates a situation allowing for a maximal interaction of mitotic spindles of binucleated cells, leading to the high rate of multipolar mitoses observed. Both multipolar mitoses and chromosome loss could also explain the consecutive detachment of hepatocytes reported for proliferating primary cultures, since the aneuploid daughter cells generated can be expected to be nonviable in most cases and eventually detach. © 1993 Wiley‐Liss, Inc.
Regulation of chicken Hsp70 and Hsp90 family gene expression by transforming growth factor‐β1
Authors: Takenaka Ivone M. , Hightower Lawrence E. 
Published: February 2005

AbstractTransforming growth factor‐β1 (TGFβ) is a regulator of protein synthesis in cultured chicken embryo cells (CEC). Preceding a gradual increase in overall protein synthesis, members of the Hsp70 family (Hsp70, Hsc70, and Grp78) and the Hsp90 family (90‐2 and 90‐3) of molecular chaperones are induced rapidly and represent a new class of TGFβ‐inducible proteins (I.M. Takenaka and L.E. Hightower, J. Cell. Physiol., 152:568–577, 1992). Herein, 32P‐labeled cDNA probes encoding Hsc70 and Hsp90 were used to show that levels of the corresponding mRNAs increased as a fraction of total RNA and in polysomes within five hours of treatment of CEC with TGFβ. This cytokine did not increase rates of hsc70 and hsp90 gene transcription as measured by run‐on transcription assays of isolated nuclei. However, the Hsp RNA inductions were inhibited by dactinomycin, indicating a requirement for newly synthesized RNA. Both Hsc70 and Hsp90 mRNAs had relatively short half‐lives, measured by Northern blot analyses of dactinomycin chases, which were not altered substantially in TGFβ‐treated cells. In contrast, Hsp mRNA half‐lives increased in heat shocked CEC exposed to dactinomycin during recovery, revealing a difference in regulation of these genes in stressed cells compared with TGFβ‐treated cells. Our results support the conclusion that hsc70 and hsp90 gene expression is regulated posttranscriptionally in TGFβ‐treated CEC, and the mechanism likely involves a nuclear event such as increasing the half‐lives of nuclear RNA transcripts, processing, or transport into the cytoplasm. © 1993 Wiley‐Liss, Inc.
TGFβ induction of extracellular matrix associated proteins in normal and transformed human mammary epithelial cells in culture is independent of growth effects
Authors: Stampfer Martha R. , Yaswen Paul , Alhadeff Myriam , Hosoda Junko 
Published: February 2005

AbstractWe have previously characterized a human mammary epithelial cell (HMEC) culture system for the effects of TGFβ1 on cell growth. In the current report, the effects of TGFβ 1 on synthesis and secretion of proteins associated with the extracellular matrix and proteolysis were examined. In particular, we compared the TGFβ responses of normal finite lifespan HMEC, which are growth inhibited by TGFβ, to two immortally transformed cell lines derived from the normal HMEC. One of these lines maintains active growth in the presence of TGFβ and the other shows partial growth inhibition. In contrast to the differing effects of TGFβ on cell growth, we found that all these cell types showed strong induction of most of the mRNA and protein species examined, including fibronectin, collagen IV, laminin, type IV collagenase, urokinase type plasminogen activator (uPA), and plasminogen activator inhibitor 1 (PAI‐1). The profile of TGFβ 1 binding proteins was the same in HMEC that were, and were not growth suppressed by TFGβ. Therefore, the effects of TGFβ on cell growth could be dissociated from its effects on specialized responses, indicating that within this one cell type there must be at least two independent pathways for TGFβ activity, one which leads to cessation of proliferation and one which induces a specific set of cellular responses. This cell system may be useful for examining the pathway of TGFβ induced growth inhibition using closely matched cells which vary in their growth‐induced response but retain similar specialized responses to TGFβ. © 1993 Wiley‐Liss, Inc.
Mechanism of retinoid‐induced activation of latent transforming growth factor‐β in bovine endothelial cells
Authors: Kojima Soichi , Rifkin Daniel B. 
Published: February 2005

AbstractCell‐associated plasmin is a putative physiological activator of latent transforming growth factor‐β (LTGF‐β). Since retinoids enhance the production of plasminogen activator (PA) and thereby increase cell‐associated plasmin activity, we tested the possibility that retinoids might induce the activation of LTGF‐β using bovine endothelial cells (ECs) as a model system. ECs treated with physiological concentrations of retinol or retinoic acid formed active TGF‐β in the culture media in a dose‐ and time‐dependent fashion. Cells were treated with 2 μM retinol for 24 h, and the amount of TGF‐β produced during a subsequent 12‐h incubation period was measured. Out of a total of 14 pM LTGF‐β secreted, 0.7 pM was converted to active TGF‐β. Northern blot analyses showed that mRNA levels for TGF‐β2 but not for TGF‐β1 increased in cells treated with retinol. Inclusion of either inhibitors of PA or of plasmin or antibody against PA in the culture medium as well as depletion of plasminogen from the serum blocked the formation of TGF‐β, suggesting that PA, plasminogen, and the resulting plasmin are essential for activation of LTGF‐β in retinoid‐stimulated cells. Antibody against the LTGF‐β binding protein blocked activation implying that localization of LTGF‐β through its binding protein may be important. However, inhibition of binding of LTGF‐β to the cell surface mannose 6‐phosphate receptor did not prevent activation. These data indicate that retinoids up‐regulate the production of LTGF‐β in ECs and induce activation of LTGF‐β, perhaps, by increasing PA and plasmin levels. Thus, TGF‐β might be a local mediator of some of the biological activities of retinoids both in vivo and in vitro. © 1993 Wiley‐Liss, Inc.
Different signal transduction by epidermal growth factor may be responsible for the difference in modulation of amino acid transport between fetal and adult hepatocytes
Authors: Leoni S. , Spagnuolo S. , Marino M. , Terenzi F. , Massimi M. , Devirgiliis L. Conti 
Published: February 2005

Abstract[1‐14C]‐2‐aminoisobutyric acid (AIB) uptake and signal transduction pattern after epidermal growth factor (EGF) stimulation were examined in freshly isolated hepatocytes from 20‐day‐old fetuses and 3‐month‐old rats. EGF induced a transient increase of AIB transport after 10 min only in adult animals; the observed unresponsiveness of fetal liver is not dependent on a lack of EGF receptors which are present though to a lesser extent on the plasma membrane in this period. As far as the production of the second messengers, inositol trisphosphate (IP3) and calcium, is concerned, substantial differences were found: EGF increased IP3 production in adult hepatocytes, whereas it had no effect in fetal ones. Moreover, the addition of EGF induced a calcium transient in hepatocytes from adult animals, while there was no increase in fetal cells. The lack of EGF effect on amino acid transport in fetal cells could be due to its inability to produce both IP3 and calcium transients, suggesting that this transduction pathway is not activated during fetal life.
Mechanism of action of angiostatic steroids: Suppression of plasminogen activator activity via stimulation of plasminogen activator inhibitor synthesis
Authors: Blei Francine , Wilson E. Lynette , Mignatti Paolo , Rifkin Daniel B. 
Published: February 2005

AbstractRecently, a novel class of angiostatic steroids which block angiogenesis in several systems has been described. Since the elaboration of proteases is believed to be an important component of angiogenesis, we tested whether these steroids blocked the fibrinolytic response of endothelial cells to the angiogenic protein, basic fibroblast growth factor [bFGF]). Cultured bovine aortic endothelial (BAE) cells were incubated with bFGF and/or medroxyprogesterone acetate (MPA), an angio‐static steroid which has been shown to inhibit vascularization, collagenolysis, and tumor growth. When bFGF (3 ng/ml) was added to confluent monolayers of BAE cells, plasminogen activator (PA) activity in the medium was increased threefold. In contrast, MPA at 10−6 M, 10−7 M, 10−8 M, and 10−9 M decreased PA levels in the medium by 83%, 83%, 75%, and 39%, respectively. The stimulation of PA levels in BAE cells by bFGF (3 ng/ml) was abrogated by the presence of 10−6 M MPA. This decrease in PA activity was found to be mediated by a significant increase in plasminogen activator inhibitor type‐1 (PAI‐1) production. MPA, therefore, negated one of the important enzymatic activities associated with the angiogenic process. In contrast to the decreased levels of secreted PA in cultures exposed simultaneously to MPA and bFGF, cell‐associated PA levels remained high, consistent with earlier observations indicating that PAI‐1 does not inhibit cell‐associated PA. Thus, angiostatic steroids may exert their inhibitory effects on angiogenesis by increasing the synthesis of PAI‐1. This, in turn, inhibits PA activity and, therefore, plasmin generation, which is essential for the invasive aspect of angiogenesis. © 1993 Wiley‐Liss, Inc.
Alterations in nuclear matrix ultrastructure of G1 mammalian cells following heat shock: Resinless section electron microscopy, biochemical, and immunofluorescence studies
Authors: Wachsberger Phyllis R. , Coss Ronald A. 
Published: February 2005

AbstractHeat shock is known to inhibit vital nuclear functions associated with DNA and RNA metabolism. It has been proposed that the reported heat‐induced excess protein accumulation in the nuclear matrix (NM) fraction may alter NM sites crucial for DNA and RNA processing. To test this hypothesis, we examined the fine structure of the NM in synchronous populations of G1 Chinese hamster ovary cells before and after heating by using the technique of resinless section electron microscopy. Heat did induce morphological alterations in the NM. The NM of control cells contained a honeycomb‐like arrangement of fibers after chromatin removal. Following heat shock, NMs appeared as more highly anastomosing networks of polymorphic fibers and an overall increase in electron density was observed. Residual nucleoli from heated NMs underwent alterations in distributions of electron density both internally and at their peripheries. The increase in electron density observed in heated NMs was accompanied by an increase in protein mass and a relatively smaller increase in RNA mass as indicated by parallel sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and isotopic labeling (protein/DNA and RNA) studies. Some excess protein accumulation could also be directly localized onto NM fibers by use of antibodies to heterogeneous ribonucleoprotein complex antigens. It is concluded that alterations of NM fine structure can reflect the heat‐stressed state of the cell, may account for the heat‐induced inhibition of nucleic acid metabolism, and may be useful as an indicator of physiological or pathological stress in general. © 1993 Wiley‐Liss, Inc.
Transforming growth factor β1‐mediated inhibition of smooth muscle cell proliferation is associated with a late G1 cell cycle arrest
Authors: Reddy Kumar B. , Howe Philip H. 
Published: February 2005

AbstractThe effect of transforming growth factor β1 (TGFβ1) on the proliferative response of aortic smooth muscle cells (SMC) in vitro was investigated. TGFβ1 substantially inhibited the growth of human and bovine SMC. Rapidly growing SMC and quiescent serum‐stimulated SMC were inhibited by TGFβ1 with an ID50 of approximately 0.5 ng/ml and maximal inhibition was observed at 10 ng/ml TGFβ1. In the presence of TGFβ1, quiescent serum‐stimulated SMC progress into the G1 phase of the cell cycle, but become reversibly arrested at a point temporally located 1–2 hours from S phase. Release from this late G1 TGFβ1 arrest point results in S phase entry within 2 hours. Associated with this inhibitory effect is a decrease in the histone H1 kinase activity of p34cdc2 protein kinase while TGFβ1 has no effect on the transcription or translation of p34cdc2. Under these growth inhibitory conditions, TGFβ1 is still capable of upregulating the expression of fibronectin mRNA. These results suggest that TGFβ1 growth inhibition in SMC is associated with the regulation of p34cdc2 activity in late G1. © 1993 Wiley‐Liss, Inc.