bluuebunny/crossref_metadata_2025_split huggingface dataset:
Glutathione stimulates A549 cell proliferation in glutamine‐deficient culture: The effect of glutamate supplementation
Authors: Kang Y. James , Feng Yi , Hatcher Emiko L. 
Published: February 2005

AbstractExtracellular glutathione (GSH) is degraded by an external cell‐surface enzyme, γ‐glutamyltranspeptidase (γ‐GT). The products are transported into cells to participate in important cellular processes. In the present study, we tested the hypothesis that extracellular GSH is a source of glutamic acid for cells that express γ‐GT. Under a glutamine‐deficient culture condition, the extracellular GSH‐supplemented glutamic acid would enhance intracellular glutamine synthesis, thereby stimulating cell proliferation. Human lung carcinoma A549 cells were cultured in glutamine‐deficient Dulbecco's modified Eagle medium, and they did not proliferate unless glutamine was supplemented. Extracellular GSH, however, provoked a partial proliferation. The GSH effect correlated with a high level of γ‐GT activity and an increased intracellular level of glutamic acid. A constituent amino acid of GSH, glutamic acid but not cysteine, produced the same growth‐stimulatory effect as GSH. Furthermore, neither oxothiazolidine‐4‐carboxylate (OTC), a celluar cysteine‐delivery compound, nor cysteinylglycine, a dipeptide released from the γ‐GT reaction, stimulated cell proliferation. Moreover, buthionine sulfoximine (BSO), a selective inhibitor of γ‐glutamylcysteine synthetase, enhanced the GSH growth stimulatory effect, suggesting that increased cellular GSH synthesis does not correlate with cell growth stimulation. The results obtained demonstrated that glutamine is required for A549 cell proliferation and exogenous GSH partially substitutes for the growth stimulatory action of glutamine. It also suggests that the glutamic acid rather than the cysteine released from the GSH is responsible for the cell proliferation. © 1994 Wiley‐Liss, Inc.
Disruption of microtubules in vivo by vincristine induces large membrane complexes and other cytoplasmic abnormalities in megakaryocytes and platelets of normal rats like those in human and Wistar Furth rat hereditary macrothrombocytopenias
Authors: Stenberg Paula E. , McDonald Ted P. , Jackson Carl W. 
Published: February 2005

AbstractAbnormal organization of platelet microtubules is associated with abnormal platelet formation in hereditary macrothrombocytopenias such as the gray platelet syndrome, May‐Hegglin anomaly, and Epstein's syndrome, and that of the Wistar Furth rat, suggesting that aberrant microtubule organization may contribute to defective platelet formation in these clinical entities. Here, we examined the consequence of microtubule disruption on the organization of megakaryocyte cytoplasmic organelles using the microtubule depolymerizing agent, vincristine (VCR). Wistar rat bone marrow was fixed and processed for transmission electron microscopy after VCR administration alone, after 5‐fluorouracil (5‐FU) administration alone, or after 5‐FU followed by intravenous injection of 0.1–1.0 mg/kg VCR for intervals of 30 min to 8 hr. 5‐FU was given to increase megakaryocyte frequency to facilitate ultrastructural evaluations. VCR alone or in combination with 5‐FU caused formation of large membrane complexes in the cytoplasm of Wistar rat megakaryocytes at all dosages studied, identical to those found in megakaryocytes of human hereditary macrothrombocytopenias and the Wistar Furth rat. The proportion of megakaryocytes with these large membrane complexes increased with time after 5‐FU and VCR, and was maximal (∼two‐third of megakaryocytes) at VCR dosages of 0.75–1.0 mg/kg. The majorityof megakaryocytes displayed other abnormalities, including blebbing of plasma membranes, an increased number of dense compartments, dilated demarcation membrane (DMS) channels, which contained dense material immunocytochemically identified as secreted α‐granule proteins, and an increased incidence of emperipolesis. Rats administered 5‐FU alone did not demonstrate these abnormalities, with the exception of an increase in dense compartments. Platelets from rats treated with VCR aloene or 5‐FU and VCR also showed abnormalities including membrane complexes, rounded shape, formation of tubulin paracrystals, development of membrane blebs, and the presence of proteinaceous material within the cisternae of the surface‐connected canalicular system (SCCS). The membrane complexes in platelets of 5‐FU‐, VCR‐treated Wistar rats as well as untreated Wistar Furth rats were composed of elements of both the SCCS and dense tubular system; membrane complexes in megakaryocytes of 5‐FU‐, VCR‐treated rats were composed of both DMS and smooth endoplasmic reticulum. We conclude that intact microtubules play a major role in the organization of the megakaryocyte DMS and may contribute to the stability of megakaryocyte α‐granules. © 1995 Wiley‐Liss, Inc.
Reglation of autocrine gastrin expression by the TGFα autocrine loop
Authors: Howell Gillian M. , Ziober Barry L. , Humphrey Lisa E. , Willson James K. V. , Sun Luzhe , Lynch Mark , Brattain Michael G. 
Published: February 2005

AbstractGastrin is transcriptionally responsive to EGF stimulation (Merchant et al., 1991, Mol. Cell. Biol., 11:2686–2696). Consequently, we hypothesized that previously recognized gastrin autocrine loops (Hoosein et al., 1990, Exp. Cell. Res., 186:15–21), might be controlled by autocrine TGFα in human colon carcinoma cells. Therefore, we examined the interaction between these two autocrine growth factors in two colon carcinoma cell lines which utlizie TGFα. The FET cell line requires exogenous TGFα/EGF for optimal growth and has a classical TGFα autocrine loop which is disrupted by TGFα or epidermal growth factor receptor (EGFr) antibodies. The HCT 116 cell line is not dependent on exogenous TGFα/EGF and exhibits a nonclassical TGFα autocrine loop which is not disrupted by neutralizing antibodies to either TGFα itself or the EGFr. Basal gastrin mRNA production is significantly higher in HCT 116 than FET as measured by RNase protection assay. In the FET cells, exogenous EGF stimulates gastrin mRNA production but not in HCT 116. When the TGFα autocrine loop in HCT 116 is disrupted by constitutive expression of antisense TGFα mRNA, the gastrin mRNA level is significantly repressed. In xenografts derived from these antisense clones, TGFα reverted to high expression, and the gastrin mRNA level was again increased. This interaction between the strong TGFα loop in HCT 116 and the gastrin autocrine loop may confer a growth advantage to these colon cells. Such interactions between growth factors may promote enhanced tumorigenicity to transformed cells with these strong, nonclassical autocrine loops. © 1995 Wiley‐Liss, Inc.
Regulation of c‐fos expression in senescing Werner syndrome fibroblasts differs from that observed in senescing fibroblasts from normal donors
Authors: Oshima Junko , Campisi Judith , Tannock T. Charles A. , Martin George M. 
Published: February 2005

AbstractThe Werner syndrome (WS) is a segmental progeroid syndrome caused by a recessive mutation (WRN) mapped to 8p12. The replicative life spans of somatic cells cultured from WS patients are substantially reduced compared to age‐matched controls. Certain molecular concomitants of the replicative decline of normal fibroblast cultures have recently been defined, and it appears that multiple changes in gene expression accompany normal cell senescence. If the mechanisms by which WS cells exit the cell cycle were entirely comparable, the molecular markers of senescence should be identical in normal and WS cells. We find that this is not the case. The constitutive expression of statin, a nuclear protein associated with the nonproliferating state, was comparably expressed in normal and WS senescent cells. Likewise, the steady state levels of p53, a protein known to be involved in the G1 checkpoint of the cell cycle, were similar in early‐passage fibroblasts from normal and WS subjects. The levels of p53 were not increased in senescent fibroblasts, whether derived from normal or WS subjects. By contrast, the inducibility of mRNA and protein expression of the c‐fos protooncogene is preserved in late‐passage WS cells. This is in contrast to what is observed in late‐passage fibroblasts from normal subjects. Additional genotypes will have to be examined, however, to determine the specificity of this new aspect of the WS phenotype. © 1995 Wiley‐Liss, Inc.
Betamethasone activation of CTP: Cholinephosphate cytidylyltransferase is mediated by fatty acids
Authors: Mallampalli Rama K. , Salome Ronald G. , Li Chu Han , van Rollins Mike , Hunninghake Gary W. 
Published: February 2005

AbstractThe purpose of the present study was to determine the mechanisms by which glucocorticoids increase the activity of CTP: cholinephosphate cytidylyltransferase, a key enzyme required for the synthesis of surfactant phosphatidylcholine. Lung cytidylyltransferase exists as an inactive, light form low in lipids (L‐form) and an active, heavy form high in lipid content (H‐form). In vivo, fatty acids stimulate and aggregate the inactive L‐form to the active H‐form. In vivo, betamethasone increases the amount of H‐form while decreasing the amount of L‐form in fetal lung. There is also a coordinate increase in total free fatty acids in the H‐form. In the present study, we used gas chromatography–mass spectrometry to measure the fatty acid species associated with the H‐forms in fetal rat lung after the mothers were treated with betamethasone (1 mg/kg). In vivo, betamethasone increased the total amount of free fatty acids associated with the H‐form by 62%. Further, the hormone selectively increased the mass of myristic and oleic acids in H‐form by 52 and 82%, respectively. However, betamethasone produced the greatest increase in the amount of H‐form linoleic acid, which increased fourfold relative to control. In vitro, each of the fatty acids increased L‐form activity in a dose‐dependent manner; however, linoleic acid was the most potent. Linoleic and oleic acids also effectively increased L‐form aggregations. These observations suggest that in vivo glucocorticoids elevate the level of specific fatty acids which convert cytidylyltransferase to the active form. © 1995 Wiley‐Liss, Inc.
Laminin decreases PRL and IGFBP‐1 expression during in vitro decidualization of human endometrial stromal cells
Authors: Brar A. K. , Frank G. R. , Richards R. G. , Meyer A. J. , Kessler C. A. , Cedars M. I. , Klein D. J. , Handwerger S. 
Published: February 2005

AbstractThe expression of laminin, a major constituent of endometrial cell basement membranes, is increased during differentiation of human endometrial stromal cells (decidualization). To determine whether laminin plays a role in decidualization, we studied the effects of laminin substrate on the synthesis and release of prolactin (PRL) and insulin‐like growth factor binding protein‐1 (IGFBP‐1), two major secretory proteins of decidualized stromal cells. Endometrial stromal cells were plated on laminin as well as several other extracellular matrix (ECM) proteins (types 1 and IV collagen or fibronectin) and on plastic, and cultured in media containing medroxyprogesterone acetate (MPA) and estradiol. Cells cultured on plastic or ECM proteins displayed similar morphological changes indicative of decidualization. However, the release of PRL and IGFBP‐1 from cells cultured on plastic and ECM proteins (types 1 and IV collagen and fibronection) was approximately 2.1‐fold and 2.8‐fold greater respectively, than from cells cultured on laminin. The decrease in PRL and IGFBP‐1 expression in cells cultured on laminin was not due to differences in initial cell attachment efficiency or final DNA content. In addition, laminin had no effect on the content of laminin protein or fibronectin mRNA levels, indicating that the effects of laminin on PRL and IGFBP‐1 were specific. PGE2 stimulated the release of PRL and IGFBP‐1 from cells cultured on laminin to levels comparable to those from cells cultured on plastic or other ECM proteins. This indicates that the decrease in PRL and IGFBP‐1 release by laminin was not due to a generalized unresponsiveness. In contrast to the effects of laminin during decidualization, PRL expression was not altered by laminin in terminally differentiated decidual cells isolated at term. Our results support a role for laminin in selectively regulating PRL and IGFBP‐1 gene expression during in vitro decidualization of human endometrial stromal cells. © 1995 Wiley‐Liss, Inc.
Role of intracellular free Ca(II) and Zn(II) in dexamethasone‐induced apoptosis and dexamethasone resistance in human leukemic CEM cell lines
Authors: Adebodun Foluso , Post Jan F. M. 
Published: February 2005

AbstractThe levels of intracellular free Ca(II) and Zn(II) during dexamethasone (dex)‐induced apoptosis in CEM cell lines were determined by 19F nuclear magnetic resonance (NMR), using the fluorinated intracellular chelator 1,2‐bis‐(2‐amino‐5‐fluorophenoxy)ethane‐N, N, N′, N′‐tetraacetic acid (5‐FBAPTA). The effects of these divalent metal ions on growth rate and DNA degradation were evaluated. Measurements were done on one dex‐sensitive (CEM‐C7) and three different dex‐resistant variants (CEM‐C1, CEM‐4R4, and CEM‐ICR27). Dex caused a continuous increase in the Ca(II) level in dex‐sensitive CEM‐C7 cells, while in CEM‐C1 cells dex caused an initial increase in the Ca(II) level which in ≈︁36 h was restored to its normal value. The intracellular Ca(II) level in CEM‐4R4 cells was not significantly affected by dex, while that of CEM‐ICR27 cells decreased after dex incubation. Only the dex‐sensitive CEM‐C7 cells showed dex‐induced DNA degradation. An intracellular free Zn(II) level of ≈︁1 nM was measured for the dex‐resistant CEM‐C1 cells. No detectable level of intracellular Zn(II) was found in the other cell lines. Incubation with <100 μM Zn(II) did not inhibit dex‐induced apoptosis in CEM‐C7 cells (e.g., DNA degradation). Treatment with ≈︁250 μM Zn(II) caused significant decrease in growth rate in all cell lines and prevented dex‐induced DNA degradation in CEM‐C7 cells. A calibrated amount of Ca(II) ionophore (A23187), used to increase Ca(II) concentrations up to the dex‐induced levels, did not induce DNA degradation in CEM‐C7 or CEM‐C1 cells. While elevation of intracellular Ca(II) by itself is not sufficient to initiate apoptosis in CEM‐C7 cells, the results reported here suggest that Ca(II) is involved in the killing mechanism as a secondary factor. The combination of dex and ionophore caused significant DNA degradation in CEM‐C1 cells, which normally showed resistance to each compound individually. The combination of dex and the Zn(II) chelator phenanthroline also caused extensive DNA degradation in the normally dex‐resistant CEM‐C1 cells, suggesting that Zn(II) plays a role in the dex resistance of these cells. © 1995 Wiley‐Liss, Inc.
Phenotypic features of breast cancer cells overexpressing ornithine‐decarboxylase
Authors: Manni Andrea , Wechter Rita , Wei Lisa , Heitjan Daniel , Demers Laurence 
Published: February 2005

AbstractPolyamines (PA) have been shown to be critical mediators of estradiol‐induced breast cancer cell proliferation. This finding suggests that constitutive activation of the PA pathway may promote tumor progression, possibly leading to hormone independence. To test this hypothesis, we transfected hormone‐responsive MCF‐7 breast cancer cells with a complementary DNA coding for ornithine‐decarboxylase (ODC), the first rate‐limiting enzyme in PA biosynthesis. Marked ODC over‐expression observed in stably transfected clones was associated with a selective increase in cellular putrescine content, while spermidine and spermine levels were not altered. ODC‐overexpressing MCF‐7 cells were resistant to the antiproliferative effects of low but not high concentrations of the enzyme inhibitor, α‐difluoromethylornithine. In agreement with our hypothesis, sensitivity to the growth‐promoting action of estradiol was reduced by approximately one third (P < 0.001) in ODC‐overexpressing MCF‐7 cells compared with vector‐only transfected clones. Basal growth under anchorage‐dependent conditions was only marginally increased by ODC overexpression (P = 0.048), while clonogenicity in soft agar was actually reduced. These data suggest that activation of PA biosynthesis may contribute in part to the acquisition of estrogen independence by breast cancer cells. Since only putrescine content was increased as a result of ODC overexpression, these data may underestimate the overall influence of the PA pathway on breast cancer phenotype. © 1995 Wiley‐Liss, Inc.
Mechanical strain increases endothelin‐1 gene expression via protein kinase C pathway in human endothelial cells
Authors: Wang Danny Ling , Wung Being‐Sun , Peng Yi‐Ching , Wang Jaang Jung 
Published: February 2005

AbstractVascular endothelial cells (ECs) are constantly subjected to mechanical strain due to relaxation and contraction of vessel walls. The effects of cyclical strain on endothelin‐1 (Et‐1) secretion and Et‐1 mRNA levels in human umbilical vein ECs were examined. Cultured ECs grown on a flexible membrane base were deformed by negative pressure (16 kPa at 60 cycles/min). Cells subjected to strain showed increased Et‐1 secretion (0.54 ng/hr/106 cells) compared with unstrained control cells (0.22 ng/hr/106 cells). Northern blot analysis of cells strained for 2 hours or longer demonstrated a sustained elevated Et‐1 mRNA level at more than double the level in unstrained controls. This strain‐induced ET‐1 mRNA level returned to its basal level 2 hours after the release of strain. Cells treated with actinomycin D before or during strain treatment showed no strain‐induced gene expression. Pretreatment of ECs with a protein kinase C (PKC) inhibitor, Calphostin C, strongly inhibited the strain‐induced Et‐1 gene expression. Pretreatment of ECs with cAMP or cGMP‐dependent protein kinase inhibitors (KT5720 or KT5823) only partially inhibited the increased Et‐1 mRNA levels in strain‐treated cells. EGTA strongly inhibited the Et‐1 gene expression. The intracellular calcium chelator BAPTA/AM also showed an inhibitory effect on Et‐1 mRNA levels. We conclude that mechanical strain can stimulate the secretion of Et‐1 from ECs by increasing Et‐1 mRNA levels via transcription, and that this gene induction is mediated predominantly via the PKC pathway and requires extracellular Ca2+. This strain‐induced Et‐1 gene expression in ECs may contribute to the regulation of vascular tone and structure in normal and pathological states of the cardiovascular system. © 1995 Wiley‐Liss, Inc.
Differential effects of a heparin antagonist (hexadimethrine) or chlorate on amphiregulin, basic fibroblast growth factor, and heparin‐binding EGF‐like growth factor activity
Authors: Cook Paul W. , Ashton Nina M. , Karkaria Cyrus E. , Siess Don C. , Shipley Gary D. 
Published: February 2005

AbstractAmphiregulin (AR) and heparin‐binding EGF‐like growth factor (HB‐EGF) are two recently identified members of the EGF family. Both AR and HB‐EGF share with EGF the ability to interact with the type‐1 EGF receptor; however, AR and HB‐EGF differ from EGF in that both of these mitogens bind to heparin while EGF does not. To determine whether interactions with heparin‐like molecules on the cell surface influence binding of AR and HB‐EGF with EGF receptors and the subsequent mitogenic activity exerted by these growth factors, murine AKR‐2B and Balb/MK‐2 cells were treated with either an inhibitor of proteoglycan sulfation (chlorate) or a heparin antagonist (hexadimethrine). As expected, neither treatment significantly altered the specific binding of 125I‐EGF on AKR‐2B cells. Interestingly, treatment with either chlorate or hexadimethrine inhibited the ability of AR to compete with 125I‐EGF for cell surface binding and also attenuated AR‐mediated DNA synthesis. Thus, as has been suggested for other heparin‐binding growth factors such as basic fibroblast growth factor (bFGF), the interaction of AR with an EGF‐binding receptor appears to be facilitated by interaction with cell‐associated sulfated glycosami‐noglycans or proteoglycans. Unexpectedly, however, neither chlorate nor hexadimethrine treatment caused an inhibition of HB‐EGF‐induced mitogenic activity. Chlorate treatment did not significantly alter the ability of HB‐EGF to compete with 125I‐EGF for cell surface binding sites, however, heparin and hexadimethrine reduced the ability of HB‐EGF to compete for 125I‐EGF binding. These results suggest that, in AKR‐2B cells, HB‐EGF may mediate its mitogenic response at least in part through a receptor which appears to be selective for HB‐EGF and permits HB‐EGF‐mediated mitogenic responses in the presence of hexadimethrine or heparin. Finally, hexadimethrine inhibited the specific binding and mitogenic activity of bFGF, suggesting that this cationic polymer can function as an antagonist of heparin‐binding mitogens other than AR. © 1995 Wiley‐Liss, Inc.
Patterns of cyclic AMP‐dependent protein kinase gene expression during ontogeny of the murine palate
Authors: Greene Robert M. , Lloyd Martha R. , Uberti Michelle , Nugent Paul , Pisano M. Michele 
Published: February 2005

AbstractNormal growth and differentiation of embryonic palatal tissue depends on regulated levels of intracellular cAMP. Cyclic AMP‐dependent protein kinases (PKA) act to mediate the biological activities of cAMP. PKA isozyme protein profiles demonstrate a clear pattern of temporal alterations in embryonic palatal tissue during its development. In order to ascertain the molecular basis for changing PKA isozyme profiles during palatal ontogeny, the spatial and temporal expression of mRNAs for regulatory (Rlα, Rllα, and Rllβ) and catalytic (Cα) subunits of PKA was examined. RNA extracted, from murine embryonic palatal tissue (days 12–14 of gestation) was examined by Northern blot analysis. Significant levels of constitutively expressed Rlα and Cα mRNA were seen on all days of gestation examined. Rlα transcripts were substantially less abundant in palate mesenchymal cells in vitro than in palatal tissue in vivo. Levels of Rllα and Rllβ mRNA were highest on gestational day (GD) 12, a period characterized by pronounced palatal tissue growth. In addition, patterns of tissue distribution of Rllβ, not previously described, were examined in the developing embryonic palate. A dramatic developmental shift in tissue distribution of Rllβ was seen. The isozyme was evenly distributed between palatal epithelial and mesenchymal cells on GD 12 but by GD 14, Rllβ was predominantly localized to palatal epithelial cells. Direct activation of adenylate cyclase with forskolin in murine embryonic palate mesenchymal (MEPM) cells resulted in an increase in Rllα mRNA levels but had no effect on steady state levels of Rllβ or Cα mRNA. In addition, elevation of intracellular levels of cAMP resulted in a shift in the transcriptional profile of Rlα mRNAs. Results of this study document specific patterns of expression for the genes encoding the various cAMP‐dependent protein kinase regulatory and Cα subunits in murine embryonic palatal tissue. In addition, we have demonstrated adaptational changes of this kinase in MEPM cells in response to conditions of increased intracellular levels of cAMP. © 1995 Wiley‐Liss, Inc.