bluuebunny/crossref_metadata_2025_split huggingface dataset:
Iron and copper requirements for proliferation and differentiation of a human promyelocytic leukemia cell line (HL‐60)
Authors: Sergeant Susan , Johnson W. Thomas 
Published: February 2005

AbstractTrace mineral deficiencies tend to have profound effects on the integrity of formed blood elements. Anemia and neutropenia are commonly seen in copper (Cu) deficiency. We therefore developed a serum‐free medium to examine the trace mineral requirements, in particular iron and Cu, for proliferation and retinoic acid (RA)‐induced differentiation of HL‐60 cells. This defined medium (DFM) was composed of Iscove's Modified Dulbecco's Medium (IMDM) supplemented with insulin and human apo‐transferrin (each at 5 μg/ml) and 1.4 μM FeSO4. The iron concentration range for optimal cellular proliferation was narrow (2–3 μM). HL‐60 cells could be maintained in DFM for 15 passages with a doubling time of 38–40 hr. The Cu content of IMDM was very low. Thus, by the fourth passage in DFM, the activity of cuproenzymes (cytochrome c oxidase, CCO; and copperzinc superoxide dismutase, CuZnSOD) began to decline. Supplementation of DFM with CuSO4 (50 nM) restored enzyme activities. Treatment of cells with a Cu chelator (tetrathiomolybdate, 1 μM) rapidly reduced the activities of both CCO and CuZnSOD. Over the Cu concentration range examined (5–350 nM), Cu supplementation had little effect on HL‐60 proliferation. Cell retained the ability to differentiate along the granulocytic pathway when treated with RA, but seemed to be less sensitive to the inducing agent except at the highest concentration tested (1 μM). This decreased sensitivity to RA did not seem to be related to the Cu status of the cells but rather to the absence of a component of serum. Indeed, cells grown in DFM regained their sensitivity to RA when allowed to differentiate in IMDM with 5% serum. These data indicate that the processes of growth and terminal differentiation in HL‐60 cells are not greatly influenced by Cu. Thus, it seems likely that the insult resulting in neutropenia which is associated with Cu deficiency may occur earlier than the promyelocytic stage. However, the possibility that the mechanisms contributing to neutropenia may be unrelated to primary defects in the biochemistry of neutrophil maturation cannot be ruled out. © 1995 Wiley‐Liss, Inc.
Antiprogestins inhibit growth and stimulate differentiation in the normal mammary gland
Authors: Li Minglin , Spitzer Eva , Zschiesche Wolfgang , Binas Bert , Parczyk Karsten , Grosse Richard 
Published: February 2005

AbstractAntiprogestins possess a potent antitumor activity in hormone‐dependent experimental breast cancer models. Though the underlying mechanism is not clear, induction of functional differentiation seems to be a major event. This study attempts to test directly for antiproliferative and differentiation promoting activities of antiprogestins on the normal mammary gland. To this end, whole organ cultures of mammary glands from estradiol/progesterone‐primed virgin mice maintained in a serum‐free medium with aldosterone, prolactin, insulin, and hydrocortisone were exposed to the antiprogestin ZK114043. A 4‐day treatment of organ cultures led to a strong inhibition of epithelial DNA synthesis. In parallel, ZK114043 caused alveolar cells to acquire a more differentiated phenotype distinguished by secretory active alveoli composed of single cell layers with increased fat droplet accumulation and enhanced expression of the milk proteins b̃‐casein and whey acidic protein (WAP). Particularly strong effects were found on the expression of mammary‐derived growth inhibitor (MDGI). Both half‐maximal inhibition of epithelial DNA synthesis and stimulation of MDGI mRNA expression were found at about 5 ng/ml of ZK114043. Presence in the medium of 5 m̈g/ml hydrocortisone rendered antiglucocorticoid effects of ZK114043 highly unlikely. Furthermore, prevention of action of ZK114043 by the progesterone agonist R5020 and ZK114043 stimulated expression of b̃‐casein and MDGI mRNA in cultured glands of 10‐week‐old unprimed virgin mice suggest a progesterone receptor‐mediated mechanism of antiprogestin action. Two other antiprogestins, Mifepristone and Onapristone, likewise stimulated MDGI expression. The data provide direct evidence that antiprogestins act like a differentiation factor in the normal mammary gland. © 1995 Wiley‐Liss, Inc.
Bone marrow stromal cells selectively stimulate the rapid expansion of lineage‐restricted myeloid progenitors
Authors: Kameoka Jun‐Ichi , Yanai Nobuaki , Obinata Masuo 
Published: February 2005

AbstractBone marrow stromal cells serve hematopoietic microenvironments where different blood cells are controlled in their growth and differentiation. To characterize functions of stromal cells, 33 bone marrow stromal cells including preadipocytes, endothelial cells, and fibroblasts were established from transgenic mice harboring temperature‐sensitive SV40 T‐antigen gene and their selective stimulatory abilities to support large colony formation of lineage‐specific hematopoietic progenitor cells (erythroid, monocyte/macrophage, granulocyte, and monocyte‐granulocyte) were examined. Among established stromal cells, 27 clones showed erythropoietic stimulatory activity in the presence of erythropoietin. On myeloid progenitors, the stromal cells showed lineage‐restricted stimulatory activity and a reciprocal relationship was observed between granulocyte formation and macrophage formation, but these activities were not dependent on the amount of produced colony‐stimulating factors (CSFs). Our present study with many stromal cells established from bone marrow indicated that each stromal cell in the bone marrow may provide the preferable microenvironment for a rapid expansion of the lineage‐restricted progenitor cells in combination with CSFs. © 1995 Wiley‐Liss, Inc.
Regulation of insulin‐like growth factor (IGF) binding protein‐2 synthesis and degradation by platelet‐derived growth factor and the IGFs is enhanced by serum deprivation in vascular smooth muscle cells
Authors: Cohick Wendie S. , Gockerman Amy , Clemmons David R. 
Published: February 2005

AbstractGrowth factors such as platelet‐derived growth factor (PDGF) and insulin‐like growth factor (IGF‐1) stimulate proliferation and migration of vascular smooth muscle cells (SMC). IGF‐l bioactivity is modulated by high‐affinity binding proteins (IGFBP) which are important regulators of these processes. Procine vascular SMC synthesize IGFBP‐2 and IGFBP‐4 in vitro. In the present study, levels of IGFBP‐2 in conditioned media (CM) were increased approximately 1.6 to 2.2‐fold when cells were exposed to PDGF (20 ng.ml) or insulin (5 μg/ml) for 24 hr following a 24 hr incubation in serum‐free media, or following a 72 hr exposure to either growth factor. Similar increases in IGFBP‐2 mRNA levels were observed. Exposure of cells to PDGF for 24 hr without prior serum deprivation resulted in smaller (47 ± 11%) increases in IGFBP‐2 protein levels but failed to alter mRNA levels. IGF‐1, FGF‐b̃ and EGF failed to increase IGFBP‐2 using either experimental paradigm. In contrast, IGFBP‐2 protein levels were consistently decreased (75 ± 14%) after 72 hr of exposure to IGF‐II without corresponding decreases in IGFBP‐2 mRNA levels. Immunoprecipitation of [35S] methionine‐labeled IGFBP‐2 indicated that this decrease was not due to a decrease in synthesis of IGFBP‐2. Immunoblot analysis of CM from cells treated with IGF‐II indicated that the decrease in intact protein corresponded with an increase in two non‐IGF binding IGFBP‐2 fragments of 22 and 14 kD. Increased abundance of these fragements was also observed following IGF‐I exposure, although corresponding decreases in intact IGFBP‐2 were not usually observed. The relative abundance of these fragments did not appear to be affected by treatment with PDGF or insulin. In contrast to IGFBP‐2, regulation of the levels of IGFBP‐4 in CM did not appear to be altered by serum deprivation. Insulin consistently increased IGFBP‐4 mRNA and protein levels under all situations. PDGF tended to increase IGFBP‐4 protein levels, although this effect was less consistent and not as great as the increase observe with insulin. Treatment with IGF‐I or ‐ll consistently decreased IGFBP‐4 levels in CM but tended to increase their mRNA levels under all situations. These data indicate that insulin, PDGF, and the IGFs regulate both IGFBP‐2 and IGFBP‐4. While PDGF and insulin stimulate IGFBP‐2 and 4 synthesis, the IGFs appear to activate protease(s) which regulate IGFBP‐2 and ‐4 levels post‐translationally. The regulation of IGFBP‐2 levels by each of these mechanisms appears to be amplified by serum deprivation, but this is not observed with IGFBP‐4. © 1995 Wiley‐Liss, Inc.
Transforming growth factor‐beta isoform expression in insulin‐like growth factor stimulated myogenesis
Authors: Bosche William J. , Ewton Daina Z. , Florini James R. 
Published: February 2005

AbstractTransforming growth factor betas (TGF‐b̃s) are the defining members of a superfamily of small proteins that are involved in the regulation of development and morphogenesis in a wide array of systems. Previous studies have demonstrated that TGF‐b̃s both inhibit and, under specialized conditions, induce the differentiation of myoblasts. TGF‐b̃s have been shown to be secreted by mouse C2C12 myoblast cultures undergoing differentiation. Insulin‐like growth factors (IGFs) have also been shown to be secreted by myoblasts and to induce myogenesis. This study characterizes the effects of IGF treatment on the expression and secretion of TGF‐b̃s in the IGF‐sensitive L6A1 myoblast line. IGF downregulated the expression of TGF‐b̃3 in a concentration‐dependent manner at 24 and 48 hours; TGF‐b̃1 was not sensitive to IGF treatment at 24 hours but was downregulated by IGFs at 48 hours. This downregulation was mediated by the type I IGF receptor and modulated by IGF binding proteins secreted by the myoblasts. Some reexpression of TGF‐b̃1 and TGF‐b̃3 mRNAs was observed after extensive morphological differentiation had occurred. These results support the hypothesis that IGFs act through the IGF type I receptor as part of a concerted mechanism to modulate expression of the TGF‐b̃ genes, as part of a coordinated set of changes associated with terminal myogenic differentiation. © 1995 Wiley‐Liss, Inc.
Characterization of a signal generated by oxidation of protein thiols that activates the heat shock transcription factor
Authors: Freeman Michael L. , Borrelli Michael J. , Syed Khalid , Senisterra Guillermo , Stafford Diane M. , Lepock James R. 
Published: February 2005

AbstractThe diazenecarbonyl derivative, diamide, was used to produce nonnative protein disulfides in Chinese hamster ovary cells in order to characterize the events that occur during thiol oxidation‐induced denaturation that trigger induction of Hsp 70. We limit the term protein denaturation to a process involving a conformational rearrangement by which the ordered native structure of a protein changes to a more disordered structure. Protein thiol oxidation resulted inimmediate destabilization of proteins, as assessed by differential scanning calorimetry (DSC). The DSC profile indicated both a decrease in the onset temperature for detection of denaturation and destabilization of a class of proteins with an average transition temperature (Tm) of 60°C. Concomitant with destabilization was an increase in proteins associated with isolated nuclei. Thiol oxidation also induced heat shock transcription factor (HSF) binding activity, however, this was nearly undetectable immediately following diamide treatment: maximum activation occurred 3 hr following exposure. In contrast, heat shock denatured thermolabile proteins which exhibited a Tm of 48°C. Heat shock also resulted in a rapid increase in proteins associated with isolated nuclei and produced immediated and maximum activation of HSF binding. The accumulation of Hsp and Hsc 70 mRNA following thiol oxidation reflected the delay in HSF binding. Acquisition of HSF binding activity occurred immediately if diamide‐treated cells were subsequently exposed to a heat shock, indicating that HSF was not inactivated by the diamide treatment. Ostensibly, the cellular system for detecting denatured/abnormal proteins failed to immediately recognize the signal generated by thiol oxidation. These results suggest that at least two processes are involved in the induction of Hsp 70 by nonnative disulfide bond formation: destabilization of protein structure resulting in denaturation and recognition of denatured protein. © 1995 Wiley‐Liss, Inc.
Interaction of CD44 variant isoforms with hyaluronic acid and the cytoskeleton in human prostate cancer cells
Authors: Welsh Catherine F. , Zhu Dan , Bourguignon Lilly Y. W. 
Published: February 2005

AbstractCD44 is a glycosylated adhesion molecule which may undergo alternative splicing of 10 possible exons to generate variant isoforms. A number of CD44 variant isoforms expressed by tumor cells have been correlated with metastatic and proliferative behavior. In this study, we have characterized CD44 isoform expression on three prostate cancer cell lines: ALVA‐31, PPC‐1, and LNCaP. Using reverse transcriptase‐polymerase chain reaction, we have found that ALVA‐31 and PPC‐1 cells express multiple CD44 isoforms, including CD44s (standard form), CD44E (epithelial form), and an exon 14‐containing form. In addition, two smaller forms have been detected: one using an alternative donor splice site within exon 5, and a novel form omitting exon 5 entirely. The CD44 isoforms expressed by ALVA‐31 and PPC‐1 cells appear to be preferentially located on the cell surface. By contrast, LNCaP cells do not express any of the CD44 forms at the RNA or protein level. Both PPC‐1 and ALVA‐31 cells display tumorigenesis and invasiveness in nude mice, whereas LNCap cells exhibit a less malignant phenotype, suggesting a correlation between CD44 variant (CD44v) expression and aggressive prostate tumor behavior. Functional characterization reveals that CD44 mediates prostate cell adhesion to extracellular hyaluyronic acid (HA). In addition, the CD44 cytoplasmic domain binds specifically to ankyrin, a membrane cytoskeletal protein. Double immunofluorescence labeling and confocal microscopic analyses indicate that HA binding induces the HA receptor (i.e., CD44) to form capped structures. Importantly, intracellular ankyrin is preferentially accumulated underneath HA receptor‐capped structures. These results suggest that cytoskeletal proteins such as ankyrin are closely associated with CD44‐mediated signaling events induced by HA. Finally, HA‐mediated transmembrane interactions between CD44 isoforms and cytoskeletal proteins (i.e. ankyrin) may play a pivotal role in regulating tumor cell behavior during human prostate cancer development. © 1995 Wiley‐Liss, Inc.
Regulation of parathyroid hormone/parathyroid hormone‐related protein receptor expression by osteoblast‐deposited extracellular matrix in a human osteoblast‐like cell line
Authors: Hausmann S. , Law F. M. K. , Bonjour J.‐Ph. , Feyen J. , Rizzoli R. 
Published: February 2005

AbstractParathyroid hormone (PTH) receptors and the biological response to PTH in osteoblasts have been shown to be influenced by glucocorticoids, growth factors, cytokines or PTH itself. Furthermore, components of extracellular matrix (ECM) appear to regulate the response to PTH as well. We investigated the effects of osteoblast‐deposited ECM on PTH‐related protein (PTHrP)‐stimulated cAMP production, PTHrP binding and PTH/PTHrP receptor mRNA in the human osteoblast‐like cell line SaOS‐2. ECM was laid down by the human osteoblastic cell line MG‐63. At confluence, maximal cAMP stimulation induced by 100 nmol/l PTHrP (1–34) was decreased in SaOS‐2 cells grown on ECM as compared with cultures on plastic dishes, without any change in PTHrP concentration producing half‐maximal stimulation. In contrast, cAMP production stimulated by PGE2 was increased in cells on ECM. Saturable 125I‐PTHrP binding (as evaluated by Scatchard plot analysis) was markedly diminished in cells grown on ECM (5,600 ± 2,010 vs. 20,700 ± 1,710 binding sites/cell, × ± S.E.M., P < 0.01, n = 4 experiments), without any significant change in affinity (1.3 ± 0.4 vs. 2.5 ± 0.5 nmol/l (NS), in cells on ECM and plastic, respectively). This apparent decrease in membrane receptor density was associated with markedly lower steady state PTH/PTHrP receptor mRNA levels as assessed by Northern blot analysis (ECM/control: 0.4 ± 0.1). A difference in PTH/PTHrP receptor mRNA levels between cells on ECM or on plastic dishes was detectable by 8 hours, but not by 4 hours, after seeding the cells at high density. By 24 hours after plating, PTH/PTHrP receptor mRNA levels were maximally decreased in cells on ECM. These results in the human osteoblast‐like cell line SaOS‐2 indicate that PTH/PTHrP receptors are down‐regulated by growth on ECM. Thus, attachment of bone cells to bone surface could influence differentiation and function of osteoblasts. © 1995 Wiley‐Liss Inc.
Regulation of DNA methyltransferase during differentiation of F9 mouse embryonal carcinoma cells
Authors: Teubner Barbara , Schulz Wolfgang A. 
Published: February 2005

AbstractDNA becomes demethylated when F9 mouse embryonal carcinoma cells differentiate into parietal endoderm. DNA methyltransferase (DNA‐MTase) activity decreased by 50% during 1 week of differentiation. The level of DNA‐MTase mRNA was also diminished accordingly, but the transcription rate of the DNA‐MTase gene measured by run‐on transcription was essentially unchanged, indicating regulation of DNA‐MTase expression at a posttranscriptional step. The decline of DNA‐MTase mRNA paralleled that of histone H3 mRNA in accord with the notion that DNA‐MTase is preferentially expressed in the S phase of the cell cycle. Since DNA‐MTase expression decreases in parallel with DNA synthesis, DNA demethylation during differentiation of F9 cells appears not to be due to limited expression of DNA‐MTase. However, the plasmid pAFP7000CAT, α‐fetoprotein (AFP), which is strongly de novo methylated when transfected into F9 stem cells became only weakly methylated after transfection into the F9 parietal endoderm derivative P1, indicating that the activity of DNA‐MTase within parietal endoderm cells is more strongly diminished than is apparent from measurements of mRNA amounts and of overall DNA‐MTase activity in vitro. The discrepancy between DNA‐MTase expression and its actual activity within the cell indicates the existence of a novel mechanism controlling the activity of DNA‐MTase. © 1995 Wiley‐Liss, Inc.
Cell cycle variation of Hsp70 levels in HeLa cells at 37°C and after a heat shock
Authors: Hang Haiying , He Liusheng , Fox Michael H. 
Published: February 2005

AbstractThe expression of the 72 kD inducible heat shock protein (hsp72) has been reported to be cell cycle associated in unheated, synchronized HeLa cells. In this study, flow cytomerty was used to investigate hsp72 levels through the cell cycle in HeLa cells by dual labeling with propidium iodide and antibodies against hsp72. The entire cell cycle distribution of hsp72 could be measured in a single sample of asynchronously growing cells. For unheated cells, the level of hsp72 increased about 30% from G1 to S phase, with about a 65% increase in G2/M, probably due to cell size differences. Neither mitotic selection nor serum stimulation induced a higher level of hsp72 than in the control cells. Western blot analysis of hsp72 from Hoechst‐stained cells sorted from G1, mid‐S, or G2/M showed that G1 cells had the lowest level of hsp72, with about a 30% increase in S phase and a 60% increase in G2/M, in good agreement with the flow cytometry results. These data conflict with previous reporty by other laboratories which showed a 3‐fold higher level of hsp72 in S phase than in G1 or G2. In contrast, heat shock (both acute and chronic) led to a non‐uniform increase in hsp72 through the cell cycle. Most cells in mid S phase had high levels of hsp72, and a larger range in the levels of hsp72 were found in G1 and late S/G2/M phase cells. © 1995 Wiley‐Liss, Inc.
Thrombin activates mitogen‐activated protein kinase in primary astrocyte cultures
Authors: Bhat Narayan R. , Zhang Peisheng , Hogan Edward L. 
Published: February 2005

AbstractThrombin is known to evoke numerous inflammatory and proliferative responses in a wide variety of its target cells. Recent studies have demonstrated morphoregulatory and mitogenic effects of thrombin on astroglial cells (astrocytes). The present study deals with thrombin‐induced activation of mitogen‐activated protein (MAP) kinase in primary cultures of rat astrocytes. Treatment of serum‐starved astrocytes with thrombin resulted in a rapid activation of tyrosine (Tyr) phosphorylation of a set of proteins including a prominent one with a molecular mass of 42 kDa (p42). The identity of p42 with MAP kinase was confirmed by MAP kinase‐immunoreactivity of isolated [i.e., immunoprecipitated with anti‐phosphotyrosine (PY) antibodies] p42 and by increased myelin basic protein (MBP) kinase activity present in MAP kinase immunoprecipitates of thrombin‐treated cultures. Pertussis toxin (PTX) pretreatment failed to inhibit thrombin stimulation of p42 phosphorylation, indicating the lack of involvement of PTX sensitive G proteins in the mechanism of activation of MAP kinase by thrombin. Chronic exposure of cultures to phorbol 12‐myristate 13‐acetate to down‐regulate PKC resulted in an attenuation of thrombin‐induced p42 Tyr phosphorylation, although H‐7, a known PKC inhibitor, failed to block thrombin effect. However, staurosporine, a nonspecific protein kinase inhibitor, prevented the activation of p42 phosphorylation. It is concluded that thrombin induces MAP kinase activation in astrocytes by a mechanism involving a staurosporine‐sensitive pathway. © 1995 Wiley‐Liss, Inc.
Chemoattractant‐induced activation of vaculoar H+ pumps and of an H+‐selective conductance in neutrophils
Authors: Nanda Arvind , Grinstein Sergio 
Published: February 2005

AbstractUpon binding to their receptors on the surface of neutrophils, chemotactic peptides elicit a burst of metabolic activity. The excess acid generated by this burst must be rapidly extruded in order to maintain intracellular pH and preserve normal microbicidal responses. Recently, H+‐pumping vacuolar‐type ATPases (V‐pumps) and a H+‐selective conductance were described in the membrane of neutrophils. However, these systems are virtually quiescent in resting cells. In this report, we analyzed whether the V‐pumps and the conductance become active and contribute to pH regulation following cell activation by chemoattractants. Formyl‐Met‐Leu‐Phe (fMLP) was found to stimulate V‐pumps, as assessed by the appearance of bafilomycin‐sensitive H+ extrusion. Concomitantly, the chemoattractant also activated the H+ conductance, detected as a voltage‐dependent and Zn2+‐sensitive net H+ efflux. In both cases, activation was prevented by treatment with competing antagonistic peptides or with pertussis toxin, implying mediation by a receptor copuled to a heterotrimeric G protein. The signalling pathways downstream of the G proteins were also investigated. Stimulation of neither th V‐pump nor the conductace required activation of protein kinase C. An elevation of cytosolic calcium ([Ca2+]i) comparable to that induced by fMLP did not suffice to trigger either transporter. Moreover activation of the conductance remained unaffected when the chemoattractant‐induced increase in [Ca2+]i was precluded. In contrast, stimulation of the V‐pump was substantially (∞50%) depressed when [Ca2+]i was prevented from rising. Tyrosine phosphorylation of several polypeptides accompanies stimulation by fMLP. Prevention of phosphotyrosine accumulation resulted in a pronounced inhibition of H+‐pumping and of the H+ conductance. Together, these data indicate that engagement of surface receptors by chemotactic peptides can lead to stimulation of two voltage‐sensitive pH regulatory pathways, a pump and a conductance, by a pathway that requires tyrosine phosphorylation. Both pathways are capable of sizable H+ extrusion, thereby contributing to pH regulation during the metabolic burst. © 1995 Wiley‐Liss Inc.