bluuebunny/crossref_metadata_2025_split huggingface dataset:
Embryonic stem cells and in vitro hematopoiesis
Authors: Snodgrass H. Ralph , Schmitt Regina M. , Bruyns Eddy 
Published: May 2005

AbstractTo study hematopoietic differentiation a variety of in vitro systems have been established using hematopoietic precursors derived from various explanted adult and fetal tissues. In this prospective we describe and discuss the potential of a novel system for studying the earliest stages of hematopoietic development. In addition, some of the applications of this system as a unique in vitro model for studying other developmental systems are discussed. Murine embryonic stem cells (ESC), which are totipotent and can be maintained undifferentiated indefinitely in vitro, have the capacity to differentiate in vitro into hematopoietic precursors of most, if not all, of the colony forming cells found in normal bone marrow. This potential can be exploited to study the control of the early stages of hematopoietic induction and differentiation. Recent results have indicated that there is a strong transcriptional activation, in a well defined temporal order, of many of the hematopoietically relevant genes. Examples of the genes expressed early during the induction of hematopoiesis include erythropoietin (Epo) and its receptor as well as the Steel (SI) factor (SLF) and its receptor (c‐kit). Several other genes, including CSF‐1, IL‐1, and G‐CSF were expressed during the later stages of hematopoietic differentiation. Contrasting with these observations, IL‐3 and GM‐CSF were not expressed during the first 24 days of ES cell differentiation suggesting that neither factor is necessary for the induction of hematopoietic precursors. Although these studies are just beginning, this system is easily manipulated and gives us an approach to understanding the control of the induction and differentiation of the hematopoietic system in ways not previously possible.
Expression of glucose transporters (GLUT 1 and GLUT 4) in primary cultured rat adipocytes: Differential evolution with time and chronic insulin effect
Authors: Hajduch Eric J. , Guerre‐Millo Michèle C. , Hainault Isabelle A. , Guichard Colette M. , Lavau Marcelle M. 
Published: May 2005

AbstractWe previously reported that in cultured adipose cell lines insulin increased selectively the expression of Glut 1, in contrast to in vivo regulation where variations in insulinemia have been shown to affect only GLUT 4. We have addressed here the question of the long‐term regulation of GLUT 1 and GLUT 4 in fat cells by using primary cultures of rat adipocytes. Epididymal fat cells were isolated by collagenase and cultured 4 days in DMEM supplemented with BSA 1%, FCS 1%, and glucose 10 mM. GLUT 1 and GLUT 4 proteins were assessed in total cellular membranes by Western blotting, using specific antibodies against their respective C‐terminal peptides. GLUT 1 steadily increased over culture time to reach at day 3, a level 3‐fold higher than the initial value. In contrast, GLUT 4 decreased sharply and stabilized at day 3, at 30% of the initial value. The changes in GLUT 1 and GLUT 4 mRNAs with culture time were parallel to changes in the corresponding proteins, suggesting a pre‐translational level of regulation. The expression of the lipogenic enzyme, fatty acid synthetase (FAS), highly expressed in fat cell, decreased over time following a pattern closely parallel to that of GLUT 4. Chronic exposure to insulin added at day 2 had no effect on GLUT 4 expression but increased the expression of GLUT 1 and FAS by 70% and 36%, respectively. Glucose consumption was stable over 4 days of culture, while lactate production increased from 24 to 36% of glucose utilization, in agreement with the loss in FAS. Glucose consumption increased only slightly with insulin (160%), in good keeping with the low levels of expression of both GLUT 4 and FAS in these cultured cells. These data indicate that culture alters oppositely the expression of GLUT 1 and GLUT 4 in rat adipocytes and suggest that factor(s) other than insulin predominate in their regulation in vivo.
Prolyl endopeptidase purified from granulomatous inflammation in mice
Authors: Nozaki Yukinori , Sato Naoyoshi , Iida Toshihiro , Hara Kenji , Fukuyama Kimie , Epstein William L. 
Published: May 2005

AbstractActivity of prolyl endopeptidase (EC 3.4.21.26) which hydrolyses the Pro7‐Phe8 bond in angiotensin II has been found to elevate in experimentally produced granulomatous inflammation in liver and skin. We purified the enzyme 1,536‐fold by 6 steps from murine hepatic granulomas. The purified enzyme has a molecular weight of 79 kDa and physiocochemical properties equivalent to those previously reported for prolyl endopeptidase purified from other sources. By HPLC analysis, the cleavage of Phe8‐Leu10 and Phe8 from angiotensin I and II, respectively, was detected and quantified. Monospecific IgG was prepared from serum of rabbits injected with purified enzyme. Concentration of the enzyme was immunohistochemically detected in cells which form granulomatous organization, but not in inflammatory cells surrounding the foci. The antibody, however, cross reacted with the enzyme in adjacent liver cells and weakly stained their cytoplasm. The findings indicate that this enzyme, in addition to angiotensin converting enzyme, may serve as a useful biochemical marker for granulomatous tissue reactions.
Modification of blood group A antigen expression in a pancreatic cancer cell line (PC‐1) by inhibitors of N‐glycan processing
Authors: Hirota Masahiko , Mogaki Masatoshi , Pour Parviz M. , Chaney William G. 
Published: May 2005

AbstractPancreatic adenocarcinomas induced in Syrian hamsters by treatment with N‐nitrosobis(2‐oxopropyl) amine express blood group A antigen, which is absent in normal pancreatic cells. On membrane glycoproteins purified from tumors, blood group A antigen has been found to be expressed on multiantennary Asn‐linked complex glycans. In this study, we investigated the effect of inhibitors of Asn‐glycan processing on blood group A antigen bearing glycan structures in a cell line (PC‐1) established from a primary induced pancreatic cancer. Expression of blood group A antigen on cells and in membrane preparations was blocked by treatment with 1‐deoxymannojirimycin, an inhibitor of mannosidase I, but was retained after treatment with swainsonine, an inhibitor of mannosidase II. However, swainsonine treatment altered the glycan structure associated with blood group A antigen from an endoglycosidase H resistant type to a sensitive type, indicating that the blood group A structure might shift from a complex type to a hybr‐id type glycan by this treatment. These results demonstrate that Asn‐linked glycans carry the major blood group A antigens in PC‐1 cells. © 1992 Wiley‐Liss, Inc.
Evidence for the presence of complex high‐molecular mass N‐linked oligosaccharides in intranuclear gylcoproteins from hela cells
Authors: Codogno Patrice , Bauvy Chantal , Sève Annie‐Pierre , Hubert Michèle , Ogier‐Denis Eric , Aubery Michèle , Hubert Jean 
Published: May 2005

AbstractNonhistone proteins were extracted in 0.4 M NaCl from membrane‐depleted nuclie of HeLa cells grown in the presence or the absence of [5,6‐3H]fucose. Control experiments strongly suggest that most extracted proteins were needed nuclear components. Several protiens, present in the 0.4 M NaCl nuclear extract, with Mr ranging from 35,000 to 115,000 were identified on Western bolts as fucosylated glycoproteins owing to their binding to the fucose‐specific lectin, Ulex eruopeus agglutinin l. Results of experiments involving mild alkaline treatment and peptide N‐glycosidase F digestion showed that the carbohydrate moieties of these fucosylated nuclear glycoproteins were N‐linked to the polypeptide backbone. Analysis of the N‐glycans revealed the presence of two populations of sialylated oligosaccharides on the basis of their relative molecular masses. The sensitivity of the High‐Mr oligoaccharides to endo‐β‐galactosidase and their incorporation of [3H]glucosamine suggest that they could contain repeating N‐acetyllactosamine units [3H]Fucose incorporated into nuclei was confined to the nucleoli, as judged by autoradiography of sections cut through cells grown in the presence of [3H]fuose. Electron microscopy autoradiography showed that the fibrillar centers were never labeled, while silver grains observed on the dense and the granular components of nucleoli. Taking into account of these data most nuclear fucosylated glycoproteins extracted in 0.4 M NaCl might be nucleolar ribonucleoprotiens. © 1992 Wiley‐Liss, Inc.
Interaction of MAR‐sequences with nuclear matrix proteins
Authors: Ivanchenko Maria , Avramova Zoya 
Published: May 2005

AbstractThe recent discovery of DNA sequences responsible for the specific attachment of chromosomal DNA to the nuclear skeleton (MARs/SARs) was an important step towards our understanding of the functional and structural organization of eukaryotic chromatin [Mirkovitch et al.: Cell 44:273–282, 1984; Cockerill and Garrard: Cell 44:273–282, 1986]. A most important question, however, remains the nature of the matrix proteins involved in the specific binding of the MARs. It has been shown that topoisomerase II and histone H1 were capable of a specific interaction with SARs by the formation of precipitable complexes [Adachi et al.; EMBO J 8:3997–4006, 1989; lzaurralde et al.: J Mol Biol 210:573–585, 1989]. Here, applying a different approach, we were able to “visualize” some of the skeletal proteins recognizing and specifically binding MAR‐sequences. It is shown that the major matrix proteins are practically the same in both salt‐ and LIS‐extracted matrices. However, the relative MAR‐binding activity of the individual protein components may be different, depending on the method of matrix preparation.The immunological approach applied here allowed us to identify some of the individual MAR‐binding matrix proteins. Histone H1 and nuclear actin are shown to be not only important components of the matrix, but to be involved in a highly efficient interaction with MAR‐sequences as well. Evidence is presented that proteins recognized by the anti‐HMG antibodies also participate in Mar‐interactions. © 1992 Wiley‐Liss, Inc.
Fluorescence cytometry of microtubules and nuclear DNA during cell‐cycle and reverse‐transformation
Authors: Vergani L. , Gavazzo P. , Facci P. , Diaspro A. , Mascetti G. , Nicolini C. , Arena N. , Gaspa L. 
Published: May 2005

AbstractSynchronized CHO‐K1 cells and their dibutyryl c‐AMP treated counterparts have been characterized by means of static and flow fluorescence cytometry at the level of nuclear DNA and cytoplasmic microtubules.In order to confirm earlier findings on synchronized population, Carnoy fixed and hydrolyzed, several new findings are here reported at the level of single intact cell. The fluorescence intensity of DAPI‐stained glutaraldehyde fixed 2C cells correlates well with the average absorbance of the corresponding Feulgen‐stained cells, thereby appearing also to be a measure of chromatin condensation during the G1 phase.In the early part of G1, the drastic alteration in anti‐β tubulin immunostaining is shown to parallel microtubule depolymerization induced by calcium or colcemide.The known 1–2 h lengthening of the G1 period after reverse‐transformation appears to correlate with a similar delay in the abrupt chromatin decondensation.The above results are discussed in terms of the role of microtubules and nuclear morphometry (and their coupling) in the control of cell cycle progression of transformed vs. fibroblast‐like cells. © 1992 Wiley‐Liss, Inc.
Recombinant murine steel factor stimulates in vitro production of granulocyte–macrophage progenitor cells
Authors: Williams Douglas E. , Foxworthe Diane , Teepe Mark , Eisenman June , Lyman Stewart D , Anderson Dirk 
Published: May 2005

AbstractThe ability of murine Steel factor to promote the in vitro production of granulocyte‐macrophage progenitor cells (CFU‐GM) was examined in short‐term liquid cultures. Bone marrow from C57BL/6J or Sl/Sld mice was placed in culture for seven days with either Steel factor alone or in the presence of IL‐3. CFU‐GM responsive to GM‐CSF, IL‐3, and CSF‐1 were measured in the input population and again after 3 or 7 days in culture. Steel factor alone increased the number of all CUF‐GM types as early as 3 days after culture initiation, with further increases at day 7. This effect was protentiated by the addition of IL‐3. Production of CFU‐GM by C57BL/6J or Sl/Sld marrow was comparable except for enhanced production of CSF‐1 responsive progenitors by Sl/Sld marrow. A recombinant Sld protein was also shown to be equivalent to the wild‐type protein in its capacity to promote CFU‐GM production from normal bone marrow. © 1992 Wiley‐Liss, Inc.
Activin A‐induced diffrentiation in K562 cells is associated with a transient hypophosphorylation of RB protein and the concomitant block of cell cycle at G1 phase
Authors: Sehy David W. , Shao Li‐En , Tsai Wei‐Min , Yu John , Yu Alice L. 
Published: May 2005

AbstractThe human erythroleukemic cell line, K562, can be induced to differentiate by the addition of activin A, a newly purified protein belonging to the TGF‐β1 family. The present studies used flow cytometric cell cycle analysis, indirect immunofluorescence staining of the proliferating cell nuclear antigen (PCNA), and thymidine incorporation assay of cell proliferation to study the effects of activin A on the cell cycle during differentiation in K652 cells. Activin phase. The latter can be observed after approximately 24 hr of incubation with activin A and then disappears after this early stage of induction of differentiation. Cell cycle kinetics analysis using synchronized K562 cells also confirms that in the presence of activin A, K562 cells progress normally through various phases of cell, except that there is prolongation of the G1 phase between 10 to 24 hr of culture. Furthermore, this transient arrest in G1 is correlated with dephosphorylation of a nucleoprotein, the RB gene product, which occurs within 9–24 hr of incubation with activin A; and phosphorylation of RB protein then develops afterward. In addition, these cell cycle‐related events are observed to occur earlier than the accumulation of hemoglobins in K562 cells. It is concluded that transient dephosphorylation of RB protein and prolongation of G1 phase of cell cycle precede and accompany erythroid differentiation caused by activin A and chemical inducers, thus constituting part of the mechanism for induction of differentiation in the erythroleukemia cells. © 1992 Wiley‐Liss, Inc.
Pericellulars substrates of human mast cell tryptase: 72,000 Dalton gelatinase and fibronectin
Authors: Lohi Jouko , Kesksi‐Oja Jorma , Harvima Ilkka 
Published: May 2005

AbstractMigrating cells degrade pericellular matrices and basement membranes. For these purposes cells produces a number of proteolytic enzymes. Mast cells produce two major proteinases, chymase and tryptase, whose physiological functions are poorly known. In the present study we have analyzed the ability of purified human mast cell tryptase to digest pericellular matrices of human fibroblasts. Isolated matrices of human fibroblasts and fibroblast conditioned medium were treated with tryptase, and alterations in the radiolabeled polypeptides were observed in autoradiograms of sodium dodecyl sulphate polyacrylamide gels. It was found that an Mr 72,000 protein was digested to an Mr62,000 form by human mast cell tryptase while the plasminogen activator inhibitor PAI‐1 was not affected. Cleavage of the Mr72,000 protein could be partially inhibited by known inhibitors of tryptase but not by aprotinin, soybean trypsin inhibitor, or EDTA. Fibroblastic cells secreted the Mr 72,000 protein into their medium and it bound to gelatin as shown by analysis of the medium by affinity chromatography over gelatin‐Sepharose. The soluble form of the Mr72,000 protein was also susceptible to cleavage by tryptase. Analysis using gelatin containing polyacrylamide gels showed that both the intact Mr72,000 and Mr62,000 degraded form of the protein process gelatinolytic activity after activation by sodium dodecyl sulphate. Immunoblotting analysis of the matrices revealed the cleavage of an immunoreactive protein of Mr72,000 indicating that the protein is related to type IV collagenase. Further analysis of the pericellular matrices indicated that the protease sensitive extracellular matrix protein fibronectin was removed from the matrix by tryptase in a dose‐dependent manner. Fibronectin was also susceptible to proteolytic degradation by tryptase. The data suggest a role for mast cell tryptase in the degradation of pericellular matrices. © 1992 Wiley‐Liss, Inc.
M‐CSF (monocyte colony stimulating factor) and M‐CSF receptor expression by breast tumour cells: M‐CSF mediated recruitment of tumour infiltrating monocytes?
Authors: Tang Ruoping , Ojeda Mario , Pouillart Pierre , Scholl Susy , Beuvon Frédéric , Mosseri Véronique 
Published: May 2005

AbstractInfiltrating immune cells in 30 primary human epithelial breast tumours were studied using specific anti‐CD3 (T cells), anti‐CD68 (macrophages), anti‐CD57 (NK cells), and an anti‐pan‐B cell antibody (L26). The majority of tumour infiltrating inflammatory cell are T cells (40‐50%) and monocytes/macrophages (15–35%).The macrophage specific chemo‐attractant and growth factor CSF‐1 is detected by immunohistochemical techniques (IHC) at the level of invasive breast cancer cell in 46/50 tumours but not at the level of in‐ situ (pre‐invasive) cancer. A mosaic staining patterns was usually observed with a very high expression in areas of obvious stromal invasion (90%cells positive) and absent or trace staining in intraductal carcinoma. Macrophages and plasma cell are equally intensely positive. In‐situ‐hybridisation experiment confirm the production of CSF‐1 (mRNA) by tumour cells and show the same pattern of expression. Expression of the CSF‐1 receptor protein (fms) was also observed by IHC in 41/48 invasive tumours, albeit at weaker intensities than in tumour infiltrating monocytes/macrophages. A concomitant expression of both CSF‐1 and fms in in‐situ carcinoma was never seen (n = 14). It is therefore proposed that the associated expression of CSF‐1 and its receptor may be linked to the invasive potential of breast cancer, the monocytic infiltrate being an indication of the quantitative importance of CSF‐1 production by the tumour. © 1992 Wiley‐Liss, Inc.
Post‐Transcriptional regulationa of collagenase and stromelysin gene expression by epidermal growth factor and dexamethasone in cultured human fibroblasts
Authors: Delany Anne M. , Brinckerhoff Constance E. 
Published: May 2005

AbstractEpidermal growth factor (EGF) is a ubiquitous fibroblast mitogen which also stimulates the synthesis of the extracellular matrix degrading metalloproteinases, collagenase, and stromelysin. Using primary cultures of human skin fiibroblassts, we show that these metalloproteinase mRNAs are coordinately up‐regrated by EGF; and that dexamethasone, a potent inhibitor of collagenase and stromelysin syntehse, cordinately down‐regulates these EGF‐induced mRNAs. Nuclear run‐on assays showed that EGF increased trasncription of collagenase and stromelysin ∼ 2‐fold over the untreated control, while repression by dexamethasone was difficult to detect. However, steady state mRNA levels were induced ∼ 10‐fold by EGF and co‐treatment with dexamethasone decreased them to below control levels, suggesting modulation of mRNA stability. Thus, we measured the half‐life of these mRNAs using “pulse‐chase” methodology. Typically, the half‐life of EGF‐induced collagenase and stromelysin mRNAs was ∼ 30 h, and co‐treatment inhibitor DRB stabilized EGF‐induced metalloproteinase mRNAs by 30–50%. Additionally, we found that the transcirption inhibitor DRB stabilitzed EGF‐induced metalloproteinase mRNAs, suggesting an mRNA degradation pathway which requires transcription. Thus our data demonstrate that collagenase and stromelysin are coordinately regulated by EGF and by dexamethasone, primarily at the level of metalloproteinase mRNA stability. © 1992 Wiley‐Liss, Inc.
Localization of the ceavage sites on fibronectin following digestion by urokinase
Authors: Gold Leslie I. , Rostagno Agueda , Frangione Blas , Passalaris Tina 
Published: May 2005

AbstractUrokinase (u‐PA) proteolytically cleaves both human plasma (pFn) and cellular (cFn) dimeric fibronectin (Mr 440,000) into four major polypeptides of approximately Mr 210,000, 200,000, 25,000, and 6,000. Amino acid sequence analysis of the polypeptide fragments indicated that the enzymatic cleavage of Fn occurs at two sites: (1) between an arginine/alanine peptide bond located C‐terminal to residue 259; this cleavage liberates the N‐terminal Mr 25,000 fragment and the Mr 210,000 and Mr 200,000 polypeptides derived from the A and B chains of Fn, respectively; and (2) between an arginine/threonine peptide bond located C‐terminal to residue 2,299, thereby yielding an Mr 6,000 dimeric fragment containing the C‐terminal interchain disulfide bonds. Predigestion of Fn with u‐PA increased the molecule's vulnerability to further attack by the enzymes plasmin and cathepsin D. These data provide further biochemical evidence for the proteolytic cleavage of fibronectin by plasminogen activators and substantiate that u‐PA digestion of Fn may be an intial event in the local degradation of the extracellular matrix by malignant cells, possessing elevated levels of these enzymes. © 1992 Wiley‐Liss, Inc.
Estrogen receptor mutations in breast cancer
Authors: Fuqua S. A. W. , Chamness G. C. , McGuire W. L. 
Published: May 2005

AbstractIt is fairly well accepted that the presence of estrogen receptor (ER) identifies those breast cancer patients with a lower risk of relapse and better overall survival [Clark and McGuire, 1988], and the measurement of ER has become a standard assay in the clinical management of breast cancer. Receptor status also provides a guideline for those tumors which may be responsive to hormonal intervention [McGuire 1978; Osborne et al., 1980; Rose et al., 1985]. But only about half of ER‐positive patients will respond to the various hormonal therapies available, and of those who do initially respond, most will eventually develop hormonally unresponsive disease following a period of treatment even though ER is often still present. Loss of ER from initially ER‐positive tumors biopsied again at a later date has been estimated at only 19% [Gross et al., 1984]. Obviously the simple measurement of ER presence by ligand‐binding assays does not provide us with an adequate estimate of the functional state of the receptor.In 1985 Sluyser and Mester hypothesized that the loss of hormone dependence of certain breast tumors may be due to the presence of mutated or truncated steroid receptors that activate transcription even in the absence of hormone [Sluyser and Mester, 1985]. Based on the recent identification of several ER sequence variants in human breast cancer cell lines and tumor specimens, we would now like to propose that some of these identified mutations play a role in receptor dysfunction in vivo, and will review those ER mutations which may prove to be important in breast cancer progression. © 1993 Wiley‐Liss, Inc.