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  • 1
    Electronic Resource
    Electronic Resource
    Methylation of the 5′ flanking sequences of the ribosomal DNA in human cell lines and in a human-hamster hybird cell line (1992)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 50 (1992), S. 357-362 
    Details
    ISSN: 0730-2312
    Keywords: rDNA ; lymphoblastoid ; methylation ; hypermethylation ; DNA ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: In human lymphoplastoid cell line (Z83) in which rDNA genes on chromosomes 22 are amplified but transcribed at a low level, immunocytological studies with antibodies to 5 methylcytidine provided evidence for hypermethylation of the rDNA. The extent of methylation of the 5′ flanking sequence of the ribosomal DNA was examined by comapring the size of restriciton fragments obtained by digestion of genomic DNA with EcoRI and Hpall or EcoRI and Mpsl. Southern blots indicated hypermethylation of the 5′ flanking sequences of many copies of rRNA genes in these cells, but not in a control lymphoblastoid cell line without rDNA amplification. Results obtained with somatic hybird human-hamster cell line, in which the rRNA genes on the single human chromosomes 22 are inactive, showed that only a small fraction of the CCGG sites in the 5′ flanking sequences of the transcriptionally silent rRNA genes in this hybird were methylated. Since inactive rRNA genes can show such a minimal level of methylation, it is likely that the extreme hypermethylation of the apmlified rRNA genes in Z83 association with their inactivation rather than following it. © 1992 Wiley-Liss, Inc.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcb.240500404
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  • 2
    Electronic Resource
    Electronic Resource
    Spontaneous fusion between metastatic mammary tumor subpopulations (1988)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 36 (1988), S. 129-136 
    Details
    ISSN: 0730-2312
    Keywords: hybrid cells ; metastasis ; heterogeneity ; generation of aneuploidy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: This study describes a differential frequency of spontaneous fusion between metastatic and nonmetastatic subpopulations derived from a single mouse mammary tumor. Subpopulations 66, 66cl4 (a variant of 66 which is resistant to both thioguanine and ouabain), 410.4, and 44FTO (a thioguanine-resistant, ouabain-resistant derivative of 410.4) spontaneously metastasize from subcutaneous and mammary fatpad sites. Subpopulations 168, 168FARO (a diaminopurine-resistant, ouabain-resistant derivative of 168), 67, 68H, and 410 do not. The ability of these subpopulation lines to fuse spontaneously in vitro was determined after coculturing a drug-resistant line with a wild-type line in nonselective media. After 16-20 h of coculture, cells were plated in the appropriate media to select for fusion products - either HAT (hypoxanthine, aminopterin, thymidine) plus ouabain or AA (alanosine, adenine) plus ouabain - to determine the number of colony-forming cells (fusion products) present per 104 cells plated. When both subpopulations of the pair in the fusion mixture were metastatic, a significantly greater number of fusion products was recovered than if one or both of the subpopulations in the fusion mixture was nonmetastatic, with one exception: line 410 readily fused with both 66cl4 and 44FTO. Subline 410 was highly metastatic when originally isolated but lost its metastatic competence after a brief time in tissue culture.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcb.240360204
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  • 3
    Electronic Resource
    Electronic Resource
    Assessing tumor drug sensitivity by a new in vitro assay which preserves tumor heterogeneity and subpopulation interactions (1984)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 121 (1984), S. 105-116 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have designed an in vitro assay to assess the influence of tumor subpopulation interactions on drug response. The assay is based upon inhibition of growth of 1 mm3-pieces of tumor embedded in a collagen gel matrix. Tumor growth is quantitated by planimetry of each colony's image, formed with a split image tracing device attached to an inverted microscope. That expansion of the colonies in collagen gel represents growth through cell replication was demonstrated by releasing and counting cell nuclei. Outgrowths from pieces of tumors produced by a series of mouse mammary tumor subpopulation lines expanded in collagen gel at a rate characteristic of each cell line: the growth rate of tumor pieces was similar to that of the corresponding tumor line embedded as a cell bolus of cultured cells, indicating that growth of pieces of tumor is due to the tumor cells rather than to stromal components. When two cell lines were grown together in collagen cultures, interactions affecting growth rate were observed. Both tumor pieces and cell boluses from cultured cells of the relatively homogeneous cell lines displayed similar, characteristic sensitivities to adriamycin (ADR) in the collagen gel assay. Advantages of the collagen assay over cloning assays are (1) preservation of potential cellular interactions which may be important in assessing tumor drug sensitivity; (2) maximization of growth of all cell populations within the tumor, as compared to growth in agar; and (3) reflection of the zonal distribution of different subpopulations within tumors; and (4) simulation of the three-dimensional growth architecture found in vivo.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041210413
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  • 4
    Electronic Resource
    Electronic Resource
    Monoclonal antibodies against plant proteins recognise animal intermediate filaments (1987)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 8 (1987), S. 312-323 
    Details
    ISSN: 0886-1544
    Keywords: plant cytoskeleton ; Chlamydomonas ; anti-IFA ; onion root tip cells ; immunoflurescence ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Four monoclonal antibodies were raised against polypetides present in a highsalt detergent-insoluble fraction from cells of Chlamydomonas reinhardtii. Indirect immunofluorescence microscopy of fibroblasts and epithelial cells grown in culture using these plant antibodies revealed staining arrays identical to those obtained with well characterised antibodies to animal intermediate filaments. Immunoflurescence microscopy of Chlamydomonas with these monoclonal antibodies and a monoclonal antibody that recognises all animal intermediate filaments (anti-IFA) gave a diffuse, patchy cytoplasmic staining pattern. Both the plant antibodies and anti-IFA stained interphase onion root tip cells in a diffuse perinuclear pattern. In metaphase through to telophase, the labelling patterns colocalised with those of microtubules. Labelling of the phragmoplast was also detected but not staining of the preprophase band. On Western blots of various animal cell lines and tissues, all the antibodies labelled known intermediate filament proteins. On Western blots of whole Chlamydomonas proteins, all the antiboides labelled a broad band in the 57,000 Mr range, and three antibodies labelled bands around 66,000 and 140,000 Mr but with varibale intensites. On Western blots of whole onion root tip proteins, all the antibodies labelled 50,0000 Mr (two to three bands) polypetides and a diffuse and around 60,000 Mr and three of the antibodies also labelled several polypeptides in the 90,000-200,000 Mr range. The consistent labelling of these different bands by several different monoclonal antibodies recognising animal intermediate filaments makes these polypetides putative plant intermediate filament proteins.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970080404
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  • 5
    Electronic Resource
    Electronic Resource
    βIV is the major β-tubulin isotype in bovine cilia (1993)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 25 (1993), S. 19-29 
    Details
    ISSN: 0886-1544
    Keywords: photoreceptor ; retina ; cilium ; trachea ; microtubules ; immunocytochemistry ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Four different isotypes of β-tubulin are known to be expressed in mammalian brain. Monoclonal antibodies against βII, βIII, and βIV were used to characterize the β-tubulin isotypes in two ciliated bovine tissues: non-motile sensory cilia of retinal rod cells and motile cilia of tracheal epithelium. Retinal rod outer segment (ROS) connecting cilia and cytoskeletons were purified by density gradient centrifugation. This preparation contained more than 20 major protein protein components, as shown by dodecyl sulfate polyacrylamide gel electrophoresis. Electroblots were used to quantitate the relative amounts of βII, βIII, and βIV. The connecting cilium and cytoskeleton of the rod outer segment has less type III β-tubulin than brain and more type IV. The ratio of βIV to βII in the ROS is nearly a factor of 8 larger than in brain. Electron microscopic immunocytochemistry showed extensive labeling of cilia by anti-type IV in thin sections of retinas and trachea, and also in purified ROS cilia and cyoskeletons. Labeling of cilia by anti-βII was also observed, although in the purified ROS cilia and cytoskeleton, the anti-βII labeling was primarily on amorphous non-ciliary material. The results suggest that both motile and non-motile cilia are enriched in the type IV β-tubulin subunit. © 1993 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970250104
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  • 6
    Electronic Resource
    Electronic Resource
    Cytoskeletal alterations in cultured cardiomyocytes following exposure to the lipid peroxidation product, 4-hydroxynonenal (1994)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 28 (1994), S. 119-134 
    Details
    ISSN: 0886-1544
    Keywords: microtubules ; vinculin ; desmin ; sarcolemmal damage ; free radicals ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Damage to the cardiac myocyte sarcolemma following any of several pathological insults such as ischemia (anoxia) alone or followed by reperfusion (reoxygenation), is most apparent as progressive sarcolemmal blebbing, an event attributed by many investigators to a disruption in the underlying cytoskeletal scaffolding. Scanning electron microscopic observation of tissue cultured rat neonatal cardiomyocytes indicates that exposure of these cells to the toxic aldehyde 4-hydroxynonenal (4-HNE), a free radical--induced, lipid peroxidation product, results in the appearance of sarcolemmal blebs, whose ultimate rupture leads to cell death. Indirect immunofluorescent localization of a number of cytoskeletal components following exposure to 4-HNE reveals damage to several, but not all, key cytoskeletal elements, most notably microtubules, vinculin-containing costameres, and intermediate filaments. The exact mechanism underlying the selective disruption of these proteins cannot be ascertained at this time. Colocalization of actin indicated that whereas elements of the cytoskeleton were disrupted by increasing length of exposure to 4-HNE, neither the striated appearance of the myofibrils nor the lateral register of neighboring myofibrils was altered. Monitoring systolic and diastolic levels of intracellular calcium ([Ca2+]i) indicated that increases in [Ca2+]i occurred after considerable cytoskeletal changes had already taken place, suggesting that damage to the cytoskeleton, at least in early phases of exposure to 4-HNE, does not involve Ca2+ -dependent proteases. However, 4-HNE-induced cytoskeletal alterations coincide with the appearance of, and therefore suggest linkage to, sarcolemmal blebs in cardiac myocytes.Although free radicals produced by reperfusion or reoxygenation of ischemic tissue have been implicated in cellular damage, these studies represent the first evidence linking cardiomyocyte sarcolemmal damage to cytoskeletal disruption produced by a free radical product. © 1994 Wiley-Liss, Inc.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970280204
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  • 7
    Electronic Resource
    Electronic Resource
    Reproductive capacity of sea urchin centrosomes without centrioles (1989)
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989), S. 264-273 
    Details
    ISSN: 0886-1544
    Keywords: microtubules ; microtubule organizing center ; mitosis ; monaster ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: For animal cells, the relative roles of the centrioles and the pericentriolar material (the cenrosomal microtubule organizing center) in controlling the precise doubling of the centrosome before mitosis have not been well defined. To this end we devised an experimental system that allowed us to characterize the capacity of the centrosomal microtubule organizing center to double regularly in the absence of centrioles. Sea urchin eggs were fertilized, stripped of their fertilization envelopes, and fragmented before syngamy. Those activated egg fragments containing just the female pronucleus assembled a monaster at first mitosis. A serial section ultrastructural analysis of such monasters revealed that the radially arrayed microtubules were organized by a hollow fenestrated sphere of electrondense material, of the same appearance as pericentriolar material, that was devoid of centrioles. We followed individual fragments with only a female pronucleus through at least three cell cycles and found that the monasters did not double between mitoses. The observation that fragments with only a male pronucleus repeatedly divided in a normal fashion indicates that the assembly and behavior of monasters were not artifacts of egg fragmentation. Our results demonstrate that the activity that controls the precise doubling of the centrosome before mitosis is distinct and experimentally separable from the centrosomal microtubule organizing center. Our observations also extend the correlation between the reproductive capacity of a centrosome and the number of centrioles it contains (G Sluder and CL Rieder, 1985a: J. Cell Biol. 100:887-896). For a cell that normally has centrioles, we show that a centrosome without centrioles does not reproduce between mitoses.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/cm.970130405
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  • 8
    Electronic Resource
    Electronic Resource
    Identification and distribution of insulin receptors on cultured bovine brain microvessel endothelial cells: Possible function in insulin processing in the blood-brain barrier (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 161 (1994), S. 333-341 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The binding of 125 I-insulin to primary cultures of bovine brain microvessel endothlial cells was examined. Insulin binding was both time and temperature dependent and inhibited by excess unlabeled insulin. Furthermore, the specific binding of insulin was polarized to the apical side of the cell monolayers. Upon binding, the labeled insulin was internalized, with approximately 70% resistant to acid wash over a 90-min period. The inhibition of insulin internalization observed with cell monolayers exposed to either phenylarsine oxide or unlabeled insulin suggests a receptor-mediated endocytic process. Furthermore, the ability of chloroquine to reduce the metabolism of insulin indicates a significant portion of the peptide iis processed through a lysosomal pathway. In contrast to the fluid-phase endocytosis marker, Lucifer yellow, as much as 65% of internalized insulin undergoes apical to basolateral trancytosis in brain microvessel endothelial cells. While most of the effluxed insulin was degraded, as assessed by trichloroacetic acid precipitation, the results of the present study suggest insulin receptors within the brain microvasculature may be involved in the processing and transport of bloodborne insulin. © 1994 Wiley-Liss, Inc.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041610218
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  • 9
    Electronic Resource
    Electronic Resource
    Multiple sites of vanadate and peroxovanadate action in Xenopus oocytes (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 162 (1995), S. 154-161 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In Xenopus laevis oocytes, the insulin mimics, vanadate and peroxovandates (PV), stimulated the uptake of 3H-2-deoxyglucose and incorporation of 35S-methionine into protein. For both hexose transport and protein synthesis, peroxovandates (produced by reacting vandate and H2O2) were at least as potent as vandate. Microinjection of peroxovandates into the oocytes stimulated 2-deoxyglucose uptake. However, methionine incorporation was not stimulated by microinjection of peroxovanadate or vanadate solutions. Consistent with these results and with the possibility that vandate and peroxovandates enter the cell on a phosphate transporter, raising the medium phosphate concentration from 1 mM to 10 mM blocked vanadate-stimulated hexose transport and partially reduced peroxovanadates stimulation of hexose transport. Increased medium phosphate did not reduce stimulation of protein synthesis by either effector. Taken together, these data indicate that vanadate/peroxovanadates act at both intracellular and extracellular sites. Action at the former stimulates hexose uptake and action at the latter, protein synthesis. © 1995 Wiley-Liss, Inc.This artilce is a US Government work and, as such, is in the public domain in the United States of America.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041620119
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  • 10
    Electronic Resource
    Electronic Resource
    Insulin-like growth factors modulate the growth inhibitory effects of retinoic acid on MCF-7 breast cancer cells (1995)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 165 (1995), S. 212-221 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Retinoids are currently being tested for the treatment and prevention of several human cancers, including breast cancer. However, the anti-cancer and growth inhibitory mechanisms of retinoids are not well understood. All-trans retinoic acid (RA) inhibits the growth of the estrogen receptor-positive (ER+) breast cancer cell line, MCF-7, in a reversible and dose-dependent manner. In contrast, insulin-like growth factors (IGF-I,IGF-II) and insulin are potent stimulators of the proliferation of MCF-7 and several other breast cancer cell lines. Pharmacologic doses of RA (≤10-6M) completely inhibit IGF-I-stimulated MCF-7 cell growth. Published data suggest that the growth inhibitory action of RA on IGF-stimulated cell growth is linear and dose-dependent, similar to RA inhibition of unstimulated or estradiol-stimulated MCF-7 cell growth. Surprisingly, we have found that IGF-I or insulin-stimulated cell growth is increased to a maximum of 132% and 127%, respectively, by cotreatment with 10-7 M RA, and that 10-9-10-7 M RA increase cell proliferation compared to IGF-I or insulin alone. MCF-7 cells that stably overexpress IGF-II are also resistant to the growth inhibitory effects of 10-9-10-7 M RA. Treatment with the IGF-I receptor blocking antibody, αIR-3, restores RA-induced growth inhibition of IGF-I-treated or IGF-II-overexpressing MCF-7 cells, indicating that the IGF-I receptor is mediating these effects. IGFs cannot reverse all RA effects since the altered cell culture morphology of RA-treated cells is similar in growth-inhibited cultures and in IGF-II expressing clones that are resistant to RA-induced growth inhibition. These results indicate that RA action on MCF-7 cells is biphasic in the presence of IGF-I or insulin with 10-9-10-7 M RA enhancing cell proliferation and ≥ 10-6M RA causing growth inhibition. As IGF-I and IGF-II ligands are frequently detectable in breast tumor tissues, their potential for modulation of RA effects should be considered when evaluating retinoids for use in in vivo experimental studies and for clinical purposes. Additionally, the therapeutic use of inhibitors of IGF action in combination with RA is suggested by these studies. © 1995 Wiley-Liss Inc.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041650124
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