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  • 1
    ISSN: 1058-8388
    Keywords: EAP-300 ; Chick embryo ; Heart development ; Purkinje fiber ; Neural crest ; HNK-1 ; Connexin42 ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The spatiotemporal distribution of EAP-300 (embryonic avian polypeptide of 300 kDa) was analyzed in embryonic chick heart using immunohistochemistry and confocal microscopy. EAP-300 is a developmentally regulated protein initially characterized in neural cells from chick retina. Myocardial cells all along the early tubular heart were ubiquitously immunolabeled for EAP-300 by embryonic day 2 (E2, Stage 13). At E5 (Stage 24), myocardial EAP-300 expression levels remained significant in both atrial and ventricular myocardium. At E6 (Stage 28), distinct populations of EAP-300 immunolabeled cells were also observed external to the heart, in septal mesenchymal tissue and neural ganglia adjacent to the outflow tract; these cell populations were confirmed as neural crest-derived by co-localization of EAP-300 and HNK-1. At E13 (Stage 39), myocardial immunolabeling for EAP-300 was no longer ubiquitous, but increasingly restricted to conduction tissues, including the atrioventricular bundle and subendocardial Purkinje cells. This restriction of immunolabeling could be demonstrated definitively at E15 (stage 41), by which stage subendocardial and periarterial Purkinje fibers were clearly immunoreactive for EAP-300 and several known markers of chick conduction tissue, including specific myosin heavy chain isoforms and connexin42, a gap junctional protein preferentially expressed by Purkinje fibers. Just prior to hatching at E21 (Stage 46), immunolabeling of conduction tissues was reduced, although still above that of non-conductile myocardium. This spatiotemporal map of cardiac EAP-300 expression indicates that it is independently and transiently expressed in early myocardium, cardiac conduction tissue, and neural crest derivatives during development. ©1995 Wiley-Liss, Inc.
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  • 2
    ISSN: 1059-910X
    Keywords: Prostate cancer ; Benign prostatic hyperplasia ; Retrovirus ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Natural Sciences in General
    Notes: Mesenchymal-epithelial interactions are associated with growth and morphogenesis of the prostate. We have detected three isoforms of transforming growth factor β (TGF-β) in the developing mouse prostate that may mediate some of these interactions. Separation of the fetal urogenital sinus (UGS) tissue into mesenchymal and epithelial components indicated that mRNA expression of TGF-β;1, 2, and 3 was more abundant in the mesenchyme compared to the epithelium. Immunohistochemical analysis revealed accumulation of TGF-β1 in the mesenchyme surrounding ductules in the UGS and neonatal prostate. Further analysis of TGF-β1 localization in surgically removed adult human prostate tissues revealed more intense staining associated with regions of abnormal growth compared to normally appearing tissue. The percent of the total stained area with extracellular accumulation of TGF-β1 was 59% in prostate cancer, 26% in benign prostatic hyperplasia (BPH), and 8.6% in normal tissue. In additional immunohistochemical studies we observed that intracellular TGF-β1 was predominantly associated with the epithelial cells in prostate cancer (epithelial cells = 33.5% of the total stained area, stromal cells = 13.3%, and unstained = 53.2%), whereas in BPH intracellular TGF-β1 was predominantly associated with stromal cells (stromal cells = 32.2% of the total stained area, epithelial cells = 12.3%, and unstained = 55.5%). Although additional experimental and clinical studies are needed to better understand the relationships between TGF-β1 and abnormal prostatic growth, our observations thus far suggest a role for TGF-β1 in the development of benign and malignant growth abnormalities in the prostate. One approach to establishing the pathobiological significance of TGF-β1 accumulation in the prostate is by introducing and overexpressing the TGF-β1 cDNA in prostate tissue using the mouse prostate reconstitution model system. We discuss applicability of transgenic animal and organ reconstitution models for experimental studies concerning TGF-β - induced prostate growth abnormalities. © 1995 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Microscopy Research and Technique 30 (1995), S. 513-520 
    ISSN: 1059-910X
    Keywords: Polyacrylamide embedding ; Confocal microscopy ; Specimen preparation ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Natural Sciences in General
    Notes: The preparation of optically clear, thick sections of fragile embryonic tissues greatly aids the power of confocal scanning laser microscopy in imaging three-dimensional structures. We report here conditions for embedding, sectioning, and staining embryos in polyacrylamide gels for a variety of confocal imaging techniques. Infiltration of tissues in standard mixtures of 10-15% acrylamide monomer yields, upon polymerization, blocks that cut easily by vibratome between 50 and 1,000 μm. These conditions worked well for tissues previously stained or for staining gel sections with low molecular weight water-soluble fluorochromes (MW 〈 5 kD [e.g., propidium iodide, phalloidin]). For immunostaining of tissue after embedding and sectioning, the acrylamide concentration was reduced to 2-3% acrylamide to allow access of immunoglobulins to antigenic sites; such gels were supplemented with 1% agarose to facilitate sectioning and handling. Either method yielded abundant, optically clear, and easily handled sections for mounting and examination in water-miscible media. © 1995 Wiley-Liss, Inc.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 65 (1997), S. 159-171 
    ISSN: 0730-2312
    Keywords: breast cancer ; droloxifene ; estrogen replacement therapy ; apoptosis ; osteoclasts ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The incidence of postmenopausal osteoporosis is increasing as the population ages. Even though estrogen replacement therapy has proven beneficial in reducing the number of skeletal fractures, the known risks and associated side-effects of estrogen replacement therapy make compliance poor. Recent research has focused on the development of tissue specific estrogen agonist/anatagonists such as droloxifene which can prevent estrogen deficiency-induced bone loss without causing uterine hypertrophy. Furthermore, droloxifene acts as a full estrogen antagonist on breast tissue and is being evaluated for treatment of advanced breast cancer. In this report we propose a common mechanism of action for droloxifene that underlies its estrogen agonist and antagonist effects in different tissues. Droloxifene and estrogen, which have identical effects on bone in vivo, both induced p53 expression and apoptosis in cells of in vitro rat bone marrow cultures resulting in a decrease in the number of bone-resorbing osteoclasts. Droloxifene is growth inhibitory in MCF-7 human breast cancer cells and therefore acts as an antagonist, whereas estrogen is mitogenic to these cells and acts as an agonist. Droloxifene, but not estrogen, induced p53 expression and apoptosis in MCF-7 cells. These results indicate that the induction of apoptosis by droloxifene may be the common mechanism for both its estrogen agonist effects in bone and its antagonist effects in breast tissue. J. Cell. Biochem. 65:159-171. © 1997 Wiley-Liss, Inc.
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  • 5
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In C2C12 myoblasts, 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulated a phospholipase D (PLD) to degrade phosphatidylcholine (PC) as measured by the release of choline and an increase in the formation of phosphatidic acid (PA) (or phosphatidylbutanol [PtdBuOH] in the presence of 0.5% butanol). Exogenous PLD also stimulated choline release, PA and PtdBuOH formation. The protein kinase C (PKC) inhibitor, Ro-31-8220, and PKC downregulation significantly inhibited the effects of TPA but Ro-31-8220 had no effect on PLD action. Neither basic Fibroblast Growth Factor (bFGF) or Epidermal Growth Factor (EGF) increased PLD activity. All agonists stimulated protein synthesis during both a 90 min and a 6 hr incubation and increased RNA accretion after 6 hr. The response at 90 min was not inhibited by the transcription inhibitor, actinomycin D. Ro-31-8220 and PKC downregulation significantly inhibited all the effects of TPA. In contrast, Ro-31-8220 significantly inhibited the increase in RNA accretion elicited by PLD but had no effect on the ability of agonists other than TPA to enhance protein synthesis. All agonists also stimulated thymidine incorporation into DNA. The effects of EGF, bFGF, and PLD were rapid and transient whereas that of TPA was delayed and sustained. Ro-31-8220 and PKC downregulation significantly inhibited the response due to TPA. Furthermore, Ro-31-8220 also significantly inhibited the effects elicited by EGF and PLD but not that induced by bFGF. In differentiated myotubes, TPA and PLD, but not bFGF or EGF, again stimulated choline release and PtdBuOH formation. However, all agents failed to stimulate protein synthesis and RNA accretion. The data demonstrate the presence in C2C12 myoblasts, but not differentiated myotubes, of both a PLD-dependent and PLD-independent pathway(s) leading to the stimulation of protein synthesis, RNA accretion, and DNA synthesis. © 1995 Wiley-Liss, Inc.
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  • 6
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 163 (1995), S. 631-635 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Nonsedimentable lipid-protein particles have been isolated from intestinal tissue of the American cockroach, Periplaneta americana. Most of the particles were within the range 30-50 nm in diameter and appear to originate from larger structures. Lipid analysis of the particles showed them to be enriched in neutral lipid components relative to microsomal membranes. Specifically, there is a decline in the amounts of phosphatidylcholine and phosphatidylethanolamine in the nonsedimentable particles compared with the microsomal membranes. Also, in contrast to microsomal membranes, the particles have a higher content of phosphatidic acid along with 1,2- and 1,3-diacyglycerols, free fatty acids and an unidentified lipid that co-migrates with sterol ester, wax ester and hydrocarbon standards in thin layer chromatograms. The cytosol, separated from the particles by ultrafiltration, contained phosphatidic acid, free fatty acids and the unidentified lipid. By contrast, the composition of neutral lipids in the cytosol resembles that of the particles. SDS - PAGE analysis of microsomal membranes, the particles and particle free cytosol shows an enrichment of low molecular weight proteins in the particles and cytosol. The particles and cytosol appear to possess proteolytic activity that is distinguishable from that of corresponding microsomal membranes since the incubation of these components with BSA resulted in the formation of distinct polypeptides. Many characteristics of these particles resemble those of the deteriosomes that have been isolated from plant tissue. © 1995 Wiley-Liss, Inc.
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  • 7
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 163 (1995), S. 532-537 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Hepatocyte growth is regulated by various growth factors, including epidermal growth factor (EGF) and insulin. Recently, several additional peptide hormones have been shown to stimulate growth of hepatocyte only in the presence of EGF or insulin and are thus termed secondary mitogens. Gastrin regulates growth of normal and neoplastic gastrointestinal tissues, but the effect on growth of hepatocyte is unknown. We examined the effect of gastrin on growth of a normal mouse hepatocyte (NMH) line established in our laboratory. Effect of gastrin-17 (G-17) (10-8 to 10-6 M) on growth of NMH cells was examined in either the presence or absence of EGF in the culture medium. Growth of NMH cells was evaluated by incorporation of either bromodeoxyuridine (BrdU) or 3H-thymidine and by counting cells. Presence of a cell-surface receptor for G-17 was determined by Scatchard analysis using 125I-G-17. In the presence of EGF, gastrin stimulated growth of NMH cells; in the absence of EGF, gastrin did not affect growth. The stimulatory effect of gastrin on NMH cells was blocked by JMV 320, a CCK-B type receptor antagonist. NMH cells possess a single, high affinity binding site for gastrin (Kd = 1.2 nM); EGF increased the gastrin binding capacity compared to non-treated cells (3.5 ± 0.4 vs. 2.2 ± 0.6 fmol/106 cells). G-17 stimulated growth of NMH cells through a single high affinity receptor for G-17 which pharmcologically appears to be the CCK-B type only in the presence of EGF and thus can be considered a secondary mitogen. © 1995 Wiley-Liss, Inc.
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  • 8
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 165 (1995), S. 172-176 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Alterations in the activity of phospholipase C (PLC) are thought to be the primary intracellular events leading to pancreatic acinar cell exocytosis of zymogen granules. When multiple hormones, each of which may stimulate different signal transduction pathways, bind to cell surface receptors, the cell must integrate these signals into a common response through communication (cross-talk) among intracellular second messengers. We show that cholecystokinin (CCK) induces amylase secretion from AR4-2J pancreatic acinar cells via stimulation of PLC activity. Secretin indirectly stimulated the PLC pathway through cross-talk of the activated cAMP pathway to potentiate the CCK-stimulated amylase secretion. Therefore, secretin potentiated the acinar cell secretory response to CCK by cAMP-mediated cross-talk with the PLC signal transduction pathway. © 1995 Wiley-Liss Inc.
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  • 9
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    BioEssays 17 (1995), S. 341-350 
    ISSN: 0265-9247
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Over the last decade, the c-myb gene and its protein product, Myb, have undergone extensive examination and manipulation in hemopoietic tissues. Although it is rarely disputed that, as a transcription factor, Myb regulates cell cycling, proliferation and differentiation, identification of genes directly controlled by Myb has been surprisingly difficult. More recently, genes with promoter regions that contain Myb recognition sequences have been identified, but a direct proliferative response to Myb via these ‘target genes’ has yet to be demonstrated. Mutagenesis studies have defined domains of the protein which influence its transcriptional activity and transforming potential; however how the molecule interacts with itself and with other cellular factors is only beginning to be understood. A broader examination of c-myb expression in normal and malignant tissues suggests an analogous role for Myb in proliferation, differentiation and transformation of nonhemopoietic tissues.
    Additional Material: 2 Ill.
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  • 10
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Bioelectromagnetics 16 (1995), S. 119-123 
    ISSN: 0197-8462
    Keywords: EMF ; melatonin ; puberty ; sheep ; transmission line ; Life and Medical Sciences ; Occupational Health and Environmental Toxicology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Physics
    Notes: In an earlier study, we found no effects of 60 Hz electric and magnetic fields (EMF) from a 500 kV transmission line on serum melatonin patterns or on puberty in ten female Suffolk lambs (Ovis aries). We conducted a larger replicate study of 15 lambs exposed to a mean electric field of 6.3 kV/m and a mean magnetic field of 3.77 μT and 15 controls exposed to EMF two orders of magnitude weaker than in the line area. The replicate produced essentially the same results as our previous study. © 1995 Wiley-Liss, Inc.
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