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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    The @Anatomical Record 208 (1984), S. 283-289 
    ISSN: 0003-276X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: This study used acrylic resin as an intravascular marker to demonstrate functional myocardial capillaries after fixation by perfusion. Eight rat hearts were excised and allowed to function as isolated organs perfused with oxygenated Krebs-Henseleit buffer (37o 10 kPa) for 10 min. Four were fixed by perfusion (4 min) with 2.5% glutaraldehyde at the same temperature and pressure and then immersion fixed (24 hr). The other four hearts were perfused with 0.2% procaine HCl for 30 sec just prior to similar fixation. Polymerizing low viscosity acrylic resin was injected at 10 kPa pressure into the fixed vascular beds and allowed to cure, then transmural blocks of left ventricular myocardium were prepared for scanning electron microscopy. Total initial coronary flow of fixative after procaine treatment was significantly increased, while in untreated hearts the initial fixative flow rate was closely similar to that of oxygenated buffer. The pattern of capillary perfusion was assessed, and the percentage of capillary profiles filled by acrylic resin were calculated. Following procaine treatment, 95.2% of capillaries appeared functional, whereas without procaine arrest, only 62.0% of capillaries allowed the passage of resin. This study indicates that perfusion fixation with glutaraldehyde stabilizes myocardial structure so that the proportion of functional capillary pathways remains closely similar to that in the beating heart and so that such functional capillaries can be identified in morphological preparations by using a low viscosity intraluminal resin marker.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Gamete Research 24 (1989), S. 185-196 
    ISSN: 0148-7280
    Keywords: free radicals ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Catalase activity was determined in human semen by measuring the oxygen burst with a Clark electrode, after H2O2 addition. Significant catalase activities (mean ± SD) were found in migrated, motile spermatozoa (44 ± 17 nmoles O2/min/108 cells) and in seminal plasma of normozoospermic men (129 ± 59 nmoles O2/min/ml). It has been demonstrated that seminal catalase originated from prostate; however, its activity was not correlated with the usual prostatic markers (such as citric acid and zinc). Our data suggest a multiglandular function secreted by this organ. The catalase activities measured in seminal samples from asthenozo-ospermic, infertile men were found lower than those from normozoospermic subjects. The understanding of the relative contribution of the different enzyme systems against O2 toxicity (superoxide dismutase, catalase, glutathione peroxidase) seem to be a priority area of research to understand disturbances of sperm function.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The isolation and characterization of a mutant murine T-cell lymphoma (S49) with altered purine metabolism is described. This mutant, AU-100, was isolated from a mutagenized populatio of S49 cells by virtue of its resistance to 0.1 mM 6-azauridine in semisolid agarose. The AU-100 cells are resistant to adenosine mediated cytotoxicity but are extraordinarily sensitive to killing by guanosine.High performance liquid chromatography of AU-100 cells extracts has demonstrated that intracellular levels of GTP, IMP, and GMP are all elevated about 3-fold over those levels found in wild type cells. The AU-100 cells also contain an elevated intracellular level of pyrophosphoribosylphosphate (PPriboseP), which as in wild type cells is diminished by incubation of AU-100 cells with adenosine. However AU-100 cells synthesize purines de novo at a rate less than 35% of that found in wild type cells.In other growth rate experiments, the AU-100 cell line was shown to be resistant to 6-thioguanine and 6-mercaptopurine. Levels of hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) measured in AU-100 cell extracts, however, are 50-66% greater than those levels of HGPRTase found in wild type cell extracts. Nevertheless this mutant S49 cell line cannot efficiently incorporate labeled hypoxanthine into nucleotides since the salvage enzyme HGPRTase is inhibited in vivo.The AU-100 cell line was found to be 80% deficient in adenylosuccinate synthetase, but these cells are not auxotrophic for adenosine or other purines. The significant alterations in the control of purine de novo and salvage metabolism caused by the defect in adenylosuccinate synthetase are mediated by the resulting increased levels of guanosine necleotides.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 146 (1991), S. 34-42 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Incubation of cultured cells in hypertonic medium and sodium-free medium have been shown to block transport at two different stages along the endocytic pathway. To determine the effects of these treatments on the exocytic pathway, we studied the transport of the membrane glycoprotein of vesicular stomatitis virus (VSV-G) in cells infected with tsO45 mutant virus. This mutant synthesizes a VSV-G that accumulates in the endoplasmic reticulum (ER) when cells are incubated at 39.5°C. In addition, VSV-G accumulates in the post-ER pre-Golgi compartment when cells are incubated at 15°C and in the trans-Golgi network (TGN) when cells are incubated at 18°C. Upon transfer of cells to 32°C in control medium, VSV-G exits each of these compartments and is transported to the cell surface. Incubation in sodium-free medium at 32°C did not block transport from any of these three compartments. In contrast, incubation in hypertonic medium blocked export from the ER, transport from the pre-Golgi compartment to the Golgi complex, and transport from the TGN to the cell surface. Our results, in combination with previous studies, suggest that hypertonic medium blocks at least five distinct transport steps: the three exocytic steps described here, endocytosis from the cell surface, and transport of cell surface proteins into the Golgi complex. This raises the possibility that vesicular transport in different parts of the cell shares common elements that are inhibited by this treatment.
    Additional Material: 6 Ill.
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 155 (1993), S. 290-300 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Insulin-like growth factor-II (IGF-II) is highly expressed in fetal tissues and may act as an autocrine growth factor during early embryogenesis. The SH-SY5Y human neuroblastoma cell line also expresses IGF-II and its receptors and responds to exogenous IGF-II with increased DNA synthesis, cell division, and neuritic outgrowth. For this study, we tested the hypothesis that IGF-II mediates autocrine growth of SH-SY5Y cells in serum-free media. SH-SY5Y cells plated at high densities proliferated in serum-free media, whereas sparsely plated cells did not. IGF-II mRNA levels increased within 24 hours of serum deprivation and were associated with increased immunoreactive IGF-II protein. Exogenous addition of IGF-II increased 3H-TdR incorporation and cell number in a dose- and time-dependent fashion. By nuclear labelling experiments using 5-Bromo-2′ deoxyuridine (BrdU), we detected a twofold higher percentage of S phase nuclei after a 24-hour incubation in IGF-II. Treatment of SH-SY5Y cells with anti-IGF-II antibodies in serum-free media inhibited cell proliferation, and this inhibition was partially overcome by the addition of increasing concentrations of IGF-II. Collectively, our results indicate that IGF-II mediates an autocrine growth mechanism in SH-SY5Y cells that is associated with increased IGF-II expression. © 1993 Wiley-Liss, Inc.
    Additional Material: 8 Ill.
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  • 6
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 89 (1976), S. 595-602 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The number of H+ ejected during passage of 2e- through each energy-conserving site of the mitochondrial respiratory chain (the H+/site ratio) was measured in three ways. In each case transmembrane movements of endogenous phosphate were minimized. (1) Measurement of the uptake of weak acids during loading of mitochondria with Ca2+ demonstrated that 2.0 weak acid anions were accumulated per Ca2+ ion. Since 1.7 to 2.0 Ca2+ ions were taken up per site, these data correspond to an H+/site ratio of 3.5 to 4.0. (2) More direct measurement of H+ ejection using the oxygen pulse technique demonstrated that the H+/site ratio was 3.0. In these experiments phosphate movements were prevented by addition of N-ethylmaleimide to inhibit phosphate-hydroxide antiport, by washing the mitochondria to remove endogenous phosphate, or by working at 5°C to reduce the rate of phosphate transport. When phosphate movements were allowed, H+/site ratios of 2.0 were observed. (3) Measurement of the initial steady rates of oxygen consumption and H+ ejection following addition of substrate to aerobic, substrate-limited mitochondria yielded H+/site ratios of 2.0, which were elevated to 4.0 when phosphate transport was prevented as described above.Previous determinations of the H+/site ratio were thus underestimates due to the unrecognized movements of endogenous phosphate; our results show that the H+/site ratio is at least 3.0 and may be as high as 4.0.
    Additional Material: 3 Ill.
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  • 7
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Nine independently derived clones of mutagenized rat hepatoma cells selected for resistance to 6-mercaptopurine (6-MP) or 6-thioguanine (6-ThioG) have been isolated. Each has severely reduced catalytic activity of hypoxanthine-guanine phosphoribosyltransferase (HPRT) and seven of them possess significantly increased activities of phosphoribosylpyrophosphate (PRPP) synthetase. The degrees of elevations of PRPP synthetase activities do not correlate with the degrees of deficiencies of HPRT activities.The cells from one of these clones, 1020/12, possess 40% of the normal HPRT catalytic activity and overproduce purines. We have extensively examined the cells from this clone. Immunotitration studies of 1020/12 cells indicate that there is a mutation in the structural gene for HPRT. Although they possess increased specific catalytic activities of the enzyme, PRPP synthetase, the catalytic parameters, heat stability, and isoelectric pH of PRPP synthetase from 1020/12 cells are indistinguishable from those of the enzyme from wild-type cells.The cause of purine overproduction by 1020/12 cells appears to be the elevated PRPP synthetase activity, rather than a PRPP “sparing” effect stemming from reduced HPRT activity. Support for this idea is provided by the observation that the complete loss of HPRT activity in a clone derived from 1020/12 cells does not further enhance the levels of PRPP synthetase or purine overproduction.We propose that the elevated levels of PRPP synthetase activity in these HPRT deficient cells result from a mutational event in the structural gene for HPRT, and that this causes the disruption of a previously undescribed regulatory function of this gene on the expression of the PRPP synthetase gene.
    Additional Material: 5 Ill.
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