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  • 1
    ISSN: 1432-2307
    Keywords: Cholestasis ; Cytokeratin ; Cytoskeleton ; Electron microscope ; Intermediate filaments
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Intermediate filaments (IF) maintain the structural and functional integrity of cells. To investigate whether IF change as a consequence of increased mechanical pressure and what the significance of such alterations is for the integrity of hepatocytes, we investigated alterations of IF in rat liver following common bile duct ligation (CBDL). Immunofluorescence of cytokeratin 18 was performed on extracted cryostat sections which were also used for electron microscopy. Ultrathin sections of mildly extracted liver tissue were applied to reveal the relationship between IF and intercellular junctions and cytoplasmic organelles. Our results showed that hepatocellular IF underwent striking changes during CBDL. The so-called pericanalicular sheath disappeared and IF were rigidly rearranged at the cell periphery, appearing as honeycomb-like structures. Increased amounts of IF were found in close association with increased numbers of desmosomes at the lateral membranes of hepatocytes, and electron-dense desmosome-like bodies were even observed in the ectoplasm at bile canaliculi. Rearrangement of IF in the cytoplasm resulted in segregation of subcellular compartments. The increased density of the IF network and desmosomes are compensatory mechanisms of hepatocytes to resist increased mechanical load and disperse the tension. However, the intracellular rearrangement of IF leading to segregation of subcellular compartments may also have distinct effects on hepatocellular metabolic functions.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-7276
    Keywords: colon cancer ; fat storing cell ; fish oil ; immunohistochemistry ; metastasis ; myofibroblast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Recently, it was demonstrated that dietary Ω-3 polyunsaturated fatty acids (PUFAs) induce 10-fold more metastases in number and 1000-fold in volume in an animal model of colon cancer metastasis in rat liver. It was observed that tumors of rats on a fish oil diet lacked peritumoral stroma unlike tumors in livers of rats on a low fat diet or a diet containing Ω-6 PUFAs. In the present study, only one-third of the tumors in livers of rats on Ω-3 PUFA diet contained peritumoral stroma, whereas peritumoral stroma was present in 87% of the tumors in livers of rats on low fat diet. To explain these findings, we tested the hypothesis that fish oil exerts a direct inhibiting effect on the formation of extracellular matrix in tumor stroma as a consequence of blocking transformation of fat storing cells into myofibroblasts. It was found with immunohistochemical analysis of desmin as marker for fat storing cells and α-smooth muscle actin as marker for myofibroblasts that numbers of myofibroblasts were higher in tumors containing intratumoral stroma only than in tumors containing both peritumoral and intratumoral stroma. As most of the tumors in fish oil-treated rats contained intratumoral stroma only, this suggests that transformation of fat storing cells into myofibroblasts was highest in tumor stroma of fish oil-treated rats. Therefore, it is unlikely that the lack of stroma around tumors in fish oil-treated rats is due to inhibition of transformation of fat storing cells into myofibroblasts, but lack of peritumoral stroma is rather a consequence of rapid development of tumors in livers of fish oil-treated rats.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Journal of cancer research and clinical oncology 122 (1996), S. 343-350 
    ISSN: 1432-1335
    Keywords: Hyperthermia ; Radiation ; Carcinogenesis ; Vitamin A ; N-Acetylcysteine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The in vivo carcinogenic risk of hyperthermia, alone or in combination with irradiation, and the anti-carcinogenic potential of vitamin A andN-acetylcysteine (AcCys) were investigated. Starting 1 month before treatment, 160 rats were divided into four diet groups: no additives, vitamin A-enriched diet, AcCys and the combination vitamin A+AcCys. In 10 animals per diet group, the hind leg was treated with either X-irradiation alone (16 Gy), hyperthermia alone (60 min at 43°C), hyperthermia 5 h prior to irradiation or hyperthermia 5 h after irradiation. Animals were observed for 2 years after treatment with regard to the development of tumours either inside or outside the treated volume. After 16 Gy alone 12±5% of the animals developed a tumour. Tumour incidence increased to 37±9% (borderline significanceP=0.07 versus treatment with X-rays alone) when hyperthermia was applied prior to X-rays, and to 24±8% (NS) with hyperthermia after irradiation. The relative risk ratio (RRR) for tumour induction was increased to 2.4 by hyperthermia if combined with X-irradiation. Pathological characterization of induced tumours showed that these were of the fibrosarcoma, osteosarcoma and carcinoma type. Vitamin A alone or in combination with AcCys slightly protected against the induction of tumours by X-rays without or with hyperthermia (RRR of 0.4). However, morphological changes such as lipid accumulation in hepatocytes and damage to the parenchyma were noticed in livers from all animals that were given a vitamin-A-enriched diet (P〈0.0001). Data from the present and past reports show that hyperthermia alone is not carcinogenic, but that it may increase radiation carcinogenesis. Treatment temperature and time of exposure to heat in addition to the radiation dose applied are important factors in the carcinogenic process. The enhancement of radiation carcinogenesis seems to occur independently of the sequence and time interval between irradiation and hyperthermia. However, not all data are consistent with this interpretation.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Histochemistry and cell biology 100 (1993), S. 297-302 
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Freeze-substituted rat liver embedded in glycol methacrylate (GMA) has been used to demonstrate the activities of several enzymes. The following enzymes could be detected in GMA-sections by the indicated histochemical procedure(s): 5′-nucleotidase (lead salt, cerium-diaminobenzidine), alkaline phosphatase (indoxyl-tetrazolium salt), catalase (diaminobenzidine), acid phosphatase (diazonium salt), lactate dehydrogenase (tetrazolium salt) and glutamate dehydrogenase (tetrazolium salt). The activities of all these enzymes were dramatically decreased compared with the activities demonstrated in unfixed cryostat sections, with the exception of catalase. The activities of the following enzymes could not be detected in GMA-sections: glucose-6-phosphate dehydrogenase (tetrazolium salt), xanthine oxidoreductase (tetrazolium salt), d-amino acid oxidase (cerium-diaminobenzidine-cobalt-hydrogen peroxide) and glucose-6-phosphatase (cerium-diaminobenzidine). The possible role of restricted penetration of reagents into the resin was studied by measuring cytophotometrically the enzyme activities in GMA-sections of 3 and 6 μm in thickness. For all the enzymes that could be detected, the 6 μm : 3 μm ratio varied from 1.4 to 2.7. An eventual retarded penetration of reagents into the resin was investigated by measuring cytophotometrically the amount of final reaction product during incubation for acid phosphatase and glutamate dehydrogenase activities. In both cases linear relationships without a lag phase were found for the specific enzyme activities with incubation time. Chemical denaturation of proteins or masking of active sites in proteins due to embedding in the resin monomer may be considered to be the main cause of decreased enzyme activities.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Microfilaments in epithelial cells are important for the structural and functional integrity of tight junctions. In the present study, we examined the relationship between microfilaments and tight junctions in hepatocytes of rat liver following common bile duct ligation (CBDL) for up to 2 weeks. Actin filaments and tight junctions were studied by fluorescence microscopy using 7-nitrobenzene-2-oxa-1,3-diazole phallacidin (NBD-ph) and an anti-ZO-1 antibody, respectively. Double-stained sections were examined with confocal laser scanning microscopy (CLSM). Electron microscopy was applied for the assessment of structural alterations in microfilaments and in tight junctions with detergent-extraction and freeze-fracture preparations. Our results showed that F-actin was present at the entire plasma membrane of hepatocytes in control liver, whereas CBDL increased the patocytes in control liver, whereas CBDL increased the amount of F-actin mainly at the bile canalicular and lateral plasma membranes. Simultaneously, the immunofluorescence of ZO-1 underwent striking changes, i.e., from a uniform to an irregular staining pattern with various fluorescence intensities. CLSM demonstrated a colocalization of ZO-1 and F-actin in control liver and its deterioration in CBDL liver. Electron microscopy showed marked alterations of microfilaments and tight junctions due to CBDL. It is concluded that actin filaments are intimately associated with tight junctions in normal hepatocytes. CBDL impairs this association by progressively diminishing the structural interaction between F-actin and ZO-1, which may in turn lead to functional disturbances of tight junctions.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Histochemistry and cell biology 54 (1977), S. 27-37 
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The lipid dye Sudan Black B, as generally used to demonstrate lipids in the interior of the cell nucleus, was studied with regard to its staining properties for isolated nuclei in relation with its chromatographic characteristics in solution, as well as with a model system consisting of lipid containing polyacrylamide films. Isolated nuclei are stained with Sudan Black B dissolved in ethylalcohol, only when the dye-solution is at least one month old. Extraction with chloroform-methanol (2∶1) before stainig resulted in a decrease of 35% in intensity. Treatment with proteolytic enzymes and DNA-se caused a complete disappearance of the staining capacity. The binding of Sudan Black B with phospholipids enclosed in the form of liposomes in modelfilms when stained with this dye in ethylene glycol obeys the law of Lambert-Beer. Proteins were however, also coloured by the dye. The chromatographic experiments showed that the dye is built up from two main and a number of secondary products. The secondary products which increase by aging of the dye-solution, change the spectrophotometric properties of the total dye and show aspecific binding. The conclusion was reached that on the basis of a positive reaction with Sudan Black B no definite conclusions can be drawn about the presence of lipids in the interior of the cell nucleus.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The extinction coefficient is essential for the conversion of cytophotometric (mean integrated) absorbance values into absolute units of enzyme activity, for instance expressed in terms of moles of substrate converted per unit time and per unit wet weight of tissue. The extinction coefficient of polymerized diaminobenzidine (polyDAB) complexed with cobalt as the final reaction product of oxidase reactions was estimated at 575 nm by comparison of the amounts of final reaction products formed after incubation of serial unfixed cryostat sections of rat kidney to demonstrate D-amino acid oxidase activity with either the tetrazolium salt method or the cerium-DAB-cobalt-hydrogen peroxide method. Both procedures resulted in similar localization patterns of final reaction product in a granular form in epithelial cells of proximal tubules in rat kidney. The granules were peroxisomes. Linear relationships were found for both methods between the specific amounts of final reaction product generated by D-amino acid oxidase activity and incubation time. The cerium salt method gave rise to 7.4 times higher absorbance values of final reaction product generated per unit time and per unit wet weight of tissue than the tetrazolium salt procedure. The extinction coefficient of tetranitro BT-formazan is 19 000 at 557 nm. Therefore, the cytophotometric extinction coefficient of the poly DAB-cobalt complex as final reaction product of oxidase reactions was established to be 140 000.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Histochemistry and cell biology 41 (1974), S. 111-118 
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary One hour after partial hepatectomy some liver nuclei already have an increased nuclear protein content, after 15 hours all measured nuclei, and after 64 hours only a proportion of the measured nuclei. A decrease of the Fast green histon content was found. This decrease occurred in the mayority of the nuclei 15 hours after partial hepatectomy. An increase in nuclear RNA was found for all measured nuclei at 15 hours, and for a proportion of the measured nuclei at 64 hours. Tetraploid and diploid nuclei behave identically. The excess of nuclear protein after hepatectomy is resistent to elution with 0.14 M NaCl, which indicates a binding with chromosomal proteins or DNA. The percentage of nuclear sap protein on the total nuclear protein content of tetraploid, diploid parenchymal nuclei and littoral cell nuclei, is equal. These nuclei differ in the amount of chromosomal bound proteins.
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Histochemistry and cell biology 68 (1980), S. 49-53 
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The purpose of this study is to compare the protein content of parenchymal and non-parenchymal nuclei, as isolated from rat liver. The nucleic have been separated by means of a 1 g-sedimentation technique. The protein content of the separated nuclei has been determined cytophotometrically using the Naphthol Yellow S staining procedure after TCA-extraction (corresponding with the total protein content) and directly (corresponding with the non-histone proteins). The ratio of the total protein content of non-parenchymal, parenchymal diploid and parenchymal tetraploid nuclei respectively was found to be 0.65:1.00:1.90. The ratio of non-histone protein to total protein was the same for all types of nuclei investigated, namely about 55%.
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1432-2277
    Keywords: Key words Liver preservation ; UW solution ; warm perfusion ; Warm perfusion ; liver preservation ; UW solution ; Preservation ; liver ; warm perfusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Pretransplant rinse solutions have been shown to reduce reperfusion injury in cold-stored liver grafts, especially at the nonparenchymal level in sinusoidal endothelial cells (SEC). In this study, different rinse temperatures were tested in a rat liver preservation model. Livers were washed out in situ via the portal vein with cold (4 °C) University of Wisconsin (UW) solution, and after hepatectomy (t0), were stored for 8, 16, or 24 h of cold ischemia time (CIT). After storage, livers were flushed with UW solution at either 4 °C, 20 °C, or 37 °C and reperfused for 90 min (37 °C). Control livers were reperfused at t0 without preflush. Levels of hyaluronic acid (HA), purine nucleoside phosphorylase (PNP), AST, and LDH were measured in the reperfusion medium. Bile production was monitored during reperfusion. At the end of reperfusion, liver biopsies were taken for enzyme hystochemistry (5'-nucleotidase and LDH). After 8-h CIT and a flush at 4 °C, a release of endogenous HA (−7 %) was observed, whereas uptake of exogenous HA occurred after the 20 °C flush (2 %, P = NS) and after the 37 °C flush (24 %, p 〈 0.001). HA release occurred at all three preflush temperatures after the 16-h CIT but was significantly lower when flushed at 37 °C (–10 %) that at 4 °C and 20 °C (−64 % and −17 %, respectively, p = 0.05). After the 24-h CIT, the release of endogenous HA increased in the 4 °C and 20 °C preflush groups, but not in the 37 °C group. Levels of PNP and AST increased until the 24-h CIT in all groups but were significantly lower after preflush at 37 °C. Release of LDH did not increase with increasing periods of cold storage in any of the flush series. Compared to control livers, mean bile production during reperfusion was significantly decreased following preflush at 4 °C or 37 °C after all periods of CIT. No differences in mean bile production could be demonstrated in the three preflush groups after any period of CIT. LDH activity in liver tissue was best preserved after the 8 and 16-h CIT in combination with the 37 °C preflush, indicating less hepatocellular damage. In conclusion, in cold stored rat livers flushed at 37 °C before reperfusion, SEC and hepatocellular damage is attenuated.
    Type of Medium: Electronic Resource
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