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Our studies show the influence of coumarins: xanthotoxin and o,o-dimethyl-fraxetin on apoptosis in human peripheral blood lymphocytes in vitro. Lymphocytes (derived by ethical protocol) were cultured for 72 h in vivo. The cell cultures were stimulated with xanthotoxin or o,o-dimethyl-fraxetin in various concentrations. Apoptotic cells (compaction and margination of nuclear chromatin, cytoplasmic condensation, membrane blebbing and cell shrinkage) were observed in BX41 fluorescent Olympus microscope and recorded by MultiScan software. We observed the highest induction of apoptosis in cells stimulated with xanthotoxin in doses ≥50 μM in 48-hrs-cultures.
BLV is an agent of enzootic bovine leukaemia (EBL), an infectious disease affecting cattle worldwide. BLV infection has been associated with immune system disorders and discrepancies in the cytokine network. The significance of dendritic cells in the pathogenesis of BLV infection is largely unknown, but considering their fundamental role in immune response it may be crucial. Dcs precursors were isolated with the use of immunomagnetic beads from BLV-infected and BLV-free cows. From these precursors cultures of monocyte derived dendritic cells (MoDCs) were generated with the use of a cytokine cocktail (IL-4 and GM-CSF). Additionally, parallel DCs from BLV-negative animals were infected in vitro. The level of cytokines: IL-6, IL-10, IL-12(p40), IL-12(p70) was determined in DC cultures: infected in vitro, originating from naturally infected cattle and BLV-free cattle. The investigation showed significant changes in almost all analyzed populations of BLV-infected Dcs. Cytokine profiles of blood MoDCs indicated activation of these groups during infection. In the case of spleen MoDCs and lymph node MoDCs a decrease in production of IL-12(p40) and IL-12(p70) in favour of IL-6 and IL-10 was noted, suggesting promotion of BLV infection development.
The exchanges between sister chromatids were detected with the fluorescence plus Giemsa technique, in which the thymine analogue, bromodeoxyuridine, was used for DNA labelling. This method was used to examine the influence of bovine leukaemia virus on chromosomal disorders in bovine blood lymphocytes. In metaphasal chromosomes of BLV infected lymphocytes, numerous sister chromatid exchanges (SCEs) were found. In chromosomal spreads of lymphocytes of BLV infected cows there were 6.5-13.0 SCEs per metaphase. In the control animals, only a small number of spontaneous SCEs (0-2.0) were found (P<0.01). The obtained results indicated that BLV may cause chromosomal disorders in lymphocytes of leukaemic cattle.
Adriamycin is an antineoplastic antibiotic which is biodegragated mainly in the liver. The purpose of this study was to analyse the apoptotic index and the histological structure of apoptotic cells in the rat liver. Rats were examined 4 and 7 weeks after adriamycin treatment. The animals were divided into four equal groups: groups I and III, consisting of rats treated with a single intraperitoneal dose of adriamycin (5 mg/kg b.w) and decapitated after 4 or 7 weeks; groups II and IV comprising control rats treated with a single intraperitoneal dose of 0.9% NaCl (0.5 ml) and decapitated after 4 or 7 weeks. After decapitation, liver specimens were collected and embedded in paraffin. Slides were stained with haematoxylin and eosin. The degree of apoptosis was determined quantitatively using the apoptotic index. Statistical analysis was performed using the one-way ANOVA test and Student’s t-test. A single dose of adriamycin induced apoptosis of hepatocytes, which increased with time The apoptotic index was significantly higher in the group examined 7 weeks after adriamycin administration. The histological structure of the liver in the experimental groups was similar. The lesions observed were focal. The blurring of the hepatic cell membrane and a focal disintegration of architectonics concerning the shape and size of hepatocytes were visible. Chromatin in the nuclei was dispersed. Some nuclei showed peripheral chromatin condensation. The cytoplasm of some cells showed numerous vacuoles. Naked nuclei and congestion resulting from vessel damage were visible.
Telomeres are the end fragments of chromosomes formed by a number of non-coding double-stranded TTAGGG repeats in vertebrates. During cell division the number of repeats decreases, leading to cell senescence or apoptosis. In immortal cells, including cancer cells, the telomere length is stable and maintained by, among other factors, telomerase. The aim of the study is to compare telomerase activity in normal lymphocytes and in leukaemic cells. Samples of acute leukaemia cells, HL 60 cell line and the lymphocytes of healthy volunteers were examined. Telomerase analysis was performed using TeloTAGGG Telomerase PCR ELISAplus (Roche). The relative telomerase activities (RTA) in leukaemic and normal cells were analysed. A high level of RTA was observed in leukaemic cells.
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