Home About us Contact
|
Home About us Contact | ||
|
|
||
Apoptotic Morphology (apoptotic + morphology)
Selected AbstractsCold-induced apoptosis of rat liver cells in University of Wisconsin solution: The central role of chelatable ironHEPATOLOGY, Issue 3 2002Uta Kerkweg Although University of Wisconsin (UW) solution aims at the prevention of cold-induced cell injury, it failed to protect against cold-induced apoptosis of hepatocytes and liver endothelial cells: when incubated in UW solution at 4°C for 24 hours and subsequently rewarmed at 37°C, 72% ± 8% of rat hepatocytes and 81% ± 5% of liver endothelial cells lost viability. In both cell types, the observed cell damage occurred under an apoptotic morphology; it appeared to be mediated by a rapid increase in the cellular chelatable iron pool by a factor ,2 (as determined in hepatocytes) and subsequent formation of reactive oxygen species (ROS). Consequently, this cell injury was decreased by iron chelators to 6 to 25% (hepatocytes) and 4% ± 2% (liver endothelial cells). Deferoxamine nearly completely inhibited the occurrence of apoptotic morphology in both cell types. In liver endothelial cells, cold-induced apoptosis occurring during rewarming after 24 hours of cold incubation in UW solution was far more pronounced than in cell culture medium (loss of viability: 81% ± 5% vs. 28% ± 13%), but viability could even be maintained for 2 weeks of cold incubation by use of deferoxamine. In conclusion, this pathological mechanism might be an explanation for the strong endothelial cell injury known to occur after cold preservation. With regard to the extent of this iron-mediated injury, addition of a suitable iron chelator to UW solution might markedly improve the outcome of liver preservation. [source] Caspase activation correlates with the degree of inflammatory liver injury in chronic hepatitis C virus infectionHEPATOLOGY, Issue 4 2001Heike Bantel Hepatitis C virus (HCV) infection is a major cause of liver disease characterized by inflammation, cell damage, and fibrotic reactions of hepatocytes. Apoptosis has been implicated in the pathogenesis, although it is unclear whether proteases of the caspase family as the central executioners of apoptosis are involved and how caspase activation contributes to liver injury. In the present study, we measured the activation of effector caspases in liver biopsy specimens of patients with chronic HCV infection. The activation of caspase-3, caspase-7, and cleavage of poly(ADP-ribose)polymerase (PARP), a specific caspase substrate, were measured by immunohistochemistry and Western blot analysis by using antibodies that selectively detect the active truncated, but not the inactive precursor forms of the caspases and PARP. We found that caspase activation was considerably elevated in liver lobules of HCV patients in comparison to normal controls. Interestingly, the immunoreactive cells did yet not reveal an overt apoptotic morphology. The extent of caspase activation correlated significantly with the disease grade, i.e., necroinflammatory activity. In contrast, no correlation was observed with other surrogate markers such as serum transaminases and viral load. In biopsy specimens with low activity (grade 0) 7.7% of the hepatocytes revealed caspase-3 activation, whereas 20.9% of the cells stained positively in grade 3. Thus, our results suggest that caspase activation is involved in HCV-associated liver injury. Moreover, measurement of caspase activity may represent a reliable marker for the early detection of liver damage, which may open up new diagnostic and therapeutic strategies in HCV infection. [source] Activation of caspase-3 alone is insufficient for apoptotic morphological changes in human neuroblastoma cellsJOURNAL OF NEUROCHEMISTRY, Issue 6 2002Margaret M. Racke Abstract Activated caspase-3 is considered an important enzyme in the cell death pathway. To study the specific role of caspase-3 activation in neuronal cells, we generated a stable tetracycline-regulated SK-N-MC neuroblastoma cell line, which expressed a highly efficient self-activating chimeric,caspase-3, consisting of the caspase-1 prodomain fused to the caspase-3 catalytic domain. Under expression-inducing conditions, we observed a time-dependent increase of processed caspase-3 by immunostaining for the active form of the enzyme, intracellular caspase-3 enzyme activity, as well as poly(ADP-ribose) polymerase (PARP) cleavage. Induced expression of the caspase fusion protein showed predominantly caspase-3 activity without any apoptotic morphological changes. In contrast, staurosporine treatment of the same cells resulted in activation of multiple caspases and profound apoptotic morphology. Our work provides evidence that auto-activation of caspase-3 can be efficiently achieved with a longer prodomain and that neuronal cell apoptosis may require another caspase or activation of multiple caspase enzymes. [source] Evidence that bcl-2 is the Target of Three Photosensitizers that Induce a Rapid Apoptotic Response,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 2 2001David Kessel ABSTRACT We originally proposed that the subcellular target for one class of photosensitizing agents was the mitochondrion. This classification was based on effects that occur within minutes of irradiation of photosensitized cells: rapid loss of the mitochondrial membrane potential (,,m), release of cytochrome c into the cytosol and activation of caspase-3. These effects were followed by the appearance of an apoptotic morphology within 30,90 min. Fluorescence localization studies on three sensitizers initially classified as ,mitochondrial' revealed that these agents bind to a variety of intracellular membranes. The earliest detectable effect of photodamage is the selective loss of the antiapoptotic protein bcl-2 leaving the proapoptotic protein bax undamaged. Bcl-2 photodamage can be detected directly after irradiation of cells at 10°C. Subsequent warming of cultures to 37°C results in loss of ,,m, release of cytochrome c and activation of caspase-3. The latter appears to amplify the other two effects. Based on results reported here we propose that the apoptotic response to these photosensitizers is derived from selective photodamage to the antiapoptotic protein bcl-2 while leaving the proapoptotic protein bax unaffected. [source] Apoptosis and Phagocytosis of Tissue-Dwelling Eosinophils in Sinonasal Polyps,THE LARYNGOSCOPE, Issue 1 2000Åke Davidsson MD Abstract Objective: Sinonasal polyps contain numerous tissue-dwelling eosinophils, but the mechanisms causing their accumulation, functional activities, and resolution are largely unknown. Study Design: Nasal polyp tissue from 14 patients was evaluated for cellular expression of CD95, CD68, and Annexin-V, for the degree of apoptosis, and for phagocytosis of eosinophils. Material and Methods: Histological sections were immunostained as single stains for CD95, CD68, and Annexin-V, and as an immunostaining for CD68 combined with a modified Vital New Red staining. The latter staining is specific for eosinophils. Other sections were stained by terminal d-UTP nick end labeling (TUNEL) assay and routinely stained for H&E. Evaluation of the amount of stained cells was performed by counting the average number in 10 randomly chosen high-power fields. The TUNEL positivity was in all cases confirmed with apoptotic morphology. Results: The inflammatory infiltrate consisted of numerous eosinophils but also a considerable amount of lymphocytes, mast cells, and macrophage-like CD68+ cells. CD95 was frequently expressed on eosinophils, on numerous other inflammatory cells, and also on morphologically apoptotic cells. Annexin-V-positive eosinophils were not as frequent as CD95+ cells, but numerous Annexin-V-positive eosinophils were found. CD68+ cells approximately equalled the number of eosinophils. The number of cells phagocytosing eosinophils varied between polyps. Apoptosis of eosinophils (as evaluated by TUNEL combined with apoptotic morphology) was a common finding in six of the polyps. Conclusions: Previous in vitro and ex vivo findings of CD95 on eosinophils are now supported by demonstration of CD95 on eosinophils in this in vivo study. This investigation revealed a switch of the membrane-bound phosphatidylserine of apoptotic cells, which is a novel observation. The study has demonstrated apoptosis of tissue-dwelling eosinophils, and that CD68+ macrophage-like cells phagocytose eosinophils within the sinonasal polyps. [source] Expression of Cell Death-Associated Proteins in Neuronal Apoptosis Associated with Pontosubicular Neuron NecrosisBRAIN PATHOLOGY, Issue 3 2001Christine Stadelmann Expression of apoptosis-associated proteins p53, bcl-2, bax, and caspase-3/CPP32, activation of caspase-3, and modification of proteins via poly(ADP-ribosyl)ation was studied in pontosubicular neuron necrosis (PSN), a form of perinatal brain damage revealing the morphological hallmarks of neuronal apoptosis. Immunoreactivity for p53 was completely absent. The majority of cells stained with the bax and procaspase-3 antibodies did not show morphological signs of apoptosis. In contrast, an antibody against activated caspase-3 almost exclusively stained cells with apoptotic morphology. Poly(ADP-ribosyl)ated proteins were only rarely detected in cells with apoptotic morphology. The expression patterns of bax, procaspase-3, bcl-2, and p53 in PSN were similar to that found in age-matched control brains. However, activated caspase-3 and poly-ADP-ribosylated proteins were exclusively found in apoptotic cells. These data indicate that detection of active caspase-3 is a reliable marker for apoptosis in formalin-fixed human tissue, and that neuronal apoptosis in pontosubicular neuron necrosis is accompanied by a pronounced activation of caspase-3. [source] The combination of the antitumoural pyridyl cyanoguanidine CHS 828 and etoposide in vitro,from cytotoxic synergy to complete inhibition of apoptosisBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2002P Martinsson The present study was aimed at elucidating the apoptosis inhibitory properties of the cyanoguanidine CHS 828. CHS 828 exhibits impressive cytotoxic activity in vitro and in vivo. Apoptosis is not its main mode of cytotoxic effect, and we have previously proposed a dual mechanism, where CHS 828 inhibits its own cell death pathways. Etoposide on the other hand, is a well-established anticancer agent with documented effect in a number of malignancies, induces apoptosis through extensively studied caspase dependent pathways. Here we studied the combined effect of the two drugs in the human lymphoma cell line U-937 GTB. Cytotoxicity was evaluated as total viability measured by the fluorometric microculture cytotoxicity assay (FMCA). Caspase activity was assessed by colorimetric detection of specific cleavage products for caspases 3, 8 and 9, respectively. Morphology was evaluated in May-Grünwald/Giemsa stained preparations. Interaction analysis based on FMCA results of simple combination exposure revealed impressive synergistic effect on cell kill. Detailed investigations of the kinetics involved showed that short pre-exposure (0,12 h) to CHS 828 enhanced caspase activation by etoposide, while longer pre-exposure (18,48 h) inhibited both caspase activation and apoptotic morphology otherwise induced by etoposide. The present results support the theory that CHS 828 block specific cell death pathways. The synergistic results are promising for future combination trials in animals, however, different dosing schedules should be considered, in order to investigate whether the above findings translate into the in vivo setting. British Journal of Pharmacology (2002) 137, 568,573. doi:10.1038/sj.bjp.0704888 [source] Cell death in lens epithelial cells after stimulation of the sigma-2 receptorACTA OPHTHALMOLOGICA, Issue 2009JO KARLSSON Purpose The aim was to investigate the mechanisms of cell death in lens epithelial cells after administration of siramesine, a sigma-2 receptor agonist. Methods Human lens epithelial cells in culture were exposed to siramesine and examined for morphological changes using DIC or calcein as a cytoplasmic marker. Lysosomes were studied using acridine orange and MagicRed. Proteolytic activity of the proteasome, calpain, caspases and cathepsins in living cells or cell extracts were studied using different fluorogenic substrates. Results Siramesine at low concentrations increased the cytoplasmic proteolytic activity of the proteasome and the calpain system. Early effects was also observed with respect to lysosomal morphology, acidity and function. Activation of caspase-3 and the appearance of nuclei with an apoptotic morphology were also found. Conclusion Siramesine at very low concentrations affects lens epithelial cells with perturbation of the major proteolytic systems, lysosomal morphology and results in caspase activation and cell death. Siramesine may be a promising substance for clinical studies concerning the treatment of PCO. [source] | |||||||||||||