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Apoptotic Nuclei (apoptotic + nucleus)
Selected AbstractsRole of p38 MAPKs in Hypericin Photodynamic Therapy-induced Apoptosis of Nasopharyngeal Carcinoma CellsPHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 5 2009Pui S. Chan The present study aims to determine the role of mitogen-activated protein kinases (MAPKs) in hypericin-mediated photodynamic therapy (HY-PDT)-induced apoptosis of the HK-1 nasopharyngeal carcinoma (NPC) cells. HY-PDT was found to induce proteolytic cleavage of procaspase-9 and -3 in HK-1 cells. Apoptotic nuclei were observed at 6 h after PDT whereas B-cell leukemia/lymphoma-2-associated-X-protein (Bax) translocation and formation of Bax channel is responsible for the cell death. Increase in phosphorylation of p38 MAPKs and c-Jun N-terminal kinase 1/2 (JNK1/2) was detected at 15,30 min after HY-PDT. The appearance of phosphorylated form of p38 MAPKs and JNK1/2 was inhibited by the singlet oxygen scavenger l -histidine. HY-PDT-induced cell death was enhanced by the chemical inhibitors for p38 MAPKs (SB202190 and SB203580), but not by the JNKs inhibitor SP600125. Knockdown of the p38, and p38, MAPK isoforms by small interfering RNA (siRNA) are more effective than the p38, in enhancing PDT-induced cell death. Augmentation of apoptosis by p38, or p38, knockdown is also correlated with the increased proteolytic cleavage of procaspase-9 after HY-PDT treatment. Our results suggested that HY-PDT activated p38 MAPKs through the production of singlet oxygen. Inhibition of p38 MAPKs with chemical inhibitors or siRNA enhances HY-PDT-induced apoptosis of the HK-1 NPC cells. [source] Methanol exposure interferes with morphological cell movements in the Drosophila embryo and causes increased apoptosis in the CNSDEVELOPMENTAL NEUROBIOLOGY, Issue 3 2004Dervla M. Mellerick Abstract Despite the significant contributions of tissue culture and bacterial models to toxicology, whole animal models for developmental neurotoxins are limited in availability and ease of experimentation. Because Drosophila is a well understood model for embryonic development that is highly accessible, we asked whether it could be used to study methanol developmental neurotoxicity. In the presence of 4% methanol, approximately 35% of embryos die and methanol exposure leads to severe CNS defects in about half those embryos, where the longitudinal connectives are dorsally displaced and commissure formation is severely reduced. In addition, a range of morphological defects in other germ layers is seen, and cell movement is adversely affected by methanol exposure. Although we did not find any evidence to suggest that methanol exposure affects the capacity of neuroblasts to divide or induces inappropriate apoptosis in these cells, in the CNS of germ band retracted embryos, the number of apoptotic nuclei is significantly increased in methanol-exposed embryos in comparison to controls, particularly in and adjacent to the ventral midline. Apoptosis contributes significantly to methanol neurotoxicity because embryos lacking the cell death genes grim, hid, and reaper have milder CNS defects resulting from methanol exposure than wild-type embryos. Our data suggest that when neurons and glia are severely adversely affected by methanol exposure, the damaged cells are cleared by apoptosis, leading to embryonic death. Thus, the Drosophila embryo may prove useful in identifying and unraveling mechanistic aspects of developmental neurotoxicity, specifically in relation to methanol toxicity. © 2004 Wiley Periodicals, Inc. J Neurobiol 60: 308,318, 2004 [source] Effect of dopamine on rat diaphragm apoptosis and muscle performanceEXPERIMENTAL PHYSIOLOGY, Issue 4 2006Janet D. Pierce The purpose of this study was to determine whether dopamine (DA) decreases diaphragm apoptosis and attenuates the decline in diaphragmatic contractile performance associated with repetitive isometric contraction using an in vitro diaphragm preparation. Strenuous diaphragm contractions produce free radicals and muscle apoptosis. Dopamine is a free radical scavenger and, at higher concentrations, increases muscle contractility by simulating ,2 -adrenoreceptors. A total of 47 male Sprague,Dawley rats weighing 330,450 g were used in a prospective, randomized, controlled in vitro study. Following animal anaesthetization, diaphragms were excised, and muscle strips prepared and placed in a temperature-controlled isolated tissue bath containing Krebs,Ringer solution (KR) or KR plus 100 ,m DA. The solutions were equilibrated with oxygen (O2) at 10, 21 or 95% and 5% carbon dioxide, with the balance being nitrogen. Diaphragm isometric twitch and subtetanic contractions were measured intermittently over 65 min. The diaphragms were then removed and, using a nuclear differential dye uptake method, the percentages of normal, apoptotic and necrotic nuclei were determined using fluorescent microscopy. There were significantly fewer apoptotic nuclei in the DA group diaphragms than in the KR-only group diaphragms in 10 and 21% O2 following either twitch or subtetanic contractions. Dopamine at 100 ,m produced only modest increases in muscle performance in both 10 and 21% O2. The attenuation of apoptosis by DA was markedly greater than the effect of DA on muscle performance. Dopamine decreased diaphragmatic apoptosis, perhaps by preventing the activation of intricate apoptotic pathways, stimulating antiapoptotic mechanisms and/or scavenging free radicals. [source] Protective up-regulation of CK2 by mutant huntingtin in cells co-expressing NMDA receptorsJOURNAL OF NEUROCHEMISTRY, Issue 3 2008Mannie M. Y. Fan Abstract Huntington's disease is caused by a polyglutamine expansion in the huntingtin (htt) protein, and previous data indicate that over-activation of NMDA receptors (NMDARs) may be involved in the selective degeneration of cells expressing NR1/NR2B NMDARs. We used KinetworksÔ multi-immunoblotting screens to examine expression of 76 protein kinases, 18 protein phosphatases, 25 heat shock/stress proteins, and 27 apoptosis proteins in human embryonic kidney 293 cells transfected with NR1/NR2B and htt containing 15 (htt-15Q; wild-type) or 138 (htt-138Q; mutant) glutamine repeats. Follow-up experiments revealed several proteins involved in the heat-shock response pathway to be up-regulated in the soluble fraction from cells expressing htt-138Q, including protein phosphatase 5 and cyclin-dependent kinase 5. Increased expression in the soluble fraction of htt-138Q-expressing cells was also noted for the stress- and calcium-activated protein-serine/threonine kinase casein kinase 2, a change which was confirmed in striatal tissue of yeast artificial chromosome transgenic mice expressing full-length mutant htt. Inhibition of casein kinase 2 activity in cultured striatal neurons from these mice significantly exacerbated NMDAR-mediated toxicity, as assessed by labeling of apoptotic nuclei. Our findings are consistent with up-regulation of components of the stress response pathway in the presence of polyglutamine-expanded htt and NR1/NR2B which may reflect an attempt at the cellular level to ameliorate the detrimental effects of mutant htt expression. [source] Alteration of argyrophilic nucleolar organizer region associated (Ag-NOR) proteins in apoptosis-induced human salivary gland cells and human oral squamous carcinoma cellsJOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 4 2001Yasuhiro Morimoto Abstract: The level of argyrophilic nucleolar organizer regions (AgNORs) and AgNOR-associated proteins (Ag-NOR proteins) varies with cell activity, including ribosomal biogenesis occurring in proliferating cells. Proteins associated with some AgNORs are detected by a specific silver staining. To investigate a possible relationship between apoptosis and the AgNORs or Ag-NOR proteins, we examined the changes of AgNORs and Ag-NOR proteins during apoptosis in a human salivary gland cell line, HSG cells, and a human oral squamous carcinoma cell line, SCC-25 cells. Apoptosis was induced by treatment of HSG and SCC-25 cells with okadaic acid. Proteins prepared from HSG and SCC-25 cells treated with varying concentrations of okadaic acid (OA) were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by transferring to transfer membranes and staining for Ag-NOR proteins by modified Western blot analysis. Four major bands (110 kDa, 43 kDa, 39kDa, and 37 kDa) were detected in the proteins obtained from the control cells. The level of the 110-kDa protein decreased in the proteins prepared from OA-induced apoptotic cells; however, the reaction intensity of the other three bands was changed in apoptotic cells. An additional band of an 80-kDa Ag-NOR protein appeared and increased in the apoptotic cells. Cellular fractionation of HSG cells and SCC-25 cells was done with or without apoptotic induction. An 80-kDa Ag-NOR protein was detected in the nuclear fraction prepared from the apoptotic cells, while the 110-kDa protein decreased in the nuclear fraction of these cells. The 110-kDa Ag-NOR protein may be nucleolin (C23) as deduced from its AgNOR staining features, including molecular weight. The 80-kDa protein may be the cleavage product of the 110-kDa protein. In the cell-free apoptotic system, in which intact nuclei of HSG cells were incubated with the cytosol fraction of apoptotic HSG and SCC-25 cells, the 80-kDa Ag-NOR protein was detected in nuclei incubated with the cytosol fraction of apoptotic cells, while the level of the 110-kDa protein decreased. The changes of Ag-NOR proteins in nuclei prepared from SCC-25 cells incubated with cytosol fractions prepared from HSG and SCC-25 cells were identical to those of the HSG cells. The alternation of AgNORs in apoptosis-induced HSG cells was also examined using double staining with Hoechst 33342 and silver nitrate. Hoechst staining revealed typical apoptotic nuclei, which exhibited highly fluorescent condensed chromatin in OA-treated HSG cells. Silver grains representing AgNORs were not detected in the cells undergoing apoptosis. The dual-imposition view confirmed that AgNORs, which are visible as dots in nucleoli in the control cells, disappeared from the apoptotic nuclei of HSG cells. Our results indicate that the 110-kDa nucleolar Ag-NOR protein is associated with apoptosis and is cleaved during apoptosis. [source] The antiproliferative activity of melatonin in B65 rat dopaminergic neuroblastoma cells is related to the downregulation of cell cycle-related genesJOURNAL OF PINEAL RESEARCH, Issue 1 2008Javier G. Pizarro Abstract:, A potential application of melatonin is its ability to rescue many cell types from cell death, because of its antioxidant properties. Likewise, recent studies suggest that melatonin may also be used as an anti-tumor drug, due to its anti-proliferative properties in tumor cells when administered at physiologic or pharmacologic doses. In the present study, we investigated the mechanisms involved in the apoptosis induced by acute exposure to melatonin and roscovitine in the rat dopaminergic neuroblastoma B65 cell line. Cell growth studies revealed that, at 24 hr of treatment, roscovitine blocked cell growth and induced apoptosis whereas melatonin delayed cell growth and induced a slight increase in the number of apoptotic nuclei. Melatonin also increased the percentage of cells in the G1-phase of the cell cycle, whereas roscovitine blocked cells in the G2/M-phase. Both compounds significantly downregulated the transcriptional activity of cdk4, while melatonin also downregulated cdk2 and cyclin D1. Taken together, our data show that melatonin at millimolar concentrations inhibits dopaminergic B65 proliferation, induces cell apoptosis, and modulates cell cycle progression by inhibiting the transcriptional activity of cyclins and cdks related to the progression of the G1-phase. [source] Hypophosphorylation of the architectural chromatin protein DEK in death-receptor-induced apoptosis revealed by the isotope coded protein label proteomic platformPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 21 2006Anja Tabbert Abstract During apoptosis nuclear morphology changes dramatically due to alterations of chromatin architecture and cleavage of structural nuclear proteins. To characterize early events in apoptotic nuclear dismantling we have performed a proteomic study of apoptotic nuclei. To this end we have combined a cell-free apoptosis system with a proteomic platform based on the differential isotopic labeling of primary amines with N -nicotinoyloxy-succinimide. We exploited the ability of this system to produce nuclei arrested at different stages of apoptosis to analyze proteome alterations which occur prior to or at a low level of caspase activation. We show that the majority of proteins affected at the onset of apoptosis are involved in chromatin architecture and RNA metabolism. Among them is DEK, an architectural chromatin protein which is linked to autoimmune disorders. The proteomic analysis points to the occurrence of multiple PTMs in early apoptotic nuclei. This is confirmed by showing that the level of phosphorylation of DEK is decreased following apoptosis induction. These results suggest the unexpected existence of an early crosstalk between cytoplasm and nucleus during apoptosis. They further establish a previously unrecognized link between DEK and cell death, which will prove useful in the elucidation of the physiological function of this protein. [source] Expression of Apoptosis Regulatory Genes and Incidence of Apoptosis in Different Morphological Quality Groups of In Vitro-produced Bovine Pre-implantation EmbryosREPRODUCTION IN DOMESTIC ANIMALS, Issue 5 2010MG Melka Contents Apoptosis occurs during early development in both in vivo - and in vitro -produced embryos, and is considered as one of the causes of embryonic loss. The objectives of this study were, therefore, investigating stage-specific expression profiles of apoptosis regulatory genes in three quality groups of in vitro -produced bovine pre-implantation embryos; and analysing the relationship between cell number and DNA fragmentation with expressions of those genes. The relative abundance of mRNA of 9 pro- (Bax, caspase-9, Bcl-xs, P53, Caspase-3 and Fas) and anti- (Bcl-w and Mcl-1) apoptotic genes was analysed. Differential cell staining and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling were performed to analyse the variation in cell numbers and detect apoptotic nuclei respectively. Expression of Bax and Caspase-3 genes was significantly (p < 0.05) higher in poor quality pre-implantation embryos as compared with that of morphologically good quality embryos of the same developmental stages. Moreover, Mcl-1 expression was significantly higher in good quality immature oocytes than that in the poor quality group. Moreover, higher DNA fragmentation was evidenced in morphologically poor quality blastocysts. In conclusion, our study demonstrates that Bax, caspase-3 and Mcl-1 can be used as potential markers of embryo quality to evaluate in vitro -produced bovine embryos. Further studies are required to investigate specific molecular signatures that can be used in evaluating in vivo -derived embryos. [source] Apoptotic Changes in the Epithelium Germinativum of the Cat (Felis catus s. domestica, L. 1758) at Different Ages and Breeding SeasonsREPRODUCTION IN DOMESTIC ANIMALS, Issue 4 2008MJ SiemieniuchArticle first published online: 25 FEB 200 Contents Apoptosis (programmed cell death) could be considered as a physiological process that takes part in a healthy organism, which helps to maintain organism homeostasis. The visible deterioration of semen quality and the number of germ cells is accompanied by a seasonal decrease of the reproductive activity in some species. This post-effect cascade is caused by apoptosis, which is the primary mechanism responsible for the elimination of germ cells during spermatogenesis. The aim of our study was to assess apoptotic changes in the epithelium germinativum in cat testes at different ages. One hundred and two pairs of testes were obtained from domestic cats aged between 4 months and 10 years. The paraffin-embedded tissue sections were labelled using the Oncogene and Calbiochem Research Products DNA Fragmentation Detection Kit (Cat# QIA21; Darmstadt, Germany), which allows the recognition of apoptotic nuclei in tissue sections with Fragment End Labelling (FragELTM) of DNA. The activity of apoptotic processes in cat testes collected from the spring-summer period compared with the autumn-winter season revealed that, 59.42% and 51.51%, respectively, males testes were characterized by insignificant changes. The obtained data revealed a distinctive apoptotic changes in the young animal testes before spermatogenesis onset. An intensification of programmed death cells in the epithelium germinativum in the elder cats (between 3,6 and 6,10 years) was not observed. Apoptotic changes slightly intensified in cats aged between 12 and 36 months. [source] REVIEW ARTICLE: Placental Apoptosis in Health and DiseaseAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 3 2010Andrew N. Sharp Citation Sharp AN, Heazell AEP, Crocker IP, Mor G. Placental apoptosis in health and disease. Am J Reprod Immunol 2010; 64: 159,169 Apoptosis, programmed cell death, is an essential feature of normal placental development but is exaggerated in association with placental disease. Placental development relies upon effective implantation and invasion of the maternal decidua by the placental trophoblast. In normal pregnancy, trophoblast apoptosis increases with placental growth and advancing gestation. However, apoptosis is notably exaggerated in the pregnancy complications, hydatidiform mole, pre-eclampsia, and intrauterine growth restriction (IUGR). Placental apoptosis may be initiated by a variety of stimuli, including hypoxia and oxidative stress. In common with other cell-types, trophoblast apoptosis follows the extrinsic or intrinsic pathways culminating in the activation of caspases. In contrast, the formation of apoptotic bodies is less clearly identified, but postulated by some to involve the clustering of apoptotic nuclei and liberation of this material into the maternal circulation. In addition to promoting a favorable maternal immune response, the release of this placental-derived material is thought to provoke the endothelial dysfunction of pre-eclampsia. Widespread apoptosis of the syncytiotrophoblast may also impair trophoblast function leading to the reduction in nutrient transport seen in IUGR. A clearer understanding of placental apoptosis and its regulation may provide new insights into placental pathologies, potentially suggesting therapeutic targets. [source] Increased Apoptosis in Human Amnion is Associated with Labor at TermAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 5 2000CHAUR-DONG HSU PROBLEM: To characterize whether increased apoptosis in human amnion was associated with labor at term. METHOD OF STUDY: Human amnion were obtained from term patients with vaginal delivery (n=5) or who underwent elective Cesarean section (C/S) without labor (n=5). Apoptosis was performed by the TUNEL (Terminal dUTP Nuclear End Labeling) assay. All nucleated cells stained with propidium iodide in the amnion epithelial cells were identified in red fluorescence. TUNEL positive apoptotic nuclei were identified in green fluorescence. Five random fields of each specimen were blindly counted by investigators. The percentage of apoptotic nuclei of total nuclei (apoptotic index) was calculated and compared between the two groups (25 microscopic fields for each group, respectively). RESULTS: Patients with term labor had a significantly higher mean apoptotic index in amnion epithelial cells than that with elective C/S without labor (27.3±4.1% versus 3.6±1.6%, P<0.001). CONCLUSIONS: Our data indicate that apoptosis in human amnion is significantly increased and associated with labor at term. [source] Accumulation of apoptotic cells in the epidermis of patients with cutaneous lupus erythematosus after ultraviolet irradiationARTHRITIS & RHEUMATISM, Issue 3 2006Annegret Kuhn Objective To examine whether apoptosis contributes to the pathogenesis of skin lesions in patients with cutaneous lupus erythematosus (CLE) after ultraviolet (UV) irradiation. Methods In situ nick translation and TUNEL were performed to detect apoptosis in 85 skin biopsy specimens from patients with various subtypes of CLE. Specimens from normal healthy donors and patients with polymorphous light eruption were used as controls. In addition to assessment of primary lesions, provocative phototesting was carried out to investigate events occurring secondary to UV irradiation during a very early stage of lesion formation. Results A significant increase in apoptotic nuclei was found in the upper epidermal layer of primary and UV light,induced skin lesions of CLE patients compared with controls. In tissue sections obtained from control subjects at 24 hours after a single exposure to UV light, a slight increase in the count of epidermal apoptotic nuclei was present as compared with skin tissue from CLE patients obtained under the same conditions before lesion formation. In sections obtained from controls at 72 hours after irradiation, a significant decrease in the apoptotic nuclei count was observed, consistent with a proper clearance of apoptotic cells in the period between 24 and 72 hours after irradiation. In striking contrast, the number of apoptotic nuclei increased significantly within this period in tissue sections from patients with CLE. Conclusion These data support the hypothesis that apoptotic cells accumulate in the skin of patients with CLE after UV irradiation, as a result of impaired or delayed clearance. The nonengulfed cells may undergo secondary necrosis and release proinflammatory compounds and potential autoantigens, which may contribute to the inflammatory micromilieu that leads to formation of skin lesions in this disease. [source] Peptidoglycan induces loss of a nuclear peptidoglycan recognition protein during host tissue development in a beneficial animal-bacterial symbiosisCELLULAR MICROBIOLOGY, Issue 7 2009Joshua V. Troll Summary Peptidoglycan recognition proteins (PGRPs) are mediators of innate immunity and recently have been implicated in developmental regulation. To explore the interplay between these two roles, we characterized a PGRP in the host squid Euprymna scolopes (EsPGRP1) during colonization by the mutualistic bacterium Vibrio fischeri. Previous research on the squid-vibrio symbiosis had shown that, upon colonization of deep epithelium-lined crypts of the host light organ, symbiont-derived peptidoglycan monomers induce apoptosis-mediated regression of remote epithelial fields involved in the inoculation process. In this study, immunofluorescence microscopy revealed that EsPGRP1 localizes to the nuclei of epithelial cells, and symbiont colonization induces the loss of EsPGRP1 from apoptotic nuclei. The loss of nuclear EsPGRP1 occurred prior to DNA cleavage and breakdown of the nuclear membrane, but followed chromatin condensation, suggesting that it occurs during late-stage apoptosis. Experiments with purified peptidoglycan monomers and with V. fischeri mutants defective in peptidoglycan-monomer release provided evidence that these molecules trigger nuclear loss of EsPGRP1 and apoptosis. The demonstration of a nuclear PGRP is unprecedented, and the dynamics of EsPGRP1 during apoptosis provide a striking example of a connection between microbial recognition and developmental responses in the establishment of symbiosis. [source] | |||||||||||||