METHODS: The cytotoxic effect of deoxyelephantopin on A549 cells and normal lymphocytes was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and 50% inhibitory concentration (IC50) value was determined. The self-renewal and proliferating potential of A549 cells after treatment with deoxyelephantopin were examined by colony formation assay. Cellular morphology of deoxyelephantopin-treated cells was observed using phase-contrast microscopy. The induction of apoptosis was evaluated using acridine orange and ethidium bromide staining, Hoechst 33342 staining, terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end-labeling (TUNEL) assay, DNA fragmentation analysis and Annexin V-fluorescein isothiocyanate staining by flow cytometry. Activation of caspases was detected using fluorogenic substrate specific to caspases 2, 3, 8 and 9 and flow cytometric analysis. The total cellular DNA content and expression of cleaved poly (ADP-ribose) polymerase was ...
Read independent reviews on CardioTACS™ In Situ Apoptosis Detection Kits from AMS Biotechnology (Archived Products) on SelectScience
Anti-apoptotic action of polyphenols derived from red wine and green tea against beta-amyloid in hippocampal cells Author: Stéphane Bastianetto and Slavica Krantic and Rémi Quirion Background: It has been suggested that accumulation of amyloid-beta (Aß) peptides into senile plaques plays a pivotal role in neuronal cell
BioAssay record AID 541028 submitted by ChEMBL: Induction of apoptosis in bovine BL3 cells assessed as early apoptotic cells after 24 hrs by annexin V/propidium iodide staining-based FACS analysis.
During apoptosis, phosphatidylserine (PS) is translocated from the cytoplasmic face of the plasma membrane to the cell surface. Annexin V has a strong, Ca2+-dependent affinity for PS and therefore is used as a probe for detecting apoptosis., Annexin V-FITC Apoptosis Detection Reagent, GTX14082, Applications: FACS; Flow cytometry/FACS; CrossReactivity:
The Annexin V-Biotin Apoptosis Detection Kit is based on the observation that soon after initiating apoptosis, most cell types translocate the membrane phospholipid phosphatidylserine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can easily bind to Biotin-conjugated Annexin V, a protein that has a strong natural affinity for PS. Annexin V-Biotin can be detected in conjunction with conventional dye-staining using any streptavidine- or avidin-dye reagents, such as (strept)avidine-fluorescein, -peroxidase, -alkaline phosphatase (AP), and -β-gal, etc. |p>‧ Detection method- Flow cytometry (Ex = 488 nm; Em = 530 nm) using and fluorescence microscopy |br>‧ Sample type- Living cells (suspension and adherant)|br>‧ Species reactivity- Mammalian |br>‧ Applications- Detect early/middle stages of apoptosis; differentiate apoptosis from necrosis.|p>|b>Features and Benefits|/b>|br>‧ Simple one step staining procedure in 30 minutes|br>‧ Fast and
TransDetect® Annexin V-EGFP/PI Cell Apoptosis Detection Kit,Cell Detection,Cell Culture and Detection,Products,Beijing TransGen Biotech Co.Ltd,OverviewContents& storageCitations & referencesRelated ImagesDownloadOverviewDescriptionThe Annexin V
Invitrogen™ eBioscience™ Annexin V Apoptosis Detection Kit PerCP-eFluor™ 710 200 tests Invitrogen™ eBioscience™ Annexin V...
Antibodies for proteins involved in negative regulation of cardiac muscle cell apoptotic process pathways, according to their Panther/Gene Ontology Classification
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Flavonoids are naturally occurring polyphenolic compounds and are among the most promising anticancer agents. A series of flavonols and their 3-methyl ether derivatives were synthesized and assessed for cytotoxicity. It was found that 3′-hydroxy-3,4′-dimethoxyflavone (flavonoid 7a) displayed strong cytotoxicity against human SK-MEL-1 melanoma cells and blocked tubulin polymerization, but had no significant cytotoxic effects against quiescent or proliferating human peripheral blood mononuclear cells. Our analyses showed that flavonoid 7a induces G2-M cell cycle arrest and apoptosis in melanoma cells which is associated with cytochrome c release and activation of both extrinsic and intrinsic apoptotic pathways of cell death ...
TY - JOUR. T1 - Formosanin C-induced apoptosis requires activation of caspase-2 and change of mitochondrial membrane potential. AU - Lee, Jenq Chang. AU - Su, Chun Li. AU - Chen, Lin Lin. AU - Won, Shen Jeu. PY - 2009/4/30. Y1 - 2009/4/30. N2 - Formosanin C is a pure compound isolated from Paris formosana Hayata (Liliaceae). The antitumor efficacy of formosanin C has been observed in cultured cells and animal systems. However, the molecular mechanisms of formosanin C remain unknown. The results of the present study indicate that formosanin C induced apoptosis of HT-29 cells characterized by exposure of phosphatidylserine, accumulation of cells at the sub-G1 phase, fragmentation of DNA, and change of nuclear morphology in a time- and dose-related manner. The apoptotic signaling cascades may proceed via proteolytic activation of caspase-2, change of mitochondrial membrane potential (Δεm), release of cytochrome c and second mitochondria-derived activator of caspase/direct IAP binding protein with ...
This kit is designed for histochemical detection of fragmented DNA using the TUNEL method. It allows quick and easy detection of single cells at the primary stage of apoptosis. Tissue sections and fixed cells may be used for samples.
Tumor Necrosis Apoptosis Inducing Ligand (TRAIL) is a death ligand with some specificity for transformed cells. However, some cancer cells develop resistance to TRAIL allowing escape from immune surveillance. Re-sensitization of these cells to TRAIL depends on identifying the mechanism of resistance and applying a targeted corrective. We studied several possible mechanisms of TRAIL resistance beginning with FLICE-Like Inhibitory Protein (FLIP). The short isoform of FLIP (FLIPshort) is an effective inhibitor of caspase 8 activation and therefore overexpression of FLIPshort may be a mechanism of TRAIL resistance. We found that downregulation of FLIP short was sufficient to re-sensitize some prostate carcinoma cells to TRAIL. Another mechanism we investigated is mediated by Elongation Factor 2 (EF2) which acilitates ribosomal translocation along mRNA strands. EF2 can be inactivated by phosphorylation or by ADP-ribosylation, thereby inhibiting protein synthesis. During inhibition of protein ...
TY - JOUR. T1 - Reversibility of apoptosis in cancer cells. AU - Tang, H. L.. AU - Yuen, K. L.. AU - Tang, H. M.. AU - Fung, M. C.. PY - 2009/1/13. Y1 - 2009/1/13. N2 - Apoptosis is a cell suicide programme characterised by unique cellular events such as mitochondrial fragmentation and dysfunction, nuclear condensation, cytoplasmic shrinkage and activation of apoptotic protease caspases, and these serve as the noticeable apoptotic markers for the commitment of cell demise. Here, we show that, however, the characterised apoptotic dying cancer cells can regain their normal morphology and proliferate after removal of apoptotic inducers. In addition, we demonstrate that reversibility of apoptosis occurs in various cancer cell lines, and in different apoptotic stimuli. Our findings show that cancer cells can survive after initiation of apoptosis, thereby revealing an unexpected potential escape mechanism of cancer cells from chemotherapy.. AB - Apoptosis is a cell suicide programme characterised by ...
Protein expression of apoptosis-associated genes by Western blot. K562 cells were treated with different reagents for 24 h. Notes: data were normalized to β-a
세포 죽음의 평가에 대한 정확한 방법을 설명합니다. 프로토콜은 세포 라인과 동물 모델의 광범위한 범위에서 기본 세포 40 % 허위 - 긍정적인 이벤트까지 표시 기존 Annexin V / propidium 요오드화물 (PI) 프로토콜에 따라이...
The fourth meeting on dependence receptors featured descriptions of previously unknown dependence receptors. New mechanistic data were presented on the switch between the trophic, antiapoptotic response with the proapoptotic response that occurs with loss of trophic support. The possibility that the loss of trophic support may also involve the binding of an active antitrophin was also discussed. New in vivo data were presented on the roles of dependence receptors in development, angiogenesis, oncogenesis, and neurodegeneration, as well as new therapeutic approaches based on dependence receptor function. The next meeting on dependence receptors is scheduled for 2012.. ...
As soon as Peripheral Blood Mononuclear Cells (PBMC) are isolated from whole blood, some cells begin dying. The rate of apoptotic cell death is increased when PBMC are shipped, cryopreserved, or stored under suboptimal conditions. Apoptotic cells secrete cytokines that suppress inflammation while promoting phagocytosis. Increased numbers of apoptotic cells in PBMC may modulate T cell functions in antigen-triggered T cell assays. We assessed the effect of apoptotic bystander cells on a T cell ELISPOT assay by selectively inducing B cell apoptosis using α-CD20 mAbs. The presence of large numbers of apoptotic B cells did not affect T cell functionality. In contrast, when PBMC were stored under unfavorable conditions, leading to damage and apoptosis in the T cells as well as bystander cells, T cell functionality was greatly impaired. We observed that measuring the number of apoptotic cells before plating the PBMC into an ELISPOT assay did not reflect the extent of PBMC injury, but measuring apoptotic cell
Few things can be considered to be more important to a cell than its threshold for apoptotic cell death, which can be modulated up or down, but rarely in both directions, by a single enzyme. Therefore, it came as quite a surprise to find that one enzyme, glycogen synthase kinase-3 (GSK3), has the pe …
Apoptosis is a genetically regulated form of cell death that occurs when the cell is exposed to physiological, pathogenic, or cytotoxic stimuli. Unregulated apoptosis (too much or too little apoptosis) at any time from embryogenesis to adulthood, can result in a variety of disease states, such as neurodegenerative disorders, autoimmunity, cardiovascular disease, liver and kidney problems, and cancer. A reasonable estimation is that either too little or too much cell death contributes to half of the main medical illnesses for which adequate therapy or prevention is lacking. The apoptotic pathways can be initiated by reactive oxygen species (ROS) and inflammatory molecules, both of which are believed to be up-regulated in a state of obesity. In addition, multiple studies have shown that the risk of developing cardiovascular disease, type 2 diabetes mellitus, nonalcoholic fatty liver disease, and certain types of cancers increase with increasing degree of obesity in both men and women. Despite the ...
AAD positive means late apoptosis? - posted in Flow Cytometry: We have a discussion in the lab about apoptosis detection by annexin V/AAD labeling. Many people say that Annexin V+/AAD+ cells must not be considered apoptotic but necrotic. I agree, provided that the apoptotic stimulus was rather short (for example 2 hours-staurosporine). However, when the stimulus lasts for 24 hours (serum starvation for instance) I think that cells which started the apoptotic process shortly after the start...
Photodynamic therapy (PDT) is considered to be an advancing antitumor technology. PDT using hydrophilic/lipophilic tetra‑α-(4-carboxyphenoxy) phthalocyanine zinc (TαPcZn-PDT) has exhibited antitumor activity in Bel-7402 hepatocellular cancer cells. However, the manner in which p38 MAPK and caspase-9 are involved in the regulation of mitochondria-mediated apoptosis in the TαPcZn-PDT-treated LoVo human colon carcinoma cells remains unclear. Therefore, in the present study, a siRNA targeting p38 MAPK (siRNA-p38 MAPK) and the caspase‑9 specific inhibitor z-LEHD-fmk were used to examine the crosstalk between p38 MAPK and caspase-9 during mitochondria-mediated apoptosis in the TαPcZn-PDT‑treated LoVo cells. The findings revealed that the TαPcZn-PDT treatment of LoVo cells resulted in the induction of apoptosis, the formation of p38 MAPK/caspase-9 complexes, the activation of p38 MAPK, caspase-9, caspase-3 and Bid, the downregulation of Bcl-2, the reduction of mitochondrial membrane ...
This kit is designed for histochemical detection of fragmented DNA using the TUNEL method. It allows quick and easy detection of single cells at the primary stage of apoptosis. Tissue sections and fixed cells may be used for samples.
Data Availability StatementThe datasets used and/or analyzed through the present study are available from your corresponding author on reasonable request. Mouse monoclonal to Chromogranin A RNA. MTT and Annexin V-fluorescein isothiocyanate/propidium iodide assays shown that CLDN1 silencing significantly inhibits proliferation and enhances apoptosis induced by 5-FU treatment in Hep/5FU cells, compared with non-silenced Hep/5FU cells. Additionally, CLDN1 silencing attenuated the migration and invasion capabilities of Hep/5FU cells. In addition, it was recognized that CLDN1 silencing decreased drug resistance by inhibiting autophagy, which was associated with a decrease in the percentage of microtubule-associated protein 1A/1B-light chain 3 (LC3)-II/LC3-I and upregulation of P62. A cell proliferation assay exposed Indotecan the addition of autophagy inhibitor 3-methyladenine decreased drug resistance of Hep/5FU cells. By contrast, incubation with the autophagy agonist Rapamycin elevated drug ...
Apoptosis is a distinct form of cell death that is functionally and morphologically different from necrosis. Nuclear chromatin condensation, cytoplasmic shrinking, dilated endoplasmic reticulum, and membrane blebbing characterize apoptosis in general. Mitochondria remain morphologically unchanged. In 1972 Kerr et al introduced the concept of apoptosis as a distinct form of cell-death, and the mechanisms of various apoptotic pathways are still being revealed today. Proteins on this pathway have targeted assays available via the CPTAC Assay Portal ...
Apoptosis is a distinct form of cell death that is functionally and morphologically different from necrosis. Nuclear chromatin condensation, cytoplasmic shrinking, dilated endoplasmic reticulum, and membrane blebbing characterize apoptosis in general. Mitochondria remain morphologically unchanged. In 1972 Kerr et al introduced the concept of apoptosis as a distinct form of cell-death, and the mechanisms of various apoptotic pathways are still being revealed today ...
SMAC/DIABLO (second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI) is a proapoptotic mitochondrial protein that is released in response to various apoptotic stimuli. Molecular mimetics of SMAC are being investigated for use in cancer treatment, but their activities in vivo have not been fully characterized.. In a recent study, Emeagi and colleagues from Vrije Universiteit Brussel, Belgium, showed that SMAC mimetics induce cancer cell death via a proinflammatory effect that is accompanied by an adaptive antitumor immune response. Transduction with lentiviral vectors encoding a cytosolic form of a SMAC mimetic (LV-tSMAC) resulted in apoptosis of cancer cells of different histologic origins, and treatment of tumor-bearing mice with the SMAC mimetic resulted in induction of apoptosis, activation of antitumor immunity, and prolonged survival.. Immune Response. The antitumor immune response included increased levels of tumor-infiltrating ...
Hochwertige Annexin V FITC Kits für Apoptose Detektion mit Flow Cytometry. ✅ Namhafte Top-Hersteller im Vergleich. ✅ Top-Support bei Fragen.
Albany, New York, Dec 1, 2017: Apoptosis Regulator BAX (Bcl 2 Like Protein 4 or BCL2L4 or BAX) - Apoptosis regulator BAX or bcl-2-like protein 4 is a protein is encoded by the BAX gene. It accelerates programmed cell death by antagonizing the apoptosis repressor BCL2. It promotes activation of CASP3, and thereby apoptosis. It undergoes a conformation change that causes translocation to the mitochondrion membrane, leading to the release of cytochrome c that then triggers apoptosis under stress conditions.. Request For Free Sample - https://www.marketresearchhub.com/enquiry.php?type=S&repid=1389111. Apoptosis Regulator BAX (Bcl 2 Like Protein 4 or BCL2L4 or BAX) pipeline Target constitutes close to 7 molecules. Out of which approximately 3 molecules are developed by companies and remaining by the universities/institutes. The molecules developed by companies in Phase III, Phase II and Preclinical stages are 1, 1 and 1 respectively. Similarly, the universities portfolio in Preclinical and Discovery ...
Heart ischemia can rapidly induce apoptosis and mitochondrial dysfunction via mitochondrial permeability transition-induced cytochrome c release. We tested whether nitric oxide (NO) can block this damage in isolated rat heart, and, if so, by what mechanisms. Hearts were perfused with 50 μM DETA/NO (NO donor), then subjected to 30 min stop-flow ischemia or ischemia/reperfusion. Isolated heart mitochondria were used to measure the rate of mitochondrial oxygen consumption and membrane potential using oxygen and tetraphenylphosphonium-selective electrodes. Mitochondrial and cytosolic cytochrome c levels were measured spectrophotometrically and by ELISA. The calcium retention capacity of isolated mitochondria was measured using the fluorescent dye Calcium Green-5N. Apoptosis and necrosis were evaluated by measuring the activity of caspase-3 in cytosolic extracts and the activity of lactate dehydrogenase in perfusate, respectively. 30 min ischemia caused release of mitochondrial cytochrome c to the cytoplasm
FACS using DAPI for apoptosis analysis - posted in Flow Cytometry: Hello, I am treating cancer cells with a drug and want to analyse cancer cell death. The only problem is that the drug auto-fluoresce a lot, so much that I cannot use conventional viable stains like PI or 7AAD (because sometimes spillover cannot be corrected). I think using DAPI could solve my problem. However, I am not familiar with DAPI. It is often used to determine cell cycle progress, I think, by histology. However,...
Evidence suggests that the signalling events which occur after apoptotic stimulation, define two basic mechanisms for the induction of apoptosis. The first is dependent on signalling via the mitochondria and the second is dependent upon signalling directly from the death receptors. After induction of apoptosis, there is a convergence in signalling at the level of caspase activation and subsequent biochemical and morphological changes. Therefore the efficacy of various inhibitors of apoptosis is dependent upon the initiating signal. In order to understand the apoptotic pathway, the mechanisms by which these inhibitors regulate chemical- and receptor-mediated apoptosis must be understood. The anti-apoptotic oncoprotein, Bcl-2, was shown to inhibit both staurosporine and Fas-mediated apoptosis in a manner which was partially dependent upon the level of Bcl-2 protein expressed. During both staurosporine and Fas-induced apoptosis Bcl-2 acted downstream of caspase-8 activation. High levels of Bcl-2 ...
The influence of diabetes enhanced inflammation on cell apoptosis and periodontitis. . Biblioteca virtual para leer y descargar libros, documentos, trabajos y tesis universitarias en PDF. Material universiario, documentación y tareas realizadas por universitarios en nuestra biblioteca. Para descargar gratis y para leer online.
Drug resistance has been mostly explained by molecular changes, such as overexpression of p-glycoprotein or O6-methylguanine-DNA-methyltransferase, which interfere with drug actions on the targets (1 , 2) . Because most anticancer agents, regardless of their diverse targets, exert effects by inducing apoptosis via activation of apoptotic pathways common to many cellular stresses, any antiapoptotic changes disrupting the common intrinsic pathways to execute physiological cell death can also make malignant cells resistant to chemotherapy (3) . Such alterations opposing apoptosis are routinely observed in malignant tumors, including both the functional loss of tumor suppressors p53, Bax, or PTEN and deregulated hyperfunction of oncogenic proteins, such as Ras, Bcl-2, and PI3K 2 ,(3) .. In tumor cells with an aberrantly activated PI3K/Akt survival pathway, the increased antiapoptotic signals overcome apoptotic signals of anticancer drugs, confer drug resistance on the tumor cells, and result in a ...
New York - September 19, 2019 - BOC Sciences, a New York-based supplier of various bio-chemicals such as inhibitors, APIs, metabolites and impurities, announced earlier this month to launch cell apoptosis assay service for scientists engaged in drug discovery research. With expertise and well-published cell based assays from BOC Sciences, researchers now dont need to worry about apoptosis detection anymore.. Apoptosis, or alternatively referred to as programmed cell death, plays an important role in many human diseases. Controlled by multiple signaling and effector pathways, apoptosis is found to be closely linked with caspase related apoptotic enzyme cascades, mitochondrial depolarization, DNA fragmentation and ultimately cell blebbing and destruction. This offers new insights for scientists who are seeking new treatment for various diseases: through approaches of genetically modulating these apoptosis-associated pathways, new therapies might be discovered. Many human conditions like ...
Apoptosis is characterized by a series of well defined morphological and biochemical features that allow cells to initiate self-destruction in response to a variety of stimuli. CD4⁺CD8⁺ is a sub-population of immature thymocytes that are especially prone to the action of apoptosis-inducing agents and are sensitive to glucocorticoid-induced apoptosis, an event that plays a critical role in eliciting the antigen-specific thymocyte repertoire. Glucocorticoids induce apoptosis through activation of the GR, a ligand-induced transcription factor that transduces the hormonal signals into the regulated expression of target genes. While much is known about the structure and function of GR, key steroid-regulated genes believed to be required for thymocyte apoptosis have not been found. Based on the transcriptional-regulation of apoptosis by ecdysone-mediated induction of reaper gene expression in Drosophila, and p53-mediated transcriptional-activation of Bax gene expression in mammalian cells, our ...
Zeng L., Li T., Xu D.C., Liu J., Mao G., Cui M.Z., Fu X., Xu X.. Cells undergo apoptosis through two major pathways, the extrinsic pathway (death receptor pathway) and the intrinsic pathway (the mitochondrial pathway). These two pathways can be linked by caspase-8-activated truncated Bid formation. Very recently, death receptor 6 (DR6) was shown to be involved in the neurodegeneration observed in Alzheimer disease. DR6, also known as TNFRSF21, is a relatively new member of the death receptor family, and it was found that DR6 induces apoptosis when it is overexpressed. However, how the death signal mediated by DR6 is transduced intracellularly is not known. To this end, we have examined the roles of caspases, apoptogenic mitochondrial factor cytochrome c, and the Bcl-2 family proteins in DR6-induced apoptosis. Our data demonstrated that Bax translocation is absolutely required for DR6-induced apoptosis. On the other hand, inhibition of caspase-8 and knockdown of Bid have no effect on DR6-induced ...
Apoptosis is a normal physiological process which eliminates cells that do not receive adequate extracellular signals. One of the pathways signalling apoptosis is controlled by the small GTPases of the Rho family, also involved in cell proliferation, differentiation and motility. Another major apoptosis signalling pathway involves the p53 tumour suppressor which is activated by a variety of stress and mediates growth arrest or apoptosis in normal cells. We show here that upon detachment from the extracellular matrix, fibroblasts undergo rapid apoptosis that can be rescued by constitutive activation of Rac1 and Cdc42Hs GTPases. Conversely, inhibition of Rac1 and Cdc42Hs efficiently triggers apoptosis in adherent cells. Interestingly, apoptosis is not observed in p53-/- cells either cultured in suspension or inhibited for Rac1 and Cdc42Hs activity. Moreover, Rac1 and Cdc42Hs extinction in normal cells activates endogenous p53. Using specific inhibitors of MAPK pathways, we demonstrate that, in our ...
As one of the cellular death mechanisms, apoptosis, also known as programmed cell death, can be defined as the process of a proper death of any cell under certain or necessary conditions. Apoptosis is controlled by the interactions between several molecules and responsible for the elimination of unwanted cells from the body.. Many biochemical events and a series of morphological changes occur at the early stage and increasingly continue till the end of apoptosis process. Morphological event cascade including cytoplasmic filament aggregation, nuclear condensation, cellular fragmentation, and plasma membrane blebbing finally results in the formation of apoptotic bodies. Several biochemical changes such as protein modifications/degradations, DNA and chromatin deteriorations, and synthesis of cell surface markers form morphological process during apoptosis.. Apoptosis can be stimulated by two different pathways: (1) intrinsic pathway (or mitochondria pathway) that mainly occurs via release of ...
Explore EarlyTox Caspase-3/7 NucView 488 Assay Kits that enables detection of apoptosis within intact cell populations through the use of NucView 488 Caspase-3 substrate.
The detection of DNA fragmentation by the use of the TUNEL technique has become a standard technique for the detection of apoptosis in tissue sections. DNA cleavage, detected by the TUNEL technique, is the last irreversible stage of the apoptosis cascade. When the nuclear DNA is cleaved in oligonucl …
Annexin V-Biotin Apoptosis Reagent (ab14165). Quick and reliable detection of phosphatidylserine exposure by flow cytometry or microscopy.
The molecular nature of calcium (Ca(2+))-dependent mechanisms and the ion channels having a major role in the apoptosis of cancer cells remain a subject of debate. Here, we show that the recently identified Orai1 protein represents the major molecular component of endogenous store-operated Ca(2+) entry (SOCE) in human prostate cancer (PCa) cells, and constitutes the principal source of Ca(2+) influx used by the cell to trigger apoptosis. The downregulation of Orai1, and consequently SOCE, protects the cells from diverse apoptosis-inducing pathways, such as those induced by thapsigargin (Tg), tumor necrosis factor α, and cisplatin/oxaliplatin. The transfection of functional Orai1 mutants, such as R91W, a selectivity mutant, and L273S, a coiled-coil mutant, into the cells significantly decreased both SOCE and the rate of Tg-induced apoptosis. This suggests that the functional coupling of STIM1 to Orai1, as well as Orai1 Ca(2+)-selectivity as a channel, is required for its pro-apoptotic effects. We have
For a TUNEL assay, dissolve DNase I in 50 mM Tris-HCl, pH 7.5, 1 mg/ml BSA. The final concentration will depend on the sample being processed. In general, start by using a low concentration (1 U/ml for 10 min at +15 to +25 °C). This procedure is described in the pack inserts of the In Situ Cell Death Detection Kits. Follow the steps described in the troubleshooting section of the pack inserts of the In Situ Cell Death Detection Kits when having problems with positive controls ...
Bullacta exarata was hydrolyzed with trypsin to prepare peptides; Hydrolysates were isolated by ultrafiltration and purified using G-25 gel filtration. The purity of the Bullacta exarata was demonstrated by HPLC and its peptide sequence analysis was detected. The effects of BEPT II and BEPT II-1 on the proliferation of PC-3 cells were examined using a MTT assay. BEPT II and BEPT II-1 significantly inhibited the proliferation of PC-3 cells in a time- and dose-dependent manner. Annexin V/PI double staining studies showed exposing PC-3 cells to 5, or 15 mg/mL BEPT II-1 for 24 h increased the percentage of the early stage of apoptotic cells from 11.22% to 22.09%. In addition, typical morphologic changes were observed in the cells with acridine orange/ethidium bromide staining. These data support that BEPT II-1 has anticancer properties and merits further investigation to understand the mechanisms of BEPT II-1-induced apoptosis in PC-3 cells.
The BCL-2 family of proteins forms a complex interaction network that regulates cellular life and death decisions and contributes to cancer development, maintenance, and chemoresistance. BH3 only member proteins (e.g. BIM) serve as cellular stress sentinels and, when triggered, signal irreversible activation of apoptosis through their α-helical BH3 death domains. These pro-apoptotic signals are normally held in check by the multidomain anti-apoptotic proteins (e.g. BCL-XL, MCL-1) but when they are unable to do so the multidomain pro-apoptotic proteins BAX and BAK induce cell death through pore formation in the mitochondrial outer membrane. Therapeutic manipulation of the BCL-2 family with BH3 mimetics (including small molecules and synthetic peptides) is an emerging paradigm in cancer treatment and immune modulation. The design of next-generation therapeutics based on the BIM BH3 helix offers the unique advantage of recapitulating BIMs natural capacity to directly target the full complement of ...
Mice and primary neuronal cultures. Hq, Apaf1-/-, and Bax-/- mice were maintained and genotyped as described previously (Fortin et al., 2001; Klein et al., 2002). Cortical neurons and cerebellar granule neurons (CGNs) were cultured from cortices of E15.5 mice and cerebellums of postnatal day 7 mice, respectively, as described previously (Fortin et al., 2001). Recombinant adenoviral vectors carrying human AIF or LacZ expression cassettes were prepared and used at 50 multiplicity of infection as described previously (Cregan et al., 2002).. Camptothecin treatment and cell viability assays. Neurons were treated with 10 μm camptothecin with or without 10 μm Boc-aspartyl (Ome)-fluoromethylketone (BAF) (Enzyme System Products, Livermore, CA) after 2 d in vitro (DIV). Cell survival was measured by the following: live/dead staining (Molecular Probes, Eugene, OR), Hoechst staining, MTT assay (Cell Titer kit; Promega, Madison, WI), terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end ...
TY - JOUR. T1 - Molecular cloning of a human cDNA encoding a novel protein, DAD1, whose defect causes apoptotic cell death in hamster BHK21 cells. AU - Nakashima, Torahiko. AU - Sekiguchi, Takeshi. AU - Kuraoka, Akio. AU - Fukushima, Kohtarou. AU - Shibata, Yosaburo. AU - Komiyama, Sohtaro. AU - Nishimoto, Takeharu. N1 - Copyright: Copyright 2017 Elsevier B.V., All rights reserved.. PY - 1993/10. Y1 - 1993/10. N2 - The tsBN7 cell line, one of the mutant lines temperature sensitive for growth which have been isolated from the BHK21 cell line, was found to die by apoptosis following a shift to the nonpermissive temperature. The induced apoptosis was inhibited by a protein synthesis inhibitor, cycloheximide, but not by the bcl-2-encoded protein. By DNA-mediated gene transfer, we cloned a cDNA that complements the tsBN7 mutation. It encodes a novel hydrophobic protein, designated DAD1, which is well conserved (100% identical amino acids between humans and hamsters). By comparing the base sequences ...
Introduction Internucleosomal cleavage of DNA is a hallmark of apoptosis. DNA cleavage in apoptotic cells can be detected in situ in fixed cells or tissue sections using the terminal deoxynucleotidyl transferase (TdT) mediated dUTP nick-end labeling (TUNEL) method. TUNEL is highly selective for the detection of apoptotic cells but not necrotic cells or cells with DNA strand breaks resulting from irradiation or drug treatment. In the TUNEL assay, TdT enzyme catalyzes the addition of labeled dUTP to the 3 ends of cleaved DNA fragments. Hapten-tagged dUTP (e.g. digoxigenin-dUTP or biotin-dUTP) can be detected using secondary reagents (e.g. anti-digoxigenin antibodies or streptavi-din) for fluorescence or colorimetric detection. Alternatively, fluorescent dye-conjugated dUTP can be used for direct detection of fragmented DNA by fluorescence microscopy or flow cytometry. The TUNEL Andy Fluor™ 594 Apoptosis Detection Kit contains dUTP conjugated to biotin and Streptavidin conjugated to bright and ...
The purpose of this study is to determine if a preventive treatment with curcumin can protect intestinal epithelial cells from inflammatory damage induced by IFNγ. To achieve this goal we have used a human intestinal epithelial cell line (HT29) treated with IFNγ to undergo apoptotic changes that can reproduce the damage of intestinal epithelia exposed to inflammatory cytokines. In this model, we measured the effect of curcumin (curcuminoid from Curcuma Longa) added as a pre-treatment at different time intervals before stimulation with IFNγ. Curcumin administration to HT29 culture before the inflammatory stimulus IFNγ reduced the cell apoptosis rate. This effect gradually declined with the reduction of the curcumin pre-incubation time. This anti-apoptotic action by curcumin pre-treatment was paralleled by a reduction of secreted IL7 in the HT29 culture media, while there was no relevant change in the other cytokine levels. Even though curcumin pre-administration did not impact the activation of the
TY - JOUR. T1 - Mifepristone regulates expression of apoptosis related genes Fas and FasL in mouse endometrium. AU - Gao, F.. AU - Xu, F. H.. AU - Zhou, X. C.. AU - Han, X. B.. AU - Liu, Y. X.. PY - 2001/7/3. Y1 - 2001/7/3. N2 - AIM: To investigate the anti-implantation mechanism of mifepriston. METHODS: In situ hybridization and immunohistochemistry were applied to determine mRNA and protein. RESULTS: After mifepriston injection, the number of implantation sites were obviously reduced, mifepriston could inhibit the embryo implantation in mouse. The expression of apoptosis related genes, Fas and FasL, in mouse endometrium was also decreased after mifepriston treatment. CONCLUSION: The expression of apoptosis related genes Fas and FasL is regulated by mifepriston and the inhibitory effect of mifepriston on the embryo implantation may be mediated by action on the Fas/FasL system.. AB - AIM: To investigate the anti-implantation mechanism of mifepriston. METHODS: In situ hybridization and ...
title: Galpha12 Protects Vascular Endothelial Cells from Serum Withdrawal-Induced Apoptosis through Regulation of miR-155, doi: 10.3349/ymj.2016.57.1.247, category: Article
Vascular Endothelial Growth Inhibitor (VEGI) is an endothelial cell autocrine factor and a member of the tumor necrosis family of ligands. VEGI is able to specifically inhibit endothelial cell growth and is an efficient inhibitor of angiogenesis. The molecular mechanisms of VEGI activity on endothelial cells remain undefined. Here we focused on two important steps in the signal transduction of VEGI. We first determined a role of NF-κB in VEGI-induced apoptosis. We found that inhibition of the NF-κB pathway resulted in an increased apoptotic potential of VEGI. We conclude that the NF-κB pathway plays a role in suppressing the apoptotic potential of VEGI. We next investigated the receptor responsible for VEGI-induced endothelial cell apoptosis. DR3 is a receptor for VEGI and thus we first focused on confirming if DR3 is the receptor responsible for VEGI-mediated endothelial cell death. We determined VEGI had diminished apoptotic activity in endothelial cells that are depleted of DR3 by siRNA. ...
HOTAIR was known to enhance radioresistance in several cancers. However, the function of HOTAIR on radioresistance involving the regulation of HIF-1α in cervical cancer has not been reported. BALB/c nude mice were injected subcutaneously with HeLa cells and irradiated by X-ray. The tumor volume was measured and the expression of HOTAIR in tumors was detected by quantitative real-time PCR. Western blot was performed to detect the protein level of HIF-1α. MTT (3-(4,5-Dimethylthiazol-2-yl) 22,5-diphenyltetrazolium bromide) assay and the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was used to examine the cell viability and cell apoptosis of HeLa cells and C33A cells exposed to radiation. Radiotherapy inhibited the tumor growth in mice bearing HeLa cells. Radiotherapy reduced the expression of HOTAIR and HIF-1α in tumor tissues and HeLa cells or C33A cells. HOTAIR overexpression abrogated the effect of radiation on the cell viability and cell apoptosis of HeLa and C33A cells
Methods and Findings: Studies were performed using wild type (WT) and A2 knockout (A2-/-) mice exposed to Oxygen Induced Retinopathy (OIR). Neuronal injury and apoptosis were assessed using immunohistochemistry, TUNEL (terminal deoxynucleotidyl transferase dUTP nick end) labeling and Western blotting. Electroretinography (ERG) was used to assess retinal function. Neuro-glial injury in WT ROP mice was evident by TUNEL labeling, retinal thinning, decreases in number of rod bipolar cells and glial cell activation as compared with room air controls. Significant reduction in numbers of TUNEL positive cells, inhibition of retinal thinning, preservation of the rod bipolar cells and prevention of glial activation were observed in the A2-/- retinas. Retinal function was markedly impaired in the WT OIR mice as shown by decreases in amplitude of the b-wave of the ERG. This defect was significantly reduced in A2-/- mice. Levels of the pro-apoptotic proteins p53, cleaved caspase 9, cytochrome C and the ...
Pure ginsenoside standards (saponins Rh2, PD, and PT), along with an Rh2-enhanced North American ginseng (Panax quinquefolius) leaf extract (LFRh2), were tested for cytotoxic activity in cultured THP-1 leukemia cells. Thermal treatment of ginseng leaf resulted in production of both Rh2 and Rg3 content that was confirmed by liquid chromatography-mass spectrometry (LC-MS). Flow cytometry of cells stained with annexin V-fluorescein isothiocyanate and propidium iodide showed that the LFRh2 significantly (p ≤ 0.05) increased apoptosis (18% ± 0.4%) after 23 h at a concentration that inhibited cell viability by 50% (LC50 (72 h) = 52 μmg/mL. In comparison, a similar significant (p ≤ 0.05) increase in apoptotic cell numbers occurred at 41 h of exposure for pure ginsenoside standards, PD (LC50 (72 h) = 13 μg/mL), PT (LC50 (72 h) = 19 μg/mL), and Rh2 (LC 50 (72 h) = 15 μg/mL). Although no further increase in apoptosis was observed in THP-1 cells after exposure to increasing concentrations of LFRh2 ...
TY - JOUR. T1 - FADD/MORT1 is a common mediator of CD95 (Fas/APO-1) and tumor necrosis factor receptor-induced apoptosis. AU - Chinnaiyan, Arul M.. AU - Tepper, Clifford G. AU - Seldin, Michael F. AU - ORourke, Karen. AU - Kischkel, Frank C.. AU - Hellbardt, Stefan. AU - Krammer, Peter H.. AU - Peter, Marcus E.. AU - Dixit, Vishva M.. PY - 1996/3/1. Y1 - 1996/3/1. N2 - CD95 (Fas/APO-1) and tumor necrosis factor receptor-1 (TNFR-1) are related molecules that signal apoptosis. Recently, a number of novel binding proteins have been proposed to mediate the signaling of these death receptors. Here we report that an N-terminal truncation of one of these candidate signal transducers, FADD/MORT1, abrogates CD95-induced apoptosis, ceramide generation, and activation of the cell death protease Yama/CPP32. In addition, this dominant-negative derivative of FADD (FADD-DN) blocked TNF- induced apoptosis while not affecting NF-κB activation. FADD-DN bound both receptors, and in the case of CD95, it disrupted ...
In this paper, the use of (123)I-Annexin V for the detection of farnesyltransferase inhibitor (FTI)-induced apoptosis in tumour-bearing athymic mice is described. In vitro binding assays on LoVo cells show time- and dosage-dependent (125)I-Annexin V binding upon treatment with Tipifarnib (Zarnestra,
Paclitaxel is used to treat breast cancers, but the mechanisms by which it induces apoptosis are poorly understood. Consequently, we have studied the role of the FoxO transcription factors in determining cellular response to paclitaxel. Western blotting revealed that in a panel of nine breast cancer cell lines expression of ... read more FoxO1a and FoxO3a correlated with the expression of the pro-apoptotic FoxO target Bim, which was associated with paclitaxel-induced apoptosis. In MCF-7 cells, which were paclitaxel-sensitive, the already high basal levels of FoxO3a and Bim protein increased dramatically after drug treatment, as did Bim mRNA, which correlated with apoptosis induction. This was not observed in MDA-231 cells, which expressed low levels of FoxOs and Bim. Gene reporter experiments demonstrated that in MCF-7 cells maximal induction of Bim promoter was dependent on a FoxO binding site, suggesting that FoxO3a is responsible for the transcriptional up-regulation of Bim. Gene silencing ...
Objectives : We investigated the effect of Haein-tang(Hairen-tang) on short-term memory and apoptosis in dentate gyrus of the gerbils with transient global ischemia. Methods : For the induction of cerebral ischemia model in mice, common carotid arteries of gerbils were occluded with aneurysm clips for 5 min. One day after operation, Haein-tang(Hairen-tang) was administrated orally injected once a day for 15 consecutive days. Gerbils were randomly divided into four group(n=10 in each group): sham-operation group, ischemia-induction group, ischemia-induction and 50 mg/kg Haein-tang(Hairen-tang)-treated group, ischemia-induction and 100 mg/kg Haein-tang(Hairen-tang)-treated group, and ischemia-induction and 200 mg/kg Haein-tang(Hairen-tang)-treated group. The effect of Haein-tang(Hairen-tang) on memory function was investigated by using step-down avoidance task. Apoptosis was confirmed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling(TUNEL) staining and immunohistochemistry ...
We used cytoplasmic extracts from chicken DU249 cells at various stages along the apoptotic pathway to analyse the events of apoptotic exe-cution. So-called S/M extracts from morphologically normal committed-stage cells induce apoptotic morphology and DNA cleavage in substrate nuclei. These apoptotic changes appear to require the function of multiple caspases (cysteine aspar-tases, a specialized class of proteases) acting in parallel. Extracts from execution-stage apoptotic cells induce apoptotic events in added nuclei in a caspase-independent manner. Biochemical frac-tionation of these extracts reveals that a column fraction enriched in endogenous active caspases is un-able to induce DNA fragmentation or chromatin condensation in substrate nuclei, whereas a caspase-depleted fraction induces both changes. Execution-stage extracts contain an ICAD/DFF45-inhibitable nuclease resembling CAD, plus another activity that is required for the apoptotic chromatin condensation. Committed-stage S/M ...
As a process, apoptosis integrates a multitude of pro- and anti-cell death signals to a single decision for life or death. This type of all-or-none response to quantitative inputs is common in biological phenomena such as cell proliferation and differentiation and is often associated with a network topology containing positive feedback loops (28). While the ability of caspases to self-cleave provides one level of such feedback activity (17), we find that this feedback extends to cross activation between both the extrinsic and intrinsic pathways, with specific activation of either extrinsic or intrinsic apoptosis leading to near simultaneous activation of both caspase-8 and -9 in all apoptotic cells tested in this experiment. This supports a model wherein, although initiation of caspases is segregated by distinct pathways, an apoptotic signal, once having reached a threshold level, leads to a simultaneous activation across both arms of the apoptotic network. Several signaling mechanisms ...
In previous study we showed that caspase-2 plays the role of an apical caspase in cell death induction by taxanes in breast cancer cells. This study deals with the role of other caspases. We tested breast cancer cell lines SK-BR-3 (functional caspase-3) and MCF-7 (nonfunctional caspase-3). Using western blot analysis we demonstrated the activation of initiator caspase-8 and -9 as well as executioner caspase-6 and -7 in both tested cell lines after application of taxanes (paclitaxel, SB-T-1216) at death-inducing concentrations. Caspase-3 activation was also found in SK-BR-3 cells. Employing specific siRNAs after taxane application, suppression of caspase-3 expression significantly increased the number of surviving SK-BR-3 cells. Inhibition of caspase-7 expression also increased the number of surviving SK-BR-3 and MCF-7 cells. On the other hand, suppression of caspase-8 and caspase-9 expression had no significant effect on cell survival. However, caspase-9 seemed to be involved in the activation of
TY - JOUR. T1 - Calcineurin Aβ Gene Targeting Predisposes the Myocardium to Acute Ischemia-Induced Apoptosis and Dysfunction. AU - Bueno, Orlando F.. AU - Lips, Daniel J.. AU - Kaiser, Robert A.. AU - Wilkins, Benjamin J.. AU - Dai, Yan Shan. AU - Glascock, Betty J.. AU - Klevitsky, Raisa. AU - Hewett, Timothy E.. AU - Kimball, Thomas R.. AU - Aronow, Bruce J.. AU - Doevendans, Pieter A.. AU - Molkentin, Jeffery D.. N1 - Copyright: Copyright 2008 Elsevier B.V., All rights reserved.. PY - 2004/1/9. Y1 - 2004/1/9. N2 - Cardiovascular disease is the leading cause of mortality and morbidity within the industrialized nations of the world, with coronary heart disease (CHD) accounting for as much as 66% of these deaths. Acute myocardial infarction is a typical sequelae associated with long-standing coronary heart disease resulting in large scale loss of ventricular myocardium through both apoptotic and necrotic cell death. In this study, we investigated the role that the calcium calmodulin-activated ...
TY - JOUR. T1 - Nitrite generation and antioxidant effects during neutrophil apoptosis. AU - Misso, Neil L.A.. AU - Peacock, Craig D.. AU - Neil Watkins, D.. AU - Thompson, Philip J.. PY - 2000/3/15. Y1 - 2000/3/15. N2 - Neutrophil apoptosis is important for the resolution of airway inflammation in a number of lung diseases. Inflammatory mediators, endogenous reactive oxygen and nitrogen species, and intracellular and extracellular antioxidants may all influence neutrophil apoptosis. This study investigated the involvement of these factors during apoptosis of neutrophils cultured in vitro. Neutrophils undergoing spontaneous apoptosis in culture as assessed by annexin V binding generated significant amounts of nitrite. Incubation with agonistic anti-Fas monoclonal antibody or tumor necrosis factor-α (TNF-α) enhanced neutrophil apoptosis at 6 h, although it decreased nitrite accumulation. Although granulocyte-macrophage colony-stimulating factor significantly reduced neutrophil apoptosis, this ...
Caspase 3 is an essential death factor for the Fas-mediated cell death, and its inactivation in cells is initiated by an interaction with p21 on mitochondria or with IAP family member ILP. Survivin is also a member of IAP family and is specifically expressed during embryogenesis and in tumor cells and suppresses cell death signaling. In our current study, we demonstrated that Survivin translocation into the nucleus is dependent on Fas stimulation and cell proliferation. Survivin also interacts with the cell cycle regulator Cdk4, leading to Cdk2/Cyclin E activation and Rb phosphorylation. As a result of Survivin/Cdk4 complex formation, p21 is released from its complex with Cdk4 and interacts with mitochondrial procaspase 3 to suppress Fas-mediated cell death. Here, we propose that Survivin supports procaspase 3/p21 complex formation as a result of interaction with Cdk4 resulting in suppression of cell death signaling.
TY - JOUR. T1 - Pretreatment with low nitric oxide protects osteoblasts from high nitric oxide-induced apoptotic insults through regulation of c-Jun N-terminal kinase/c-Jun-mediated Bcl-2 gene expression and protein translocation. AU - Tai, Yu-Ting. AU - Cherng, Yih-Giun. AU - Chang, Chia Chen. AU - Hwang, Yi Ping. AU - Chen, Jui Tai. AU - Chen, Ruei-Ming. PY - 2007/5. Y1 - 2007/5. N2 - Nitric oxide (NO) can regulate osteoblast activity. In this study, we evaluated the effects of pretreatment with a low concentration of NO on osteoblast injuries induced by a high level of NO and its possible molecular mechanisms. Exposure of osteoblasts to 0.3 mM sodium nitroprusside (SNP), an NO donor, slightly increased cellular NO levels without affecting cell viability. SNP at 2 mM greatly increased the levels of cellular NO and reactive oxygen species, and induced osteoblast death. Thus, osteoblasts were treated with 0.3 and 2 mM SNP as the sources of low and high NO, respectively. Exposure of osteoblasts ...
Because defects in apoptosis programs, for example, high expression of antiapoptotic molecules, can cause resistance to treatment regimens including radiotherapy (8), current attempts to improve the outcome of glioblastoma patients depend on strategies to increase apoptosis sensitivity. In this study, we identify a new proapoptotic role of NF-κB in γ-irradiation-mediated apoptosis of glioblastoma cells by showing, for the first time, that NF-κB is critically required for Smac mimetic-triggered radiosensitization. This conclusion is supported by several independent pieces of evidence. First, BV6 and γ-irradiation cooperate to trigger apoptosis in glioblastoma cells. This interaction is highly synergistic in A172 glioblastoma cells (CI , 0.3). Second, BV6 stimulates NF-κB activation, which is required for the potentiation of γ-irradiation-induced apoptosis because genetic inhibition of NF-κB by overexpression of the dominant-negative superrepressor IκBα-SR significantly reduces BV6- and ...
TY - JOUR. T1 - Nitric oxide-GAPDH-Siah. T2 - a novel cell death cascade.. AU - Hara, Makoto R.. AU - Snyder, Solomon H. PY - 2006/7. Y1 - 2006/7. N2 - 1. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an extremely abundant glycolytic enzyme, and exemplifies the class of proteins with multiple, seemingly unrelated functions. Recent studies indicate that it is a major intracellular messenger mediating apoptotic cell death. This paper reviews the GAPDH cell death cascade and discusses its clinical relevance. 2. A wide range of apoptotic stimuli activate NO formation, which S-nitrosylates GAPDH. The S-nitrosylation abolishes catalytic activity and confers upon GAPDH the ability to bind to Siah, an E3-ubiquitin-ligase, which translocates GAPDH to the nucleus. In the nucleus, GAPDH stabilizes the rapidly turning over Siah, enabling it to degrade selected target proteins and affect apoptosis. 3. The cytotoxicity of mutant Huntingtin (mHtt) requires nuclear translocation which appears to be ...
3H]Thymidine Incorporation Studies. Thymidine incorporation studies were performed with cells grown in 35-mm culture dishes. The cells were labeled by the addition of methyl-[3H]thymidine (1 μCi/ml) to the medium, and the reactions were stopped after 1 h. The medium was drawn off, and the cells were rinsed twice with ice-cold phosphate-buffered saline (PBS). The cultures were then fixed with ice-cold 5% trichloracetic acid overnight at 4°C, after which the cells were extracted as previously described (Guo and Reiners, 2000). A second replicate set of nonfixed dishes was treated with 0.25% trypsin-EDTA to estimate cell numbers. [3H]Thymidine was detected by scintillation counting and expressed as disintegration per minute per 103 cells.. Assessment of Apoptosis. The effects of HET0016 to induced apoptosis in 9L cells were examined by fluorescence-activated cell sorting analysis after the cells were labeled with an annexin V-fluorescein isothiocyanate (FITC) antibody (Sigma Chemical, St. Louis, ...
Previous studies suggested that dithio-carbamates are potent apoptosis and anti-apoptosis inducing agents in various cancer cells. Here, the anti-proliferative and apoptosis inducing effects of a new derivative (2-NDC) from the dithio-carbamate family was examined in human leukemia K562 cells. We use thiazolyl blue tetrazolium bromide (MTT) to measure viability and cell growth inhibition. The 2-NDC showed effects on viability in a dose and time-dependent manner, inhibiting proliferation at concentrations of 10-30 M after 24-48 hours of treatment and increasing values after 72 hours at 40-120 M. The cytotoxic effect of the compound was calculated with an IC50 of 30 M after 24-hour. Apoptosis induction was confirmed by acridine orange-ethidium bromide (AO/EtBr) staining, DNA fragmentation assay, flow cytometric assessment and also caspase-3 activation assay. Furthermore, enzymes level such as superoxide dismutase (SOD) and catalase (CAT) involved in oxidative stress were evaluated. The results of this
Our results uncovered an unexpected role for MPO to influence the fate of neutrophils and consequently the duration of inflammation. By suppressing the constitutive cell death program, MPO prolonged the life span of neutrophils, thereby delaying the resolution of inflammation. These actions were specific for MPO, because other azurophilic granule constituents lactoferrin and elastase failed to affect neutrophil apoptosis.. Consistent with the commitment of neutrophils to apoptosis, MPO at clinically relevant concentrations delayed, rather than blocked apoptosis, resulting in prolonged survival of human neutrophils in vitro. We confirmed that increasing plasma concentrations of MPO in rats to levels comparable to those detected in patients with inflammatory vascular diseases20,21 was sufficient to retard the apoptotic machinery in neutrophils as assayed ex vivo. Furthermore, MPO also suppressed apoptosis in neutrophils that had emigrated into the airways and delayed resolution of inflammation in ...
The effects of the acute phase proteins AGP and AAT on early (2-hour) and late (24-hour) apoptosis and inflammation after renal I/R were investigated. Both AGP and AAT administered at reperfusion decrease early as well as late apoptosis, as reflected by renal internucleosomal DNA cleavage, TUNEL histology, and caspase-like activities. In line, we previously demonstrated that abrogating acute early apoptosis with selective antiapoptotic agents prevents subsequent inflammation as well as secondary apoptosis caused by inflammation, whereas antiapoptotic treatment that is initiated after the onset of apoptosis does not reduce I/R-induced inflammation.1 In contrast, in the present study, treatment after 2 hours of reperfusion inhibited inflammation, which is supported by reports of anti-inflammatory effects mediated by AGP and AAT.12 13 Early primary19 20 as well as late secondary2 21 apoptosis after I/R has been reported to be caused by various means. The present results, showing that AGP and (to a ...
TY - JOUR. T1 - Response of Apoptosis Related Proteins to Running in Fluoride-Exposed Mice. AU - Canning, M.. AU - Zohoori, Fatemeh. AU - Valentine, Ruth. AU - Khan, Z.. AU - Ahmed, A.. AU - Amaral, A.. AU - Azevedo, Liane. AU - Buzalaf, Maria. AU - Fabricio, M.. AU - Fernandes, M.. AU - Maguire, Anne. PY - 2017. Y1 - 2017. M3 - Meeting Abstract. VL - 96A. JO - Journal of Dental Research. JF - Journal of Dental Research. SN - 0022-0345. ER - ...
An in vitro ischemia model was established and the effect of the metabolic inhibitors cycloheximide (CHX) and actinomycin D (ActD) on apoptosis in astrocytes under ischemia studied. CHX decreased by 75% the number of cells dying after 6 hr of ischemia compared with control cultures. TdT-mediated dUTP nick end labelling (TUNEL) staining of comparable cultures was reduced by 40%. ActD decreased cell death by 60% compared with controls. The number of TUNEL-positive cells was reduced by 38%. The nuclear shrinkage in TUNEL-positive astrocytes in control cultures did not occur in ActD-treated astrocytes, indicating that nuclear shrinkage and DNA fragmentation during apoptosis are two unrelated processes. Expression of bcl-2 (alpha and beta), bax, and Ice in astrocytes under similar ischemic conditions, as measured by quantitative reverse transcription-polymerase chain reaction, indicated that ischemia down-regulated bcl-2 (alpha and beta) and bax. Ice was initially down-regulated from 0 to 4 hr, ...
AIMS--To compare in situ end-labelling (ISEL) of apoptosis in lung carcinoma with quantitative and semiquantitative light microscopic assessment and ultrastructural observations. METHODS--ISEL of apoptosis was evaluated in 42 lung carcinomas (24 squamous cell carcinomas, 12 adenocarcinomas and six small cell carcinomas). Results were correlated semiquantitatively with the extent of apoptosis in haematoxylin and eosin stained sections, with apoptotic indices and with ultrastructural observations (nine cases). RESULTS--In each tumour type the extent of apoptosis identified by ISEL correlated with that observed on light and electron microscopy. Tumour cells undergoing apoptosis showed either uniform nuclear staining with a surrounding halo or peripheral nuclear membrane staining. The latter pattern was more prominent in small cell carcinoma and correlated ultrastructurally with early apoptosis. A variable proportion of apoptotic cells and apoptotic bodies were unlabelled. Necrotic tumour cells ...
TY - JOUR. T1 - Increased cardiomyocyte apoptosis and changes in proapoptotic and antiapoptotic genes bax and bcl-2 during left ventricular adaptations to chronic pressure overload in the rat. AU - Condorelli, Gianluigi. AU - Morisco, Carmine. AU - Stassi, Giorgio. AU - Notte, Antonella. AU - Farina, Felicia. AU - Sgaramella, Giuseppe. AU - De Rienzo, Assunta. AU - Roncarati, Roberta. AU - Trimarco, Bruno. AU - Lembo, Giuseppe. PY - 1999/6/15. Y1 - 1999/6/15. N2 - Background - Left ventricular hypertrophy (LVH) represents both an adaptive response to increased cardiac work load and a precursor state of heart failure. Recent evidence linked cardiac myocyte death by apoptosis with LVH and heart failure. It remained unclear, however, whether apoptosis participated in the transition from LVH to left ventricular dysfunction (LVD). Methods and Results - Cardiac myocyte apoptotic events and changes in apoptosis-specific genes were studied in a rat model of chronic pressure overload induced by ...
Animals and plants diverged over one billion years ago and evolved unique mechanisms for many cellular processes, including cell death. One of the most well-studied cell death programmes in animals, apoptosis, involves gradual cell dismantling and engulfment of cellular fragments, apoptotic bodies, through phagocytosis. However, rigid cell walls prevent plant cell fragmentation and thus apoptosis is not applicable for executing cell death in plants. Furthermore, plants are devoid of the key components of apoptotic machinery, including phagocytosis as well as caspases and Bcl-2 family proteins. Nevertheless, the concept of plant
When hypertonicity is imposed with sufficient intensity and acuteness, cells die. Here we investigated the cellular pathways involved in death using a cell line derived from renal epithelium. We found that hypertonicity rapidly induced activation of an intrinsic cell death pathway- release of cytochrome c and activation of caspase-3 and caspase-9-and an extrinsic pathway-activation of caspase-8. Likewise, a lysosomal pathway of cell death characterized by partial lysosomal rupture and release of cathepsin B from lysosomes to the cytosol was also activated. Relationships among the pathways were examined using specific inhibitors. Caspase inhibitors did not affect cathepsin B release into the cytosol by hypertonicity. In addition, cathepsin B inhibitors and caspase inhibitors did not affect hyper-tonicity-induced cytochrome c release, suggesting that the three pathways were independently activated. Combined inhibition of caspases and cathepsin B conferred significantly more protection from ...
Differentiation refers to the ability of cancer cells to revert to their normal counterparts, and its induction represents an important noncytotoxic therapy for leukemia, and also breast, prostate, and other solid malignancies. Flavonoids are a group of differentiation-inducing chemicals with a potentially lower toxicology profile than retinoids. Flavonoid-rich polyphenol fractions from the pomegranate (Punica granatum) fruit exert anti-proliferative, anti-invasive, anti-eicosanoid, and pro-apoptotic actions in breast and prostate cancer cells and anti-angiogenic activities in vitro and in vivo.. The results highlight an important, previously unknown, mechanism of the cancer preventive and suppressive potential of pomegranate fermented juice and pericarp extracts.. Journal of Medicinal Food, Volume: 7 Issue 1: July 7, 2004 ...
TY - JOUR. T1 - Reduced wall compliance suppresses Akt-dependent apoptosis protection stimulated by pulse perfusion. AU - Li, Manxiang. AU - Chiou, Kuan Rau. AU - Bugayenko, Artem. AU - Irani, Kaikobad. AU - Kass, David A.. PY - 2005/9/16. Y1 - 2005/9/16. N2 - Reduced arterial compliance and increased pulse pressure are common and major risk factors for cardiovascular disease. Here, we reveal a novel mechanism whereby loss of wall distensibility blunts endothelial cell protection to oxidant stress-induced apoptosis. Bovine aortic endothelial cells cultured in compliant or stiff silastic tubes were pulse perfused by arterial pressure/flow waveforms generated by a servo-pump. Pulse perfusion induced time-dependent Akt activation peaking ,6-fold after 2 hours in compliant tubes and a similar time course but half the magnitude in stiff tubes. This was accompanied by quantitatively similar disparities in phosphoinositide-3 kinase activation and in Akt-stimulated suppressors of apoptosis: glycogen ...
Worldwide Market Reports added Latest Research Report titled Global Apoptosis Assays Market Is Expected To Reach Around USD 6.96 Billion By 2025″ to its Large Report database. The report, the global Apoptosis Assays Market, demonstrates an all-encompassing investigation of the global Apoptosis Assays Market that appraises the market magnitude and evaluates the market assessment above the predicted time. The ruling players of the Apoptosis Assays Market are visible inside the report in company with thorough details referring to their revenue break-up, synopsis of the business, latest developments, product segmentation, and so on. The operative market players in the global Apoptosis Assays Market are further discussed in the report.. This Research will help you grow your Business: [Download free Sample PDF of This Research Report]. Some of the Major Apoptosis Assays Market Players Are:. Merck KGaA, Thermo Fischer Scientific, Bio-Rad Laboratories, GE Healthcare, Geno Technology, BioTek ...
The basal machinery controlling PCD appears to have been substantially conserved throughout metazoan evolution. The mammalian bcl‐2 gene family encodes cell death regulator proteins that exhibit structural and functional homology with the product of the death‐suppressing ced‐9 gene of C.elegans. Likewise, the ced‐3 killer gene of C.elegans encodes a member of the conserved caspase family of cysteine proteases, members of which play critical proapoptotic roles in mammalian apoptosis.. However, despite this conservation of the basal machinery between the nematode and mammals, neither ced‐9/bcl‐2 nor caspase homologues has yet been identified in the fruit fly, D.melanogaster. Indeed, the proapoptotic machinery so far defined in Drosophila via genetic screening comprises three genes, rpr, hid and grim that share little if any homology with known components of the mammalian death machinery and none at all with the nematode. The only clear link thus far between the apoptotic machinery in ...
In this paper we used a multiparametric approach to analyze extensively the events occurring during apoptotic cell death of thymocytes, and furthermore, we asked whether alterations in mitochondrial structure and function are occurring in early stages of apoptosis. A multiparametric quantitative analysis was performed on normal or apoptotic thymocytes emerging from a few-hour culture performed in culture medium or in the presence of dexamethasone. Simultaneous detection of light scattering properties, integrity of plasma membrane (trypan blue exclusion), chromatin condensation (AO/EB staining of entire cells or PI staining of nuclei), and DNA fragmentation (in situ nick-translation in apoptotic cells) allowed a precise analysis of the preapoptotic and apoptotic stages. Moreover a thorough study of mitochondrial transmembrane potential (delta psi m) assessed following in a time course study the uptake by apoptotic cells of the cationic lipophilic dye DiOC6(3) or the J-aggregate-forming cation ...
Successful wound closure is mainly the result of two cellular processes: migration and proliferation. Apoptosis has also been suggested to play a role in the mechanisms of wound healing. The fast calcium wave (FCW), triggered immediately after a wound is produced, has been proposed to be involved in determining healing responses in epithelia. We have explored the effects of the reversible inhibition of FCW on the apoptotic and proliferative responses of healing bovine corneal endothelial (BCE) cells in culture. The most important findings of this study are that caspase-dependent apoptosis occurs during the healing process, that the amount of apoptosis has a linear dependence on the migrated distance, and that FCW inhibition greatly increases the apoptotic index. We have further been able to establish that FCW plays a role in the control of cell proliferation during BCE wound healing. These results indicate that one of the main roles of the wave is to inhibit an excessive apoptotic response of the
CCAR1; cell division cycle and apoptosis regulator 1; cell division cycle and apoptosis regulator protein 1; CARP 1; CARP1; FLJ10590; death inducer with SAP domain; cell cycle and apoptosis regulatory protein 1; MGC44628; RP11-437A18.1; novel protein similar to vertebrate cell division cycle and apoptosis regulator 1 (CCAR1 ...
Cell apoptosis assay PaTu8988 cell apoptosis was Inhibitors,Modulators,Libraries detected by the Annexin V Apoptosis Detection Kit in accordance on the producers protocol. Briefly, 1 million cells with indicated therapies were stained with FITC Annexin V and PI. Each early and late apoptotic cells had been sorted by fluorescence activated cell sorting. Cell morphologic examination A total of 4 104 PaTu8988 cells were seeded on glass cover slips during the six very well plate and taken care of using the indicated concentration of SAHA for 48 h. Cells were fixed and stained with Wright Giemsa stain. The slides had been photographed using oil microscopy. In vitro tube formation assay or vasculogenic mimicry assay The tumor cell formation of capillary structure in vitro was examined as we previously described.. Cellular immuno fluorescence staining PaTu8988 cells were seeded on glass cover slips in merely six nicely plates and taken care of with described dosage of SAHA for 48 h. Cells about the ...
The development and growth of aneurysms involve the complex events of arterial wall cells remodeling (38). Previous studies have demonstrated that apoptosis is involved in the pathogenesis of aneurysms (6,7). However, the molecular mechanisms underlying the stimulation of apoptosis in the mouse IA model remains unknown.. To investigate the apoptotic events of IA mice model, the present study investigated the mRNA expression of caspases associated with the mitochondrial apoptotic pathway, including caspase-3, −8 and −9. The activation of caspases serves an essential role during the process of apoptosis (39), which may be caused by an extrinsic or intrinsic pathway, which lead to a terminal common pathway (40). Caspase-8 activation is involved in the extrinsic pathway and caspase-9 is involved in the intrinsic pathway. In the mitochondrial pathway, the release of Cyt-c constitutes an apoptotic complex, which subsequently results in the activation of caspase-9. The activation of caspase-8 and ...
The nuoG gene of Mycobacterium tuberculosis (Mtb) has the ability to inhibit host cell apoptosis. This ability is a virulence factor and does not exist in facultative pathogenic and non-pathogenic mycobacterial species. NuoG is part of the NDH-1 complex, and this study addressed the potential link between the role of NuoG in apoptosis inhibition and the biochemical activity of the NDH-1 complex. Different mycobacterial species were tested for their NDH-1 activities. Among the bacteria tested were bacteria transformed with the Mtb nuoG plasmid, or with the almost entire NDH-1 coding region. Surprisingly, the levels of NDH-1 activity did not correlate with apoptosis levels, suggesting a potential independent, novel mechanism by which NuoG inhibits host cell apoptosis. ...
Abstract: PURPOSE: The clinical use of the zoledronic acid has been increasing recently, and especially for the treatment of bone metastases. The synergistic effects of zoledronic acid with other chemotherapeutic agents have been shown. However it is not known whether a similar synergistic apoptotic effects exist for a combination therapy of zoledronic acid with radiation. METHODS: The MCF-7 human breast cancer cell lines were treated with 10 micrometer, 50 micrometer or 100 micrometer of zoledronic acid, irradiated with 5 Gy, and they were also treated with a combination of both treatments. The results of the synergistic effect on apoptosis were identified by performing XTT assay, DNA fragmentation assay, FACS analysis and western blot analysis at 24, 48, 72 hours after treatment. RESULTS: Zoledronic acid afftects anti-proliferative and apoptotic effects on MCF-7 breast cancer cell line in a dose-dependent manner. The synergistic cytotoxic effect of zoledronic acid and radiation was noted. The ...
Apoptotic signaling is altered at many loci in cancer cells. Although many tumors develop resistance to cytochrome c-induced apoptosis, we have discovered that breast cancer cells exhibit a unique hypersensitivity to cytochrome c-induced apoptosis. Interestingly, this sensitivity is not due to changes in core apoptosomal proteins ( 44) but is due to a PHAPI-mediated posttranslational event that enhances the recruitment of caspase-9 to the Apaf-1 CARD. These findings have the potential to affect breast cancer chemotherapy through the development of apoptosome activators or cytochrome c mimetics as shown, in principle, by the fact that malignant mammary epithelial cells could be more easily killed by cytosolic cytochrome c than their normal counterparts.. PHAPI-mediated increase in caspase activation in breast cancer. Whereas many inhibitory signaling pathways converge on the apoptosome, there are very few physiologic/pathologic examples of enhanced apoptosome activation. Our data suggest that ...