Notes: The effects of hydrogen sulfide on inflammasome activation were investigated. Lactate dehydrogenase levels and caspase activation were monitored using the CytoTox-ONE™ and Caspase-Glo® assays. Hydrogen sulfide was shown to decrease caspase activation and inflammasome activity both in vitro and in vivo. (5147)

Notes: These authors investigated silicon dioxide nanoparticles (SiNPs) as a potential cancer treatment and identified the mechanism mediating cancer cell death. The Caspase-Glo® 3/7, Caspase-Glo® 9, CellTiter-Glo® luminescent cell-viability, ROS-Glo™, and Caspase-Glo® 1 assays were used to monitor cellular response to SiNPs. Together, SiNPs were identified to activate the intrinsic apoptosis pathway and lead to cell death.

  (5165)

Notes: These authors investigated the mechanism of action of the synthesized resveratrol analogue, (E)-N-(2-(4-methoxystyryl) phenyl) furan-2-carboxamide (CS) on colorectal cancer cells. They report that CS exerts a potent suppressive effect on HCT116 colorectal cancer cells, but not on normal colon cells. CS caused apoptosis via activation of an extrinsic pathway leading to caspase activation and cell cycle arrest from activated p53. The Caspase-Glo® 3/7, 8 and 9 Assays were used to detect caspase activation. (5045)

Notes: The role of the carcinogen Ochratoxin A in cellular inflammation was examined. HEK293 cells were treated with Ochratoxin A and cellular inflammatory markers were monitored. ATP levels and caspase activation were analyzed using the CellTiter Glo® and Caspase-Glo® assays. (5155)

Notes: A novel target of non-steroidal anti-inflammatory drugs (NSAIDs) was identified using the Caspase-Glo® -1, Caspase-Glo® 3/7, and Caspase-Glo® 9 assays. The authors observed that an inhibition of caspase activity lead to decreased cell death and inflammation. The CellTiter-Fluor™ and CellTiter-Glo®  assays were using to monitor cell viability. (5163)

Notes: Chagas disease is caused by the parasite Trypanosoma cruzi and can lead to Chronic Chagas disease cardiomyopathy (CCC). The ability of N,N-dimethylsphingosine (DMS) to decrease inflammation and function as a therapeutic for CCC was determined. As part of this study, Caspase-Glo® assays were used to detect inflammatory (caspase 1 and 8), or apoptotic (caspase 9 and 3/7) activities. (5166)

Notes: The authors utilized phosphoproteomics to identify host factors that are activated upon Influenza A infection. In total, 1116 proteins showed changes in regulation including cyclin-dependent kinases. The authors tested the effects of cyclin-dependent kinase inhibitors on cell viability and caspase activation using the CellTiter Glo® and Caspase-Glo® assays. (5154)

Notes: The authors of this review article highlight the use of bioluminescence as a readout for high-throughput ADME/Tox assays. They discuss three strategies for designing bioluminescent assays, using either luciferase, ATP or luciferin substrates as the limiting reagents for a luciferase-catalyzed reaction. Reporter gene assays limit the production of luciferase by tying it to a promoter or DNA regulatory region of interest. Such assays can be used to study genes that are regulated by drugs and other xenobiotics. Bioluminescent assays in which ATP is the limiting reagent of the luciferase reaction can be designed to monitor cell viability or the activity kinases. Bioluminescent assays in which the substrate is limiting can be designed so that the activity of a particular enzyme results in the production of a luciferin substrate that can, in turn, be acted upon by luciferase. (3926)

Notes: The authors of this paper review bioluminescent assay technologies, discussing HTS reporter, cell-based and luciferase biosensor assays. They divide luminescent assays into three basic categories: assays that measure ATP concentration (cell viability and kinase assays), assays that measure changes in luciferase levels (reporter assays, GPCR assays), and assays that measure changes in luciferin levels (protease [including caspase], P450 and MAO assays). (3737)

Notes: The authors sought to determine the mechanism by which first-trimester trophoblasts resist FAS ligand-induced apoptosis but remain sensitive to TNFα-mediated apoptosis. First trimester trophoblasts express XAF1 [X-linked inhibitor of apoptosis (XIAP)-associated factor 1], which may be involved in regulating their response to proapoptotic signals. The authors created HaloTag™-XAF1 fusion constructs and transiently transfected the first trimester trophoblast cell line (3A). Cells were labeled with the HaloTag™ TMR ligand, and XAF1 was shown to localize to the cytoplasm. 3A cells transiently transfected with the fusion construct were also separated into cytoplasmic and mitochondrial fractions. The fractions were labeled with HaloTag™ TMR ligand. Expression of the fusion peaked at 48 hours after transfection in both mitochondrial and cytoplasmic fractions. TNFα-treatment of 3A cells induced translocation of endogenous XAF1 to the mitochondria. The authors used the Caspase-Glo® Assays to demonstrate activation of caspase-3 and caspase-9 in response to expression of XAF-1. They also show that caspase-3 activation and XIAP cleavage correlate with translocation of endogenous XAF1 to mitochondria. Viability of 3A and primary trophoblasts over expressing XAF1 was evaluated using the CellTiter® 96 AQueous One-Solution Assay. (3760)

Notes: In this study, the Caspase-Glo^® 8 and Caspase-Glo^® 9 Assays were used to assess caspase activation in Zebrafish embryos after radiation exposure. After irradiation, caspase activation, morphological abnormalities, and DNA fragmentation were observed, all three of which could be partially reversed by treatment with the radiomodifier amifostine. The Caspase-Glo^® 9 Assay was shown to be sensitive enough to detect the effects of radiation on as few as 2 embryos. The authors concluded that the Caspase-Glo^® Assays provided an effective and convenient means for rapidly assessing the lethal effects of radiation on Zebrafish embryos, and for assaying the ability of the radiomodifier to counteract these effects. The assay could be performed within hours of radiation exposure directly on the embryos in a 96-well plate format. (3571)

Notes: Bacterial cytolethal distending toxin (CDT) is known to induce apoptosis of immune cells, but little is known of the toxin from Campylobacter jejuni. In this study, the authors examined the effects of C. jejuni CDT on cultured monocytes. The human monocyte line 28SC was inoculated with 2µg Campylobacter membrane proteins per milliliter of culture. After 8 hours, the cells were harvested and assessed for caspase-8 or caspase-9 activity using the Caspase-Glo^® 8 and Caspase-Glo^® 9 Assays, respectively. The 28SC cells were pulsed with 6.2 µM camptothecin for 2 hours as a positive control for apoptosis. (3288)

Notes: In this paper, researchers examined the effect of activating mammalian homologues of the Drosophila Toll receptor gene (TDR) on apoptosis in the trophoblast cell lines HTR8 and 3A. Bacterial peptidoglycan (PDG) was used to indirectly activate caspase-3/7, -8 and -9 through TDRs. Caspase activity was measured using the Capase-Glo™ 3/7, 8 and 9 Assays. Caspase-3/7, -8 and -9 activities decreased in both the HTR8 and 3A cell lines transiently transfected with a Fas Associated Death Domain (FADD-DN). Data were expressed as relative light units and were normalized to protein levels in lysates. (3171)

Notes: The Caspase-Glo^® 3/7, -8 and -9 Assays were used to  measure Caspase-3, -8, and -9 activities in mouse liver homogenates. The authors describe a modified procedure for making liver homogenates in a hypotonic extraction buffer.  Homogenates were cleared with a 15 minute spin at 13,000 x g and normalized to a 1mg/ml concentration before use the assays. Data comparing control and cSN50 peptide-treated animals at various time points are presented.  (3213)