thapsigargin and cypermethrin

The endoplasmic reticulum (ER) serves as a critical site of protein synthesis and processing. The temperature-sensitive hamster fibroblast cell line (tsBN7) displays deficient N-linked glycosylation activity at the restrictive temperature and activates cellular apoptosis. Temperature-shifted tsBN7 cells display induction of Grp78 and Gadd153, genes known to be induced by ER stress, and activate apoptosis via the release of cytochrome c from the mitochondria. Cyclosporin A (CsA), a potent blocker of the mitochondrial permeability transition pore (PTP), was sufficient to block cytochrome c release and to rescue tsBN7 cells from apoptosis. CsA-treated cells displayed Grp78 induction at the restrictive temperature, consistent with an ER stress signal being carried to the nucleus, while the apoptosis-associated transcription factor, Gadd153, displayed only a mild induction. Cypermethrin, a type II pyrethroid known to perturb Ca(2+) signaling in neuronal cells, was sufficient to arrest apoptosis under these conditions. This work identifies type II pyrethroids as a valuable new tool in the characterization of cellular stress signaling pathways.

Topics: Animals; Apoptosis; Blotting, Western; Caspase 12; Caspase Inhibitors; Caspases; CCAAT-Enhancer-Binding Proteins; Cell Line; Cell Survival; Cricetinae; Cyclosporine; Cytochromes c; Endoplasmic Reticulum; Endoplasmic Reticulum Chaperone BiP; Fibroblasts; Glycosylation; Heat-Shock Proteins; In Situ Nick-End Labeling; Insecticides; Ion Channels; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Mitogen-Activated Protein Kinase Kinases; Molecular Chaperones; Nitriles; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Pyrethrins; Signal Transduction; Tacrolimus; Temperature; Thapsigargin; Transcription Factor CHOP; Transcription Factors; Tunicamycin; Ultraviolet Rays

While effector molecules produced by activated macrophages (including nitric oxide, tumor necrosis factor alpha, interleukin 1, etc.) help to eliminate pathogens, high levels of these molecules can be deleterious to the host itself. Despite their importance, the mechanisms modulating macrophage effector functions are poorly understood. This work introduces two key negative regulators that control the levels and duration of macrophage cytokine production. Vacuolar-type H+-ATPase (V-ATPase) and calcineurin (Cn) constitutively act in normal macrophages to suppress expression of inflammatory cytokines in the absence of specific activation and to inhibit macrophage cytokine responses induced by bacterial lipopolysaccharide (V-ATPase), interferon gamma (V-ATPase and Cn), and calcium (Ca2+) flux (Cn). Cn and V-ATPase modulate effector gene expression at the mRNA level by inhibiting transcription factor NF-kappaB. This negative regulation by Cn is opposite to its crucial positive role in T cells, where it activates NFAT transcription factor(s) leading to expression of interleukin 2, tumor necrosis factor alpha, and other cytokine genes. The negative effects of V-ATPase and Cn on NF-kappaB-dependent gene expression are not limited to the macrophage lineage, as similar effects have been seen with a murine fibroblast cell line and with primary astrocytes.

Topics: Animals; Astrocytes; Calcineurin; Cell Line; Cells, Cultured; Cytokines; DNA-Binding Proteins; Gene Expression Regulation; Insecticides; Interferon-gamma; Interleukin-12; L Cells; Lipopolysaccharides; Macrophages, Peritoneal; Mice; Mice, Inbred Strains; NF-kappa B; NFATC Transcription Factors; Nuclear Proteins; Proton-Translocating ATPases; Pyrethrins; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes; Tacrolimus; Thapsigargin; Transcription Factors; Tumor Necrosis Factor-alpha; Vacuolar Proton-Translocating ATPases