• The Korean Journal of Physiology and Pharmacology
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• v.9 no.6
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• pp.315-322
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• 2005

In this study, the authors investigated whether death of vascular smooth muscle cell (VSMC) had a pathological pertinence. Conditioned media obtained from rat aorta smooth muscle cell (SMC) that were induced death by expressing FADD in the absence of tetracycline (FADD-SMC) triggered death of normal SMC. DNA fragmentation and caspase-3 activation were observed in dying SMC by conditioned media. FADD-SMC showed transcriptional activation of tumor necrosis factor $(TNF)-{\alpha}$. Conditioned medium contained $TNF-{\alpha}$, indicating secretion of the cytokine from dying FADD-SMC. It was investigated if secreted $TNF-{\alpha}$ was functional. Conditioned medium activated ERK and p38 MAPK pathways and induced MMP-9 expression, whereas depletion of the cytokine with its soluble receptor (sTNFR) remarkably inhibited induction of MMP-9 by conditioned medium. These findings suggest that $TNF-{\alpha}$ in conditioned medium seems to be active. Then, contribution of $TNF-{\alpha}$ on death-inducing activity of conditioned medium was examined. Depletion of $TNF-{\alpha}$ with soluble $TNF-{\alpha}$ receptor decreased the death activity of conditioned medium by 35%, suggesting that $TNF-{\alpha}$ play a partial role in the death activity. Boiling of medium almost completely abolished the death-inducing activity, suggesting that other heat labile death inducing proteins existed in conditioned medium. Taken together, these results indicate that SMC undergoing death could contribute to inflammation by expressing inflammatory cytokines and pathological complications by inducing death of neighboring cells.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.16
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• pp.6939-6944
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• 2014

Sulfasalazine (SSZ) is an anti-inflammatory drug that has been used to treat inflammatory bowel disease and rheumatoid arthritis for decades. Recently, some reports have suggested that SSZ also has anti-cancer properties against human tumors. However, little is known about the effects of SSZ on oral cancer. The aim of this study was to investigate the anti-cancer effects of SSZ in oral squamous cell carcinoma (OSCC) cells and to elucidate the mechanisms involved. The authors investigated the anti-proliferative effect of SSZ using the MTT method in HSC-4 cells (an OSCC cell line). Cell cycle analysis, acidic vesicular organelle (AVO) staining, monodansylcadaverine (MDC) staining and Western blotting were also conducted to investigate the cytotoxic mechanism of SSZ. SSZ significantly inhibited the proliferation of HSC-4 cells in a dose-dependent manner. In addition, SSZ induced autophagic cell death, increased microtubule-associated protein 1 light chain (MAP1-LC; also known as LC) 3-II levels, as well as induced punctate AVO and MDC staining, resulted in autophagic cell death. Furthermore, these observations were accompanied by the inhibition of the Akt pathway and the activation of ERK pathway. These results suggest that SSZ promotes autophagic cell death via Akt and ERK pathways and has chemotherapeutic potential for the treatment of oral cancer.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.2
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• pp.115-121
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• 2009

Functional defects in mitochondria are involved in the induction of cell death in cancer cells. We assessed the toxic effect of camptothecin against the human cervical and uterine tumor cell line SiHa with respect to the mitochondria-mediated cell death process, and examined the combined effect of camptothecin and anticancer drugs. Camptothecin caused apoptosis in SiHa cells by inducing mitochondrial membrane permeability changes that lead to the loss of mitochondrial membrane potential, decreased Bcl-2 levels, cytochrome c release, caspase-3 activation, formation of reactive oxygen species and depletion of GSH. Combination of camptothecin with other anticancer drugs (carboplatin, paclitaxel, doxorubicin and mitomycin c) or signaling inhibitors (farnesyltransferase inhibitor and ERK inhibitor) did not enhance the camptothecin-induced cell death and caspase-3 activation. These results suggest that camptothecin may cause cell death in SiHa cells by inducing changes in mitochondrial membrane permeability, which leads to cytochrome c release and activation of caspase-3. This effect is also associated with increased formation of reactive oxygen species and depletion of GSH. Combination with other anticancer drugs (or signaling inhibitors) does not appear to increase the anti-tumor effect of camptothecin against SiHa cells, but rather may reduce it. Combination of camptothecin with other anticancer drugs does not seem to provide a benefit in the treatment of cervical and uterine cancer compared with camptothecin monotherapy.

• Molecules and Cells
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• v.45 no.4
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• pp.257-272
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• 2022

In addition to inducing apoptosis, caspase inhibition contributes to necroptosis and/or autophagy depending on the cell type and cellular context. In macrophages, necroptosis can be induced by co-treatment with Toll-like receptor (TLR) ligands (lipopolysaccharide [LPS] for TLR4 and polyinosinic-polycytidylic acid [poly I:C] for TLR3) and a cell-permeable pan-caspase inhibitor zVAD. Here, we elucidated the signaling pathways and molecular mechanisms of cell death. We showed that LPS/zVAD- and poly I:C/zVAD-induced cell death in bone marrow-derived macrophages (BMDMs) was inhibited by receptor-interacting protein kinase 1 (RIP1) inhibitor necrostatin-1 and autophagy inhibitor 3-methyladenine. Electron microscopic images displayed autophagosome/autolysosomes, and immunoblotting data revealed increased LC3II expression. Although zVAD did not affect LPS- or poly I:C-induced activation of IKK, JNK, and p38, it enhanced IRF3 and STAT1 activation as well as type I interferon (IFN) expression. In addition, zVAD inhibited ERK and Akt phosphorylation induced by LPS and poly I:C. Of note, zVAD-induced enhancement of the IRF3/IFN/STAT1 axis was abolished by necrostatin-1, while zVAD-induced inhibition of ERK and Akt was not. Our data further support the involvement of autocrine IFNs action in reactive oxygen species (ROS)-dependent necroptosis, LPS/zVAD-elicited ROS production was inhibited by necrostatin-1, neutralizing antibody of IFN receptor (IFNR) and JAK inhibitor AZD1480. Accordingly, both cell death and ROS production induced by TLR ligands plus zVAD were abrogated in STAT1 knockout macrophages. We conclude that enhanced TRIF-RIP1-dependent autocrine action of IFNβ, rather than inhibition of ERK or Akt, is involved in TLRs/zVAD-induced autophagic and necroptotic cell death via the JAK/STAT1/ROS pathway.

• Molecules and Cells
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• v.46 no.6
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• pp.387-398
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• 2023

Microtubule acetylation has been proposed as a marker of highly heterogeneous and aggressive triple-negative breast cancer (TNBC). The novel microtubule acetylation inhibitors GM-90257 and GM-90631 (GM compounds) cause TNBC cancer cell death but the underlying mechanisms are currently unknown. In this study, we demonstrated that GM compounds function as anti-TNBC agents through activation of the JNK/AP-1 pathway. RNA-seq and biochemical analyses of GM compound-treated cells revealed that c-Jun N-terminal kinase (JNK) and members of its downstream signaling pathway are potential targets for GM compounds. Mechanistically, JNK activation by GM compounds induced an increase in c-Jun phosphorylation and c-Fos protein levels, thereby activating the activator protein-1 (AP-1) transcription factor. Notably, direct suppression of JNK with a pharmacological inhibitor alleviated Bcl2 reduction and cell death caused by GM compounds. TNBC cell death and mitotic arrest were induced by GM compounds through AP-1 activation in vitro. These results were reproduced in vivo, validating the significance of microtubule acetylation/JNK/AP-1 axis activation in the anti-cancer activity of GM compounds. Moreover, GM compounds significantly attenuated tumor growth, metastasis, and cancer-related death in mice, demonstrating strong potential as therapeutic agents for TNBC.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.118-118
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• 2002

Programmed cell death (PCD) is thought as a well-controlled process by which unwanted cells are selectively eliminated. During the last decade many researches have elucidated molecules and their interactions involved in cell death by using largely in vitro induction of cell death or survival signals in a more defined manner, While these critical information and novel findings provide us with clearer understanding of mechanisms underlying cell death, it does by no means explain how PCD occurs and which cells or tissues are affected during normal embryonic development in vivo. In this study, we used zebrafish to examine whether the PCD is occurring selectively or randomly in developing embryos by whole mount in situ TUNEL analysis with specific markers for neural cells. The result revealed that the degree and distribution of TUNEL staining varied considerably throughout gastrulation stage, and there was also a number of TUNEL-negative embryos. Most of TUNEL-positive cells were scattered randomly throughout the blastoderm. During the gastrulation stage about 75 % of the embryos analyzed exhibited more than 5 TUNEL-positive cells. As the dorsal epiblast begins to thicken rather abruptly near the end of gastrulation, TUNEL-positive cells were mainly located along the dorsal side. Although there were some variations in TUNEL staining during segmentation and pharyngeal stages, TUNEL staining continued to be localized to the central nervous system, and was also detected in the sensory organs, trigeminal ganglions, and the primary sensory neurons. High levels of the cell death in developing brain between 20-somite and prim-6 stages are thought to play a role in the morphogenesis and organization of the brain. At prim-16 stage, cell death is considerably reduced in the brain region. Dying cells are mainly localized to the prospective brain region where ectodermal cells are about to initiate neurogenesis. As development progressed, high levels and more reproducible patterns of cell death were observed in the developing nervous system. Intensive TUNEL staining was restricted to the trigeminal ganglions, the primary sensory neurons, and sensory organs, such as olfactory pits and otic vesicles. Thus, PCD patterning in zebrafish embryos occurs randomly at early stages and becomes restricted to certain region of the embryos. The spatio-temporal pattern of PCD during the early embryonic development in zebrafish will provide basic information for further studies to elucidate genes involved in. regulation of PCD largely unknown in vivo during vertebrate embryogenesis.

• Korean Journal of Clinical Laboratory Science
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• v.52 no.3
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• pp.245-252
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• 2020

Macrophage cell death contributes to the formation of plaque, leading to the development of atherosclerosis. The accumulation of triglyceride (TG) is also associated with the pathogenesis of atherosclerosis. A previous study reported that TG induces the cell death of macrophages. This study examined whether the cytoplasmic release of cathepsin B from lysosome is associated with the TG-induced cell death of macrophage. The release of cathepsin B was increased in the TG-treated THP-1 macrophages, but the TG treatment did not affect cathepsin B expression. Furthermore, the inhibition of cathepsin B by its inhibitor, CA-074 Me, partially inhibited the TG-induced cell death of macrophage. TG-triggered macrophage cell death is mediated by the activation of caspase-1, -2, and apoptotic caspases. Therefore, this study investigated whether cathepsin B is implicated in the activation of these caspases. The inhibition of cathepsin B blocked the activation of caspase-7, -8, and -1 but did not affect the activity of caspase-3, -9, and -2. Overall, these results suggest that TG-induced cytoplasmic cathepsin B causes THP-1 macrophage cell death by activating caspase-1, leading to subsequent activation of the extrinsic apoptotic pathway.

• The Journal of Korean Medicine
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• v.22 no.2
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• pp.64-74
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• 2001

Objectives : The effects of aqueous extract of Backhapgogumtanggami-bang (BGTG, a newly devised herb medicine) on the induction of apoptotic cell death were investigated in human lymphoid origin leukemia cell lines, HL-60. Methods : Cells were treated with various concentrations and $400{\;}\mu\textrm{g}/ml$ BGTG for 12 hr. Genomic DNA was isolated and separated on 1.8% agarose gels. Lysates from the cells were used to measure the activity of caspase-2, -3, -8, and -9 protease by using fluorogenic peptide. Cells were preincubated with SB-203580 for 30 min. Nuclear protein from the cells was incubated with oliginucleotide probe of AP-l and NF-kB. Nuclear extracts from the cells were isolated and reacted with antibodies. Results : The viability of HL-60 cells were markedly decreased by BGTG extract in a dose- and time-dependent manner. BGTG extract induced the apoptotic death of HL-60 cells which was characterized by the DNA fragmentation. The activations of Caspase-2, 3, and 9 were induced by BGTG. However, selective inhibition of the p38 mitogen-activated protein kinase pathways by SB-203580 did not affect the extent of BGTG extract-induced cell death. Furthermore, we observed the transient activations of transcriptional factors such as AP-l and NF-kB. Conclusions : These results suggest that BGTG extract induced apoptotic death of HL-60 cells and caspase activations as well as the modulation of transcriptional factors such as AP-1 and NF-kB.

• The Korean Journal of Physiology and Pharmacology
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• v.12 no.6
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• pp.287-291
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• 2008

Ampicillin, a $\beta$-lactam antibiotic, has been reported to induce astrocytic glutamate transporter-l which plays a crucial role in protecting neurons against glutamate excitotoxicity. We investigated the effect of ampicillin on neuronal damage in the mouse hippocampus and neostriatum following transient global forebrain ischemia. Male C57BL/6 mice were anesthetized with halothane and subjected to bilateral occlusion of the common carotid artery for 40 min. Ampicillin was administered post-ischemically (for 3 days) and/or pre-ischemically (for $3{\sim}5$ days until one day before the onset of ischemia). Pre- and post-ischemic treatment with ampicillin (50 mg/kg/day or 200 mg/kg/day) prevented ischemic neuronal death in the medial CAI area of the hippocampus as well as the neostriatum in a dose-dependent manner. In addition, ischemic neuronal damage was reduced by pre-ischemic treatment with ampicillin (200 mg/kg/day). In summary, our results suggest that ampicillin plays a functional role as a chemical preconditioning agent that protects hippocampal neurons from ischemic insult.

• Journal of Acupuncture Research
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• v.19 no.2
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• pp.28-38
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• 2002

Objective : This study is aimed to investigate the effects of bee venom and purified bee venom on cell death in synovial cell line. Methods : It was evaluated by using MTT assay, morphological method, flow cytometry, immunocytochemistry analysis, RT-PCR. Results : The result obtained is as follows. 1. The MTT assay demonstrated that synovial cell viability was significantly inhibitted dose-dependently by treatment with BV and PBV in comparison with control. And the inhibitory effect of BV and PBV was almost same. 2. The morphologic study demonstrated that synovial cell showed apoptotic body resulted from apoptosis after treatment with BV and PBV for 6 hours using microscope. 3. The Flow cytometry demonstrated that apoptosis of synovial cell treated with BV and PBV was related with stop of cell cycle in stage of G0/G1. 4. Immunocytochemistry assay demonstrated that COX-II and iNOS were slightly expressed by treatment with BV and PBV in comparison with control group. 5. RT-PCR analysis demonstrated that COX-II were almost down-regulated by high dose treatment with BV and PBV in comparison with control group. iNOS were well down-regulated by treatment with $5{\mu}g/ml$ BV and PBV whereas it was well expressed in control group. Conclusion : These results suggest that bee venom and purified bee venom have significant effect on cell death in synovial cell line and further study is needed in vivo.