• Bulletin of the Korean Chemical Society
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• v.27 no.1
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• pp.87-92
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• 2006

Fas-associated death domain protein (FADD) recruits and activates procaspase-8 through interactions between the death effector domains of these two proteins. Cellular FLICE-inhibitory protein (c-FLIP) was identified as a molecule with sequence homology to caspase-8. It has been postulated that c-FLIP prevents formation of the competent death-inducing signaling complex in a ligand-dependent manner, through its interaction with FADD and/or caspase-8. However, the interaction of FADD and $c-FLIP_s$ (short form) in apoptosis signaling has been controversially discussed. We show the purification and the characterization of human full-length FADD and $c-FLIP_s$ expressed in Escherichia coli. The purified FADD and $c-FLIP_s$ are shown as homogeneity, respectively, in SDS-PAGE analysis and light-scattering measurements. The folding properties of the $\alpha$-helical structure of FADD and the super-secondary structure of $c-FLIP_s$ proteins were characterized by circular dichroism spectroscopy. Furthermore, we report here a series of biochemical and biophysical data for FADD-$c-FLIP_s$ binding in vitro. The binding of both FADD and $c-FLIP_s$ proteins was detected by BIAcore biosensor, fluorescence measurement, and size-exclusion column (SEC).

• BMB Reports
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• v.57 no.2
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• pp.92-97
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• 2024

Elevated blood glucose is associated with an increased risk of atherosclerosis. Data from the current study showed that glucosamine (GlcN), a normal glucose metabolite of the hexosamine biosynthetic pathway (HBP), promoted lipid accumulation in RAW264.7 macrophage cells. Oleic acid- and lipopolysaccharide (LPS)-induced lipid accumulation was further enhanced by GlcN in RAW264.7 cells, although there was no a significant change in the rate of fatty acid uptake. GlcN increased acetyl CoA carboxylase (ACC), fatty acid synthase (FAS), scavenger receptor class A, liver X receptor, and sterol regulatory element-binding protein-1c (SREBP-1c) mRNA expression, and; conversely, suppressed ATP-binding cassette transporter A1 (ABCA-1) and ABCG-1 expression. Additionally, GlcN promoted O-GlcNAcylation of nuclear SREBP-1 but did not affect its DNA binding activity. GlcN stimulated phosphorylation of mammalian target of rapamycin (mTOR) and S6 kinase. Rapamycin, a mTOR-specific inhibitor, suppressed GlcN-induced lipid accumulation in RAW264.7 cells. The GlcN-mediated increase in ACC and FAS mRNA was suppressed, while the decrease in ABCA-1 and ABCG-1 by GlcN was not significantly altered by rapamycin. Together, our results highlight the importance of the mTOR signaling pathway in GlcN-induced macrophage lipid accumulation and further support a potential link between mTOR and HBP signaling in lipogenesis.

• Journal of Life Science
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• v.29 no.5
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• pp.514-523
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• 2019

We investigated the gene expression patterns and the mechanisms of action of the apoptotic response by microarray analysis of human keratinocyte HaCaT cells treated with ginsenoside Rb1, a saponin of Panax ginseng C. A. Meyer. Genes related to apoptosis, the G2/M transition of the mitotic cell cycle, cell division, mitotic nuclear division, and intracellular protein transport were 2-fold up-regulated in HaCaT cells treated with the ginsenoside Rb1, whereas genes related to DNA repair, regeneration fission, and extracellular matrix organization were 2-fold down-regulated. Apoptosis signaling may be mediated by FAS and PLA2G4A, and pathway analysis indicated that STAT3 might be an upstream regulator of these genes. The activity of FAS and PLA2G4A was verified by qPCR, which showed that FAS was increased about 2-fold in HaCaT cells treated with $10{\mu}g/ml$ of ginsenoside Rb1 for 24 hr, PLA2G4A was increased about twice after 6 hours, and gene expression was increased more than 2-fold after 24 hr. Knockdown of STAT3 with siRNA decreased FAS expression and increased PLA2G4A expression but only FAS was passed from the upstream regulator STAT3. These results indicate that STAT3, which is an upstream regulator, induces apoptosis via FAS during treatment with ginsenoside Rb1.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.12
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• pp.7069-7074
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• 2013

Programmed cell death is a basic cellular process that is critical to maintaining tissue homeostasis. In contrast to apoptosis, necrosis was previously regarded as an unregulated and uncontrollable process. However, as research has progressed, necrosis, also known as necroptosis or programmed necrosis, is drawing increasing attention, not least becasu of its possible impications for cancer research. Necroptosis exhibits a unique signaling pathway that requires the involvement of receptor interaction protein kinases 1 and 3 (RIP1 and RIP3), mixed lineage kinase domain-like (MLKL), and phosphoglycerate mutase 5 (PGAM5) and can be specifically inhibited by necrostatins. Not only does necroptosis serve as a backup cell death program when apoptosis is inhibited, but it is now recognized to play a pivotal role in regulating various physiological processes and the pathogenesis of a variety of human diseases such as ischemic brain injury, immune system disorders and cancer. The control of necroptosis by various defined trigger factors and signaling pathways now offers the opportunity to target this cellular process for therapeutic purposes. The purpose of this paper is to review current findings concerning the connections between various trigger factors and the RIP1/RIP3 signaling pathway as it relates to necroptosis.

• BMB Reports
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• v.46 no.4
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• pp.207-212
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• 2013

The main purpose of this study is to examine the effect of caffeine on lipid accumulation in human hepatoma HepG2 cells. Significant decreases in the accumulation of hepatic lipids, such as triglyceride (TG), and cholesterol were observed when HepG2 cells were treated with caffeine as indicated. Caffeine decreased the mRNA level of lipogenesis-associated genes (SREBP1c, SREBP2, FAS, SCD1, HMGR and LDLR). In contrast, mRNA level of CD36, which is responsible for lipid uptake and catabolism, was increased. Next, the effect of caffeine on AMP-activated protein kinase (AMPK) signaling pathway was examined. Phosphorylation of AMPK and acetyl-CoA carboxylase were evidently increased when the cells were treated with caffeine as indicated for 24 h. These effects were all reversed in the presence of compound C, an AMPK inhibitor. In summary, these data indicate that caffeine effectively depleted TG and cholesterol levels by inhibition of lipogenesis and stimulation of lipolysis through modulating AMPK-SREBP signaling pathways.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.9
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• pp.1300-1308
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• 2012

The 5'-adenosine monophosphate-activated protein kinase (AMPK) is a key part of a kinase-signaling cascade that acts to maintain energy homeostasis. The objective of this experiment was to investigate the possible effects of fasting and refeeding on the gene expression of hypothalamic AMPK, some appetitive regulating peptides and lipid metabolism related enzymes. Seven-day-old male broiler (Arbor Acres) chicks were allocated into three equal treatments: fed ad libitum (control); fasted for 24 h; fasted for 24 h and then refed for 24 h. Compared with the control, the hypothalamic gene expression of $AMPK{\alpha}2$, $AMPK{\beta}1$, $AMPK{\beta}2$, $AMPK{\gamma}1$, Ste20-related adaptor protein ${\beta}$ ($STRAD{\beta}$), mouse protein $25{\alpha}$ ($MO25{\alpha}$) and agouti-related peptide (AgRP) were increased after fasting for 24 h. No significant difference among treatments was observed in mRNA levels of $AMPK{\alpha}1$, $AMPK{\gamma}2$, LKB1 and neuropeptide Y (NPY). However, the expression of $MO25{\beta}$, pro-opiomelanocortin (POMC), corticotropin-releasing hormone (CRH), ghrelin, fatty acid synthase (FAS), acetyl-CoA carboxylase ${\alpha}$ ($ACC{\alpha}$), carnitine palmitoyltransferase 1 (CPT-1) and sterol regulatory element binding protein-1 (SREBP-1) were significantly decreased. The present results indicated that 24 h fasting altered gene expression of AMPK subunits, appetite regulation peptides and lipometabolism related factors in chick's hypothalamus; the hypothalamic FAS signaling pathway might be involved in the AMPK regulated energy homeostasis and/or appetite regulation in poultry.

• Journal of Biomedical Engineering Research
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• v.41 no.6
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• pp.235-246
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• 2020

This study aimed to evaluate the inhibitory effect of micro-current stimulation(MCS) on adipogenesis regarding with Wnt/β-catenin pathway using the ob/ob mouse and 3T3-L1 cell line. 6-week old ob/ob male mice were equally assigned to four groups: obese group(ob), obese with MCS groups(50 μA, 200 μA, and 400 μA). 6-week old C57BL/6J male mice were assigned to the control group(CON). We analyzed abdominal adipose tissue volume by using in vivo micro-CT and measured the body weight, feed intake, liver weight and triglycerides in serum. All the MCS groups showed that significantly reduced body weight and triglycerides in serum. In the case of liver weight and abdominal adipose tissue volume, the inhibitory effect of adipogenesis was shown in the 200 μA and 400 μA groups. To elucidate the anti-obesity effect of MCS, β-catenin, C/EBPα and FAS protein expressions were analyzed by western blotting. β-catenin expression was upregulated, C/EBPα and FAS expression were down-regulated in the relatively high-intensity groups(200 μA and 400 μA). Thus, the 200 μA and 400 μA for the intensity of MCS were chosen for cell experiments. In the 3T3-L1 cell line, Wnt/β-catenin pathway including Wnt10b, Wnt3a, β-catenin and Cyclin D1 was activated in all MCS groups. Accordingly, the expression level of C/EBPα was decreased during the differentiation and lipid droplet was significantly reduced in Oil red O staining results. These results suggest that the Wnt/β-catenin signaling might be activated by MCS with current intensities between 200-400 μA and it may lead to anti-obesity effects.

• Journal of Microbiology and Biotechnology
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• v.25 no.3
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• pp.343-352
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• 2015

H9 is an ethanol extract prepared from nine traditional/medicinal herbs. This study was focused on the anticancer effect of H9 in non-small-cell lung cancer cells. The effects of H9 on cell viability, apoptosis, mitochondrial membrane potential (MMP; ${\Delta}\psi_{m}$), and apoptosisrelated protein expression were investigated in A549 human lung cancer cells. In this study, H9-induced apoptosis was confirmed by propidium iodide staining, expression levels of mRNA were determined by reverse transcriptase polymerase chain reaction, protein expression levels were checked by western blot analysis, and MMP (${\Delta}\psi_{m}$) was measured by JC-1 staining. Our results indicated that H9 decreased the viability of A549 cells and induced cell morphological changes in a dose-dependent manner. H9 also altered expression levels of molecules involved in the intrinsic signaling pathway. H9 inhibited Bcl-xL expression, whereas Bax expression was enhanced and cytochrome C was released. Furthermore, H9 treatment led to the activation of caspase-3/caspase-9 and proteolytic cleavage of poly(ADP-ribose) polymerase; the MMP was collapsed by H9. However, the expression levels of extrinsic pathway molecules such as Fas/FasL, TRAIL/TRAIL-R, DR5, and Fas-associated death receptor were downregulated by H9. These results indicated that H9 inhibited proliferation and induced apoptosis by activating intrinsic pathways but not extrinsic pathways in human lung cancer cells. Our results suggest that H9 can be used as an alternative remedy for human non-small-cell lung cancer.

• The Journal of Korean Medicine
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• v.22 no.4
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• pp.58-68
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• 2001

Objectives : This study was designed to investigate the protective mechanisms of Samul-tang (SMT) on $H_2O_2$-induced toxicity in H9c2 cardiomyoblast cells. Methods : The cultured cells were pretreated with SMT and exposed to $H_2O_2$. The cell damage was assessed by using MTT assay. Also, we used Hoechst staining, Western blotting analysis. Results : SMT significantly reduced both $H_2O_2$-induced cell death and chromatin fragmentation. The decrease of Bcl2 expression by $H_2O_2$ was inhibited by SMT. In addition, the increase of Bax expression was also inhibited by SMT. In particular, Fas expression, which is generally recognized as cell death inducing signal by Fas/FasL interaction, was markedly decreased by $H_2O_2$ in a time-dependent manner, whereas this decrease was completely prevented by SMT. The cotreatment of SMT and $H_2O_2$ in H9c2 cells also induced the phosphorylation of ERK in a time-dependent manner. Moreover, PD098059, a specific inhibitor of ERKl/2, attenuated the protective effect of SMT on $H_2O_2$-induced toxicity in H9c2 cardiomyoblast cells. Furthermore, the protective effect of SMT was significantly blocked by treatment of SB203580, a specific inhibitor of p38. Conclusions : Taken together, this study suggests that the protective effects of the water extract of SMT against oxidative damages may be mediated by the modulation of Bel2 and Bax expression via the regulation of ERK and p38 signaling pathway.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.2
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• pp.205-213
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• 2017

Quercetin, a plant-derived flavonoid found in fruits, vegetables and tea, has been known to possess bioactive properties such as anti-oxidant, anti-inflammatory and anti-cancer. In this study, anti-cancer effect of quercetin and its underlying mechanisms in triple-negative breast cancer cells was investigated. MTT assay showed that quercetin reduced breast cancer cell viability in a time and dose dependent manner. For this, quercetin not only increased cell apoptosis but also inhibited cell cycle progression. Moreover, quercetin increased FasL mRNA expression and p51, p21 and GADD45 signaling activities. We also observed that quercetin induced protein level, transcriptional activity and nuclear translocation of Foxo3a. Knockdown of Foxo3a caused significant reduction in the effect of quercetin on cell apoptosis and cell cycle arrest. In addition, treatment of JNK inhibitor (SP 600125) abolished quercetin-stimulated Foxo3a activity, suggesting JNK as a possible upstream signaling in regulation of Foxo3a activity. Knockdown of Foxo3a and inhibition of JNK activity reduced the signaling activities of p53, p21 and GADD45, triggered by quercetin. Taken together, our study suggests that quercetin induces apoptosis and cell cycle arrest via modification of Foxo3a signaling in triple-negative breast cancer cells.