• Proceedings of the Korean Society of Applied Pharmacology
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• 2002.07a
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• pp.249-249
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• 2002

• Proceedings of the PSK Conference
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• 2001.10a
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• pp.182.2-182.2
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• 2001

• Proceedings of the PSK Conference
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• 2001.10a
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• pp.184.1-184.1
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• 2001

• Biomolecules & Therapeutics
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• v.18 no.3
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• pp.262-270
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• 2010

CD95 receptor is a member of tumor necrosis factor receptor family that mediates apoptosis in many cell types when bound by CD95 ligand or cross-linked by agonistic anti-CD95 antibodies. To determine the role of neutral sphingomyelinase (nSMase) on CD95-mediatd apoptosis, human Jurkat T lymphocytes were exposed to recombinant human CD95 ligand. Treatment with CD95 ligand induced cell death in a concentration and time-dependent manner. CD95-induced cell death was suppressed by inhibitors of SMase such as AY9944 or desipramine. Transfection with human nSMase1 siRNA plasmid into CD95 ligand-treated cells significantly prevented CD95-mediated cell death. CD95-mediated elevation of intracellular ceramide level detected by FACS analysis with anti-ceramide antibody was also decreased by nSMase1 siRNA. Knock-down of nSMase1 expression also blocked cytochrome c release into cytosol and caspase-3 cleavage in CD95-treated cells. Taken together, these results suggest that nSMase1 may play an important role in CD95-mediated apoptotic cell death in Jurkat T cells.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.103.2-103.2
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• 2003

The hydrolysis of sphingomyelin (SM), known as the SM pathway, is induced by the activation of sphingomyelinase (SMase) to generate the second messenger ceramide, which plays a key role in cellular responses such as apoptosis, differentiation, senescence, and inflammation. Here, we identified a 60 kDa membrane-associated, neutral and Mg$\^$2+/-dependent SMase, termed N-SMase $\varepsilon$, from mammalian brains, which was revealed as the heat shock protein 60 (HSP60) through cDNA cloning and mass spectrometrical analysis. (omitted)

• Nutrition Research and Practice
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• v.18 no.5
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• pp.602-616
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• 2024

BACKGROUND/OBJECTIVES: Inflammation and ferroptosis are implicated in various diseases and lipopolysaccharides (LPS) have been linked with these disorders. Recently, many edible insects, such as Gryllus bimaculatus, Protaetia brevitarsis larvae (PB) and Tenebrio molitor larvae, have been recommended as alternative foods because they contain lots of nutritional sources. In this study, we explored the potential of PB extract in preventing LPS-induced inflammation and ferroptosis in Hep3B cells. MATERIALS/METHODS: PB powder was extracted using 70% ethanol and applied to Hep3B cells. Co-treatment with LPS was conducted to induce ferroptosis and inflammation. The anti-inflammatory and anti-ferroptosis mechanisms of the PB extract were confirmed using Western blot, enzyme-linked immunosorbent assay, and real-time polymerase chain reaction analysis. RESULTS: PB extract effectively prevented LPS-induced cell death and restored LPS-induced inflammatory cytokine production, NF-κB signaling, endoplasmic reticulum (ER) stress and ferroptosis. Interestingly, PB extract reduced LPS-induced ceramide increase and acid sphingomyelinase (ASMase) expression. The use of the ASMase inhibitor, desipramine, also demonstrated a reduction in these pathways, highlighting the pivotal role of ASMase in inflammation and ferroptosis. Treatment with each inhibitor revealed that ferroptosis causes ER stress and that NF-κB and MAP kinase pathways are involved in inflammation. CONCLUSION: PB emerges as a potential functional food with inhibitory effects on LPS-induced inflammation and ferroptosis, making it a promising candidate for nutritional interventions.

• Molecules and Cells
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• v.46 no.9
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• pp.545-557
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• 2023

Sphingomyelinase (SMase) catalyzes ceramide production from sphingomyelin. Ceramides are critical in cellular responses such as apoptosis. They enhance mitochondrial outer membrane permeabilization (MOMP) through self-assembly in the mitochondrial outer membrane to form channels that release cytochrome c from intermembrane space (IMS) into the cytosol, triggering caspase-9 activation. However, the SMase involved in MOMP is yet to be identified. Here, we identified a mitochondrial Mg²⁺-independent SMase (mt-iSMase) from rat brain, which was purified 6,130-fold using a Percoll gradient, pulled down with biotinylated sphingomyelin, and subjected to Mono Q anion exchange. A single peak of mt-iSMase activity was eluted at a molecular mass of approximately 65 kDa using Superose 6 gel filtration. The purified enzyme showed optimal activity at pH of 6.5 and was inhibited by dithiothreitol and Mg²⁺, Mn²⁺, Ni²⁺, Cu²⁺, Zn²⁺, Fe²⁺, and Fe³⁺ ions. It was also inhibited by GW4869, which is a non-competitive inhibitor of Mg²⁺-dependent neutral SMase 2 (encoded by SMPD3), that protects against cytochrome c release-mediated cell death. Subfractionation experiments showed that mt-iSMase localizes in the IMS of the mitochondria, implying that mt-iSMase may play a critical role in generating ceramides for MOMP, cytochrome c release, and apoptosis. These data suggest that the purified enzyme in this study is a novel SMase.

• Proceedings of the Korean Society of Toxicology Conference
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• 2003.05a
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• pp.13-14
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• 2003

Ceramide, a potential cell death marker formed by sphingomyelinase, is involved in the expression of cyclooxygenase-2 (COX-2). This study examines the effect of C2-ceramide (C2), a cell-permeable ceramide analog, on the LPS-inducible COX-2 expression and signaling pathways. C2 did not induce COX-2, but potentiated LPS-inducible COX-2 expression in Raw264.7 cells, whereas dihydro-C2 was inactive.(omitted)

• The Korean Journal of Physiology and Pharmacology
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• v.6 no.6
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• pp.281-286
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• 2002

To understand the cytotoxic mechanism of $MPP^+,$ we examined the involvement of ceramide in $MPP^+-induced$ cytotoxicity to human neuroblastoma SH-SY5Y cells. When SH-SY5Y cells were exposed to $MPP^+,\;MPP^+$ induced dose-dependent cytotoxicity accompanied by 2-fold elevation of intracellular ceramide levels in SH-SY5Y cells. Three methods were used to test the hypothesis that the elevated intracellular ceramide is related to $MPP^+-induced$ cytotoxicity: $C_2-ceramide$ was directly applied to cells, sphingomyelinase (SMase) was exogenously added, and oleoylethanolamine (OE) was used to inhibit degradation of ceramide. Furthermore, inhibition of ceramide-activated protein phosphatase (CAPP), the effector of ceramide, using okadaic acid (OA) attenuated cell death but treatment of fumonisin $B_1,$ the ceramide synthase inhibitor, did not alter the cytotoxic effect of $MPP^+.$ Based on these, we suggest that the elevation of intracellular ceramide is one of the important mediators in $MPP^+-induced$ cell death.

• Archives of Pharmacal Research
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• v.28 no.3
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• pp.311-318
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• 2005

Oxidative stress has been reported to elevate ceramide level during cell death. The purpose of the present study was to modulate cell death in relation to cellular glutathione (GSH) level and GST (glutathione S-transferase) expression by regulating the sphingolipid metabolism. LLC­PK1 cells were treated with H$_2$O$_2$ in the absence of serum to induce cell death. Subsequent to exposure to H$_2$O$_2$, LLC-PK1 cells were treated with desipramine, sphingomyelinase inhibitor, and N-acetylcysteine (NAC), GSH substrate. Based on comparative visual observation with H202-treated control cells, it was observed that 0.5 $\mu$M of desipramine and 25 $\mu$M of NAC exhibited about 90 and $95\%$ of cytoprotection, respectively, against H$_2$O$_2$-induced cell death. Desipramine and NAC lowered the release of LDH activity by 36 and $3\%$ respectively, when compared to $71\%$ in H$_2$O$_2$-exposed cells. Cellular glutathione level in 500 $\mu$M H202-treated cells was reduced to 890 pmol as compared to control level of 1198 pmol per mg protein. GST P1-1 expression was decreased in H$_2$O$_2$-treated cells compared to healthy normal cells. In conclusion, it has been inferred that H$_2$O$_2$-induced cell death is closely related to cellular GSH level and GST P1-1 expression in LLC-PK1 cells and occurs via ceramide elevation by sphingomyelinase activation.