• Toxicological Research
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• 제35권1호
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• pp.83-91
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• 2019

Nanoparticles (NPs) have been recognized as both useful tools and potentially toxic materials in various industrial and medicinal fields. Previously, we found that zinc oxide (ZnO) NPs that are neurotoxic to human dopaminergic neuroblastoma SH-SY5Y cells are mediated by lipoxygenase (LOX), not cyclooxygenase-2 (COX-2). Here, we examined whether human bone marrow-derived mesenchymal stem cells (MSCs), which are different from neuroblastoma cells, might exhibit COX-2- and/or LOX-dependent cytotoxicity of ZnO NPs. Additionally, changes in annexin V expression, caspase-3/7 activity, and mitochondrial membrane potential (MMP) induced by ZnO NPs and ZnO were compared at 12 hr and 24 hr after exposure using flow cytometry. Cytotoxicity was measured based on lactate dehydrogenase activity and confirmed by trypan blue staining. Rescue studies were executed using zinc or iron chelators. ZnO NPs and ZnO showed similar dose-dependent and significant cytotoxic effects at concentrations ${\geq}15{\mu}g/mL$, in accordance with annexin V expression, caspase-3/7 activity, and MMP results. Human MSCs exhibited both COX-2 and LOX-mediated cytotoxicity after exposure to ZnO NPs, which was different from human neuroblastoma cells. Zinc and iron chelators significantly attenuated ZnO NPs-induced toxicity. Conclusively, these results suggest that ZnO NPs exhibit both COX-2- and LOX-mediated apoptosis by the participation of mitochondrial dysfunction in human MSC cultures.

• Journal of Microbiology and Biotechnology
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• 제26권10호
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• pp.1701-1707
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• 2016

Lipoxygenase (LOX) is an industrial enzyme with wide applications in food and pharmaceutical industries. The available structure information indicates that eukaryotic LOXs consist of N terminus β-barrel and C terminus catalytic domains. However, the latest crystal structure of Pseudomonas aeruginosa LOX shows it is significantly different from those of eukaryotic LOXs, including the N-terminal helix domain. In this paper, the functions of this N-terminal helix domain in the soluble expression and catalysis of P. aeruginosa LOX were analyzed. Genetic truncation of this helix domain resulted in an insoluble P. aeruginosa LOX mutant. The active C-terminal domain was obtained by dispase digestion of the P. aeruginosa LOX derivative containing the genetically introduced dispase recognition sites. This functional C-terminal domain showed raised substrate affinity but reduced catalytic activity and thermostability. Crystal structure analyses demonstrate that the broken polar contacts connecting the two domains and the exposed hydrophobic substrate binding pocket may contribute to the insoluble expression of the C terminus domain and the changes in the enzyme properties. Our data suggest that the N terminus domain of P. aeruginosa LOX is required for its soluble expression in E. coli, which is different from that of the eukaryotic LOXs. Besides this, this N-terminal domain is not necessary for catalysis but shows positive effects on the enzyme properties. The results presented here provide new and valuable information on the functions of the N terminus helix domain of P. aeruginosa LOX and further improvement of its enzyme properties by molecular modification.

• 생명과학회지
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• 제5권2호
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• pp.14-14
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• 1995

The effect of lipoxygenase (LOX) on the oxidation and co-oxidation of lipid fraction was studied in the model system of rainbow trout. For the reaction in model system 1 g of lipid fraction and 50mL of enzyme extract(LOX, 140 unit in 50mL phosphate buffer solution at pH 7,4)), which were obtained from rainbow trout, were homoginized in the presence of Tween 20 and kept at 23$\circ$C for 3 days. The activity of LOX was decreased to 43% of initial level during the reaction in the model system. The initial composition of rainbow trout lipid was showed to be consisted of trigliceride(TG;82%) and free fatty acid(FFA;0.1%), while this converted to 59% of TG and 20% of FIFA, respectively after reaction in model system. Change of fatty acid composition was also observed and the content of linoleic acid, one of the major fatte acids, was decreased to 13% from 54% in the content of total fatty acids after reaction. The carotenoids in rainbow trout were composed of 0.4% $\alpha$-carotene, 1.6% $\beta$ -carotene, 80% canthaxanthin, 7% lutein and 11% zeaxanthin, thus the canthaxanthin was the major component. This canthaxanthin was the most degraded carotenoid by lipoxygenase catalyzed co-oxidation during the reaction. On the other hand the tocopherol isomers found in the rainbow trout were $\alpha$ and $\beta$ -tocopherol, and $\alpha$-tocopherol had a higher degradation rate by the lipoxygenase catalyzed co-oxidation than of $\beta$-tocopherol in the reaction of model system.

• Archives of Pharmacal Research
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• 제21권6호
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• pp.664-670
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• 1998

The role of intracellular $Ca^{2+}$, in the regulation of tumor cell proliferation by products of arachidonic acid (AA) metabolism was investigated using U-373 MG human as trocytoma cells. Treatment with nordihydroguaiaretic acid (NDGA), a lipoxygenase (LOX) inhibitor, or caffeic acid (CA), a specific 5-LOX inhibitor, suppressed proliferation of the tumor cells in a dose-dependent manner. However, indomethacin (indo), a cyclooxygenase (COX) inhibitor, did not significantly alter proliferation of the tumor cells. At anti-proliferative concentrations, NDGA and CA significantly inhibited intracellular $Ca^{2+}$ release induced by carbachol, a known intracelluar $Ca^{2+}$ agonist in the tumor cells. Exogenous administration of leukotriene $B_4(LTB_4)$, an AA metabolite of LOX pathway, enhanced proliferation of the tumor cells in a concentration-dependent fashion. In addition, $LTB_4$, induced intracelluar $Ca^{2+}$ release. Intracellular $Ca^{2+}$-inhibitors, such as an intracellular $Ca^{2+}$ chelator (BAPTA) and intracellular $Ca^{2+}$-release inhibitors (dantrolene and TMB-8), significantly blocked the LTB4-induced enhancement of cell proliferation and intracellular $Ca^{2+}$ release. These results suggest that LOX activity may be critical for cell proliferation of the human astrocytoma cells and that intracelluar $Ca^{2+}$ may play a major role in the mechanism of action of LOX.

• 한국식품영양과학회지
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• 제45권1호
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• pp.35-43
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• 2016

LOX 결핍콩(진양, 05C4 및 LS)의 가공적성을 평가하기 위하여 재래 된장을 제조하여 일반 콩(태광) 된장과 품질 특성 및 항산화 활성을 비교하였다. 된장에서 수분과 조단백질 함량은 LS 된장이 유의적으로 높았으며, 조지방 함량은 7.92~8.22%로 콩 품종에 따른 유의차가 없었다. 탄수화물 함량은 LS 된장이 가장 낮았다. 된장의 환원당은 태광 된장에 비해 진양 된장이 유의적으로 높았다. 아미노태 질소 함량은 태광 된장에 비해 3종의 LOX 결핍콩 된장에서 유의적으로 높았다. 된장의 색깔은 태광 된장에 비해 05C4 및 LS 된장은 차이가 없었으며, 진양 된장은 두드러진 색차를 보였다. 된장의 구수한 맛은 태광 된장이 다소 높았으며, 짠맛은 LOX 결핍콩 된장이 다소 높게 평가되었다. 향미는 LS 된장이 가장 강한 것으로 인지되었다. 전반적인 기호도는 시료간에 차이가 미미하였다. 총 아미노산 함량은 태광 된장에 비해 진양 및 05C4 된장에서 다소 높았으며, 필수 아미노산은 태광 된장에 비해 3종의 LOX 결핍콩 된장이 높게 정량되었다. 이소플라본의 총량은 태광 된장($374.79{\mu}g/g$)에 비해 3종의 LOX 결핍콩 된장($522.16{\sim}684.46{\mu}g/g$)에서 유의적으로 높았다. 총 페놀 함량은 태광 된장에 비해 05C4 및 LS 된장이 유의적으로 높았으며, 플라보노이드 함량은 LS 된장만 태광 된장에 비해 유의적으로 높았다. DPPH 라디칼소거 활성과 FRAP법에 의한 환원력은 진양 된장이 우수하였으나 ABTS 라디칼 소거 활성은 LOX 결핍콩 된장이 태광 된장에 비해 유의적으로 높았다. LOX 결핍콩으로 제조한 된장은 아미노산과 이소플라본의 함량 및 항산화 활성 측면에서 된장 가공적성에 적합한 것으로 판단된다.

• Biomolecules & Therapeutics
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• 제17권1호
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• pp.86-91
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• 2009

In our ongoing search for biological components from the Korea endemic plants, the MeOH extract of Ailanthus altissima leaves (Simaroubaceae) showed cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) dual inhibitory activity by assessing their effects on the production of prostaglandin $D_2$ ($PGD_2$) and leukotriene $C_4$ ($LTC_4$) in mouse bone marrow-derived mast cells (BMMCs). In further study, eight compounds, squalene (1), ${\beta}$-sitosterol (2), scopoletin (3), quercetin (4), luteolin (5), astragalin (6), scopolin (7), and daucosterol (8) were isolated, the chemical structures were elucidated on the basis of physicochemical and spectroscopic data and by comparison with those of published literatures. Among the compounds, 2, 4, and 5 strongly inhibited both the COX-2-dependent PGD2 generation with $IC_{50}$ values of 2.6, 7.3 and 2.5 ${\mu}M$, respectively and the generation of $LTC_4$ in the 5-LOX dependent phase with $IC_{50}$ values of 2.0, 5.1 and 1.8 ${\mu}M$, respectively, which suggest that the anti-inflammatory activity of A. altissima might occur in part via the inhibition of both $PGD_2$ and $LTC_4$ generation by 2, 4 and 5.

• Nutritional Sciences
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• 제9권3호
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• pp.152-158
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• 2006

Helicobacter pylori (H. pylori) infection rapidly stimulated either COX-2 or 5-LOX and released arachidonic acid metabolites that have been considered as pivotal mediators in H. pylori-induced inflammatory responses. To determine whether red ginseng extract (RGE) can suppress the biosynthesis of 5(S)-hydroxyeicosatetraenoic acids (HETE), a precursor metabolite of leukotrienes B4 (LTB4) in H. pylori-provoked inflammatory responses in gastric epithelial cells, the biosynthesis of monohydroxy fatty acids was measured using radioactive arachidonic acid and validated by RP-HPLC using non-radioactive AA as substrate in AGS cells cocultured with H. pylori (ATCC 43504) with or without pretreatment of RGE. Among three known major HETEs, H. pylori infection specifically induced the biosynthesis of $^{14}C-5(S)-HETE$ rather than the complex of $^{14}C-15S-/^{14}C-12(S)-HETE$ from $^{14}C-AA$, concomitantly obtained by HPLC(p<0.01). RGE, 1 to $100{\mu}g/ml$, selectively suppressed H. pylori-stimulated $^{14}C-5(S)-HETE$ production implying the attenuation of 5-lipoxygenase activity, of which was similar to known LOX inhibitor NDGA $(10{\mu}M)$ (p<0.01). However, the amount of 5(S)-HETE was significantly reduced by higher dose of RGE $(100{\mu}g/ml)$ (p<0.05). These results indicated that LOX pathway might be one of principle pathogenic mechanisms of H. pylori and red ginseng could be a nutraceutical against H. pylori infection through inhibiting action of LOX activity.

• Biomolecules & Therapeutics
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• 제20권1호
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• pp.113-117
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• 2012

The fruits of Foeniculum vulgare (Foeniculi Fructus) have been widely used in Chinese medicine as an antiemetic, ameliorating stomach ailments and as an analgesic. In order to establish its potential for antiallergic use, inhibitory actions of the fruit on 5-lipoxgenase (5-LOX) and ${\beta}$-hexosaminidase release were evaluated. The 70% ethanol extract of this plant material (FR) considerably inhibited 5-LOX-catalyzed leukotriene production from A23187-induced rat basophilic leukemia (RBL)-1 cells. The $IC_{50}$ was $3.2{\mu}g/ml$. From this extract, 12 major compounds including sabinene, fenchone, ${\gamma}$-terpinene, ${\alpha}$-pinene, limonene, p-anisylacetone, panisylaldehyde, estragole (4-allylanisole), trans-anethole, scopoletin, bergapten and umbelliferone were isolated. And it was found that several terpene derivatives including ${\gamma}$-terpinene and fenchone as well as phenylpropanoid, trans-anethole, showed considerable inhibitory action of 5-LOX. In particular, the $IC_{50}$ of trans-anethole was $51.6{\mu}M$. In contrast, FR and the isolated compounds did not show considerable inhibitory activity on the degranulation reaction of ${\beta}$-hexosaminidase release from antigen-treated RBL-2H3 cells. Against arachidonic acid-induced ear edema in mice, FR and trans-anethole showed significant inhibition by oral administration at doses of 100-400 mg/kg. In conclusion, FR and several major constituents are 5-LOX inhibitors and they may have potential for treating 5-LOX-related disorders.

• Biomolecules & Therapeutics
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• 제18권2호
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• pp.197-204
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• 2010

To establish the anti-inflammatory activity of the total flavonoid fraction of the root barks of Broussonetia papyrifera (EBP) and a new formula, the ethanol extract of the root barks of B. papyrifera was fractionated with ethylacetate, yielding the hydrophobic prenylated flavonoid-enriched fraction. EBP and the ethanol extract of the whole Lonicera japonica (ELJ) plant were then mixed at a ratio of 1:1 (w/w) to give a new preparation (BL) in the hope of obtaining an optimal formula with a higher anti-inflammatory activity. Evaluation of the effects of these preparations on A23187-treated rat basophilic leukemia (RBL-1) cells revealed that EBP potently inhibited 5-lipoxygenase (5-LOX), while ELJ showed weak inhibition. Additionally, the mixture (BL) clearly showed stronger inhibitory effects against 5-LOX than either preparation alone. These preparations also inhibited cyclooxygenase-2-catalyzed $PGE_2$ and inducible nitric oxide (NO) synthase-catalyzed NO production by lipopolysaccharide-treated RAW 264.7 cells. When tested against arachidonic acid-induced mouse ear edema, EBP showed strong inhibitory activity at doses of 5-200 mg/kg when administered orally, but BL had obviously stronger inhibitory effects. When tested against ${\lambda}$-carrageenan-induced paw edema in mice, BL showed a potent and synergistic anti-inflammatory effect. In addition, in the acetic acid-induced writhing test, BL was found to have strong analgesic activity at 50-400 mg/kg. Taken together, these results indicate that each of these preparations exert anti-inflammatory activity in vitro and in vivo. In particular, BL showed stronger anti-inflammatory activity than EBP, and these anti-inflammatory effects were partially related to the inhibition of eicosanoid and NO production. BL may be useful for the treatment of human inflammatory disorders.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2010년도 정기총회 및 추계학술발표회
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• pp.18-18
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• 2010

Molecular insights on the role of plant growth promoting rhizobacteria (PGPR) in potato tuberization are reported in the present study. The PGPRwere isolated from the soil collected from potato fields of Highland Agricultural Research Centre, Pyeongchang, Korea and they were identified to the genus level based on the 16S rRNA sequence analysis. These PGPR were heat-killed, filtered and the filtrates were addedindividually at a concentration of $10^7\;cfu\;mL^{-1}$ in MS (Murashige and Skoog's) medium supplemented with 7% (w/v) sucrose to study their influence on in vitro potato tuberization. Tuber initiation occurred early in untreated control, while tuber growth was pronounced in case of PGPR treatments. The control explants showed tuber formation as a result of sub-apical swelling of stolons while several sessile tubers formed directly in the axils of nodal cuttings in case of PGPR treatments, which is an indication of strong induction for tuberization. Theexplants cultured on MS medium supplemented with bacterial isolate 6 (Bacillus firmus strain 40) showed highest average tuber yield (Ca. 12.56 g per treatment) after 30 days of culture, which was 3 folds increase over the untreated control. A significant increase in lipoxygenase (LOX1) mRNA expression and activity of LOX enzyme were also detected in the tubers induced on PGPR treatments as compared to untreated control. This LOX expression level correlated with increased tuber growth and tuber yield. Further studies focused on the role of bacteria cell wall components, growth regulators and signal molecules released by PGPR are under investigation to elicit clues for PGPR-mediated signal pathway controlling potato tuberization.