• 대한의생명과학회지
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• 제27권4호
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• pp.223-230
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• 2021

Tumor necrosis factor alpha (TNF-α) is a principal regulator of inflammation and immunity. The proinflammatory properties of TNF-α can be attributed to its ability to activate the enzyme cytosolic phospholipase A₂ (cPLA₂), which generates potent inflammatory lipid mediators, eicosanoids. L-glutamine (Gln) plays physiologically important roles in various metabolic processes. We have reported that Gln has a potent anti-inflammatory activity via rapid upregulation of mitogen-activated protein kinases (MAPKs) phosphatase (MKP)-1, which preferentially dephosphorylates the key proinflammatory enzymes, p38 MAPK and cytosolic phospholipase A₂ (cPLA₂). In this study, we have investigated whether Gln could inhibit TNF-α-induced cPLA₂ activation. Gln inhibited TNF-α-induced increases in cPLA₂ phosphorylation in the lungs and blood levels of the cPLA₂ metabolites, leukotrine B4 (LTB4) (lipoxygenase metabolite) and prostaglandin E2 (PGE2) (cyclooxygenase metabolite). TNF-α increased p38 and cPLA₂ phosphorylation and blood levels of LTB4 and PGE2, which were blocked by the p38 inhibitor SB202190. Gln inhibited TNF-α-induced p38 and cPLA₂ phosphorylation and production of the cPLA₂ metabolites. Such inhibitory activity of Gln was no longer observed in MKP-1 small interfering RNA-pretreated animals. Our data indicate that Gln inhibited TNF-α-induced cPLA₂ phosphorylation through MKP-1 induction/p38 inhibition, and suggest that the utility of Gln in inflammatory diseases in which TNF-α plays a major role in their pathogenesis.

• 대한의생명과학회지
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• 제15권4호
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• pp.273-279
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• 2009

Cordycepin (3'-deoxyadenosine) is an adenosine analogue isolated from Cordyceps militaris, and it has been used as an anti-cancer and anti-inflammation ingredient in traditional Chinese medicine. We investigated the effects of cordycepin on human platelet aggregation induced by thapsigargin, and determined the cytosolic free $Ca^{2+}$ levels ($[Ca^{2+}]_i$), an aggregation-stimulating factor. Cordycepin significantly inhibited thapsigargin-induced platelet aggregation. Its inhibitory effect was continually sustained at the maximal aggregation concentration of thapsigargin. The thapsigargin-induced $[Ca^{2+}]_i$ were clearly reduced by cordycepin in the presence of exogenous $CaCl_2$ or extracellular $Ca^{2+}$-chelator (EDTA). These results suggest that cordycepin inhibited thapsigargin-induced $Ca^{2+}$-influx from extracellular domain and thapsigargin-induced $Ca^{2+}$-mobilization from intracellular $Ca^{2+}$ storage. Accordingly, our data demonstrated that cordycepin may have a beneficial effect on platelet aggregation-mediated thrombotic diseases by inhibiting a $[Ca^{2+}]_i$-elevation.

• 한국미생물·생명공학회지
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• 제33권4호
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• pp.260-266
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• 2005

본 실험에서는 MRF가 다량 존재하는 RBL-2H3 세포주에 다양한 PKC isotype별 억제제를 처리하여 in vitro상에서 Na, K-ATPase $\alpha$1L3를 이용한 pull-down assay와 RBL-2H3 세포를 이용한 membrane fractionation을 실시하였다. 그 결과 HRF는 in vitro에서 $\alpha$1L3와 결합한다는 사실을 재확인 할 수 있었고 실제 세포주 내에서 Na,K-ATPase와 결합한다는 것을 알 수 있었다. 사용한 약물로부터 PKC $\alpha,\;\beta,\;\delta$뿐 아니라 protein phosphatase 2B(PP2B)도 HRF와 $\alpha$1L3의 결합에 관여한다는 사실을 알 수 있었다. 또한 이들 PKC, PP2B에 의해 인산화된 HRF 분자는 cytosolic fraction으로 이행할 수 있으며 이러한 결과는 탈인산화된 HRF가 Na,K-ATPase와 결합하여 Na, K-ATPase의 기능을 조절한다고 추정할 수 있다. 그러나 약물자체가 histamine 분비에 영향을 미칠 수 있으며 cytosolic HRF보다 exocytosis된 HRF가 histamine를 더 분비하도록 할 수 있으므로, 약물을 전처리한 세포에 외부에서 HRF를 첨가하여 histamine이 유리되는 정도가 어떻게 변화하는지를 HRF를 가하지 않은 결과와 비교해야 할 것이다.

• Food Science and Biotechnology
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• 제15권6호
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• pp.975-979
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• 2006

The extracts of Oenanthe javanica were evaluated for their effects on the expression of cyclooxygenase-2 (COX-2), which is mediated by the translocation of the p65 subunit into the nucleus. Fractions of ethyl acetate and chloroform from 80% ethanol extracts of O. javanica exhibited inhibitory effects on the secretion of tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) from lipopolysaccharide (LPS)-stimulated peritoneal macrophages; however, the aqueous- and hexane-fractions showed no significant effect. The ethyl acetate- and chloroform-fractions also reduced the COX-2 enzyme levels after 24-hr treatment. RT-PCR showed that the mRNA levels of COX-2 decreased following treatment with these fractions, suggesting that COX-2 expression is transcriptionally regulated by these extracts. We examined the effects of the chloroform- and ethyl acetate-fractions on the cytosolic activation of nuclear factor-${\kappa}B$ ($NF-{\kappa}B$, p65 subunit) and on the degradation of inhibitor-${\kappa}B{\alpha}$ ($I-{\kappa}B{\alpha}$) in order to determine the mechanism of COX-2 regulation. The LPS-stimulated activation of the p65 subunit was significantly blocked upon the addition of $50\;{\mu}g/mL$ of these fractions, and the cytosolic $I-{\kappa}B{\alpha}$ degradation process was simultaneously inhibited. These findings suggest that the inhibition of COX-2 expression by the ethyl acetate-and chloroform-fractions may result from the inhibition of p65 translocation by blocking the degradation of $I-{\kappa}B{\alpha}$; this may be the mechanistic basis for the anti-inflammatory effects of O. javanica.

• 한국어병학회지
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• 제22권2호
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• pp.137-146
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• 2009

The black rockfish Sebastes schlegelii neutrophil cytosolic factor components p47 phox (phagocyte oxidase) cDNA was cloned. The sequence of the cDNA showed that rockfish p47 phox cDNA consisted of 1,952 bp contained open reading frame encoding predicted polypeptide of 420 amino acids. Additionally analysis of the p47 phox amino acid sequence showed two potential SH3 domains. The functional domains are highly conserved in many animals, though the sequence of the components of the black rockfish showed low homology with that of mammals. The deduced amino acid sequence of the black rockfish p47 phox was similar to those of the carp (60.4%), zebrafish (59,2%), rainbow trout (68.5%), xenopus (55.2%), mouse (54.2%), rabbit (54.5%), rat (53.7%), and chicken (50.9%). The expression of the rockfish p47 phox molecule was induced in peripheral blood leukocytes (PBLs) from 1 to 12 h following LPS stimulation, with a peak at 6 h after the stimulation, and which increased at 1, 3, and 12 h after treated with Poly I:C compared with the control. The rockfish p47 phox gene was expressed in various tissues of healthy fish. The level of p47 phox expression was high in the PBLs, kidney and spleen.

• Journal of Microbiology and Biotechnology
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• 제18권3호
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• pp.490-496
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• 2008

Symbiobacterium toebii is a commensal symbiotic thermophile that cannot grow without support from a partner bacterium. We investigated the properties of Symbiobacterium growth-supporting factors (SGSFs) produced by the partner bacterium Geobacillus toebii. SGSFs occurred in both the cell-free extract (CFE) and culture supernatant of G. toebii and might comprise multifarious materials because of their different biological properties. The heavy SGSF contained in the cytosolic component exhibited heat- and proteinase-sensitive proteinaceous properties and had a molecular mass of >50 kDa. In contrast, the light SGSF contained in the extracellular component exhibited heat-stable, proteinase-resistant, nonprotein properties and had a molecular mass of <10 kDa. Under morphological examination using light microscopy, S. toebii cultured with the culture supernatant of G. toebii was filamentous, whereas S. toebii cultured with the CFE of G. toebii was rod-shaped. These results strongly suggest that the SGSFs produced by G. toebii comprise two or more types that differ in their growth-supporting mechanisms, although all support the growth of S. toebii. Upon the examination of the distribution of SGSFs in other bacteria, both cytosolic and extracellular components of Geobacillus kaustophilus, Escherichia coli, and Bacillus subtilis had detectable growth-supporting effects for S. toebii, indicating that common SGSF materials are widely present in various bacterial strains.

• Biomolecules & Therapeutics
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• 제23권5호
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• pp.421-427
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• 2015

Imperatorin has been known to exert many biological functions including anti-inflammatory activity. In this study, we investigated the inhibitory effects of imperatorin on the production of inflammatory mediators in mouse bone marrow-derived mast cells (BMMC). Imperatorin inhibited degranulation and the generation of eicosanoids (leukotriene $C_4$ ($LTC_4$) and prostaglandin $D_2$ ($PGD_2$) in IgE/antigen (Ag)-stimulated BMMC. To elucidate the molecular mechanism involved in this process, we investigated the effect of imperatorin on intracellular signaling in BMMC. Biochemical analyses of the IgE/Ag-mediated signaling pathway demonstrated that imperatorin dramatically attenuated degranulation and the production of 5-lipoxygenase-dependent $LTC_4$ and cyclooxygenase-2-dependent $PGD_2$ through the inhibition of intracellular calcium influx/phospholipase $C{\gamma}1$, cytosolic phospholipase $A_2$/mitogen-activated protein kinases and/or nuclear factor-${\kappa}B$ pathways in BMMC. These results suggest that the effects of imperatorin on inhibition of degranulation and eicosanoid generation through the suppression of multiple steps of IgE/Ag-mediated signaling pathways would be beneficial for the prevention of allergic inflammation.

• BMB Reports
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• 제28권6호
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• pp.499-503
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• 1995

In our previous studies (Chae et al., 1990; Chae et a1., 1993), we found that a phytochrome signal was clearly connected with the change in cytosolic free $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) in oat cells. It was determined that the $[Ca^{2+}]_i$ change occured both by mobilization out of the intracellular $Ca^{2+}$ store and by influx from the medium. The specific aim of this work is to elucidate the processes connecting $Ca^{2+}$ mobilization and influx. The cells treated with thapsigargin (increasing $[Ca^{2+}]_i$ by inhibition of the $Ca^{2+}$-ATPase in the calcium pool) in the presence of external $Ca^{2+}$ showed the same increasing pattern (sustained increasing shape) of $[Ca^{2+}]_i$ as that measured in animal cells. Red light irradiation after thapsigargin treatment did not increase $[Ca^{2+}]_i$ These results suggest that thapsigargin also acts specifically in the processes of mobilization and influx of $Ca^{2+}$ in oat cells. When the cells were treated with TEA ($K^+$ channel blocker), changes in $[Ca^{2+}]_i$ were drastically reduced in comparison with that measured in the absence of TEA. The results suggest that the change in $[Ca^{2+}]_i$ due to red light irradiation is somehow related with $K^+$ channel opening to change membrane potential. The membrane potential change due to $K^+$ influx might be the critical factor in opening a voltage-dependent calcium channel for $Ca^{2+}$ influx.

• BMB Reports
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• 제30권3호
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• pp.182-187
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• 1997

The NADPH oxidase of phagocytes catalyzes the reduction of oxygen to $O_2^-$ at the expense of NADPH. The enzyme is dormant in resting neutrophils and becomes activated on stimulation. During activation, $p47^{phox}\;(\underline{ph}agocyte\;\underline{ox}idase\;factor)$, a cytosolic oxidase subunit, becomes extensively phosphorylated at a number of serines located between S303-S379. Oxidase activation can also be achieved by the addition of phosphorylated recombinant $p47^{phox}$ by protein kinase C in the cell-free system in the presence of $GTP{\gamma}S$. The cell-free activation is inhibited by wortmannin and LY294002. specific inhibitors of phosphatidylinositol 3kinase (PI 3-kinasel) These results indicate that PI 3-kinase may playa pivotal role in the activation of NADPH oxidase.

• Applied Microscopy
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• 제23권1호
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• pp.25-34
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• 1993

Prolactin has been reported to be present in the tear film of humans and prolactin-like immunoreactivity has been detected by immunofluorescence in acinar cells of the lacrimal glands of humans and rats. The present study was aimed at clarifying the intracellular distribution of the prolactin-like immunoreactivity, using the electron microscope immunogold technique. The lacrimal gland acinar cells have two types of secretory granules: 1) Secretory granules containing flocculent materials irregularly shaped and are often coalesced. 2) Secretory granules are fairly round and contain homogenous materials of a moderate electron density. The density of the granular content varies even within a single cell. We found prolactin-like reactivity in secretory granules, some smaller cytosolic vesicles, Golgi cisternae and nuclei in acinar cells from intact glands of rat. Our present results are consistent with the conclusion that prolactin is present in lacrimal cells. The presence of prolactin reactivity in the nucleus suggests that prolactin may be a regulatory factor modulating gene expression.