• Proceedings of the Korean Society of Applied Pharmacology
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• 2003.11a
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• pp.98-98
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• 2003

${\beta}$-(1,3)-D-Glucans have been known to exhibit antitumor and antimicrobial activities. The presence of dectin-1,${\alpha}$, ${\beta}$-glucan receptor of dendritic cell, on macrophage has been controvertial. RT-PCR analysis led to the detection of dectin-1${\alpha}$ and ${\beta}$ in murine macrophage Raw264.7 cell line. Among the various organs of mouse, dectin-1${\alpha}$ and ${\beta}$ were detected in the thymus, lung, spleen, stomach and intestine. To analyze gene expression modulated by ${\beta}$-glucan treated murine Raw264.7 macrophage, total mRNA was applied to cDNA microarray to interrogate the expression of 7,000 known genes. cDNA chip analysis showed that ${\beta}$-glucan of P. osteatus increased gene expressions of immunomodulating genes, membrane antigenic proteins, chemokine ligands, complements, cytokines, various kinases, lectin associated genes and oncogenes in Raw 264.7 cell line. When treated with ${\beta}$-glucan of P. osteatus and LPS, induction of gene expression of TNF-${\alpha}$ and IFN-R1 was confirmed by RT-PCR analysis. Induction of TNF-R type II expression was confirmed by FACS analysis. IL-6 expression was abolished by EDTA in ${\beta}$-glucan and LPS treated Raw264.7 cell line, indicating that ${\beta}$-glucan binds to dectin-l in a Ca$\^$＋＋/ -dependent manner. To increase antitumor efficacy of ${\beta}$-glucan, ginsenoside Rh2 (GRh2) was co-treated with ${\beta}$-glucan in vivo and in vitro tests. IC$\sub$50/ values of GRh2 were 20 and 25 $\mu\textrm{g}$/$m\ell$ in SNU-1 and B16 melanoma F10 cell line, respectively. Co-treatment with ${\beta}$-glucan and GRh2 showed synergistic antitumor activity with cisplatin and mitomycin C both in vitro and in vivo. Single or co-treatment with ${\beta}$-glucan and GRh2 increased tumor bearing mouse life span. Co-treatment with ${\beta}$-glucan and GRh2 showed more increased life span with mitomycin C than that with cisplatin. Antitumor activities were 67% and 72 % by co-injection with ${\beta}$-glucan and GRh2 in the absence or presence of mitomycin C, respectively.

• Journal of Life Science
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• v.27 no.6
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• pp.632-639
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• 2017

Mature haptoglobin (Hp) is a plasma glycoprotein and acts as an antioxidant by scavenging cell-free hemoglobin (Hb). Prohaptoglobin (proHp) is an unprocessed Hp precursor which is present a little in circulation. However, the biological function of proHp remains unknown. To investigate the structural and functional differences between proHp and Hp, we prepared recombinant proHp isoforms and compared their sialic acid content and Hb-binding capacity with those of mature isoforms. When proHp samples were analyzed by Western blot under non-reducing conditions, proHp1 was detected as one band of approximately 130 kDa and proHp2 as multiple bands >200 kDa, in the manner of mature Hp1-1 and Hp2-2, respectively. On the native polyacrylamide gel under non-reducing and non-denaturing conditions, both proHp isoforms migrated more slowly than their mature Hp counterparts. In addition, the lectin-based ELISA assay demonstrated that the content of sialic acid in proHp1 and proHp2 was much less than in Hp1-1 and Hp2-2. The Hb-binding capacity of proHp was also lower than those of mature Hp. These findings indicate that proHp and Hp are similar in the size and polymerization pattern, but different in sialic acid content and Hb-binding activity. It suggests precursor proHp may exert different functions in circulation than does mature Hp.

• The Korean Journal of Malacology
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• v.28 no.4
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• pp.377-384
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• 2012

The importance of biological resources has been gradually increasing, and mollusks have been utilized as main fishery resources in terrestrial ecosystems. But little is known about genomic and transcriptional analysis in mollusks. This is the first report on the transcriptomic profile of Meretrix lusoria. In this study, we constructed cDNA library and determined 542 of distinct EST sequences composed of 284 singletons and 95 contigs. At first, we identified 180 of EST sequences that have significant hits on protein sequences of the exclusive Mollusks database through BLASTX program and 343 of EST sequences that have significant hits on NCBI NR database. We also found that 211 of putative sequences through local BLAST (blastx, E < e-10) search against KOG database were classified into 16 functional categories. Some kinds of immune response related genes encoding allograft inflammatory factor 1 (AIF-1), B-cell translocation gene 1 (BTG1), C-type lectin A, thioester-containing protein and 26S proteasome regulatory complex were identified. To determine phylogenetic relationship, we identified partial sequences of four genes (COX1, COX2, 12S rRNA and NADH dehydrogenase) that significantly matched with the mitochondrial genomes of 3 species-Ml (Meretrix lusoria), Mp (Meretrix petechialis) and Mm (Meretrix meretrix). As a result, we found that there was a little bit of a difference between sequences of Korean isolates and other known isolates. This study will be useful to develop breeding technology and might also be helpful to establish a classification system.

• Biomedical Science Letters
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• v.11 no.3
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• pp.259-266
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• 2005

Mast cells and goblet cells have been known to protect the host against parasites. In this study, we examined the response of the mast cells and goblet cells over a period of 6 weeks in the duodenum, jejunum and ileum of C3H/HeN and C57BL/6 mice infected with Echinostoma hortense (E. hortense). In addition, we investigated whether the worm recovery rate of uninfected mice (the control group) or E. hortense-infected mice (the experimental group) was associated with the number of mast cells and goblet cells. The worm recovery rate was higher in the C3H/HeN mice than in the C57BL/6 mice. The number of goblet cells significantly increased in the experimental group of the C3H/HeN and C57BL/6 mice compared with the control group of both strains (P<0.005). Worm recovery peaked 3 weeks after the infection of the C57BL/6 mice and at 2 weeks after the infection of the C3H/HeN mice, and it was higher in the duodenum than in the jejunum and ileum. However, the infected site in the intestine had no relation with worm expulsion. In the C3H/HeN and C57BL/6 mice, the number of goblet cells in the experimental group was significantly higher than that in the control group (P<0.005). The number reached a peak 2 weeks after the infection and it even increased in duodenum, jejunum and ileum. The increased number of goblet cells was retained 6 weeks after infection. The number of goblet cells was higher in the C3H/HeN mice than in the C57BL/6 mice (P<0.01). These results indicate that goblet cells are related with the worm expulsion. Furthermore, immunohistostaining of the antral intestinal walls for lectin showed the significant increase of the number of goblet cells in the experimental group (P<0.001). The high infection rate in the duodenum was found during the early infection. An increased infection rate in the jejunum and ileum was found 3 weeks after infection and the infection rate was higher in the C3H/HeN mice than in the C57BL/6 mice. Taken together, the present study indicates that goblet cells, rather than mast cells, may play critical roles in parasite expulsion.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.390-394
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• 2004

The genetically modified glyphosate-tolerant soybean contains the following introduced DNA sequences: the EPSPS (5-enol-pyruvylshikimate-3-phosphate synthase) gene from Agrobacterium sp. strain CP4, the 35S promoter from the cauliflower mosaic virus, and the NOS terminator from Agrobacterium tumefaciens. In the present study, detection of these introduced DNAs was performed by amplification using the polymerase chain reaction (PCR). A multiplex PCR method was also applied to prevent false positive results. When primers for 35S promoter, nos3', CTP(chloroplast transit peptide), and CP4 EPSPS (EPSPS from Agrobacterium sp. CP4) were used, positive results were obtained in PCR reactions using DNA from genetically modified glyphosate-tolerant soybeans. There were no false positive results when using DNA from non-genetically modified soybeans. The CP4 EPSPS gene was detected when less than 125 pg glyphosate-tolerant soybean DNA was amplified. Lectin Lel and psb A were amplified from both non-genetically modified and genetically modified glyphosate-tolerant soybean DNA. Multiplex PCR was performed using different primer sets for actin Sacl, 35S promoter and CP4 EPSPS. The actin gene was detectable in both non-genetically modified and glyphosate-tolerant soybeans as a constant endogenous gene. Target DNAs for the 35S promoter, and CP4 EPSPS were detected in samples containing 0.01-0.1% glyphosate-tolerant soybean, although there were variations depending on primers by multiplex PCR. Soybean seeds from five plants of non-genetically modified soybean were co-cultivated for six months with those of genetically modified soybean, and they were analyzed by PCR. As a result, they were not positive for 35S promoter, nos3' or CP4 EPSPS. Therefore, these results suggest there was no natural crossing of genes between glyphosate-tolerant and non-genetically modified soybean during co-cultivation, which indicates that gene transfer between these plants is unlikely to occur in nature.

• Korean Journal of Pharmacognosy
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• v.36 no.2 s.141
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• pp.129-135
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• 2005

Lectins from Allomyrina dichotoma (ADL) and Bombyx mori (BML) were partially purified by physiological saline extraction, ammonium sulfate fractionation, anion exchange column chromatography on DEAE Sephadex A-50 and gel filtration column chromatography on Sephadex G-200. An assay for cytokine expression was carried out by using reverse transcription polymerase chain reaction(RT-PCR). mRNA isolated from PBMC(human peripheral blood mononuclear cells) were stimulated with ADL(O.D.=0.2) and BML(O.D.=0.1) for various times(1,4,8,24,48 and 72 h) and various cytokine mRNA assessed by RT-PCR were shown as follows: The patterns of bands for IL-1 mRNA of BML were very similar with those from ADL and these bands were decreased along the increasing reaction times after showing a strong band at 1 h. However mRNA expressions for IL-2, IL-6, $IFN{\gamma}$ and $TNF{\alpha}$ showed different patterns between ADL and BML. With the effect of ADL, the expression of IL-2 and IL-6 mRNA were continuously detected until 72 h with the strongest band of IL-2 mRNA at 24 h. The strong bands of $IFN{\gamma}$ mRNA were observed from 4 to 8 h but the strongest one of $TNF{\alpha}$ was just observed at 1 h. Meanwhile with BML, the bands for IL-2 and $IFN{\gamma}$ were increased along the increasing reaction times until 72 h. The strongest bands were showed from 4 to 8 h with IL-6 and at 8 h with $TNF[\alpha}$. To verify quantitatively ELISA was used for assay of protein secretions of the cytokine gene with IL-2 and $IFN{\gamma}$ expressed markedly different in RT-PCR. The highest cytokine secretion for IL-2 was demonstrated at 48 h. The production of $IFN{\gamma}$ was markedly increased at 24 h and secreted highest at 72 h. These result suggest that ADL and BML, as inducers of cytokines, can elicit detectable cytokine mRNA from PBMC within the first few hours of stimulation and maintain the production of cytokines for a few days by the methods of RT-PCR and ELISA.

• Biomolecules & Therapeutics
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• v.26 no.2
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• pp.121-129
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• 2018

Oxidized low-density lipoprotein (ox-LDL)-induced macrophage foam cell formation and apoptosis play critical roles in the pathogenesis of atherosclerosis. Thioredoxin-1 (Trx) is an antioxidant that potently protects various cells from oxidative stress-induced cell death. However, the protective effect of Trx on ox-LDL-induced macrophage foam cell formation and apoptosis has not been studied. This study aims to investigate the effect of recombinant human Trx (rhTrx) on ox-LDL-stimulated RAW264.7 macrophages and elucidate the possible mechanisms. RhTrx significantly inhibited ox-LDL-induced cholesterol accumulation and apoptosis in RAW264.7 macrophages. RhTrx also suppressed the ox-LDL-induced overproduction of lectin-like oxidized LDL receptor (LOX-1), Bax and activated caspase-3, but it increased the expression of Bcl-2. In addition, rhTrx markedly inhibited the ox-LDL-induced production of intracellular reactive oxygen species (ROS) and phosphorylation of p38 mitogen-activated protein kinases (MAPK). Furthermore, anisomycin (a p38 MAPK activator) abolished the protective effect of rhTrx on ox-LDL-stimulated RAW264.7 cells, and SB203580 (a p38 MAPK inhibitor) exerted a similar effect as rhTrx. Collectively, these findings indicate that rhTrx suppresses ox-LDL-stimulated foam cell formation and macrophage apoptosis by inhibiting ROS generation, p38 MAPK activation and LOX-1 expression. Therefore, we propose that rhTrx has therapeutic potential in the prevention and treatment of atherosclerosis.

• Journal of Embryo Transfer
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• v.18 no.1
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• pp.51-59
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• 2003

This study was carried out to evaluate the effect of different freezing and thawing rates on the viability, motility and acrosomal changes of frozen canine spermatozoa. The ejaculated semen was extended with Tris-egg yolk buffer containing 8% glycerol and equilibrated for 60 min after cooled to 4$^{\circ}C$ for 58 min. The straws were cryopreserved gradually by slow-cooling at different distance(6, 10 and 17 cm, respectively) from the liquid nitrogen (L$N_2$) to achieve temperature rate of 3, 8.9 and 19$^{\circ}C$ /min. Thawing of the straws was performed in a water bath fur 2 min at 37$^{\circ}C$ and 55$^{\circ}C$ , respectively. The motility of frozen-thawed spermatozoa was assessed by phase-contrast microscopy. To assess their viability and acrosome content, spermatozoa were stained with a vital stain and Fluorescence conjugated lectin Pisum Savitum Agglutinin (FITC/PAS), respectively. Concentration of the ejaculated fresh semen was normal range of 3.44 $\times$ 10$^{8}$ /ml. Freezing temperature were reduced to -110, -70 and -35$^{\circ}C$, as higher distance from liquid nitrogen, 6, 10 and 17 cm, respectively. Freezing at 3$^{\circ}C$/min in distance of 17 cm from liquid nitrogen yielded better motility, viability and rate of intact acrosome than 8.9 or 19$^{\circ}C$/min and the optimal thawing was 37$^{\circ}C$ for 2 min.

• Korean Journal of Veterinary Research
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• v.51 no.2
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• pp.139-149
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• 2011

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease in the murine central nervous system (CNS) and has long been used as an animal model for human multiple sclerosis. Development of EAE requires coordinated expression of a number of genes that are involved in the activation and effector functions of inflammatory cells. Galectin-3 (Gal-3) is a member of the betagalactoside- binding lectin family and plays an important role in inflammatory responses through its functions on cell activation, cell migration or inhibition of apoptosis. We investigated the functional role of Gal-3 in EAE mice following immunization with myelin oligodendrocyte glycoprotein $(MOG)_{35-55}$ peptide. During the peak stage of EAE, the localization of Gal-3 in inflammatory cells markedly increased in subarachnoid membranes and perivascular regions of CNS. In contrast, Gal-3 was weakly detected in cerebrum and spinal of the recovery stage of EAE. Consistent with this finding, western blot analysis revealed that Gal-3 expression was significantly increased at the peak stage while it was slightly decreased at the recovery stage in the CNS. In addition, the population of $CD11b^{+}$ macrophage expressing Gal- 3 in spleen of EAE mice was markedly increased compared with control mice. In fact, most of activated macrophages isolated from spleen of EAE mice expressed Gal-3. Taken together, our results demonstrate that the over-expression of Gal-3 in activated macrophages may play a key role in promoting inflammatory cells in the CNS during EAE.

• The Journal of Internal Korean Medicine
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• v.22 no.2
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• pp.145-160
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• 2001

Objectives : The purpose of this investigation is to evaluate the effect of Geupoongjibo-dan Extracts on Reversible Forebrain Ischemia in Mongolian Gerbils. Methods : The change rate of water content in cerebral tissues, the numercal change of the CA1 pyramidal neuron in the hippocampus, the change of delayed neuronal death(necrosis apoptosis) through light microscopy, the reactivity change of glycoprotein in neuronal membrane and the ultrastructural change of pyramidal neuron through electron microscopy caused by dalayed neuronal death were investigated. Results : 1. The change rate of water content in the normal group showed 78.90% on the third day, and 79.12% on the seventh day after an attack of ischemia. The rate in the control group showed 82.25% and 85.13%, respectively. The rate in the sample group showed a significant decrease: 81.72% and 83.66%. 2. Light microscopy revealed that the cells, continuous and systematic forms in the pyramidal cells of hippocampus, changed into discontinuous and unsystematic forms in the normal group when compared with the control group. The cells were less damaged in the sample group. 3. The mean of the numerical change of the CA1 pyramidal neurons in the hippocampus was 104 in the normal group. The mean of the control group was decreased to 27. The mean of the sample group was 44. 4. TUNEL staining examination reveals that the whole part of the hippocampus of the normal group had negative reactivity. As far as CA1 pyramidal neurons in the hippocampus, the control group had positive reactivity. The sample group was more positive than the control group. 5. Electron microscopy reveals that the ischemic injury of the control group had both necrotic and apoptotic morphology. The sample group was less necrotic, and more apoptotic morphology than the control group. 6. Lectin histochemisrical examination reveals that the normal group had positive reactivity to PNA and SBA in interneuron, and weak positive reactivity to WGA Con A LCA in intercelluar space. The reactivity to PNA and WGA decreased in the control group. The reactivity to PNA and WGA tended to increase in the sample group. Conclusions : The data shows that the effect of Geupoongjibo-dan Extracts on Reversible Forebrain Ischemia in MG is a significant result.