• Title, Summary, Keyword: Lectin

Search Result 349, Processing Time 0.094 seconds

Detection of Recombinant Marker DNA in Genetically Modified Glyphosate- Tolerant Soybean and Use in Environmental Risk Assessment

  • Kim, Young-Tae;Park, Byoung-Keun;Hwang, Eui-Il;Yim, Nam-Hui;Lee, Sang-Han;Kim, Sung-Uk
    • Journal of Microbiology and Biotechnology
    • /
    • v.14 no.2
    • /
    • pp.390-394
    • /
    • 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.

Optimization of Culture Conditions of Bacillus pumilus JB-1 for Chungkook-jang Fermentation in Soybean Boiling-Waste Liquor Medium (대두 열수 침출액을 이용한 청국장 발효균주 Bacillus pumilus JB-1의 배양 최적화)

  • Kwon, Ha-Young;Ryn, Hee-Young;Kwon, Chong-Suk;Lee, Sang-Han;Sohn, Ho-Yong
    • Microbiology and Biotechnology Letters
    • /
    • v.35 no.4
    • /
    • pp.304-309
    • /
    • 2007
  • Soybean is useful source of protein, especially in Asia. But soybean needs heat inactivation or fermentation process before consumption, since it contains the toxic lectin and various protease inhibitors. Therefore, production of soybean boiling-waste liquor (SBWL) as a byproduct is inevitable. In this study, the chemical composition of SBWL and the optimization of culture conditions for Bacillus pumilus JB-1, a selected strain for functional chungkuk-jang fermentation, using SBWL were investigated. The SBWL contains 88% water, 9.5% free sugar, 1.6% crude protein, 0.3% crude fat, 0.1% crude fiber and 2.1% ash, respectively. The contents of total polyphenol, total flavonoids and free amino acid in SBWL were 55%, 76%, and 30% of those of raw soybean, respectively. Culture conditions for B. pumilus JB-1 in SBWL were optimized. The 1/10-diluted, 0.1 % of $(NH_4)_2SO_4$ added SBWL without pH adjustment and carbon source addition was cultured at $37^{\circ}C$ for 48 h with agitation (120 rpm). The 0.5% of inoculation was enough. The large scale fermentation in 5-L jar fermentor showed that the SBWL is a good resource for production of chungkuk-jang starter and functional ingredients.

Reaction of Mast Cells and Goblet Cells in the Small Intestine of C57BL/6 and C3H/HeN Mice Infected with Echinostoma hortense

  • Park Kyeong-Yeol;Lee Kyu-Jae;Kim In-Sik;Yang Eun-Ju;Lim Su-jung;Lim Byung-Hyuk;Ryang Yong-Suk
    • Biomedical Science Letters
    • /
    • v.11 no.3
    • /
    • pp.259-266
    • /
    • 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.

  • PDF

Cytokine Expressions with Lectins from Allomyrina dichotoma and Bombyx mori (장수풍뎅이(Allomyrina dichotoma)와 누에(Bombyx mori) 렉틴의 사이토카인 생성 양상 비교)

  • Kim, Se-Jin;Chung, See-Ryun;Jeune, Kyung-Hee
    • Korean Journal of Pharmacognosy
    • /
    • v.36 no.2
    • /
    • pp.129-135
    • /
    • 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.

Overexpression of Galectin-3 in Macrophages of C57BL/6 mice with Experimental Autoimmune Encephalomyelitis (자가면역성 뇌척수염을 유도한 C57BL/6 마우스 큰포식세포에서의 Galectin-3의 과발현)

  • Kim, Dae Seung;Hwang, Insun;Park, Suk-jae;Ahn, Ginnae;Park, Sang-Joon;Park, Hyun Jeong;Joo, Hong-Gu;Jee, Youngheun
    • Korean Journal of Veterinary Research
    • /
    • v.51 no.2
    • /
    • pp.139-149
    • /
    • 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.

Changes of the Structural and Biomechanical Properties of the Bovine Pericardium after the Removal of ${\alpha}$-Gal Epitopes by Decellularization and ${\alpha}$-Galactosidase Treatment

  • Nam, Jinhae;Choi, Sun-Young;Sung, Si-Chan;Lim, Hong-Gook;Park, Seong-Sik;Kim, Soo-Hwan;Kim, Yong Jin
    • The Korean Journal of Thoracic and Cardiovascular Surgery
    • /
    • v.45 no.6
    • /
    • pp.380-389
    • /
    • 2012
  • Background: Bovine pericardium is one of the most widely used materials in bioprosthetic heart valves. Immunologic responses have been implicated as potential causes of limited durability of xenogenic valves. This study aimed to determine the effectiveness of decellularization and ${\alpha}$-galactosidase (${\alpha}$-gal) to remove major xenoreactive antigens from xenogenic tissues. Materials and Methods: Recombinant Bacteroides thetaiotaomicron (B. thetaiotaomicron) ${\alpha}$-gal or decellularization, or both were used to remove ${\alpha}$-gal from bovine pericardium. It was confirmed by ${\alpha}$-gal-bovine serum albumin-based enzyme-linked immunosorbent assay (ELISA), high-performance anion exchange chromatography, flow cytometry, 3,3'-diaminobenzidine-staining, and lectin-based ELISA. The mechanical properties of bovine pericardium after decellularization or ${\alpha}$-gal treatment were investigated by tests of tensile-strength, permeability, and compliance. Collagen fiber rearrangement was also evaluated by a 20,000${\times}$ transmission electron microscope (TEM). Results: Recombinant B. thetaiotaomicron ${\alpha}$-gal could effectively remove ${\alpha}$-gal from bovine pericardium B. thetaiotaomicron (0.1 U/mL, pH 7.2) while recombinant human ${\alpha}$-gal removed it recombinant human ${\alpha}$-gal (10 U/mL, pH 5.0). There was no difference in the mechanical properties of fresh and recombinant ${\alpha}$-gal-treated bovine pericardium. Furthermore, the TEM findings demonstrated that recombinant ${\alpha}$-gal made no difference in the arrangement of collagen fiber bundles with decellularization. Conclusion: Recombinant B. thetaiotaomicron ${\alpha}$-gal effectively removed ${\alpha}$-gal from bovine pericardium with a small amount under physiological conditions compared to human recombinant ${\alpha}$-gal, which may alleviate the harmful xenoreactive immunologic responses of ${\alpha}$-gal. Recombinant ${\alpha}$-gal treatment had no adverse effects on the mechanical properties of bovine pericardium.

Study of Rat Mammary Epithelial Stem Cells In Vivo and In Vitro (생체 및 시험관에서 유선 상피 모세포의 분리와 동정)

  • Nam Deuk Kim;Kee-Joo Paik
    • Journal of the Korean Society of Food Science and Nutrition
    • /
    • v.24 no.3
    • /
    • pp.470-486
    • /
    • 1995
  • Mammary epithelial cells contain a subpopulation of cells with a large proliferativ potential which are responsible for the maintenance of glandular cellularity and are the progenitor cells of mammary cancer. These clonogens give rise to multicellular clonal alveolar or ductal units(AU or DU) on transplantation and hormonal stimulation. To isolate putative mammary clonogens, enzymatically monodispersed rat mammary epithelial cells from organoid cultures and from intact glands are sorted by flow cytometry according to their affinity for FITC labeled peanut lectin(PNA) and PE labeled anti-Thy-1.1 antibody(Thy-1.1) into four subpopulations : cells negative to both PNA and Thy-1.1(B-), PNA+cells, Thy-1.1+cells, and cells positive to both reagents(B+). The in vivo transplantation assays indicate that the clonogenic fractions of PNA+cells from out-growths of organoids in primary cultures for three days in complete hormone medium(CHM) are significantly higher than those of cells from other subpopulations derived from cultrues or from intact glands. Extracellular matrix(ECM) is a complex of several proteins that regulated cell function ; its role in cell growth and differentiation and tissue-specific gene expression. It can act as a positive as well as a negative regulator of cellular differentiation depending on the cell type and the genes studied. Regulation by ECM is closely interrelated with the action of other regulators of cellular function, such as growth factors and hormones. Matrigel supports the growth and development of several different multicellular colonies from mammary organoids and from monodispersed epithelial cells in culture. Several types of colonies are observed including stellate colonies, duct-like structures, two- and three-dimensional web structures, squamous organoids, and lobulo-duct colonies. Organoids have the greatest proliferative potential and formation of multi-cellular structures. Phase contrast micrographs demonstrate extensive intracellular lipid accumulation within the web structures and some of duct-like colonies. At the immunocytochemical and electron micrograph level, casein proteins are predominantly localized near the apical surface of the cells or in the lumen of duct-like or lobulo-duct colonies. Squamous colonies are comprised of several layers of squamous epithelium surrounding keratin pearls as is typical fo squamous metaplasia(SM). All-trans retinoic acid(RA) inhibits the growth of SM. The frequency of lobulo-ductal colony formation increased with the augmentation of RA concentration in these culture conditions. The current study models could provide powerful tools not only for understanding cell growth and differentiation of epithelial cells, but also for the isolation and characterization of mammary clonogenic stem cells.

  • PDF

Macrophage and Anticancer Activities of Feed Additives on β-Glucan from Schizophyllum commune in Breast Cancer Cells (치마버섯균 유래의 베타글루칸에 대한 사료첨가제로서의 대식세포 기능 활성 및 유방암 세포주에서의 항암효능 효과)

  • Lee, Jin-Seok;Lee, Seung-Ho;Jang, Yong-Man;Lee, Jong-Dae;Lee, Byoung-Hee;Jung, Ji-Youn
    • Journal of the Korean Society of Food Science and Nutrition
    • /
    • v.40 no.7
    • /
    • pp.949-955
    • /
    • 2011
  • [ ${\beta}$ ]Glucan is a polysaccharide expressed on the cell walls of fungi. It is known that ${\beta}$-glucan is recognized by a family of C-type lectin receptors, dectin-1, which is expressed mainly on myeloid immune cells, including macrophages, neutrophils and dendritic cells. Raw 264.7 cells were treated with ${\beta}$-glucan from Schizophyllum commune. ${\beta}$-Glucan was not cytotoxic up to 400 ${\mu}g$/mL as measured by MTT assay. To measure the activity of macrophages, NO and TNF-${\alpha}$ assays were performed in Raw 264.7 cells. Treatment with ${\beta}$-glucan for 24 hr significantly increased production of NO and TNF-${\alpha}$ compared with control groups (p<0.05), indicating activation of macrophages. To measure inhibition of breast cancer cell proliferation, MTT assay was performed in MDA-MB-231 cells. Cell viability was significantly decreased in the group treated with 400 ${\mu}g$/mL of ${\beta}$-glucan for 48 hr (p<0.05) compared to the control group. However, tumor volume was decreased in the groups administered 200 ${\mu}g$ of ${\beta}$-glucan/mouse compared to the control group. These results indicate that ${\beta}$-glucan inhibits breast cancer cell growth through the induction of apoptosis.

Effect of Clonorchis sinensis Excretory-secretory Product on the Cultured SD Rat Bile Duct Fibroblast (배양된 흰쥐 담관 섬유모세포에 대한 간흡충 분비배설 물질의 영향)

  • Kwon, Jung-Nam;Min, Byoung-Hoon;Lee, Haeng-Sook;Kim, Soo-Jin;Joo, Kyoung-Hwan
    • Applied Microscopy
    • /
    • v.39 no.2
    • /
    • pp.117-124
    • /
    • 2009
  • Clonorchis sinensis is the most important widely distributed parasite of the human bile duct in East Asia and the most prevalent parasitic helminth in Korea. The prevalence rate of human clonorchiasis has remained at about 2.9% in Korea. C. sinensis induces dilatation of the duct, hyperplasia of the mucosa, metaplasia or neoplasia of the mucosal epithelium, periductal inflammation and fibrosis, and thickening of the ductal wall. Fibroblast are the most common cells in connective tissue and are responsible for the synthesis of extracellular matrix components. The fibrosis associated with chronic inflammation and injury may also contribute to cholangiocarcinoma pathogenesis, particularly through an increase in extracellular matrix components, which participate in the regulation of bile duct differentiation during development. In this study, ultrastructural changes, the distribution of lectin receptors and actin protein in cultured SD rat bile duct fibroblast after infection of C. sinensis were observed. Experimental group had been divided into four groups: normal bile duct fibroblast cultured in basal media (G1); C. sinensis infected bile duct fibroblast cultured in basal media (G2); normal bile duct fibroblast cultured in basal media containing excretory-secretory product (ESP) (G1-1); C. sinensis infected bile duct fibroblast cultured in basal media containing ESP (G2-1). Overall, once a host is infected by C. sinensis, it affects the host to the extent that sialic acid of ductal fibroblast is increased. Number of cytoplasmic process of SD rat bile duct fibroblast was increased. Actin protein and sialic acid were located in cell surface. Fibroblast induced by C. sinensis was not recovered to normal fibroblast. The cytoplasm bulk and cytoplasmic process were increased whereas the growth rate of the fibroblast of infected SD rat was reduced rather than that of normal fibroblast. In result, it inhibits fibroblast proliferation and increases actin protein on fibroblast cytoplasm, and so causes fibroblast metamorphosis and cellular mutation.