Nuclear Factor - kappa B Activation and Signal Transduction Pathway in Human Intestinal Epithelial Cells Stimulated with Bacteroides fragilis Enterotoxin

No abstract available.

Inflammatory Reaction in Intestinal Mucosa by Bacteroides fragilis Enterotoxin Stimulation: Regulation of Chemokine Gene Expression in Human Intestinal Epithelial Cells

No abstract available.

Basolaterial Secretion of CXC Chemokines by Human Intestinal Epithelial Cells in Response to Bacteroides fragilis Enterotoxin Via NF-kB Pathway

No Abstract Available.

Basolaterial Secretion of CXC Chemokines by Human Intestinal Epithelial Cells in Response to Bacteroides fragilis Enterotoxin Via NF-kB Pathway

No Abstract Available.

Cytokine gene expression of peritoneal tissues in response to mixed infection of Bacteroides fragilis and Escherichia coli

Bacteroides fragilis and Escherichia coli, normal colonic inhabitants, are the most frequently isolated bacteria in infected tissues, particularly in intraabdominal abscesses. This study was designed to determine whether enteric bacteria may alter the B. fragilis-induced expression of proinflammatory cytokines in mouse peritoneal tissue (MPT). After C57BL/6 mice were inoculated with abscess-forming mixture containing B. fragilis in the presence or absence of E. coli, RNA was extracted from MPT. Expression of interleukin (IL)-1alpha and tumor necrosis factor (TNF)alpha mRNA was assessed using RT-PCR and standard RNA. Each cytokine protein was also measured by ELISA. The co-inoculation of E. coli into mouse peritoneal cavity advanced the onset of abscess development by B. fragilis infection. When mouse was co-infected with E. coli and B. fragilis intraperitoneally, there was a synergistic increase in the expression of IL-1alpha and TNFalpha mRNA in MPT and this was paralleled by increased cytokine protein secretion. Mixed inoculation of heat-killed E. coli and B. fragilis did not cause a synergistic increase in those cytokine mRNA expression. These results suggest that enteric bacteria may significantly affect proinflammatory cytokine signal produced by host peritoneal cavity in response to B. fragilis infection.

Production and Characterization of Human CD27lg, CD40fg and CD95lg Fusion Proteins in Chinese Hamster Ovary Cell / 대한면역학회지

No abstract available.

Expression of Proinflammatory Cytokine Genes in Mouse Peritoneal Tissue Infected with Bacteroides fragilis

OBJECTIVE: Bacteroides fragilis, normal colonic inhabitant, is the most frequently isolated anaerobes in infected tissues, particularly in intraabdominal abscesses. In the acute infection model with abscesses, the response to B. fragilis infection is characterized by infiltration of neutrophils and macrophages. This study was designed to determine whether proinflammatory cytokines could be upregulated in peritoneal tissue of B. fragilis-infected mouse model. METHODS: After C57BL/6 mice were infected with abscess-inducing encapsulated B. fragilis, RNA was extracted from the intraperitoneal tissues. Cellular RNA was also extracted from mouse peritoneal macrophages (MPM) and human peripheral blood mononuclear cells (PBMC) after infection with B. fragilis. Expression of various cytokine mRNA was assessed using RT-PCR and standard RNA. Each cytokine protein was also measured by ELISA. RESULTS: B. fragilis-infected intraperitoneal tissues showed upregulated expression of IL-1u, IL-6 and TNFu mRNA. Expression of IL-1u and TNFu mRNA and protein was significantly higher in MPM or PBMC infected with B. fragilis than in those without infection. However, expression of IL-6 mRNA and protein was not increased in MPM or PBMC infected with B. fragilis compared with those without infection. CONCLUSION: These results suggest that the cytokines can be involved in immunopathologic reactions of the peritoneal tissue infected with B. jragilis.

Expression of Cyclooxygenase - 2 in Intestinal Epithelial Cells in Response to Invasive Bacterial Infection and its Role of Epithelial Cell Apoptosis

Invasion of enteric bacteria, such as Salmonella and invasive E. coli, into intestinal epithelial cells induces proinflammatory gene responses and finally epithelial cell apoptosis. In this study, we asked whether invasive bacterial infection of human intestinal epithelial cells could upregulate cyclooxygenase-2 (COX-2) gene expression and whether increased COX-2 expression could influence intestinal epithelial cell apoptosis. Expression of COX-2 mRNA and prostaglandin (PG) E production were upregulated in HT-29 colon epithelial cells which were infected with S. dublin or invasive E. coli, as examined by quantitative RT-PCR and radioimmunoassay. Inhibition of COX-2 expression and PGE2 production using NS-398, a specific COX-2 inhibitor, showed a significant increase af epithelial cell apoptosis and caspase-3 activation in HT-29 cells infected with invasive bacteria. However, the addition of valerylsalicylate, a specific COX-1 inhibitor, did not change apoptosis in S. dublin-infected HT-29 cells. These results suggest that upregulated COX-2 expression and PGE2 production in response to invasive bacterial infection could contribute to host defense by inhibiting apoptosis of intestinal epithelial cells.

Electron Microscopic Analysis of Apoptosis of HGPRT- Mouse Myeloma Cell Induced by Aminopterin, a de novo Pathway Blocking Agent / 대한면역학회지

Programrned cell death (PCD), or apoptosis, is a process by which cells die in response to specific physiological and toxicological signals. This genetically programmed form of cellular suicide is intirnately involved in many biological processes including growth, metamorphosis, embryogenesis, and oncogenesis. Cells undergoing PCD in normal and neoplasmic tissues display the following biochemical and morphological features: internucleosomal DNA fragmentation, reduced cell volume, condensed chromatin in nucleus, zeiosis and generation of apoptotic bodies containing intact organelles and plasma rnembrane. Hybridoma cell production, resulting from the fusion of myeloma cells with antibody producing spleen cells, is widely used in various fields of life science. This technique requires hypoxanthine guanine phosphoribosyl transferase (HGPRT) deficient mutant myeloma cell line as a fusion partner. When these mutants cell is treated with aminopterin plus hypoxanthine-thymidine (HAT) after the cell fusion they are selectively and efficiently eliminated remaining fused hybridoma celis. But there hasn't been any report regarding the selective elimination mechanism of this HGPRT mutant myeloma cell. By using HGPRT myeloma P3-X 63-Ag8.653 (V653) as a model system, this study demonstrated that PCD was induced by aminopterin treatment of this V653 cell line. And the sequential ultrastructural changes during this death process were observed by using electron microscope. When V653 cells were incubated with 0.4 uM aminopterin, DNA fragmentation was detectable after 3 hours and peaked between 12 and 18 hours of aminopterin treatment and the cell viability decreased in a time dependant manner. V653 cells incubated with amiopterin showed following ultrastructural changes during the death process. Dilatation of rough endoplasmic reticulum (RER) and detachment of ribosomes were the earliest ultrastructural changes and first seen after 30 minute incubation. Dilatation of perinuclear cisternae began to appear after 1 hour and deformation of nucleoplasm such as decreased electron density of perinuclear heterochromatin and increased electron density of euchromatin were seen after 3 hours. Increased electron density of cytoplasm, decreased cell volume, condensation of chromatin and apoptotic bodies were observed in many cells after 9 hours but vacuolation by severe dilatation of RER was seen in some cells. 24 hours after incubation with aminopterin, many cells showed typical form of apoptosis characterized by cell shrinkage, increased electron density of cytoplasm and apoptotic bodies. On the contrary, some cells showed different type of cell death characterized by increased cell volume, decreased electron density of cytoplasm, severely dilated RER and apoptotic bodies. In both types of cells, mitochondrial cristae and limiting membrane of mitochondria are comparatively well preserved. In other cells, nuclei did not show significant changes but there were deformations of mitochondria such as markedly increased electron density and formation of lamella bodies. The death process of V653 cell was not synchronized among cells. The results of this study proved that aminopterin-induced selective elimination of fusion partner V653 myeloma cell is due to PCD. The earliest ultrastructural changes observed in this process were dilatation of RER and detachment of ribosomes. And there were two distinct morphological types in the PCD.

Expression of Monocyte Chemotactic Protein-1 and Macrophage Inflammatory Protein-1alpha in Intraabdominal Abscess Induced by Bacteroides fragilis Infection

OBJECTIVE: Bacteroides fragilis is the most frequently isolated anaerobes in tissue of intraabdominal infection, particularly in intraabdominal sepsis or abscess. In acute experimental model with an intraabdominal infection, the response to B. fragilis is characterized by infiltration of neutrophils and monocytes. To understand the pathogenesis of B. fragilis infection, it is important to explore the mechanism for inflammatory signals such as chemokines induced by this bacteria. The goal of this study was to determine whether peritoneal monocytes or fibroblasts surrounding with intraabdominal abscess induced by B. fragilis could express chemokines such as monocyte chemotactic protein-1 (MCP-1) or macrophage inflammatory protein-1a (MIP-1a). METHODS: 1) After C57BL/6 mice were intraperitoneally inoculated with abscess-inducing agents containing B. fragilis, RNA was extracted from the intraperitoneal tissues of the mice using Ottawa sand and the guanidinium thiocyanate-phenol-chloroform method in 3 days later. 2) After C57BL/6 mouse peritoneal monocytes were infected with B. fragilis for 1, 4 and 9 hours, cellular RNA was extracted from the cells. 3) Fibroblasts isolated from intraabdominal abscess nodules induced by B. fragilis infection were growth in tissue culture for 3 to 4 weeks. After the fibroblasts were stimulated with IL-1alpha (0.1-10 ng/ml) or TNFalpha (0.1-10 ng/ml) for 24 hours, total cellular RNA was extracted. MCP-1 or MIP-1alpha mRNA expression was assessed using RTPCR. MCP-1 or MIP-1alpha proteins in cluture supernatants or tissue extracts were also measured by ELISA. RESULTS: 1) MCP-1 or MIP-1alpha mRNA was highly expressed in peritoneal tissue of C57BL/6 mice bearing with intraabdominal abscess induced by B. fragilis. 2) Expression of MCP-1 mRNA increased at 9 hours in mouse peritoneal monocytes infected with B. fragilis. MIP-1alpha mRNA was initially expressed and perisisted in the monocytes infected with B. fragilis for 9 hours. MCP-1 or MIP-1alpha proteins was also parallel to the expression of those chemokines. 3) The fibroblasts isolated from intraabdominal abscess nodules by B. fragilis infection constitutively expressed MCP-1, MIP-1alpha, production in the fibroblasts was significantly upregulated in response to proinflammatory cytokines produced in the monocytes, including IL-1alpha and TNFalpha, but MCP-1 production were not. The normal fibroblasts from uninfected mice didnot show significant production of MCP-1 or MIP-1a in response to IL-1a or TNFalpha. CONCLUSION: These results suggest that peritoneal monocytes and fibroblasts surrounding with abscesses induced by B. fragilis produce MCP-1 or MIP-1a. Furthermore, it could be extrapolated that those effects may play a role in the formation of intraabdominal abscess nodules.

Alteration of Phospholipase D Activity in the Rat Tissues by Irradiation / 대한치료방사선과학회지

PURPOSE: Phospholipase D (PLD) catalyzes the hydrolysis of phosphatidyl choline to phosphatidic acid (PA) and choline. Recently, PLD has been drawing much attentions and considered to be associated with cancer process since it is involved in cellular signal transduction. In this experiment, oleate-PLD activities were measured in various tissues of the living rats after whole body irradiation. MATERIAL AND METHODS: The reaction mixture for the PLD assay contained 0.1microCi 1,2-di[1-14C]palmitoyl phosphatidylcholine, 0.5mM phosphatidylcholine, 5mM sodium oleate, 0.2% taurodeoxycholate, 50mM HEPES buffer (pH 6.5), 10mM CaCl2, and 25mM KF. phosphatidic acid, the reaction product, was separated by TLC and its radioactivity was measured with a scintillation counter. The whole body irradiation was given to the female Wistar rats via Cobalt 60 Teletherapy with field size of 10cm x 10cm and an exposure of 2.7Gy per minute to the total doses of 10Gy and 25Gy. RESULTS: Among the tissues examined, PLD activity in lung was the highest one and was followed by kidney, skeletal muscle, brain, spleen, bone marrow, thymus, and liver. Upon irradiation, alteration of PLD activity was observed in thymus, spleen, lung, and bone marrow. Especially PLD activities of the spleen and thymus revealed the highest sensitivity toward gamma-ray with more than two times amplification in their activities. In contrast, the PLD activity of bone marrow appears to be reduced to nearly 30%. Irradiation effect was hardly detected in liver which showed the lowest PLD activity. CONCLUSION: The PLD activities affected most sensitively by the whole-body irradiation seem to be associated with organs involved in immunity and hematopoiesis. This observation strongly indicates that the PLD is closely related to the physiological function of these organs. Furthermore, radiation stress could offer an important means to explore the phenomena covering from cell proliferation to cell death on these organs.

Characterization of cytoplasmic Form of Human CTLA - 4 Molecule / 대한면역학회지

CTLA-4 (=CD152), a T cell activation antigen, has been known to be homologous to CD28 in its molecular and genomic structure. Both of these two molecules are sharing their counterreceptors, B7 (CDSO) and B7-2 (CD86) and are known to play a crucial role in T cell activation. In previous our study it was reported that there are 2 forms of CTLA-4 antigen in activated human T cells, 30 kD membrane-bound form and 34 kD cytosolic-sequestered form and the former was less than 5 % of total of this antigen induced. Aims of this study are to confirm previous finding by using flow cytometry and to characterize the cytoplasmic form of human CTLA-4 by using ultrafiltration and immunoprecipitation techniques. In PHA stimulated peripheral blood lymphocyte surface expression of CTLA-4 was less than 2.1% of any of CD4+, CD8+ and CD56+ subsets. And the 34 kD form of CTLA-4 was detected in CDS+ subset only. This discrepancy confirms that 34 kD antigen is the cytoplasmic form of human CTLA-4. In ultrafiltration and subsequent Western blot analysis study this 34 kD antigen was detected in >100 kD fraction only. And in non-reducing condition this antigen formed high molecular weght complex (MW > 350 kD). In immunoprecipitation study using anti-peptide A antibody it was found that this high molecular weight complex consists of the 34 kD cytoplasmic form of CTLA-4 and previously unknown 54 kD antigen and 46 kD antigen at 1:1:8-10 ratio. And none of these 3 molecules were identified in membrane fraction of activated human T cell. The result of this study implies that CTLA-4 molecule induced upon T cell activation mainly sequestered in cytoplasrn and another signal is necessary to target this antigen on the activated T cell surface.

Development of Monoclonal Antibodies Recognizing Human Peripheral Blood T Lymphocytes Cytoplasmic Proteins Induced upon Activation / 대한면역학회지

Antigen-specific T cell activation requires interaction of the T cell with specialized antigen-presenting cells. Signaling through the TCR is necessary but not sufficient to induce antigen-specific T cell activation and cytokine secretion. This first signal, termed signal 1, is both antigen-specific and MHC-restricted. Signal 2, which is neither antigen-specific nor MHC-restricted, is necessary to induce cytokine secretion, cellular proliferation, and effector function. Recently immunological studies in T cell activation area are mainly focused on biological and molecular biological characterization of TCR/CD3 complex and accessary molecules providing costimulatory signal (signal 2). If signal 2 is not delivered, T cell enter a state of long term un-responsiveness to specific antigen-termed anergy. Monoclonal antibody technique has been especially involved in recognizing novel inducible cell surface antigens on T cell activation. This study was aimed to develop monoclonal antibodies recognizing novel cytoplasmic proteins present in activated T cells. We make 6 monoclones involved in changing pattern of T cell activated cytoplasmic proteins. Using these 6 monoclonal antibodies analyze to find novel molecules involved in T cell activation associated response, apoptosis, and/or heat shock response of the T cells in early T cell activation.

Seroprevalence of Norwalk-Like Viruses in Seoul: Detection Using Baculovirus-Expressed Norwalk Virus Capsid Antigen

The present study was designed to estimate the seroprevalence of NLVs among diarrheagenic children and in healthy adults in Seoul and its vicinity with the use of an EIA and an Western blot (WB) based on recombinant Norwalk virus capsid protein (rNV) and crude virus preparations as antigen. Seroconversion was observed in 34 (83%) of 41 tested using the EIA and in 21 (54%) of 39 using the WB, suggesting that the NLVs with epitopes common to rNV are prevalent in Seoul area. Diarrheal children who were known to have been infected with several other strains of the NLVs showed no significant antibody response to the rNV. Infection with rNV occurred earlier in life: primary infections with rNV were common before the age of 6 months and over 91% of children had evidence of infection by that age by the EIA. Since the amount of the NLV antigens available for seroepidemiologic surveys is limited, we tried to detect NLV antibody by using crude virus preparations as antigen. One crude virus preparation of a child whose stool yielded genetically distinct NLV revealed the presence of the plural number of bands upon SDS-PAGE, but precipitated only one band (62 kDa) after the WB with a serum (collected 10 days after the onset of symptoms) of another diarrheal child. The WB assay we present in this report revealed that the NLVs are prevalent among Korean population and that the sera contained antibody to a single major structural protein, with molecular sizes of 58 to 62 kDa, compatible with the sizes reported for the Norwalk virus and Snow Mountain agent proteins, respectively. When the results of the WB were compared with those obtained by the EIA, the EIA antibody assay was sensitive enough to detect an antibody rise of as much as 4096-fold but not as specific as the WB. The WB assay presented in this paper will provide a powerful tool to elucidate not only antigenic structures of the NLVs but also seroepidemiology of the NLV infection. The availability of an unlimited source of antigen will enable a large scale serologic studies that will greatly increase our understanding of the role of NLVs in human enteric illness.

Respiratory Syncytial Virus Infections and Application of Nested Reverse Transcription-Polymerase Chain Reaction

No abstract available.

Expression of P, type 1 fimbriae, HEp-2 cell adherence, and colonization factor antigens of uropathogenic escherichia coli

No abstract available.

Expression of human CTL?4 extracellular domain in escherichia coli / 대한면역학회지

No abstract available.

CD4+T lymphocyte, beta2-microglobulin and p24 antigen level in HIV infected persons / 감염

No abstract available.

Detection of human CTLA-4 by using anti-peptide antibody

No abstract available.

Inhibition of IL-2 dependent DTLL-2 proliferation by immune complex from patient with ovarian cancer

No abstract available.