• The Journal of Korean Society of Virology
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• v.26 no.1
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• pp.47-58
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• 1996

Histopathological vascular changes in hemorrhagic fever with renal syndrome (HFRS) caused by Hantaan virus include increased vascular permeability, disseminated intravascular coagulation, thrombocytopenia and changes in coagulation activity. Although vascular endothelial cells of main target organs such as kidney infected with Hantaan virus are not damaged but swelling of endothelial cells, perivascular exudates and infiltration of mononuclear cells and fresh interstitial hemorrhages are common. However, the pathogenesis of cell infiltration and hemorrhages around vascular endothelial cells are not well understood. Some endothelial cell molecules or vascular adhesins that acts as adhesion moleulces for leukocyte are expressed on endothelial cells close to site of inflammation. However, whether the expression of endothelial adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule (ICAM-1) and endothelial leukocyte adhesion molecule (ELAM) on vascular endothelial cells are increased by infection with Hantaan virus has not been studied. In this study, the relationship between the expression of VCAM-1, ICAM-1 and ELAM and adhesion of mononuclear cells on endothelial cells of human blood vessels infected with Hantaan virus was investigated. The endothelial cells of umbilical vein was passaged three times in culture medium and the monolayered cells were infected with $10^5\;pfu/ml$ of Hantaan virus grown in Vera E6 cell cultures. The multiplication of virus in cultured endothelial cells was monitored by immunohistochemistry and the expression of adhesion molecules was demonstrated by immunohistochemistry using monoclonal antibodies against VCAM-1, ICAM-1 and ELAM. And in situ hybriditation against ICAM-1 was also performed. The endothelial adhesion molecules, VCAM and ICAM, were expressed after 6 hours postinfection, respectively, and their expressions lasted for 72 hours. Similar expression of VCAM and ICAM appeared on endothelial cells by infection with virus, but the expression of ELAM was not recognized up to 72 hours postinfection. Microscopically, it was noted that many monocuclear cells adhered on endothelial cells infected with viruses. In an electronmicroscopic study, the transendothelial migration of mononuclear cells was observed on monolayered endothelial cells infected with virus. This results suggested that the endothelial adhesion molecules, particulary VCAM and ICAM, might be expressed on endothelial cells by infection with Hantaan virus and these molecules play a key role in the adhesion and extravasation of inflammatory cells around blood vessels.

• Bulletin of the Korean Chemical Society
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• v.30 no.12
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• pp.2999-3005
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• 2009

Among the more than 120 different types of human papillomavirus (HPV), types 16 and 18 have been known to be high risk agents that cause cervical cancer. We examined, in an immuno-chromatographic analysis, the potential of using the early gene product, E7 protein, as a diagnostic marker of cervical cancer caused by HPV. We developed monoclonal antibodies specific to HPV-16 and 18 E7 proteins that were produced from bacterial cells using gene recombinant technology. For each E7 protein, the optimal antibody pair was selected using the immuno-chromatographic sandwichtype binding system based on the lateral flow through membrane pores. Under these conditions, this rapid testing assay had a detection capability as low as 2 ng/mL of E7 protein. Furthermore, since viral analysis required the host cell to be lysed using chemicals such as detergents, it was possible that the E7 protein was structurally damaged during this process, which would result in a decrease in detection sensitivity. Therefore, we examined the detrimental effects caused by different detergents on the E7 protein using HeLa cells as the host. In these experiments, we found that the damage caused by the detergent, nonylphenylpolyethylene glycol (NP-40), was minimal relative to Triton X-100 commonly used for the cell lysis. Temperature also affected the stability of the E7 protein, and we found that the E7 protein was stabilized at 4$^{\circ}C$ for about 2 h, which was 4 times longer than at room temperature. Finally, a HPV-infected cervical cancer cell line, which was used as a real sample model, was treated using the optimized conditions and the presence of E7 proteins were analyzed by immuno-chromatography. The results of this experiment demonstrated that this rapid test could specifically detect HPV-infected samples.

• Biomedical Science Letters
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• v.1 no.1
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• pp.27-35
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• 1995

Several studies showed that the immunological factors such as CD4+ cell number, CD4%, CD8+ cell number and CD4/CD8 ratio and the serological factors such as, ${\beta}^2$-microglobulin(${\beta}^2$-MG), neopterin, soluble CD4, and soluble CD8 are related to the risk of development of AIDS. Especially, the CD4+ cell counts have been used to monitor progresson of HIV disease, to stratify, and to follow patients in clinical trials. Recently, the Centers for Disease Control and Prevention(CDCP) in USA has made the CD4+ cell count as a part of the classification of HIV disease. It is composed of 3 categories such as 1, 2, and 3 which asr $\geq$ 500/$mm^3$, 200/$mm^{3} $\geq$ and < 500/$mm^3$, and < 200/$mm^3$, respectively. In this study, to estimate the differences of immunological factors between HIV-infected and normal human groups in Korea, CD4+ T and CD8+ T cells, and the CD4/CD8 ratio were measured in 185 HIV-infected subjects and 140 healthy adult subjects. The lymphocyte subsets such as CD4+ T and CD8+ T were analysed by flow cytometer(FACStar) with two-color immunofluorescent stain using monoclonal antibodies such as anti-CD4 and anti-CD8 antibodies. The absolute numbers and percentages of CD4+ T and CD8+ T and the CD4/CD8 ratio of HIV infected persons were $462\pm{277}/mm^3$, $18.2\pm7.7%$, $1,170\pm{534}/mm^3$, $47.0\pm10.6%$ and $0,43\pm0.26 whereas those of uninfected persons were $886\pm{299}mm^3$, $32.9\pm{7.0%}, 730{\pm}259/mm^3$, $26.8\pm6.4%$ and $1.31\pm0.46$(P<0.01). In addition, estimating the reference values of peripheral blood lymphocyte subsets of Korean, the absolute numbers and percentages of CD4+ T and CD8+ T and the CD4/CD8 ratio of 140 healthy adults persons were measured and compared with those of foreigners. The reference ranges of CD4+ T cells, CD8+ T cells, CD4％, CD8%, and the CD4/CD8 ratio and 1.31$\pm$0.46, respectively. The significant differences were not observed when compared with those of foreigners. However a little difference was observed in the percentages of CD4+ T and the absolute numbers of CD8+ T between the normal values of Korean and those of foreigners were $43.6\pm8.9%$, $560\pm{230}/mm^3$. This result can also be useful as a basic data for the treatment and surveillance of HIV-infected patients in Korea.

• Tuberculosis and Respiratory Diseases
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• v.42 no.6
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• pp.823-830
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• 1995

Background: It has been found that Helper T cells in the peripheral blood are decreased in the cell mediated immunity in the pulmonary tuberculosis. But it has not been confirmed yet that only decrease in number of cells which has phenotype in the peripheral blood is defined to decrease in cell mediated immunity. The immunocytochemical study was performed to observe the change of the percentage of T-lymphocytes with their subsets and activated T cells in the peripheral blood of pulmonary tuberculosis and to know how many T cells would be activated, relative to resting cells in the peripheral blood. Methods: The peripheral blood obtained from twenty two patients and ten healthy controls were smeared on the gelatin coated slide glass prepared for of mononuclear cells. The double bridge technique of alkaline phosphatase-antialkaline phosphatase(APAAP) method was used. As the primary antibodies, $T_1$(anti-human T cell), $T_4$(anti-human helper/inducer T cells) and $T_8$(anti-human supressor/cytotoxic T cell) antibodies and interleukin-2 receptor (for early activated T cell), very late activation antigen (for activated cytotoxic T cell), T cell lineage specific activation antigen monoclonal actibodies were used. Results: 1) There were significantly decrease in the absolute number of $T_4$(+) cells but significantly increase of $T_8$(+) cells in the peripheral blood of pulmonary tuberculosis (p<0.05). 2) The percentage of $T_4$(+) cells showed significantly decrease in pulmonary tuberculosis but $T_8$(+)cells significantly increase(p<0.05). $T_4(+)/T_8(+)$ ratio showed significantly decrease in the peripheral blood of the pulmonary tuberculosis(p<0.05). 3) There were significantly increase in the absolute number of variable stages of activated T cells in the peripheral blood of the pulmonary tuberculosis(p<0.05). 4) The percentage of IL-2R, VLA-1, TLiSA were 6.45+1.56%, $7.64+1.34^*$, 10.45+1.16% in order which showed significantly increase in the peripheral blood of the pulmonary tuberculosis(p<0.05). Conclusion: We speculate that only a few percentage of T lymphocyte is activated in cell mediated immunity in pulmonary tuberculosis.

• IMMUNE NETWORK
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• v.2 no.3
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• pp.142-149
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• 2002

Background: Our previous study showed that purified protein derivative (PPD)-stimulated pleural mononuclear cells (PMC) from tuberculous pleurisy (Tbp) produced significantly more $IFN-{\gamma}$ (10- to 70-fold) after in vitro PPD stimulation than freshly isolated pleural cells from malignant pleurisy. The present study was designed to determine whether blocking the CD40-CD40 ligand (CD40L) interaction decreases $IFN-{\gamma}$ production by altering IL-12 levels. Methods: IL-12 and $IFN-{\gamma}$ production after neutralizing anti-CD40L antibody treatment was compared to the efficacy of anti-CD80, anti-CD86, and a combination of anti-CD80 and CD86 (CD80+86) monoclonal antibodies (mAb). These activities were measured by enzyme-linked immunosorbent assays (ELISAs) and reverse transcription-polymerase chain reaction (RT-PCR), after in vitro stimulation with PPO antigen (Ag). Results: Neutralization of CD80, CD86 and CD80+86 did not decrease $IFN-{\gamma}$ and IL-12 production in Tbp-PMC, whereas neutralization of CD40L significantly depressed IL-12 p40 and $IFN-{\gamma}$. In addition, neutralization of CD40L completely inhibited IL-12 p40 and $IFN-{\gamma}$ mRNA expression. Conclusion: The CD40-CD40L interaction might play a major role in IL-12 and $IFN-{\gamma}$ production in Tbp-PMC, thus contributing to protective immunity in human tuberculosis.

• IMMUNE NETWORK
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• v.1 no.1
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• pp.77-86
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• 2001

Background: Hepatitis C virus(HCV), a family of Flaviviridae, has a host cell-derived envelope containing a positive-stranded RNA genome, and has been known as the maj or etiological agent for chronic hepatitis, hepatic cirrhosis, and hepatocellular carcinoma. There remains a need to dissect a molecular mechanism of pathogenesis for the development of therapeutic and effective preventive measure for HCV. Identification of cellular receptor is of central importance not only to understand the viral pathogenesis, but also to exploit strategies for prevention of HCV. This study was aimed at identifying peptide mimotopes inhibiting the binding of E2 protein of HCV to MOLT-4 cell. Methods: In this study, phage peptide library displaying a random peptides consisting of 7 or 12 random peptides was employed in order to pan against E2 protein. Free HCV particles were separated from the immune complex forms by immunoprecipitation using anti-human IgG antibody, and used for HCV-capture ELISA. To identify the peptides inhibiting E2-binding to MOLT-4 cells, E2 protein was subj ect to bind to MOLT-4 cells under the competition with phage peptides. Results: Several phage peptides were selected for their specific binding to E2 protein, which showed the conserved sequence of SHFWRAP from 3 different peptide sequences. They were also able to recognize the HCV particles in the sera of HCV patients captured by monoclonal antibody against E2 protein. Two of them, showing peptide sequence of HLGPWMSHWFQR and WAPPLERSSLFY respectively, were revealed to inhibit the binding of E2 protein to MOLT-4 cell efficiently in dose dependent mode. However, few membrane-associated receptor candidates were seen using Fasta3 programe for homology search with these peptides. Conclusion: Phage peptides containing HLGPWMSHWFQR and WAPPLERSSLFY respectively, showed the inhibition of E2-binding to MOLT-4 cells. However, they did not reveal any homologues to cellular receptors from GenBank database. In further study, cellular receptor could be identified through the screening of cDNA library from MOLT-4 or hepatocytes using antibodies against these peptide mimotopes.

• Journal of Periodontal and Implant Science
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• v.29 no.2
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• pp.333-350
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• 1999

The modulation of leukocyte cell surface adhesion molecules may influence the development of cellular events that determine the course of the inflammatory process. Direct interaction between activated T cells and monocytes resulted in a large production of $IL-1{\beta}$ by monocytes. In this reactions, adhesion molecules play an important part, yet the role of them in Tmonocytes interaction remain unclear. This study was undertaken in an effort to elucidate, 1) the influence of 1.25(OH)$_2D_3-induced$ differentiation on the monocyte responsiveness to direct contact with T lymphocytes, and 2) the role of adhesion molecules on the T-monocyte direct interaction. Initially, I observed that direct contact of monocyte cell line THP-1 with stimulated fixed T cell line HuT78 markedly induces IL-1${\beta}$ production by THP-1. $IL-1{\beta}$ production was higher when THP-1 had been previously exposed to 1.25(OH)$_2D_3$ as compared to control, with ${\alpha}$- 1.25(OH)$_2D_3$ dose-dependent and exposure time-dependent manner. It was shown that 1.25(OH)$_2D_3$ also increased the expression of ${\beta}_2$ integrin adhesion receptor Mac-1(CD11b/CD18) dose- and timedependently, but did not increase the expression of human leukocyte antigen- D(HLA-D) and intercellular adhesion molecule-1(ICAM-1). The $IL-1{\beta}$ producing activity of THP-1 cells correlated well with the ability to induce the Mac-1 expression on THP-1 surface. Monoclonal antibody raised against relevant cell surface glycoproteins on THP-1 were tested for their ability to block the response of THP-1 to T cells. Antibody to Mac-1 only partially blocked $IL-1{\beta}$ production by THP-1, whereas antibodies to ICAM-1 and HLA-D did not. These data indicate that regulation of Mac-1 expression on THP-1 cells can alter the responsiveness of these cells to contact by activated T cells, however other unknown structures on the THP-1 cells may be involved in this process also.

• Korean Journal of Biological Psychiatry
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• v.3 no.2
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• pp.170-180
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• 1996

Lowered immune function in the senile dementia patients may be related to the abnormal metabolism of amyloid precursor protein(APP). To investigate the passibility of an abnormal metabolism of APP in lymphocytes and the possible role of APP in the activation of lymphocytes in senile dementia patients, immunohistochemical study of rat spleen and fluorescence activated cell sorter analysis(FACS) of human lymphocytes with the specific antigen far each lymphocyte and double fluorescent marker with antibody to APP were performed. After stimulating lymphocyte with phytohemagglutinin(PHA), APP mRNA and protein were extracted and quantitfied and the influence of ${\beta}$-amyloid protein($A{\beta}$) specific antibody on lymphocyte division was investigated. In spleen, the majority of cells showing $A{\beta}$ immunoreactivity was found in the T-sell dependent zone. FACS indicated that around 90% $CD_4(+)$ T-cells and 60% of $CD_8(+)$ T-sell were immunoreactive to $A{\beta}$ specific antibody(mAb 4G8). Northern blot analysis shows that lymphocyte APP mRNA was gradually increased to reach a maximum at 3 days after activation with lectin mitogen PHA. However, the $A{\beta}$ immunoreactivity an cell surface remained constant during stimulation with PHA, indicating that the release of APP(secreted farm of APP) might be increased. A very large increase in soluble APP secretion was observed in T-lymphocyte upon activation, but only law levels in the resting stale. Immunoblot was carried out an the protein obtained from cell lysate after stimulating lymphocyte by applying PHA to the cultured lymphocyte, and the result was that $A{\beta}$ band of immature farm under 116 KDa marker decreased as the duration of culture was increased after PHA stimulation. The monoclonal $A{\beta}$ specific(4G8) and polyclonal APP antibodies did not inhibit the [$^3H$]-thymidine uptake of mitogen-treated lymphocytes significantly, suggesting that mitogenesis can not be inhibited by specific $A{\beta}$ and polyclonal APP antibody. These results suggest that APP is expressed in T-cell and might be closely associated with the function of T-cells.

• The Journal of Korean Society of Virology
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• v.26 no.1
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• pp.9-22
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• 1996

Respiratory Syncytial virus (RSV) is an important cause of acute lower respiratory tract infections in human, with infants and young children being particularly susceptible. In the temperate zones, sharp annual outbreaks of RSV occur during the colder months, in both the northern and the southern hemisphere. RSV is unusual in that it can repeatedly reinfect individuals throughout life and infect babies in the presence of maternal antibody. RSV isolates can be divided into two subgroups, A and B, on the basis of their reactions with monoclonal antibodies, and the two subgroups are also distinct at the nucleotide sequence level. The specific diagnosis of RSV infection was best made by isolation of virus in tissue culture, identification of viral antigen, or by specific serologic procedures. Recently, rapid detection of RSV and analysis of RSV strain variation became possible by development of methods of reverse transcription and polymerase chain reaction amplification. In this study, to determine the genetic diversity of RSV found in Korea, 173 bp and 164 bp spanning selected regions of the RSV F and SH genes were enzymatically amplified and sequenced, respectively. Eight for F gene and three for SH gene were detected in 66 nasopharyngeal swap samples tested. Two major antigenic subgroups, A and B were confirmed from Korean samples (seven for subgroup A and one for subgroup B). At the nucleotide level of the F gene region, Korean subgroup A strains showed 95-99% homologies compared to the prototype A2 strain of subgroup A and 93-100% homologies among Korean subgroup A themselves. For the SH gene region, Korean subgroup A strain showed 97.5% homology compared to the prototype A2 strain of subgroup A, and Korean subgroup B strain showed 97% homology compared to the prototype 18537 strain of subgroup B. Most of base changes were transition and occured in codon position 3, which resulted in amino acid conservation. Using the maximum parsimony method, phylogenetic analysis indicated that Korean RSV strains formed a group with other RSV strains isolated from the United States, Canada, the Great Britain and Australia.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.10
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• pp.5973-5981
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• 2013

At present, the most common cause of cancer-related death in women is breast cancer. In a large proportion of breast cancers, there is the overexpression of human epidermal growth factor receptor 2 (HER2). This receptor is a 185 KDa growth factor glycoprotein, also known as the first tumor-associated antigen for different types of breast cancers. Moreover, HER2 is an appropriate cell-surface specific antigen for passive immunotherapy, which relies on the repeated application of monoclonal antibodies that are transferred to the patient. However, vaccination is preferable because it would stimulate a patient's own immune system to actively respond to a disease. In the current study, several bioinformatics tools were used for designing synthetic peptide vaccines. PEPOP was used to predict peptides from HER2 ECD subdomain III in the form of discontinuous-continuous B-cell epitopes. Then, T-cell epitope prediction web servers MHCPred, SYFPEITHI, HLA peptide motif search, Propred, and SVMHC were used to identify class-I and II MHC peptides. In this way, PEPOP selected 12 discontinuous peptides from the 3D structure of the HER2 ECD subdomain III. Furthermore, T-cell epitope prediction analyses identified four peptides containing the segments 77 (384-391) and 99 (495-503) for both B and T-cell epitopes. This work is the only study to our knowledge focusing on design of in silico potential novel cancer peptide vaccines of the HER2 ECD subdomain III that contain epitopes for both B and T-cells. These findings based on bioinformatics analyses may be used in vaccine design and cancer therapy; saving time and minimizing the number of tests needed to select the best possible epitopes.