• IMMUNE NETWORK
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• v.4 no.1
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• pp.31-37
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• 2004

Background: 1-8D gene is a member of human 1-8 interferon inducible gene family and is shown to be overexpressed in fresh colon cancer tissues. Three peptides 1-6, 3-5 and 3-7 derived from 1-8D gene were shown to have immunogenicity against colon cancer. Methods: To study tumor immunotherapy of these peptides we established an adoptive transfer model. $D^{b-/-}{\times}{\beta}2$ microglobulin (${\beta}2m$) null mice transgenic for a chimeric HLA-A2.1/$D^b-{\beta}2m$ single chain (HHD mice) were immunized with irradiated peptide-loaded RMA-S/HHD/B7.1 transfectants. Spleens were removed after last immunization, and splenocytes were re-stimulated in vitro. Lymphocytes from vaccinated HHD mice were transferred together with IL-2 to the tumor bearing nude mice that were challenged S.C. with the HCT/HHD/B7 colon carcinoma cell line that was found to grow in these mice. Results: Peptide 3-5 was found to be highly effective in CTL activity. Adoptively transferred anti-peptide 3-5 cytolytic T lymphocytes caused significant retardation in tumor growth. Conclusion: This study shows that peptide 3-5 can be the most effective candidate for the vaccine of adoptive immunotherapy against colon cancer.

• IMMUNE NETWORK
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• v.8 no.1
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• pp.7-12
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• 2008

Background: Members belonging to the interferon-lambda (IFN-${\lambda}$) family exert protective action against viral infection; however, the mechanisms of their action have remained elusive. To study IFN-${\lambda}$ biology, such as endocytosis of IFN-${\lambda}$, we produced monoclonal antibodies (Abs) against human IFN-${\lambda}$ and examined their usefulness. Methods: We purified recombinant human IFN-${\lambda}$1 expressed in Escherichia coli by using affinity columns. Then, we generated hybridoma cells by fusing myeloma cells with splenocytes from IFN-${\lambda}$1-immunized mice. For evaluating the neutralizing activity of the monoclonal Abs against IFN-${\lambda}$1, we performed RT-PCR for the MxA transcript. In order to study the binding activity of IFN-${\lambda}$ and the monoclonal Ab complex on HepG2 cells, we labeled the monoclonal Ab with rhodamine and determined the fluorescence intensity. Results: Four hybridoma clones secreting Abs specific to IFN-${\lambda}$1 were generated and designated as HL1, HL2, HL3, and HL4. All the Abs reacted with IFN-${\lambda}$1 in the denatured form as well as in the native form. Abs produced by HL1, HL3, and HL4 did not neutralize the induction of the MxA gene by IFN-${\lambda}$1. We also demonstrated the binding of the HL1 monoclonal anbitody and IFN-${\lambda}$ complex on HepG2 cells. Conclusion: Monoclonal Abs against IFN-${\lambda}$1 were produced. These Abs can be used to study the cellular binding and internalization of IFN-${\lambda}$.

• IMMUNE NETWORK
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• v.1 no.3
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• pp.187-195
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• 2001

The expression of recombinant proteins fused to 26 kDa glutathione S-transferase (GST) extracted from Schistosoma japonicum represents an attractive system for purifiying proteins of interest in a single step using GST-affinity chromatography. In addition, the GST-tag is used conveniently for detecting fused proteins since its high solubility as well as its relatively small size rarely interferes with the biological activity of the fused protein. In this regard, the GST system is frequently applied for tracing fusion proteins in both prokaryotic and eukaryotic cells to elucidate the physiological interactions and functional compartments of proteins. To provide a further tool in analyzing GST-fusion proteins, a new monoclonal antibody, with a high specificity to the S. japonicum GST was produced. Methods: BALB/c mice were immunized both with recombinant S. japonicum GST proteins, and by the fusion of splenocytes from these mice with myeloma cells. From this, a new anti -GST monoclonal antibody, termed SARAH, was generated. The specificity and reactivity of this antibody was confirmed by ELISA and by Western blot analysis. Results: SARAH showed a high reactivity to recombinant GST and GST fusion protein but not with native mammalian GST proteins as derived from other species including humans, cows, rabbits and rats. The applicability of SARAH was further demonstrated by confocal laser scanning microscopy, where GST proteins that were expressed transiently in mouse fibroblast cells, were specifically detected without interference of endogenous GST. Conclusion: SARAH is new monoclonal antibody with a high specificity to recombinant GST proteins but not to endogenous GST in mammalian cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.21 no.5
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• pp.1233-1242
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• 2007

Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation characterized by hyperplasia of synovial cells in affected joints, which might be mediated by the altered activation of Immune system, ultimately leading to the destruction of cartilage and bone. To examine effects of GHS on rheumatoid arthritis DBA/1J mice were immunized with bovine type II collagen to induced arthritis and then treated with GHS once a day for 7 weeks. Oral administration of GHS (200 mg/Kg) significantly suppressed the progression of CIA, which extend is comparable to that of methotrexate (MTX, 0.3 mg/Kg), a positive control. The severity of arthritis within the knee joints, which was evaluated by histological assessment of cartilage destruction and pannus formation, was also lowered by GHS. The production of TNF-and IL-6 in serum was significantly suppressed. The levels of IFN-g in the culture supernatant of splenocytes stimulated with CD3/CD28 or collagen were dramatically decreased, while those of IL-4 was increased. The levels of IgG and IgM RA factor were also decreased in the serum. FACS analysis indicated that B cells (in DLN), CD3+ T cells (in spleen, and paw joint), CD11b+Gr-1+ cells (in paw joint), CD3+CD49b(DX5) (in PBMC) were decreased and there was increased proportion of CD3+, CD4+, CD8+, CD4+CD25+ T cells in DLN. In conclusion, our results demonstrates that GHS significantly suppressed the progression of CIA and this action was characterized by the decreased production of TNF-a, IL-6, and rheumatoid factors, and modulations of immune cell populations.

• Toxicological Research
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• v.16 no.4
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• pp.269-274
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• 2000

Epoxidised soya bean oil (ESBO, 1000, 2000 or 4000 mg/kg) was orally administered to BALB/c mice daily for 28 consecutive days, and the control mice were exposed to vehicle (corn oil). Mice were immunized and challenged with sheep red blood cells (SRBC) or bovine serum albumin (BSA). In groups exposed to ESBO, the body weight gains and the relative lymphoid organ weights were not significantly changed as compared with control group. Secondary IgG antibody response to BSA was not significantly changed by ESBO, but plaque-forming cell (PFC) response to SRBC was significantly suppressed in mice treated with 4000 mg ESBO/kg/day. The mitogenic response of splenic B cells induced by LPS was not effected by ESBO in any of the groups. These results indicate that ESBO did not induce significant humoral immune response at a dose less than 2000 mg/kg/day in mice.

• Herbal Formula Science
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• v.21 no.1
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• pp.161-176
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• 2013

Objectives : To clarify the anti-arthritic activity of Ostericum koreanum Maxim. (OS) plus Aralia continentalis Kitagawa (AC) in vivo. Methods : All mice were immunized with bovine type II collagen. After a second collagen immunization, mice were treated with OS plus AC once a day for 7 weeks. Oral administration of OS plus AC (200 or 50 mg/kg) significantly suppressed the progression of CIA, which extend is comparable to that of methotrexate (MTX, 0.3 mg/kg), a positive control. The severity of arthritis within the knee joints was evaluated by histological assessment of cartilage destruction and pannus formation. Results : Administration of OS plus AC significantly suppressed the progression of CIA and inhibited the production of TNF-${\alpha}$ and IL-6 in serum. The erosion of cartilage was dramatically reduced in mouse knees after treatment with OS plus AC. In conclusion, our results demonstrates that OS plus AC significantly suppressed the progression of CIA and that this action was characterized by the decreased production of IL-6, IFN-${\gamma}$ and collagen II specific antibody in serum, CD3+CD69+ T cells, MHC class II+/CD11c+ (in DLN), CD11b+Gr-1+ cells (in PBMC), CD11b +Gr-1+ cells, B220+/CD23+ (in paw joint). Conclusions : The the levels of IFN-${\gamma}$ in the culture supernatant of splenocytes stimulated with CD3/CD28 or collagen were dramatically decreased, while those of IL-4 was increased. In the serum of OS and AC-treated mice, the levels of IgM RA factor were decreased.

• The Plant Pathology Journal
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• v.19 no.5
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• pp.260-265
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• 2003

Gummy stem blight caused by Didymella bryoniae occurs exclusively in cucurbits. This fungus has been known not to produce its pycnidium in vitro unless irradiated. In this study, cultural conditions for the mass-production of pycnidiospore by Metal Halide (MH) lamp irradiation were maximized. The mycelia were cultured at $26^{\circ}C$ on PDA for 2 days under dark condition, and then the plate was illuminated with MH lamp continuously for 3-4 days at $26^{\circ}C$. Results show that a great number of pycnidia were simultaneously formed. The pycnidiospores produced were then used as immunogen. Fusions of myeloma cell (v-653) with splenocytes from immunized mice were carried out. Two hybridoma cell lines that recognized the immunogen D. bryoniae were obtained. One monoclonal antibody (MAb), Dbl, recognized the supernatant while another MAb, Db15, recognized the spore. Two clones were selected which were used to produce ascite fluid of the two MAb, Dbl and Db15, the immunotypes of which were identified as IgG1 and IgG2b, respectively. Titers of MAb Dbl and MAb Db15 were measured and the absorbance exceeded 0.5 even at a $10^{-5}$ dilution. The MAbs reacted positively with D. bryoniae but none reacted with other viral isolates, Cucumber mosaic virus and Cucumber green mottle mosaic virus. Sensitivity of MAb was precise enough to detect spore concentration as low as $10^{-3}$/well by indirect ELISA. Characterization of the MAbs Dbl, Db15 antigen by heat and protease treatments, which suggests that the epitope recognized by these two MAbs was glycoprotein.

• BMB Reports
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• v.33 no.1
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• pp.92-95
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• 2000

The HIV-1 p24(202-221) sequence ETINNEEEWDRVHPV HAGP contains a B-cell epitope with the earliest immune response and the highest antibody titer against anti-mouse sera obtained by immunization with p24 antigens. A novel mouse monoclonal antibody (mAb) was generated against the immunodominant B-cell epitope of the HIV-1 p24 capsid protein, p24(202-221). BALB/c mice were immunized with the four branched multiple antigenic peptide (MAP) containing the HIV-1p24(202-221) sequence, and antibody-secreting hybridoma were produced by fusion of mouse splenocytes with P3X63Ag8.653, mouse myeloma cells. One clone which produced the antigen-specific mAb named KI-24 (Isotype IgG1, light chain: ${\kappa}$) was identified. mAb KI-24 was highly specific for both the p24(202-221) and p24 proteins when analyzed by ELISA and Western blotting. Since p24(202-221) also contains a cytotoxic T-lymphocyte epitope, this specfic peptide epitope and the monoclonal antibody with specific reactivity against the p24 protein and p24(202-221) can be used in peptide vaccine development and p24 antigen detection from HIV patients.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.2
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• pp.987-991
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• 2013

Aim: To study anti-tumor effects of exosomes from class II transactivator (CIITA) gene transfected CT26 cells. Methods: In this study, we established an MHC class II molecule-expressing murine colon cancer cell line (CT26-CIITA) by transduction of the CIITA gene. Immune effects in vitro and tumor protective results in vivo were tested and monitored. Results: Exosomes from CT26-CIITA cells were found to contain a high level of MHC class II protein. When loaded on dendritic cells (DCs), exosomes from CT26-CIITA cells significantly increased expression of MHC class II molecules, CD86 and CD80, as compared to exosomes from CT26 cells. In vitro assays using co-culture of immunized splenocytes and exosome-loaded DCs demonstrated that CIITA-Exo enhanced splenocyte proliferation and IFN-${\gamma}$ production of CD4+T cells, while inhibiting IL-10 secretion. In addition, compared to exosomes from CT26 cells, CT26-CIITA-derived exosomes induced higher TNF-${\alpha}$ and IL-12 mRNA levels. A mouse tumour preventive model showed that CT26-CIITA derived exosomes significantly inhibited tumour growth in a dose-dependent manner and significantly prolonged the survival time of tumour-bearing mice. Conclusion: Our findings indicate that CT26-CIITA-released exosomes are more efficient to induce anti-tumour immune responses, suggesting a potential role of MHC class II-containing tumour exosomes as cancer vaccine candidates.

• The Journal of the Korean Society for Microbiology
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• v.35 no.2
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• pp.117-127
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• 2000

The non-ELR-containing CXC chemokines Mig and IP-10 have been shown to function as chemotactic cytokines for activated T lymphocytes. In this study, we examined the potential involvement of Mig and IP-10 in antimycobacterial response of mice immunized or infected with M. bovis BCG. The accumulation of Mig and IP-10 mRNA in resident peritoneal monocytes ($RPM{\Phi}$) was slightly reduced by stimulation with vBCG, and the degree was greater for 24 hr culture even though IFN-${\gamma}$ was added. Expression of Mig, IP-10, and IFN-${\gamma}$ in 24 hr delayed-type hypersensitivity (DTH) response was stronger in vBCG-immune mice than in the non-immune. The increase of DTH measured by foot-pad thickness appears to be clearly related to the levels of chemokines Mig and IP10 messages and those of IFN-${\gamma}$ and IL-12. Stimulation with vBCG for 2 days decreased or completely dropped the levels of Mig message in non-immune or immune splenocytes, respectively, whereas IP-10 message was slightly decreased in 2 days culture. Moreover, messages for IL-12 (p40) showed similar kinetics for Mig. The levels of Mig and IP-10 mRNA during the course of infection with BCG were not readily changed in lungs, livers, and spleens from BCG-infected mice. Although there was no obvious changes of Mig and IP-10 messages in the target organs during infection process, we found that the infection progressed over the first 3 wk before being contained by the emerging immune response suggested from detectable amount of IFN-${\gamma}$ mRNA around this time. In view of selectivity of chemokines Mig and IP-10 for activated T cells, these data suggest that chemokine Mig and IP-10, especially in collaboration with IL-12 and IFN-${\gamma}$, may playa role as T cell recruiters in immune response against mycobacterial infection.