B 림프구의 면역반응 조절과 자가면역

이혜영,백상기 충남대학교 생물공학연구소 2003 생물공학연구지 Vol.9 No.1

The immune system of any organism must preserve a well balance between activation and inhibition. It must raise an effective immune response to target non-self molecules while not hampering the organism itself. Failure to maintain this balance will result in either immunodeficiency or autoimmunity. In many systemic autoimmune diseases, the production of pathogenic autoantibodies, released from B lymphocytes, has been the causes of autoimmunity, although the nature of the mechanism remains unclear. In recent years, several factors involved in the survival and inhibition of B lymphocytes were uncovered and found to be related to the several autoimmune diseases. Those are B lymphocytes inhibitory receptors and cytokines such as FcrRⅡ, CD22, PD-1, BAFF and BAFF-R. Investigating the genetic alterations and signalling components in these molecules and the consequences for the regulation of apoptosis/survival could show distinct ways to cure autoimmune diseases.

Silencing of BNIP3 Results from Promoter Methylation by DNA Methyltransferase 1 Induced by the Mitogen-Activated Protein Kinase Pathway

안현정,Hayyoung Lee,백상기 한국분자세포생물학회 2011 Molecules and cells Vol.31 No.6

We have previously shown that Ras mediates NO-induced BNIP3 expression via the MEK-ERK-HIF-1 pathway in mouse macrophages, and that NO-induced death results at least in part from the induction of BNIP3. In the present study, we describe another aspect of Ras regulation of BNIP3 expression in pancreatic cancer cells. Human BNIP3 promoter-driven luciferase activity was efficiently induced by activated Ras in AsPC-1, Miapaca-2, PK-1 and PANC-1 cells. However, expression of endoge-nous BNIP3 was not induced, and BNIP3 up-regulation by hypoxia was also inhibited. Treatment of the cells with the DNMT inhibitor, 5-aza-2-deoxycytidine, restored BNIP3 induction, indicating that DNA methylation of the BNIP3 promoter was respon-sible for the inhibition of BNIP3 induction. Furthermore, inhibition of the MEK pathway with U0126 reduced DNMT1 expression, but not that of DNMT3a and 3b, and restored the hypoxia-inducibility of BNIP3, suggesting that the DNA methylation of the BNIP3 promoter was mediated by DNMT1 via the MEK pathway.

대식세포주 RAW264.7에서 Nitric oxide에 의해 과발현되는 CD53의 기능분석

김태림,이혜영,김인규,백상기 충남대학교 생물공학연구소 2004 생물공학연구지 Vol.10 No.2

The CD53 antigen is a member of the tetraspanin membrane protein family that is expressed in the lymphoid-myeloid lineage. It is highly expressed in Radio-resistant tumor cells. Recently, it was reported that CD53 associates with the GSH-metabolizing protein γ-glutamyl transpeptidase. Its biological roles however remains unknown. Macrophages activated by microbial lipopolysaccharides (LPS) produce a burst of nitric oxide and reactive oxygen species. Nitric oxide plays important roles in macrophage activation as a toxic agent towards infectious organisms, an inducer or suppressor of apoptosis or an immunoregulator, whether nitric oxide can induce or inhibit apoptosis is highly dependent on different cell types at different stages of the inflammatory process. Using cDNA microarrays, we identified CD53 as one of the principal genes up-regulated by exposure of macrophages to LPS. We found that mRNA and protein levels of CD53 were increased by nitric oxide as well as LPS treatment in macrophages. Cells stably transfected with sense CD53 cDNA had lower levels of peroxide, and were more resistant to γ-irradiation. The stably transfected RAW264.7 cells with antisense CD53 recombination had the opposite properties. Activation of CD53 by cross-linking with monoclonal anti-CD53 antibody (OX-79) showed the expression of the inducible nitric oxide synthase. We propose that the induction of CD53 is one of major self-protection mechanisms of macrophages against LPS induced-oxidative stress and irradiation.

Kinetics of Binding of LPS to Recombinant CD14, TLR4, and MD-2 Proteins

Han Jae Shin,Hayyoung Lee,Jong Dae Park,Hak Chul Hyun,Hyung Ok Sohn,Dong Wook Lee,Young Sang Kim 한국분자세포생물학회 2007 Molecules and cells Vol.24 No.1

The Membrane-Bound Form of IL-17A Promotes the Growth and Tumorigenicity of Colon Cancer Cells

Van Anh Do Thi,박상민,Hayyoung Lee,김영상 한국분자세포생물학회 2016 Molecules and cells Vol.39 No.7

Interleukin-17A is a member of the IL-17 family, and is known as CTLA8 in the mouse. It is produced by T lym-phocytes and NK cells and has proinflammatory roles, inducing cytokine and chemokine production. However, its role in tumor biology remains controversial. We investigated the effects of locally produced IL-17A by transferring the gene encoding it into CT26 colon cancer cells, either in a secretory or a membrane-bound form. Expression of the membrane-bound form on CT26 cells dramatically enhanced their proliferation in vitro. The enhanced growth was shown to be due to an increased rate of cell cycle progression: after synchronizing cells by adding and withdrawing colcemid, the rate of cell cycle progression in the cells expressing the membrane-bound form of IL-17A was much faster than that of the control cells. Both secretory and membrane-bound IL-17A induced the expression of Sca-1 in the cancer cells. When tumor clones were grafted into syngeneic BALB/c mice, the tumor clones expressing the membrane-bound form IL-17A grew rapidly; those expressing the secretory form also grew faster than the wild type CT26 cells, but slower than the clones expressing the membrane-bound form. These results indicate that IL-17A promotes tumorigenicity by enhancing cell cycle progression. This finding should be considered in treating tumors and immune-related diseases.

Ectopically Expressed Membrane-bound Form of IL-9 Exerts Immune-stimulatory Effect on CT26 Colon Carcinoma Cells

Van Anh Do Thi,Sang Min Park,Hayyoung Lee,Young Sang Kim 대한면역학회 2018 Immune Network Vol.18 No.1

Inhibitory effect of a phosphatidyl ethanolamine derivative on LPS-induced sepsis

Lee, Chunghyun,An, Hyun-Jung,Kim, Jung-ln,Lee, Hayyoung,Paik, Sang-Gi Springer-Verlag 2009 Molecules and cells Vol.27 No.2

<P>Sepsis is the leading cause of death in critically ill patients. Today, around 60% of all cases of sepsis are caused by Gram-negative bacteria. The cell wall component lipopoly-saccharide (LPS) is the main initiator of the cascade of cellular reactions in Gram-negative infections. The core receptors for LPS are toll-like receptor 4 (TLR4), MD-2 and CD14. Attempts have been made to antagonize the toxic effect of endotoxin using monoclonal antibodies against CD14 and synthetic lipopolysaccharides but there is as yet no effective treatment for septic syndrome. Here, we describe an inhibitory effect of a phosphatidylethanolamine derivative, PE-DTPA (phosphatidylethanolamine diethyl-enetriaminepentaacetate) on LPS recognition. PE-DTPA bound strongly to CD14 (K ( d ), 9.52 x 10(-8) M). It dose dependency inhibited LPS-mediated activation of human myeloid cells, mouse macrophage cells and human whole blood as measured by the production of tumor necrosis factor-a (TNF-alpha) and nitric oxide, whereas other phospho-lipids including phosphatidylserine and phosphatidylethanolamine had little effect. PE-DTPA also inhibited transcription dependent on NF-kappaB activation when it was added together with LPS, and it rescued LPS-primed mice from septic death. These results suggest that PE-DTPA is a potent antagonist of LPS, and that it acts by competing for binding to CD14.</P>

Interleukin-9 Inhibits Lung Metastasis of Melanoma through Stimulating Anti-Tumor M1 Macrophages

Park, Sang Min,Do-Thi, Van Anh,Lee, Jie-Oh,Lee, Hayyoung,Kim, Young Sang Korean Society for Molecular and Cellular Biology 2020 Molecules and cells Vol.43 No.5

Interleukin-9 (IL-9) is well known for its role in allergic inflammation. For cancer, both pro- and anti-tumor effects of IL-9 were controversially reported, but the impact of IL-9 on tumor metastasis has not yet been clarified. In this study, IL-9 was expressed as a secretory form (sIL-9) and a membrane-bound form (mbIL-9) on B16F10 melanoma cells. The mbIL-9 was engineered as a chimeric protein with the transmembrane and cytoplasmic region of TNF-α. The effect of either mbIL-9 or sIL-9 expressing cells were analyzed on the metastasis capability of the cancer cells. After three weeks of tumor implantation into C57BL/6 mice through the tail vein, the number of tumor modules in lungs injected with IL-9 expressing B16F10 was 5-fold less than that of control groups. The percentages of CD4⁺ T cells, CD8⁺ T cells, NK cells, and M1 macrophages considerably increased in the lungs of the mice injected with IL-9 expressing cells. Among them, the M1 macrophage subset was the most significantly enhanced. Furthermore, peritoneal macrophages, which were stimulated with either sIL-9 or mbIL-9 expressing transfectant, exerted higher anti-tumor cytotoxicity compared with that of the mock control. The IL-9-stimulated peritoneal macrophages were highly polarized to M1 phenotype. Stimulation of RAW264.7 macrophages with sIL-9 or mbIL-9 expressing cells also significantly increased the cytotoxicity of those macrophages against wild-type B16F10 cells. These results clearly demonstrate that IL-9 can induce an anti-metastasis effect by enhancing the polarization and proliferation of M1 macrophages.

The Membrane-Bound Form of IL-17A Promotes the Growth and Tumorigenicity of Colon Cancer Cells

Thi, Van Anh Do,Park, Sang Min,Lee, Hayyoung,Kim, Young Sang Korean Society for Molecular and Cellular Biology 2016 Molecules and cells Vol.39 No.7

Interleukin-17A is a member of the IL-17 family, and is known as CTLA8 in the mouse. It is produced by T lymphocytes and NK cells and has proinflammatory roles, inducing cytokine and chemokine production. However, its role in tumor biology remains controversial. We investigated the effects of locally produced IL-17A by transferring the gene encoding it into CT26 colon cancer cells, either in a secretory or a membrane-bound form. Expression of the membrane-bound form on CT26 cells dramatically enhanced their proliferation in vitro. The enhanced growth was shown to be due to an increased rate of cell cycle progression: after synchronizing cells by adding and withdrawing colcemid, the rate of cell cycle progression in the cells expressing the membrane-bound form of IL-17A was much faster than that of the control cells. Both secretory and membrane-bound IL-17A induced the expression of Sca-1 in the cancer cells. When tumor clones were grafted into syngeneic BALB/c mice, the tumor clones expressing the membrane-bound form IL-17A grew rapidly; those expressing the secretory form also grew faster than the wild type CT26 cells, but slower than the clones expressing the membrane-bound form. These results indicate that IL-17A promotes tumorigenicity by enhancing cell cycle progression. This finding should be considered in treating tumors and immune-related diseases.

Cell-Based IL-15:IL-15Rα Secreting Vaccine as an Effective Therapy for CT26 Colon Cancer in Mice

Thi, Van Anh Do,Jeon, Hyung Min,Park, Sang Min,Lee, Hayyoung,Kim, Young Sang Korean Society for Molecular and Cellular Biology 2019 Molecules and cells Vol.42 No.12

Interleukin (IL)-15 is an essential immune-modulator with high potential for use in cancer treatment. Natural IL-15 has a low biological potency because of its short half-life and difficulties in mass-production. IL-15Rα, a member of the IL-15 receptor complex, is famous for its high affinity to IL-15 and its ability to lengthen the half-life of IL-15. We have double-transfected IL-15 and its truncated receptor IL-15Rα into CT26 colon cancer cells to target them for intracellular assembly. The secreted IL-15:IL-15Rα complexes were confirmed in ELISA and Co-IP experiments. IL-15:IL-15Rα secreting clones showed a higher anti-tumor effect than IL-15 secreting clones. Furthermore, we also evaluated the vaccine and therapeutic efficacy of the whole cancer-cell vaccine using mitomycin C (MMC)-treated IL-15:IL-15Rα secreting CT26 clones. Three sets of experiments were evaluated; (1) therapeutics, (2) vaccination, and (3) long-term protection. Wild-type CT26-bearing mice treated with a single dose of MMC-inactivated secreted IL-15:IL-15Rα clones prolonged survival compared to the control group. Survival of MMC-inactivated IL-15:IL-15Rα clone-vaccinated mice (without any further adjuvant) exceeded up to 100%. This protection effect even lasted for at least three months after the immunization. Secreted IL-15:IL-15Rα clones challenging trigger anti-tumor response via CD4<SUP>+</SUP> T, CD8<SUP>+</SUP> T, and natural killer (NK) cell-dependent cytotoxicity. Our result suggested that cell-based vaccine secreting IL-15:IL-15Rα, may offer the new tools for immunotherapy to treat cancer.