• Development and Reproduction
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• v.23 no.2
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• pp.79-92
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• 2019

Humanized mice, containing engrafted human cells and tissues, are emerging as an important in vivo platform for studying human diseases. Since the development of Nod scid gamma (NSG) mice bearing mutations in the IL-2 receptor gamma chain, many investigators have used NSG mice engrafted with human hematopoietic stem cells (HSCs) to generate functional human immune systems in vivo, results in high efficacy of human cell engraftment. The development of NSG mice has allowed significant advances to be made in studies on several human diseases, including cancer and graft-versus-host-disease (GVHD), and in regenerative medicine. Based on the human HSC transplantation, organ transplantation including thymus and liver in the renal capsule has been performed. Also, immune reconstruction of cells, of the lymphoid as well as myeloid lineages, has been partly accomplished. However, crosstalk between pluripotent stem cell derived therapeutic cells with human leukocyte antigen (HLA) mis/matched types and immune CD3 T cells have not been fully addressed. To overcome this hurdle, human major histocompatibility complex (MHC) molecules, not mouse MHC molecules, are required to generate functional T cells in a humanized mouse model. Here, we briefly summarize characteristics of the humanized mouse model, focusing on development of CD3 T cells with MHC molecules. We also highlight the necessity of the humanized mouse model for the treatment of various human diseases.

• Toxicological Research
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• v.16 no.4
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• pp.317-323
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• 2000

The immunosuppressive effects of thirty nine chemicals chosen by their potential toxicity were evaluated using a three-step testing method. The immunotoxicity test method developed in this study consisted of three simple assays of lymphoproliferation, mixed leukocyte response, and interleukin (IL)-2 production. The first step was mitogen-induced proliferation assay. Ten chemicals showed the inhibitory effects on the mitogen (lipopolysaccharide or concanavalin A)-induced proliferation in dose-dependent manners. The second step was mixed lymphocyte response. This step crosschecked the growth-suppressive effects detected at the first step. All of 10 chemicals, which showed suppression of lymphoproliferation, also exhibited the suppressive effects on the mixed lymphocyte response in the similar range of chemical concentration. The third step was planned to determine whether or not this growth suppression was mediated through an early activation of T-cell, which could be represented with IL-2 production. Six out of 10 chemicals decreased the interleukin-2 production in the similar concentration range used in the step 1 and 2. These results suggest that those 6 chemicals might have their targets on the signal transduction path-way toward the IL-2 production. In the meantime the other 4 chemicals might have their targets after the IL-2 production signal. Taken all together, the three-step test would be simple, fast, and efficient to deter-mine whether or not the chemical has immunosuppressive effects.

• IMMUNE NETWORK
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• v.13 no.1
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• pp.16-24
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• 2013

CTLA-4Ig is regarded as an inhibitory agent of the T cell proliferation via blocking the costimulatory signal which is essential for full T cell activation. To improve applicability, we developed the CTLA-4Ig-CTKC in which the c-terminal lysine had been replaced by cysteine through single amino acid change. The single amino acid mutation of c-terminus of CTLA-4Ig was performed by PCR and was checked by in vitro transcription and translation. DNA construct of mutant form was transfected to Chinese hamster ovary (CHO) cells by electroporation. The purified proteins were confirmed by Western blot and B7-1 binding assay for their binding ability. The suppressive capacity of CTLA-4Ig-CTKC was evaluated by the mixed lymphocyte reaction (MLR) and in the allogeneic pancreatic islet transplantation model. CTLA-4Ig-CTKC maintained binding ability to B7-1 molecule and effectively inhibits T cell proliferation in MLR. In the murine allogeneic pancreatic islet transplantation, short-term treatment of CTLA-4Ig-CTKC prolonged the graft survival over 100 days. CTLA-4Ig-CTKC effectively inhibits immune response both in MLR and in allogeneic islet transplantation model, indicating that single amino acid mutation does not affect the inhibitory function of CTLA-4Ig. CTLA-4Ig-CTKC can be used in vehicle-mediated drug delivery system such as liposome conjugation.

• Tuberculosis and Respiratory Diseases
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• v.43 no.1
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• pp.22-29
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• 1996

Background: Abnormalities of the peripheral blood are frequent and varied in patients with miliary tuberculosis. Anemia, leukopenia, thrombocytopenia, pancytopenia, monocytosis, basophilia, eosinophilia and leukemoid reactions have been reported. These abnormalities are more frequent in patients with positive bone marrow study. In this report, we evaluated clinical, hematological and immunological features in patients with miliary tuberculosis in order to know whether difference is existed between "bone marrow biopsy positive group(pathologically proven to miliary tuberculosis)" and "negative group". Method: Clinical evaluation, serum ADA, sIL-2R, and T-lymphocyte subsets were measured in 40 patients with miliary tuberculosis who received bone marrow biopsy. Results: 1) The average age of patients was 39 year-old. There were 23 male and 17 female patients. Associated extrapulmonary tuberculosis are 9 CNS tuberculosis, 6 joint tuberculosis, and 2 tuberculous pleurisy. 2) Sixteen of the 40 patients were positive bone marrow biopsy(60%). 3) Sixteen of the 40 patients(60%) had anemia(11 positive patients: 13 negative patients). Leukopenia occurred in 12 per cent(4:1). Thrombocytopenia was noted in 10%(3:1). 4) The mean value of serum ADA was 83 U/L(90 U/L: 70.6 U/L, p=0.23). 5) The mean activity of Soluble IL-2 receptor was 4,643 pmol/L($6840{\pm}7446\;pmol/L$: $1,897{\pm}1,663\;pmol/L$, p=0.06). 6) In the T lymphocyte subsets, the percent of T-lymphocytes was 64%(62%:73%, p=0.2). In some patients(9), $T_4$ and $T_8$ ratio in BAL fluid($1.97{\pm}1.2$) was higher than that in the peripheral blood($1.16{\pm}0.5$). Conclusion: Bone marrow examination are diagnostic in 60% of cases of miliary tuberculosis. Percents of the total T lymphocyte and helper T cell in BAL are more elevated than in peripheral blood. There was no significant difference in peripheral blood abnormalities and marker of T lymphocyte activation between the bone marrow biopsy positive and negative group.

• Journal of Physiology & Pathology in Korean Medicine
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• v.36 no.6
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• pp.235-241
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• 2022

The purpose of this study was to investigate the immune activation effect of Sasamsaengmaek-san (SSSMS) consisted of a mixture of Adenophora triphylla var. japonica, Liriope platyphylla, Panax ginseng C. A. Meyer and Schisandra chinensis. in Balb/c mice. Measuring alanine aminotransferase (ALT) and aspartic acid transaminase (AST) levels in Balb/c mice was performed to analyze the cytotoxicity. Cytokines (IFN-γ, IL-2, IL-12) which regulate the immune activation in Balb/c mice were measured by enzyme-linked immunosorbent assay (ELISA). Activated T lymphocytes in peripheral blood mononuclear cell (PBMC), spleen, lymph nodes were analyzed by flow cytometry using percentages. All tests were compared with red ginseng 100 ㎍/mL (RG 100), which is the most used for immune activity. As a result, cytokine activity was significantly increased at SSSMS 300 group. Activated T lymphocytes in PBMC, spleen, lymph nodes were significantly increased at SSSMS 300 group. These results suggest that there is a possibility of SSSMS activating an immune system by activating the cytokines, and it is confirmed that SSSMS also effective for generation and differentiation of T, B lymphocytes which activate the immune response.

• Journal of Life Science
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• v.16 no.6
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• pp.904-910
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• 2006

Dendritic cells (DCs) are the only antigen presenting cells (APCs) capable of initiating immune responses, which is crucial for priming the specific cytotoxic T lymphocyte (CTL) response and tumor immunity. Upon activation by DCs, CD4+ helper T cells can cross-prime CD8+ CTLs via IL-12. However, recently activated DCs were described to prime in vitro strong T helper cell type 1 $(Th_1)$ responses, whereas at later time points, they preferentially prime $Th_2$ cells. Therfore, we examined in this study the optimum kinetic state of DCs activation impacted on in vivo priming of tumor-specific CTLs by using ovalbumin (OVA) tumor antigen model. Bone-marrow-derived DCs showed an appropriate expression of surface MHC and costimulatory molecules after 6 or 7-day differentiation. The 6-day differentiated DCs pulsed with OVA antigen for 8 h (8-h DC) and followed by restimulation with LPS for 24 h maintained high interleukin (IL)-12 production potential, accompanying the decreased level in their secretion by delayed re-exposure time to LPS. Furthermore, immunization with 8-h DC induced higher intracellular $interferon(IFN)-{\gamma}+/CD8+T$ cells and elicited more powerful cytotoxicity of splenocytes to EG7 cells, a clone of EL4 cells transfected with an OVA cDNA, than immunization with 24-h DC. In the animal study for the evaluation of therapeutic or protective antitumor immunity, immunization with 8-h DC induced an effective antitumor immunity against tumor of EG7 cells and completely protected mice from tumor formation and prolonged survival, respectively. The most commonly used and clinically applied DC-based vaccine is based on in vitro antigen loading for 24 h. However, our data indicated that antigen stimulation over 8 h decreased antitumor immunity with functional exhaustion of DCs, and that the 8-h DC would be an optimum activation state impacted on in vivo priming of tumor-specific CTLs and subsequently lead to induction of strong antitumor immunity.

• Journal of Periodontal and Implant Science
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• v.24 no.3
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• pp.472-482
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• 1994

Cytokines expressed specifically in leukocytes subsets and in activated cells, which are involved in chemotaxis and activation of leukocytes, are recently defined as chemokines. Macrophage inflammatory $protein-1{\alpha}(MIP-1{\alpha})$ and $MIP-1{\beta}$ are members of C-C chemokine subfamily which produces wide immunomodulatory, proinflammatory, and hematopoietic modulatory actions. We have studied their gene expression by using Northern blot analysis in various blood cells such as cytolytic T lymphocyte(CTL), helper T lymphocyte(HTL), macrophage, and B lymphocyte. Resting CTL line CTLL-R8 expressed $MIP-1{\alpha}$ mRNA which was downregulated by ConA stimulation. Both of resting and ConA stimulated HTL line Hut78 and Jurkat did not express $MIP-1{\alpha}$ mRNA. There was detectable $MIP-1{\alpha}$ transcript in HTL hybridoma 2B4.11 which was a little upstimulated by ConA stimulation. B cell line 230, and macrophage cell line RAW264.7 and WR19M.1 showed distinct $MIP-1{\alpha}$ message which were induced after LPS stimulation. Expression pattern of $MIP-1{\beta}$ in all cell lines or cell were almost identical to that of $MIP-1{\alpha}$. Also strategies employed to identify and characterize the biological functions was preceded by receptor cloning to trace the shorcut to the final goal of cytokine research. For the cloning of $MIP-1{\alpha}$ receptor(R), we used synthetic oligonucleotides of transmembrane(T) conserved sequences of already cloned human(h) IL-8-R, and performed reverse transcription-polymerase chain reaction(RT-PCR) amplification using murine(m) macrophage cell line mRNA. Among 5RT-PCR products, we isolated a homologous cDNA with hIL-8-R which were shown to be putative mIL-8-R cDNA.

• IMMUNE NETWORK
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• v.20 no.1
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• pp.5.1-5.15
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• 2020

The γδ T cells are unconventional lymphocytes that function in both innate and adaptive immune responses against various intracellular and infectious stresses. The γδ T cells can be exploited as cancer-killing effector cells since γδ TCRs recognize MHC-like molecules and growth factor receptors that are upregulated in cancer cells, and γδ T cells can differentiate into cytotoxic effector cells. However, γδ T cells may also promote tumor progression by secreting IL-17 or other cytokines. Therefore, it is essential to understand how the differentiation and homeostasis of γδ T cells are regulated and whether distinct γδ T cell subsets have different functions. Human γδ T cells are classified into Vδ2 and non-Vδ2 γδ T cells. The majority of Vδ2 γδ T cells are Vγ9δ2 T cells that recognize pyrophosphorylated isoprenoids generated by the dysregulated mevalonate pathway. In contrast, Vδ1 T cells expand from initially diverse TCR repertoire in patients with infectious diseases and cancers. The ligands of Vδ1 T cells are diverse and include the growth factor receptors such as endothelial protein C receptor. Both Vδ1 and Vδ2 γδ T cells are implicated to have immunotherapeutic potentials for cancers, but the detailed elucidation of the distinct characteristics of 2 populations will be required to enhance the immunotherapeutic potential of γδ T cells. Here, we summarize recent progress regarding cancer immunology of human γδ T cells, including their development, heterogeneity, and plasticity, the putative mechanisms underlying ligand recognition and activation, and their dual effects on tumor progression in the tumor microenvironment.

• Biomolecules & Therapeutics
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• v.30 no.6
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• pp.529-539
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• 2022

Rheumatoid arthritis (RA) is a multifactorial immune-mediated disease, the pathogenesis of which involves different cell types. T-cell activation plays an important role in RA. Therefore, inhibiting T-cell activation is one of the current therapeutic strategies. Cytotoxic T-lymphocyte antigen 4-immunoglobulin (CTLA4-Ig), also known as abatacept, reduces cytokine secretion by inhibiting T-cell activation. To achieve a homeostatic therapeutic effect, CTLA4-Ig has to be administered repeatedly over several weeks, which limits its applicability in RA treatment. To overcome this limitation, we increased the number of sialic acid-capped antennas by genetically engineering the CTLA4 region to increase the therapeutic effect of CTLA4-Ig. N-acetylglucosaminyltransferase (GnT) and α2,6-sialyltransferase (α2,6-ST) were co-overexpressed in Chinese hamster ovary (CHO) cells to generate a highly sialylated CTLA4-Ig fusion protein, named ST6. The therapeutic and immunogenic effects of ST6 and CTLA4-Ig were compared. ST6 dose-dependently decreased paw edema in a mouse model of collagen-induced arthritis and reduced cytokine levels in a co-culture cell assay in a similar manner to CTLA4-Ig. ST6- and CTLA4-Ig-induced T cell-derived cytokines were examined in CD4 T cells isolated from peripheral blood mononuclear cells after cell killing through irradiation followed by flow- and magnetic-bead-assisted separation. Interestingly, compared to CTLA4-Ig, ST6 was substantially less immunogenic and more stable and durable. Our data suggest that ST6 can serve as a novel, less immunogenic therapeutic strategy for patients with RA.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.342-347
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• 2011

T cells play a key role in viral infection. However, in patients with chronic hepatitis C virus (HCV) infection, HCV-specific T cells are dysfunctional and impaired in the liver, which is the primary site for HCV replication. There are multiple potential mechanisms for HCV-specific T cell dysfunction including induction of immune inhibitory pathways (program death-1; PD-1, cytotoxic t lymphocyte associated antigen-4; CTLA-4) and immune tolerance induced specific for the liver. However, the interaction between hepatocytes and HCV-specific CD8 T cells has not clearly established. In this study, we confirmed huh (human hepatoma) 7.5 cells expressing HLA (human leukocyte antigen) A2 presented antigen to activate HCV-specific CD8 T cells in HLA A2-restricted manner and expression of PD-L (program death ligand) 1 on huh7.5 cells reduced HCV-specific CD8 T cell activation, suggesting an immune modulatory activity. Loss of HCV-specific tetramer responses following antigenic stimulation correlated with increased caspase-3 activity. In addition, PD-L1 on huh7.5 cells rescued HCV-specific CD8 T cells from apoptosis. Our results suggest that the interaction between PD-L1 and PD-1 can recover the function of HCV-specific CD8 T cells in the liver, which could be applied in therapy of HCV chronic infection.