• 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.

• The Journal of Internal Korean Medicine
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• v.28 no.4
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• pp.681-693
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• 2007

Background and Objective : Atractylodes japonica (AJ) is a commonly-used herbal medicine in Asian countries such as Korea, China and Japan. The present study was designated to evaluate the direct effects of AJ on helper T cell activities and on Th1/Th2 lineage development in vitro. Materials and Methods : Spleen cells from 8-week BALB/c mice were cultured in CR extracts containing medium without activation for 24 hours and with activation for 48 hours. CD4+ T cells were isolated and analyzed for mRNA expression levels of INF-$\gamma$, IL-4, T-bet and GATA-3 by RT-PCR and secretion cytokines levels of INF-$\gamma$, IL-2, IL-4, IL-5 and IL-10 by ELISA. Results : The results demonstrated that AJ had no mitogenic effects on unstimulated CD4+ T cells, but augmented CD4+T-cell proliferation upon activation with anti-CD3/anti-CD28 antibodies in a dose-dependent manner. AJ treatment significantly increased CD4+ T cell population and IFN-$\gamma$ expression was significantly enhanced, while IL-4 expression significantly decreased. In addition, in vitro Th1/Th2 polarization experiments revealed that AJ enhanced IFN-$\gamma$ secretion in Th1 cells, but reduced the IL-4 in Th2 cells in dose-dependent manner. Conclusion : These results suggest that AJ treatment could be a desirable alternative therapy for the prevention or correction of Th2 dominant pathological disorders, such as allergy and asthma.

• IMMUNE NETWORK
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• v.3 no.4
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• pp.287-294
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• 2003

Background: In kidney transplantation, donor specific transfusion may induce tolerance as a result of some immune regulatory cells against the graft. In organ transplantation, the immune state arises from a relationship between the immunocompromised graft and the immunocompetent host. However, a reverse immunological situation exists between the graft and the host in hematopoietic stem cell transplantation (HSCT). In addition, early IL-2 injections after an allogeneic murine HSCT have been shown to prevent lethal graft versus host disease (GVHD) due to CD4+ cells. We investigated the induction of the regulatory CD4+CD25+ cells after a transfusion of irradiated recipient cells with IL-2 into a donor. Methods: The splenocytes (SP) were obtained from 6 week-old BALB/c mice ($H-2^d$) and irradiated as a single cell suspension. The donor mice (C3H/He, $H-2^k$) received $5{\times}10^6$ irradiated SP, and 5,000 IU IL-2 injected intraperitoneally on the day prior to HSCT. The CD4+CD25+ cell populations in SP treated C3H/He were analyzed. In order to determine the in vivo effect of CD4+CD25+ cells, the lethally irradiated BALB/c were transplanted with $1{\times}10^7$ donor BM and $5{\times}10^6$ CD4+CD25+ cells. The other recipient mice received either $1{\times}10^7$ donor BM with $5{\times}10^6$ CD4+ CD25- cells or the untreated SP. The survival and GVHD was assessed daily by a clinical scoring system. Results: In the MLR assay, BALB/c SP was used as a stimulator with C3H/He SP, as a responder, with or without treatment. The inhibition of proliferation was $30.0{\pm}13%$ compared to the control. In addition, the MLR with either the CD4+CD25+ or CD4+CD25- cells, which were isolated by MidiMacs, from the C3H/He SP treated with the recipient SP and IL-2 was evaluated. The donor SP treated with the recipient cells and IL-2 contained more CD4+CD25+ cells ($5.4{\pm}1.5%$) than the untreated mice SP ($1.4{\pm}0.3%$)(P<0.01). There was a profound inhibition in the CD4+CD25+ cells ($61.1{\pm}6.1%$), but a marked proliferation in the CD4+CD25- cells ($129.8{\pm}65.2%$). Mice in the CD4+CD25+ group showed low GVHD scores and a slow progression from the post-HSCT day 4 to day 9, but those in the control and CD4+CD25- groups had a high score and rapid progression (P<0.001). The probability of survival was 83.3% in the CD4+CD25+ group until post-HSC day 35 and all mice in the control and CD4+CD25- groups died on post-HSCT day 8 or 9 (P=0.0105). Conclusion: Donor graft engineering with irradiated recipient SP and IL-2 (recipient specific transfusion) can induce abundant regulatory CD4+CD25+ cells to prevent GVHD.

• IMMUNE NETWORK
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• v.5 no.2
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• pp.78-88
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• 2005

Background: Collagen-induced arthritis (CIA) in mice is animal model of autoimmune disease known as rheumatic arthritis in human. We investigated CII-specific CD4+ T cell receptor usage in CIA mice. Methods: In CIA model, draining lymph node (dLN) CD4+ T cells and splenocytes at $3^{rd},\;5^{th},\;8^{th}$ week, we investigated CII-specific T cell proliferation, production of IL-17, IFN-${\gamma}$, TNF-${\alpha}$, IL-4 and IL-10. And we also performed anti-CII IgG Ab measurements in serum level, TCRV ${\beta}$ usage and T cell clonality with RT-PCR-SSCP analysis. Also, we performed proliferative response against CII when CII-specific T cell subset is deleted. Results: CIA mice showed more increase in the serum level of anti-CII IgG than normal mice after induction of arthritis. And the level of anti-CII IgG2a in CIA mice was increased after $3^{rd}$ week after primary immunization, while anti-CII IgG1 was decreased. Draining LN CD4+ T cells have proliferated against CII stimulation at $3^{rd}$ week after $1^{st}$immunization. CD4+T cells derived from dLN of CIA mice produced proinflammatory cytokine IFN-${\gamma}$, IL-17 etc. Draining LN CD4 T cells of CIA presented higher proportion of CD4+V ${\beta}3$+subset compared to those of normal mice at $3^{rd}$ week after $1^{st}$ immunization, and they were increased in proportion by CII stimulation. Draining LN CD4+ T cells without TCRV ${\beta}3+/V{\beta}8.1/8.2+/V{\beta}$10b+cells were not responsive against CII stimulation. But, CII-reactive response of TCRV ${\beta}3-/V{\beta}8.1/8.2-/V{\beta}$10b- T cells was recovered when $V{\beta}3+$ T cells were added in culture. Conclusion: Our results indicate that CD4+$V{\beta}3+$ T cells are selectively expanded in dLN of CIA mice, and their recovery upon CII re-stimulation in vitro, as well as the production Th1-type cytokines, may play pivotal role in CIA pathogenesis.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.4
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• pp.754-761
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• 2008

The purpose of this study was to evaluate the effect of Baekhasuoyijung-Tang(BHSYJT)on mouse T cell cytokines. The proliferation of mouse CD4 T cells under the influence of BHSYJT extract was measured. When mouse CD4 T cell were stimulated with anti-CD3 and anti-CD28 in various concentrations of BHSYJT extract, it increased proliferation of CD4 cells by 28% in $10{\mu}g/m{\ell}$ concentration and by 32% in $100{\mu}g/m{\ell}$ concentration. Treatment of CD4+ T cells stimulated by anti-CD3e and anti-CD28 with BHSYJT resulted in reduction of $IFN-{\gamma}$,but IL-4 levels is not changed. Oral administration of BHSYJT resulted in increase of both CD4+ and CD8+ T cell population in Balb/c mice by 11%. Oral administration of BHSYJT resulted in reduction of serum $IFN-{\gamma}$ level by 27% but, IL-4 level is not changed. CD4+ T cells under Th1/Th2 polarizing conditions for 3 days with BHSYJT resulted in decrease of $IFN-{\gamma}$ level in TH1 cells. Experimental results of this study show that BHSYJT helps to reduce secretion of $IFN-{\gamma}$ by mouse T helper cell in vitro and it had the same effect in vivo. Thus, it can be concluded that use of BHSYJT is an effective treatment for correcting immune imbalance in immune disorders and autoimmune diseases by reducing secretion of cytokine by Th1 cells.

• Journal of Physiology & Pathology in Korean Medicine
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• v.24 no.3
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• pp.446-451
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• 2010

We investigated whether oral administration with capsicum extract (Capsicum annuum var. cheongyang) would affect the immune system by examining the immune cells of Peyer's patch (PP), a gut associated lymphoid tissue, ex vivo. The mice were orally administrated with capsicum extract (100 mg/kg/day), capsaicin (10 mg/Kg), and the vehicle for four consecutive days, and PPs were isolated from intestines 2 days later. When the PP cells were cultured in the presence of Concanavalin A for 72 hr, the levels of cytokines, including IL-2 and IFN-${\gamma]$, were dramatically increased, while the levels of IL-4 remained unchanged compared with the control. Data from the FACS analysis of PP cells indicated that capsicum extract significantly increased the number of CD3+ and CD4+ T cells as well as CD 19+ B cells compared with the control but not CD11b+ cells. Furthermore, the percentages of IL-2+ /CD4+ cells and IFN-${\gamma}+$/CD4+ were greatly increased. These data suggested that oraladministration with capsicum extract might activate the CD4+ T cells leading to cytokine production as well as CD19+ B cells in Peyer's patches. As such, capsicum extract might have potential as an immune modulating agent.

• Proceedings of the Membrane Society of Korea Conference
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• 2004.05a
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• pp.55-58
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• 2004

Cell separation from peripheral blood was investigated using surface-modified polyurethane (PU) membranes with different functional groups. Both red blood cells and platelets could pass through unmodified PU and PU-SO$_3$H membranes, while the red blood cells preferentially passed through PU-N(C$_2$H$_{5}$ )$_2$ and PU-NHC$_2$H$_4$OH membranes. The permeation ratio of T and B cells was less than 25% for the surface-modified and unmodified PU membranes. CD34$^{+}$ cells have been recognized as various kinds of stem cells including hematopoietic and mesenchymal stem cells. The adhesiveness of CD34$^{+}$ cells on the PU membranes was found to be higher than that of red blood cells, platelets, T cells or B cells. Overall, the adhesiveness of blood cells on the PU membranes increased in the following order: red blood cells $\leq$ platelets ＜ T cells $\leq$ B cells ＜ CD34$^{+}$ cells. Treatment of PU-COOH membranes with a human albumin solution to detach adhered blood cells, allowed recovery of mainly CD34$^{+}$ cells in the permeate, while both red blood cells and platelets could be isolated in the permeate using unmodified PU membranes. The PU membranes showed different permeation and recovery ratios of specific cells depending on the functional groups attached to the membranes.mbranes.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.3815-3819
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• 2012

The CD4+CD25+ regulatory T cell (Treg) is a special kind of T cell subset. Studies have showed that Treg cells are involved in a number of physiological processes and pathologic conditions such as autoimmune diseases, transplantation tolerance and cancer. Tregs with unique capacity for immune inhibition can impair anti-tumour immunity and help tumor cells to escape from immune surveillance. The aim of our study was to investigate whether Tregs are involved in hepatocellular carcinoma (HCC). A BABL/C mouse with HCC in situ model was established to evaluate the Treg existence in carcinoma tissues and the changes of Tregs in spleen using flow cytometry and immunohistochemistry methods. Granzyme B expression in carcinoma tissues was analyzed by immunohistochemistry to investigate the tumor local immune status.The proportion of CD4+CD25+/CD4+ spleen lymphocytes of tumor bearing mice ($18.8%{\pm}1.26%$) was found to be significantly higher than that in normal mice ($9.99%{\pm}1.90%$) (P<0.01 ). Immunohistochemistry of spleen tissue also confirmed that there was an increase in Treg in tumor-bearing mice, while in carcinomas it showed Treg cells to be present in tumor infiltrating lymphocyte areas while Granzyme B was rarely observed. Anti-tumour immunity was suppressed, and this might be associated with the increase of Tregs. Our observations suggest that the CD4+CD25+Treg/CD4+ proportion in spleen lymphocytes can be a sensitive index to evaluate the change of Tregs in hepatocellular carcinoma mice and the Treg may be a promising therapeutic target for cancer.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.11
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• pp.5909-5913
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• 2012

Aims: We aimed to analyze the phenotype of tumor-infiltrating lymphocytes (TILs) and non-tumor infiltrating lymphocytes (NILs) in HCC and non-tumor tissues, and evaluate relationships between changes in these cells and the prognosis of HCC. Methods: Lymphocytes were isolated from HCC and corresponding non-tumor tissues and tested by flow cytometry. For comparison, clinical parameters were analyzed. Results: Compared with the non-tumor tissue, tumor tissue had a lower intensity of NK, NKT andCD8+T cell infiltration. TILs had higher intensity of CD4+CD25+Foxp3+regulatory T cell (Treg cells) infiltration compared with that in NILs. The prevalence of Treg cells was associated with fewer CD8 + T lymphocytes in the HCC immune microenvironment. The frequencies of NK cells and CD8+T cells in TILs of HCC patients with metastasis less than 12 months were lower than those without metastasis. However, the frequency of Treg cells was higher than those without metastasis. Conclusion: These results suggest that the frequencies of CD8+T, NK and NKT cells as well as Treg cells in the tumor tissue of HCC are significantly associated with patient survival, and could be applied as predictive indicators for HCC prognosis.

• 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.