• Title/Summary/Keyword: FACs Analysis

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Effects of Green Tea Extract on the p53 Pathway in the MCF-7 Breast Cancer Cell Line (유방암 세포 주 MCF-7에서의 녹차 추출물이 p53 경로에 미치는 영향)

  • Kwak, Inseok
    • Journal of Life Science
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    • v.28 no.11
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    • pp.1316-1320
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    • 2018
  • The effects of a green tea extract (GTE) were examined using the MCF-7 human breast cancer cell line. Cell viability assays using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays revealed that GTE had a significant cytotoxic effect on MCF-7 cells, depending on the concentration of GTE. Western blotting of p53 and its related proteins, p21/cip1 and CDK2, after GTE treatment revealed that a significant and concentration dependent increase in p53 protein in response to GTE. The levels of p21/cip1 proteins were also increased at low GTE concentrations were significantly increased even at the highest GTE concentrations. However, the level of CDK2 was significantly decreased by treatment with high concentrations of GTE. These results indicate that treatment with GTE increased the p53 level in MCF-7 cells, and this activation of p53 markedly elevated the levels of p21/cip1proteins, which, in turn, inhibited CDK2 expression in the MCF-7 cells. The inhibition of CDK2 expression might then affect cell cycle progression. Subsequent FACS analysis indicated that GTE treatment the gradually increased progression of the MCF-7 to the G1 phase. These results clearly demonstrate that the anti-tumor effect of GTE in MCF-7 cells is regulated by p53 arrest of the MCF-7 cells at the G1 stage of cell cycle.

Evaluation on Immunopotentiation Activities of Combined Extract of Silkworm and Cinnamomum cassia in vivo (누에와 육계 복합 추출물의 in vivo 면역증진 기능성 연구)

  • Kim, Kyeong Jo;Park, Hae Jin;Kim, Il Gyu;Kim, Min Ju;Shin, Mi-Rae;Roh, Seong-Soo
    • The Korea Journal of Herbology
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    • v.33 no.4
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    • pp.19-26
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    • 2018
  • Objectives : The aim of this study was to investigate the immunopotentiating activity of combine extract that Silkworm and Cinnamomum cassia. Recently, acute epidemic diseases such as cold and viral respiratory diseases have been emerging. So, interested in immunity enhancement has been increasing, and research on natural products to promote immunity activity has been actively conducted. Methods : To confirm the immunopotentiating activity effect, Silkworm (SW), Cinnamomum cassia (CC), and SWCC combined extracts were treated 14 days at 300 mg/kg/day. The changes of glutamic oxalacetic transaminase (GOT), glutamic pyruvate transaminase (GPT) in serum were analyzed after experiment. The changes in the total spleen cell number were measured. Immune cells in spleen were analyzed using fluorescence activated cell sorter (FACS). also, analyzed the expression of cytokines in spleen. Results : Total number of cells in the spleen and FACS analysis of T lymphocytes activated in the spleen showed that the SWCC combined treated group had much higher frequency of active cells than both single groups. The ratio of CD4+CD8+, CD4+CD69+ and CD4+CD25+ T cells in spleen, SWCC is higher than other groups except Nor in CD4+, CD4+CD69+, CD4+CD25+ T cells. The results of this study suggest that SWCC can help immune function via IL-2, IL-10, IL-12, IFN-${\gamma}$ cytokine production, increased T lymphocytes and splenocyte proliferation. Conclusion : Therefore, these results suggested that the SWCC combined extracts administration increase stronger immunity enhancement than when SW and CC adminstration.

Cytotoxic effects of ethanol extracts of Saussurea lappa mediated by mitochondrial apoptotic pathway

  • Koh Seung-Hee;Ko Seong-Gyu;Jun Chan-Yong;Park Chong-Hyeong
    • The Journal of Korean Medicine
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    • v.25 no.4
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    • pp.79-89
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    • 2004
  • Saussurea lappa and Taraxacum mongolicum have been used for herbal medicinal treatments against cancers in East Asia. We performed this study to understand the molecular basis underlying the anti-tumor effects of two herbs and analyzed the effects of these medicinal herbs on proliferation and on expression of cell growth/apoptosis related molecules by using an AGS gastric cancer cell line. The treatments of Saussurea lappa dramatically reduced cell viabilities in a dose and time-dependent manner, but Taraxacum mongolicum did not. FACS analysis and Annexin V staining assay also showed that Saussurea lappa induces apoptotic cell death of AGS. Expression analyses via RT-PCR and Western blots revealed that Saussurea lappa increased expression of the p53 and its downstream effector p21/sup Waf1/, and that the both increased expression of apoptosis related Bax and cleavage of active caspase-3 protein. We also confirmed the translocation of Bax to mitochondria Collectively, our data demonstrate that Saussurea lappa induces growth inhibition and apoptosis of human gastric cancer cells, and these effects are correlated with down- and up-regulation of growth-regulating apoptotic and tumor suppressor genes, respectively.

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Experimental Studies on Antimetastatic and Immunomodulating Effects of Patriniae Radix Herbal-acupuncture (패장약침(敗醬藥鍼)의 암전이 억제 및 면역 조절 효과에 관한 실험적 연구)

  • Park, Hee-Soo;Park, Jai-Young
    • Journal of Acupuncture Research
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    • v.23 no.4
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    • pp.187-203
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    • 2006
  • Objectives : This study was designed to investigate the antimetastatic and immunomodulating effects of Patriniae Radix. Methods : Acute and subacute cytotoxicity experiment of Patriniae Radix was performed. Antimetastatic experiment was administered in vitro and in vivo. To observe the immunomodulating effects of Patriniae Radix, FACS analysis and ELISA assay were performed. Results : There was no acute and subacute toxicity responses in mouse treated with Patriniae Radix. Antimetastatic experiment in vitro and in vivo showed that Patriniae Radix has antimetastatic effects. This research revealed that Patriniae Radix mediate cellular immunity response. As compared with control, the population of total T cell, helper T cell, cytotoxic T cell and macrophage were increased. The production of Th 1 type cytokines from splenocyte and cytokines which is associated with anti-tumor activity form macrophage were increased significantly. Conclusion Patriniae Radix Herbal-acupuncture appears to have considerable activity on the treatment of liver metastasis from colon26-L5 carcinoma cell line, and deserves further evaluation in this setting.

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Chondrogenesis of Periosteum-derived Progenitor Cells on Hyaluronic Acid Fiber Scaffold (Hyalograft $3D^{\circledR}$)

  • Shin, Hyun-Chong;Choi, Yong-Soo;Lim, Sang-Min;Lee, Chang-Woo;Kim, Dong-Il
    • 한국생물공학회:학술대회논문집
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    • 2005.04a
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    • pp.239-243
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    • 2005
  • Periosteum-derived progenitor cells (PDPCs) were isolated and characterized by flow cytometric analysis using fluorescence-activated cell sorter (FACS). The chondrogenesis of PDPCs was performed on hyaluronic acid fibers ($Hyalrograft^{\circledR}$) 3D) in chondrogenic induction medium. PDPCs showed the chondrogenic potential when cultured on hyaluronic acid fibers. These results showed that the characterized PDPCs were the chondrogenic progenitor cells and $Hyalrograft^{\circledR}$ 3D served as a useful carrier for PDPCs in transplantation proliferation, matrix synthesis and differentiation. Therefore, it could be used as a matrix for healing the defected cartilage.

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Contribution of TLR2 to the Initiation of Ganglioside-triggered Inflammatory Signaling

  • Yoon, Hee Jung;Jeon, Sae Bom;Suk, Kyoungho;Choi, Dong-Kug;Hong, Young-Joon;Park, Eun Jung
    • Molecules and Cells
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    • v.25 no.1
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    • pp.99-104
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    • 2008
  • Gangliosides, sialic acid-containing glycosphingolipids, are implicated in many neuronal diseases, but the precise molecular mechanisms underlying their pathological activities are poorly understood. Here we report that TLR2 participates in the initiation of ganglioside-triggered inflammatory signaling responses. Using FACS analysis and immunofluorescence microscopy, we found that gangliosides rapidly enhanced the cell surface expression of TLR2 in microglia, while reducing that of TLR4. The ganglioside-dependent increase of TLR2 expression was also observed at the messenger and protein levels. We also showed that gangliosides stimulate the interaction of TLR2 with Myd88, an adaptor for TLRs, and obtained evidence that lipid raft formation is closely associated with the ganglioside-induced activation of TLR2 and subsequent inflammatory signaling. These results collectively suggest that TLR2 contributes to the ability of gangliosides to cause inflammatory conditions in the brain.

Study on Antitumor and Immunomodulatory Effects of Cambodian Phellinus linteus (캄보디안 상황버섯의 항암 및 면역조절작용에 대한 연구)

  • Lee Hyo Jeong;Park Jung Min;Song Gyu Yong;Kang Kyung Sun;Kim Sung Hoon
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.16 no.2
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    • pp.332-337
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    • 2002
  • Phellinus Iinteus from Cambodia was confirmed to have a homologous DNA sequencec to Phellinus Iinteus. Antitumor and immunomodulatory activities were evaluated with aquous extract of Cambodian Phellinus Iinteus(CPL). CPL didn't show any significant cytotoxicity on HT1080, Sarcoma 180 and B16BL6, whereas it inhibited the relaxation of DNA topoisomerase I from the concentration of 250ug/ml. In the pulmonary colonization assay it inhibited pulmonary metastasis by B16BL6 in C57BL6 mice to 36%, 36.9% and 55.5% at various doses of 2 mg, 20 mg and 50 mg. From FACS analysis with splenocytes pretreated with CPL, it significantly increased lymphoblast and induced production of IL-2. These results indicate Cambodian Phellinus Iinteus has antitumor and immunomodulatory activities still suggesting more study on its mechanism and effective compound in detail.

Roles of Neutral Sphingomyelinase 1 on CD95-Mediated Apoptosis in Human Jurkat T Lymphocytes

  • Lee, Hyun-Min;Surh, Bo-Young;Chun, Young-Jin
    • Biomolecules & Therapeutics
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    • v.18 no.3
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    • pp.262-270
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    • 2010
  • CD95 receptor is a member of tumor necrosis factor receptor family that mediates apoptosis in many cell types when bound by CD95 ligand or cross-linked by agonistic anti-CD95 antibodies. To determine the role of neutral sphingomyelinase (nSMase) on CD95-mediatd apoptosis, human Jurkat T lymphocytes were exposed to recombinant human CD95 ligand. Treatment with CD95 ligand induced cell death in a concentration and time-dependent manner. CD95-induced cell death was suppressed by inhibitors of SMase such as AY9944 or desipramine. Transfection with human nSMase1 siRNA plasmid into CD95 ligand-treated cells significantly prevented CD95-mediated cell death. CD95-mediated elevation of intracellular ceramide level detected by FACS analysis with anti-ceramide antibody was also decreased by nSMase1 siRNA. Knock-down of nSMase1 expression also blocked cytochrome c release into cytosol and caspase-3 cleavage in CD95-treated cells. Taken together, these results suggest that nSMase1 may play an important role in CD95-mediated apoptotic cell death in Jurkat T cells.

Adenovirus-Mediated Antisense Expression of Telomerase Template RNA Induces Apoptosis in Lung Cancer Cells

  • Song, Joon-Seok;Kim, Sang-Bae;Lee, Young-Ho;Lee, Kyu-Wan;Jung, Hak-Huyn;Kim, Mee-Hye;Kim, Kyung-Tai;Brown, Robert;Kim, Young-Tae
    • Journal of Microbiology and Biotechnology
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    • v.12 no.1
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    • pp.89-95
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    • 2002
  • Telomerase is a ribonucleoprotein complex, whose function is to add telomeric repeats $(TTAGGG)_n$ to chromosomal ends and is also known to play an important role in cellular immortalization. Telomerase is highly active in most tumor cells, yet not in normal cells. Therefore, it may have possible applications in cancer gene therapy. Telomerase consists of two essential components; a telomerase RNA template (hTR) and a catalytic subunit (hTERT). The current study attempted to inhibit the "open" part of the human telomerase RNA (hTR) with an antisense sequence-expressing adenovirus. It was found that the antisense telomerase adenovirus suppressed the telomerase activity, tumor cell growth, and survival in vitro. Furthermore, FACS analysis and TUNEL assay suggested that the reduce viability was mediated through the induction of apoptosis, indicating that this approach might be a useful method for suppressing cancer growth in targeted cancer gene therapy.

Immunomodulatory Effects of Eckol, a Pure Compound of Ecklonia Cava, on Dendritic Cells

  • Kim, Mi-Hyoung;Joo, Hong-Gu
    • IMMUNE NETWORK
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    • v.6 no.4
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    • pp.199-203
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    • 2006
  • Background: Eckol purified from Ecklonia cava, a brown alga has been known to have cytoprotective effects on some cell lines against oxidants and ionizing radiation. However, there is no study about the effects of eckol on immune cells. Methods: Bone marrow (BM)-derived dendritic cells (DCs) were used to demonstrate the immunomodulatory effects of eckol on DCs, such as viability, the expression of surface markers, allogeneic stimulating capacity using MTI, flow cytometric, $^3H$-thymidine incorporation assay. Results: Eckol did protect DCs against cytokine withdrawal-induced apoptosis in a concentration dependent manner based on MTT assay. And also, it increased the expression of MHC class II and CD86 (B7.2) molecules, maturation markers, on the surface of viable DCs gated in FACS analysis. Furthermore, eckol-treated DCs stimulated the proliferation of allogeneic $CD4^+$ T lymphocytes compared to imDCs in $^3H$-thymidine incorporation assay. $CD4^+$ T lymphocytes activated with eckol-treated DCs produced the larger amount of IFN-${\gamma}$ and IL-4 than those cells with imDCs. Conclusion: Taken together, we demonstrate in this study that eckol, a pure compound of Ecklonia cava, may modulate the immune responses through the phenotypic and functional changes of DCs.