• Title/Summary/Keyword: flow cytometry

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MTHFR 3'-untranslated region polymorphisms contribute to recurrent pregnancy loss risk and alterations in peripheral natural killer cell proportions

  • Kim, Eun Sun;Kim, Jung Oh;An, Hui Jeong;Sakong, Jung Hyun;Lee, Hyun Ah;Kim, Ji Hyang;Ahn, Eun Hee;Kim, Young Ran;Lee, Woo Sik;Kim, Nam Keun
    • Clinical and Experimental Reproductive Medicine
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    • v.44 no.3
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    • pp.152-158
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    • 2017
  • Objective: To identify the associations between polymorphisms of the 3'-untranslated region (UTR) of methylenetetrahydrofolate reductase (MTHFR) gene, which codes for an important regulatory enzyme primarily involved in folate metabolism, and idiopathic recurrent pregnancy loss (RPL) in Korean women. Methods: The study population comprised 369 RPL patients and 228 controls. MTHFR 2572C > A, 4869C > G, 5488C > T, and 6685T > C 3'-UTR polymorphisms were genotyped by polymerase chain reaction-restriction fragment length polymorphism analysis or by TaqMan allelic discrimination assays. Natural killer cell proportions were determined by flow cytometry. Results: The MTHFR 2572-5488-6685 (A-C-T) haplotype had an adjusted odds ratio of 0.420 (95% confidence interval, 0.178-0.994; p= 0.048) for RPL. Analysis of variance revealed that MTHFR 4869C > G was associated with altered $CD56^+$ natural killer cell percentages (CC, $17.91%{\pm}8.04%$; CG, $12.67%{\pm}4.64%$; p= 0.024) and folate levels (CC, $12.01{\pm}7.18mg/mL$; CG, $22.15{\pm}26.25mg/mL$; p= 0.006). Conclusion: Variants in the 3'-UTR of MTHFR are potential biomarkers for RPL. However, these results should be validated in additional studies of ethnically diverse groups of patients.

Effect of sun ginseng potentiation on epirubicin and paclitaxel-induced apoptosis in human cervical cancer cells

  • Lin, Yingjia;Jiang, Dan;Li, Yang;Han, Xinye;Yu, Di;Park, Jeong Hill;Jin, Ying-Hua
    • Journal of Ginseng Research
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    • v.39 no.1
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    • pp.22-28
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    • 2015
  • Background: Sun ginseng (SG), a specific formulation of quality-controlled red ginseng, contains approximately equal amounts of three major ginsenosides (RK1, Rg3, and Rg5), which reportedly has antitumor-promoting activities in animal models. Methods: MTT assay was used to assess whether SG can potentiate the anticancer activity of epirubicin or paclitaxel in human cervical adenocarcinoma HeLa cells, human colon cancer SW111C cells, and SW480 cells; apoptosis status was analyzed by annexin V-FITC and PI and analyzed by flow cytometry; and apoptosis pathway was studied by analysis of caspase-3, -8, and -9 activation, mitochondrial accumulation of Bax and Bak, and cytochrome c release. Results: SG remarkably enhances cancer cell death induced by epirubicin or paclitaxel in human cervical adenocarcinoma HeLa cells, human colon cancer SW111C cells, and SW480 cells. Results of the mechanism study highlighted the cooperation between SG and epirubicin or paclitaxel in activating caspase-3 and -9 but not caspase-8. Moreover, SG significantly increased the mitochondrial accumulation of both Bax and Bak triggered by epirubicin or paclitaxel as well as the subsequent release of cytochrome c in the targeted cells. Conclusion: SG significantly potentiated the anticancer activities of epirubicin and paclitaxel in a synergistic manner. These effects were associated with the increased mitochondrial accumulation of both Bax and Bak that led to an enhanced cytochrome c release, caspase-9/-3 activation, and apoptosis. Treating cancer cells by combining epirubicin and paclitaxel with SG may prove to be a novel strategy for enhancing the efficacy of the two drug types.

Influencing Factors of Peripheral Blood Lymphocyte subsets in Workers with Chronic Obstructive Pulmonary Disease Exposed to Inorganic Dust (만성폐쇄성폐질환을 동반한 무기분진 노출 이직근로자의 혈중 림프구 아형분포에 미치는 영향요인)

  • Baek, Jin Ee;Shin, Jae Hoon;Hwang, Joo Hwan;Lee, Youlim;Lee, Jong Seong;Choi, Byung-Soon
    • Journal of Korean Society of Occupational and Environmental Hygiene
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    • v.31 no.3
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    • pp.286-293
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    • 2021
  • Objectives: Inorganic dust is known to be a risk factor for chronic obstructive pulmonary disease (COPD) regardless of smoking and pneumoconiosis. Adaptive and innate immunity, including lymphocyte infiltrate, are involved in the pathogenesis of COPD. The purpose of this study was to analyze the lymphocyte subsets in the blood of workers exposed to inorganic dust and confirm the influencing factors. Methods: The general characteristics of the subjects (n=107) were analyzed through a personal questionnaire. Diagnosis of COPD was established according to pulmonary function tests with FEV1/FVC post bronchodilator lower than 70%, according to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) guidelines. For lymphocyte analysis, blood was stained with a fluorescent CD marker and analyzed by flow cytometry. Results: The increase in CD4+ T lymphocytes was associated with a decrease in age (𝛽=-0.273, p=0.008) and an increase in the cumulative smoking amount (𝛽=0.205, p=0.034). The increase in NK cells was associated with an increase in age (𝛽=0.325, p=0.001) and a decrease in cumulative smoking (𝛽=-0.220, p=0.019). The period of exposure to dust, %FVC predicted and %FEV1/FVC, and the relative population of peripheral blood lymphocytes did not show a statistically significant relationship. Conclusions: CD4+ T lymphocytes and CD56+CD16+ NK cells in peripheral blood were more related to age and cumulative smoking than the duration of dust exposure. Age and smoking are major risk factors for the development of COPD, so it can be predicted that peripheral blood CD4+ T lymphocytes and CD56+CD16+ NK cells are related to the development of COPD in workers exposed to inorganic dust.

Effect of Marine Environment Changes on the Abundance and Community Composition of Cyanobacteria in the South Sea of Korea (남해 해역의 해양환경변화가 시아노박테리아 개체수와 군집 조성에 미치는 영향)

  • Won, JongSeok;Lee, Yeonjung;Lee, Howon;Noh, Jae Hoon
    • Ocean and Polar Research
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    • v.43 no.4
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    • pp.279-293
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    • 2021
  • To investigate the effect of seasonal marine environment conditions on the cyanobacteria abundance and diversity in the South Sea, four-seasonal surveys were conducted along the 127.5°E survey transect line in the central South Sea using flow cytometry and 16S-23S ITS on the Miseq platform from August 2016 to May 2017. The average abundance of Synechococcus varied from 3.3 × 103 to 7.4 × 104 cells ml-1. The abundance was the highest in the summer and the lowest in the winter, and the abundance fluctuated according to water temperature. The abundance was high in the outer sea affected by TWC. However, in summer, the Coastal areas affected by the Yangtze River were more populated than the outer sea. Prochlorococcus was rare and could not penetrate into coastal areas due to the fronts, but showed its dominance in the waters influenced by the TWC. Synechococcus clades II, VII, IX, CRD1, and CRD2 were predominant in the outer sea area affected by the TWC. In the coastal area, clades I and IV showed higher dominance whereas clades V, VI, WPC1, and 5.3-MS3 with euryhaline characteristics, showed a high dominance rate in the water masses affected by the low-salinity water of the Yangtze River in the summer. Clade XVI, XVII, CB1, CB5, and 5.3-I/II showed high dominance in nutrient-rich waters in the summer with increased water temperature. The abundance and community composition of cyanobacteria changed in the South Sea due to the influence of the TWC and stratification. In the summer, the abundance and the community composition differed, and were mainly affected by the general influence of the TWC in addition to the influence of the Yangtze River low-salinity water.

Professional Certification of Medical Technologists in Korea, Japan, and United States of America (한국, 일본, 미국 임상병리사 전문자격)

  • Koo, Bon-Kyeong
    • Korean Journal of Clinical Laboratory Science
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    • v.51 no.1
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    • pp.1-14
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    • 2019
  • The names of medical technologist in Korea, Japan, and the United State of America vary depending on the medical system, educational system, and work areas. Conventionally, jobs equal or similar to medical technologist (commonly known as clinical laboratory technologist, medical laboratory technologist, clinical laboratory scientist, and medical laboratory scientist), histotechnologist, cytotechnologist, and genetic technologist. Jobs, such as extracorporeal technologist, medical electrophysiology technologist, and medical sonographer, allow other allied health professionals including medical technologists, to obtain their qualification. The work areas of medical technologists are classified into clinical pathologic technology, pathologic technology, clinical genetics technology, nuclear medicine in vitro technology, and clinical physiologic technology. New titles of Korean specialized medical technologists refer to certification systems, such as CLMJ, CMAJ, ASCP, IAC, ABRET, and ARMDS. Currently, there are eight types of specialized medical technologists under the cooperation of specialized academic societies. They are hematology, transfusionology, chemistry, immunology, microbiology, histology, cytology, nuclear medicine. It can be further divided into 20 categories according to the work areas such as mass spectrometry, infection control, Pathologists' Assistant, flow cytometry, HLA, cytogenetics, molecular genetics, cardiopulmonary physiology, neurophysiology (EEG/EMG/PSG/etc.), cardiac sonography, neurosonography (TCD/carotid), and medical device quality manager.

Isolation and Identification of Pheophytin, a Photosensitizer from Nostoc commune that Induces Apoptosis in Leukemia and Cancer Cells (Nostoc commune으로부터 백혈병세포와 간암세포에 대한 apoptosis 유도 광과민성물질 pheophytin a의 분리 및 구조동정)

  • Park, Jae-Eun;Lee, Jun-Young;Lee, Min-Woo;Jang, Eun-Jin;Hong, Chang-Oh;Kim, Keun Ki
    • Journal of Life Science
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    • v.28 no.11
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    • pp.1321-1331
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    • 2018
  • The aim of this study was to separate the photosensitizer that induces apoptosis of U937 and SK-HEP-1 cells from Nostoc commune. Dried N. commune was extracted with $CH_2Cl_2/MeOH$ (1:1) to separate the photosensitizer using various chromatographic techniques. The isolated compound was identified as pheophytin a ($C_{55}H_{74}N_4O_5$) with a molecular weight of 870. Its photodynamic activities were assessed under different irradiation conditions (light and non-light) at the same concentration range of $1.15-23.0{\mu}M$. The apoptosis inducing activity in U937 or SK-HEP-1 cells appeared only in the light. The mechanisms underlying the pheophytin a-mediated photodynamic inhibition of cancer cells were further investigated by examining cell morphology changes, cytotoxicity, caspase-3/7 activity, fluorescence staining, flow cytometry analysis, and DNA fragmentation in these two cell lines. The positive control and the light irradiation group showed typical apoptotic responses, including morphological changes, cytotoxicity, caspase activity, nucleus shrinkage owing to chromatin condensation, DNA laddering, and the presence of apoptotic bodies. Cytotoxicity markedly increased in a dose-dependent manner after a 12 hr exposure. Caspase-3/7 activity was higher in U937 cells than in SK-HEP-1 cells. Apoptosis induction therefore appeared to be both concentration- and light-dependent. In conclusion, pheophytin a, isolated from the blue green alga N. commune, had a photodynamic apoptosis-inducing effect on U937 and SK-HEP-1 cells. The findings reported here can be used as basic data for the development of next-generation photosensitizers from N. commune.

Cancer Chemoprevention Effects of Geldanamycin and 17-AAG in Human Oral Squamous Cell Carcinoma (Geldanamycin과 17-AAG가 구강편평세포암종 세포주에 미치는 암예방 효과)

  • Lee, Eun Ju
    • Korean Journal of Clinical Laboratory Science
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    • v.50 no.4
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    • pp.462-469
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    • 2018
  • HSP90 regulates various proteins involved in differentiation and cell survival. Levels of HSP90 tend to increase during development of squamous cell carcinoma in the head and neck including the mouth. Thus, many studies have been conducted to treat these cancers through suppression of HSP90. This study investigated the effect of two HSP90 inhibitors, geldanamycin and 17-AAG, on the proliferation, apoptosis, and invasion of human oral squamous cell carcinoma cells. Cell survival and cell cycle analyses, as well as western blot analysis, were performed with oral cancer cell lines, YD-10B and YD-38. After treatment with HSP90 inhibitors, cell proliferation was significantly inhibited. When YD-10B and YD-38 cells were treated with various concentrations of geldanamycin and 17-AAG (0, 0.1, 0.3, 1 and $10{\mu}M$) for 24 hr, the growth of YD-10B cells was markedly reduced compared to that of YD-38 cells. Thereafter, the cells were subjected to flow cytometry, which revealed G2 arrest. These results demonstrated that geldanamycin induced G2 arrest and inhibited cell proliferation through the $p-GSK-3{\beta}$ pathway in YD-10B and YD-38 cells, thus inhibiting cell survival. HSP90 inhibitors are therefore expected to have a therapeutic effect on various cancer cell lines.

Induction of Apoptosis and Growth-Inhibition by Thymoquinone in ACHN and GP-293 Cell Lines in Comparable with Cis-Platinum

  • Shahraki, Samira;Mohebbati, Reza;Shafei, Mohammad Naser;Mahmoudi, Mahmoud;Hosseinian, Sara;Parhizgar, Soghra;Yazd, Zohreh Naji Ebrahimi;Heravi, Nazanin Entezari;Abadi, Reza Nejad Shahrokh;Khajavirad, Abolfazl
    • Journal of Pharmacopuncture
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    • v.22 no.3
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    • pp.176-183
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    • 2019
  • Objective: In the current work, we investigated the cytotoxic and apoptotic effects of Thymoquinone (TQ), an active compound of Nigella sativa (N. sativa) and Cis-platinum, on normal renal epithelial (GP-293) and human renal adenocarcinoma cell lines (ACHN). Methods: GP-293 and ACHN cell lines were cultured in Dulbecco's modified Eagle's medium (DMEM) with 10% Fetal bovine serum (FBS) and 1% penicillin plus streptomycin antibiotic. The MTT assay was used for cellular viability assessment. Viability of cells was observed using inverted light microscope 24, 48 and 72 h after exposure of the cells to various concentrations of TQ (1, 2.5, 5, 10, 50 and $100{\mu}g/ml$) and Cis-platinum (0.5, 1, 1.5, 2, 3, 6 and $12.5{\mu}g/ml$). Moreover, apoptosis was analyzed with a flow-cytometry method. The untreated cells were considered as control group. Results: Morphological changes such as reduced cell number and increased intercellular distance and reduced cell viability in ACHN and GP-293cell lines were observed in both TQ and Cis- platinum groups; however, Cis-platinum had greater effect on ACHN cell line than GP-293 cell line. In addition, GP-293 cell line was more sensitive to TQ compared to ACHN cell line. Furthermore, TQ and Cis-platinum had apoptotic effects on both ACHN and GP-293 cell lines. Conclusion: Our findings demonstrated that TQ and Cis-platinum had cytotoxic and apoptotic effects on both cell lines, However, GP-293 cell line was more sensitive to TQ. Additionally, Cis-platinum was more effective on ACHN cell line than on GP-293 cell line.

Immunomodulatory Properties of Lactobacillus plantarum NC8 Expressing an Anti-CD11c Single-Chain Fv Fragment

  • Liu, Jing;Yang, Guilian;Gao, Xing;Zhang, Zan;Liu, Yang;Yang, Xin;Shi, Chunwei;Liu, Qiong;Jiang, Yanlong;Wang, Chunfeng
    • Journal of Microbiology and Biotechnology
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    • v.29 no.1
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    • pp.160-170
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    • 2019
  • The lactic acid bacteria species Lactobacillus plantarum (L. plantarum) has been used extensively for vaccine delivery. Considering to the critical role of dendritic cells in stimulating host immune response, in this study, we constructed a novel CD11c-targeting L. plantarum strain with surface-displayed variable fragments of anti-CD11c, single-chain antibody (scFv-CD11c). The newly designed L. plantarum strain, named 409-aCD11c, could adhere and invade more efficiently to bone marrow-derived DCs (BMDCs) in vitro due to the specific interaction between scFv-CD11c and CD11c located on the surface of BMDCs. After incubation with BMDCs, the 409-aCD11c strain harboring a eukaryotic vector pValac-GFP could lead to more efficient expression of GFP compared with wild-type strains shown by flow cytometry analysis, indicating the enhanced translocation of pValac-GFP from L. plantarum to BMDCs. Similar results were also observed in an in vivo study, which showed that oral administration resulted in efficient expression of GFP in both Peyer's patches (PP) and mesenteric lymph nodes (MLNs) within 7 days after the last administration. In addition, the CD11c-targeting strain significantly promoted the differentiation and maturation of DCs, the differentiation of $IL-4^+$ and $IL-17A^+$ T helper (Th) cells in MLNs, as well as production of $B220^+$ $IgA^+$ B cells in the PP. In conclusion, this study developed a novel DC-targeting L. plantarum strain which could increase the ability to deliver eukaryotic expression plasmid to host cells, indicating a promising approach for vaccine study.

Inhibition of miR-128 Abates Aβ-Mediated Cytotoxicity by Targeting PPAR-γ via NF-κB Inactivation in Primary Mouse Cortical Neurons and Neuro2a Cells

  • Geng, Lijiao;Zhang, Tao;Liu, Wei;Chen, Yong
    • Yonsei Medical Journal
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    • v.59 no.9
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    • pp.1096-1106
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    • 2018
  • Purpose: Alzheimer's disease (AD) is the sixth most common cause of death in the United States. MicroRNAs have been identified as vital players in neurodegenerative diseases, including AD. microRNA-128 (miR-128) has been shown to be dysregulated in AD. This study aimed to explore the roles and molecular mechanisms of miR-128 in AD progression. Materials and Methods: Expression patterns of miR-128 and peroxisome proliferator-activated receptor gamma ($PPAR-{\gamma}$) messenger RNA in clinical samples and cells were measured using RT-qPCR assay. $PPAR-{\gamma}$ protein levels were determined by Western blot assay. Cell viability was determined by MTT assay. Cell apoptotic rate was detected by flow cytometry via double-staining of Annexin V-FITC/PI. Caspase 3 and $NF-{\kappa}B$ activity was determined by a Caspase 3 Activity Assay Kit or $NF-{\kappa}B$ p65 Transcription Factor Assay Kit, respectively. Bioinformatics prediction and luciferase reporter assay were used to investigate interactions between miR-128 and $PPAR-{\gamma}$ 3'UTR. Results: MiR-128 expression was upregulated and $PPAR-{\gamma}$ expression was downregulated in plasma from AD patients and $amyloid-{\beta}$ $(A{\beta})-treated$ primary mouse cortical neurons (MCN) and Neuro2a (N2a) cells. Inhibition of miR-128 decreased $A{\beta}-mediated$ cytotoxicity through inactivation of $NF-{\kappa}B$ in MCN and N2a cells. Moreover, $PPAR-{\gamma}$ was a target of miR-128. $PPAR-{\gamma}$ upregulation attenuated $A{\beta}-mediated$ cytotoxicity by inactivating $NF-{\kappa}B$ in MCN and N2a cells. Furthermore, $PPAR-{\gamma}$ downregulation was able to abolish the effect of anti-miR-128 on cytotoxicity and $NF-{\kappa}B$ activity in MCN and N2a cells. Conclusion: MiR-128 inhibitor decreased $A{\beta}-mediated$ cytotoxicity by upregulating $PPAR-{\gamma}$ via inactivation of $NF-{\kappa}B$ in MCN and N2a cells, providing a new potential target in AD treatment.