• Title/Summary/Keyword: flow cytometric analysis

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Use of Flp-Mediated Cassette Exchange in the Development of a CHO Cell Line Stably Producing Erythropoietin

  • Kim, Min-Soo;Lee, Gyun-Min
    • Journal of Microbiology and Biotechnology
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    • v.18 no.7
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    • pp.1342-1351
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    • 2008
  • The feasibility of the use of Flp-mediated cassette exchange in the development of a CHO cell line, which produces erythropoietin (EPO) stably and largely, was investigated. A stable, high enhanced green fluorescence protein (EGFP)-producing clone was screened by extensive flow cytometric analysis. An EPO expression unit was targeted into the premarked locus of the stable parental clone by Flp-mediated cassette exchange and a correctly targeted clone (FC28T7) was obtained. The EPO production of FC28T7 was proven to be stable in long-term culture. Furthermore, the Flp-mediated cassette exchange did not alter the stable parental clone's characteristics concerning transgene expression level and stability. Taken together, the data obtained here indicated that the establishment of CHO cell lines stably producing a desired protein is achievable using Flp-mediated cassette exchange.

Intracellular $Ca^{2+}$ release mediates apoptosis induced by ascorbic acid (vitamin C) in HepG2 human hepatoma cells

  • Kang, Young-Shin;Lee, Yong-Soo
    • Proceedings of the PSK Conference
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    • 2003.10b
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    • pp.88.2-88.2
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    • 2003
  • Ascorbic acid (vitamin C) has been shown to have anti-cancer actions. However, the exact mechanism of this action is not fully understood. In this study we investigated the possible mechanism of anti-cancer action of ascorbic acid in HepG2 human hepatoblastoma cells. Ascorbic acid induced apoptotic cell death in a dose-dependent manner in the HepG2 cells, assessed by the flow cytometric analysis of hypodiploid nuclei stained with propodium iodide. In addition, ascorbic acid increased intracellular Ca$\^$2+/ concentration, whereas the level of reactive oxygen species was not significantly changed, suggesting that ascorbic acid may not alter cellular redox potential in the cells. (omitted)

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Cytogenetic Analysis of the Triploid Pacific Abalone, Haliotis discus hannai (북방전복, Haliotis discus hannai 3배체의 세포유전학적 연구)

  • Jee, Young-Ju;Chang, Young-Jin
    • The Korean Journal of Malacology
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    • v.28 no.1
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    • pp.37-43
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    • 2012
  • In this study, we invesgated a cytogenetic analysis of the Pacific triploid abalone, Haliotis discus hannai induced by low temperature treatment. We got a lot of mitotic metaphase chromosome spreads from the triploid and diploid Pacific abalones' hatched larvae (trochophores). The chromosome number of diploid abalone was 2n = 36 and that of triploid abalone was 3n = 54, so the chromosome number of triploid abalone was 1.5 times higher than that of diploid abalone. We developed a modified flow cytometric method for Pacific abalone from the existing methods. We uesd 51 months aged triploid and diploid Pacific abalones' hemolymph to get the DNA contents by flow cytometry. The DNA content of diploid abalone was 1.743 pg/cell and the DNA content of triploid abalone was 1.49 times higher than that of diploid one. It proved that triploid abalone was consisted with two sets of maternal diploid and one set of paternal genome.

Effect of ${\alpha}-spinasterol$ Extracted from Phytolacca americanna on the Apoptosis of U937 cell line (상륙에서 추출한 ${\alpha}-spinasterol$의 백혈병세포주(U937) 자멸사 유도 효능)

  • Yang, Jun-Seok;Jeong, Sang-Hun;Kim, Ho;Han, Ung;Jin, Jae-Ho;Jung, Il-Kook;Kim, Dae-Keun;Jeong, Seung-Il;Jeong, Han-Sol;Lee, Kwang-Gyu
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.21 no.5
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    • pp.1108-1117
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    • 2007
  • To investigate the possible mechanism of ${\alpha}-spinasterol$ as a candidate of anti-cancer drug, I examined the effects of ${\alpha}-spinasterol$ on the apoptosis of U937 cells MTT assay, flow cytometric analysis, SDS-polyacrylamide gel electrophoresis, Western blot analysis, and RT-PCR were performed. ${\alpha}-spinasterol$ treatment reduced the cell viablilty of U937 cells in a dose-dependent manner, which was associated with the induction of apoptotic cell death. ${\alpha}-spinasterol$ treatment also reduced the levels of Bcl-xL anti-apoptotic protein expression and increased the levels of caspase-3, p53, pro-apoptotic protein, in U937 cells. After treatment the level of Bcl-xL, anti-apoptotic gene expression was decreased and the level of ICE pro-apoptotic gene expression was increased. These findings suggest that ${\alpha}-spinasterol$ induced the apoptotic cell death via regulation of several growth regulatory gene products. The abbreviations used are: FBS, fetal bovine serum; PBS, phosphate buffered saline; PI, propidium iodide; OD, optical density; DiOC6, 3,3-dihexyloxa carbcyanine iodide; MTT, 3 [4-5-dimethylthiazol-2-yl] -2-diphenyltetrazolium bromide.

The Effects of Artemisia capillaries Herbal Acupuncture on Ethanol-induced Apoptosis in Neuroblastoma Cell Line (인진 약침액이 신경아세포주에서 에탄올에 의해 유발된 아폽토시스에 미치는 영향)

  • Ee-Hwa, Kim;Youn-Hee, Kim;Youn-Jung, Kim;Mi-Hyun, Jang;Joo-Ho, Chung;Chang-Ju, Kim
    • The Journal of Korean Medicine
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    • v.22 no.1
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    • pp.90-95
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    • 2001
  • 목적:인진 약침액이 SK-N-MC 신경아세포주에서 에탄올에 의해 유발된 아폽토시스에 미치는 영향을 조사하였다. 방법 : SK-N-MC cell line에서의 아폽토시스 변화를 관찰하기 위해서 MTT assay, DAPI staining 및 flow cytometric analysis 방법을 이용하였다. 결과: MTT assay를 이용하여 분석한 결과 농도에 따른 세포 독성의 효과가 에탄올 투여로부터 관찰되었다. 또한 인진 약침액으로 전처치하고 에탄올을 처치하였을 때 세포 독성이 크게 감소되었다. DAPI staining에서 인진 약침액 투여군은 에탄올 투여군에 비해서 fragmentation이 억제되었다. Flow cytometry를 통하여 인진 약침액 투여군은 에탄올 투여군에 비하여 세포주기 중 sub $G_1$ 분획의 증가가 억제되었다. 결론 : SK-N-MC 신경아세포주에서 에탄올에 의해서 유발된 아폽토시스는 전형적인 세포사별 형태를 나타내었다. 또한 인진 약침액은 에탄올에 의해서 유발된 아폽토시스에서 세포보호 효과가 있음이 확인되었다.

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Immunomodulatory Effect of Bio-Q, Bokiikhyul Prescription (보기익혈 복합방, 바이오 큐의 면역조절 효과)

  • Lee, Hyo-Jung;Khil, Jae-Ho;Baek, Nam-In;Ra, Jeong-Chan;Kim, Sung-Hoon
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.19 no.4
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    • pp.908-912
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    • 2005
  • Bio-Q is a modified prescription with the activities of supplementing Qi and blood in human body. In the present study, immunomodulatory effect of Bio-Q was examined. After oral administration of Bio-Q for 7 days to Balb/c mice, splenocytes were isolated and immunological experiments were performed. Bio-Q significantly increased the proliferation of splenocytes exposed to concanavalin A (Con A), while it did not in case of lipopolysaccharide (LPS) stimulation. Bio-Q also significantly increased CD3/CD19, CD4/CDB and NK cells by flow cytometric analysis. In addition, Bio-Q significantly enhanced the level of $INF-\gamma$ in splenocytes, but not $TNF-\alpha$ by ELISA. These results strongly suggest that Bio-Q has immunomodulatory activity through the regulation of T cell mediated immune pathway.

Immunoactivities of the Protein-Polysaccharides of the Tips of the Growing Carpophores of Ganoderma lucidum (영지버섯(Ganoderma lucidum) 생장점으로부터 분리한 단백다당류의 면역활성)

  • Chung, Kyeong-Soo;Kim, Sang-Bum;Chung, Soo-Hyun
    • YAKHAK HOEJI
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    • v.41 no.1
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    • pp.105-110
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    • 1997
  • Two protein-polysaccharide fractions, GLA and GLB, respectively, were prepared from the pileus of the fully grown carpophores and the tips of the growing carpophores of Ganod erma lucidum. At a dose of 100mg/kg/day ip, GLA and GLB inhibited the growth of sarcoma 180 solid tumor in ICR mice by 56.3% and 81.8%, respectively. In a flow cytometric (FCM) analysis, GLA and GLB enhanced the formation of lymphoblasts of BALB/c, splenic leukocytes at a concentration of 100 ${\mu}$g/ml, by 38.3% and 61, 3%, respectively. When ip injected into ICR mice, GLB exerted anti-leukopenic effect against cyclophospamide (75mg/kg, ip) in that the leukocyte counts of the peripheral blood of the normal and the cyclophosphamide-treated mice. respectively. was (11.1 ${\pm}$ 3.8) ${\times}$ 10$^3$ and (4.0 ${\pm}$ 1.8) ${\times}$ 10$^3$, while the GLB-cyclophosphamide treated mice showed a leukocyte count of (10.8 ${\pm}$ 5.1) ${\times}$ 10$^3$ CELLS/${\mu}$l. These results suggest that GLB is a promising candidate for an effective, cancer immunotherapeutic agent.

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Growth Inhibition and Apoptosis Induction of Trichosanthis Radix Extract on Human Uterine Cervical Carcinoma Cells (자궁경부암세포에 대한 천화분(天花粉)의 성장억제 및 세포사멸효과)

  • Lim, Eun-Mee;Lee, Hyun-Hee
    • The Journal of Korean Obstetrics and Gynecology
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    • v.18 no.3
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    • pp.77-91
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    • 2005
  • Purpose : Trichosanthis Radix is traditional medical herb which has been shown to inhibit tumor cell proliferation. In this study, the effects of Trichosanthis Radix extract were investigated on inducing growth inhibition and apoptosis of human uterine cervical carcinoma cells. Methods : Human uterine cervical carcinoma cells line, ME-180, was used for the study. The cells were treated with varying concentrations of Trichosanthis Radix extract. Cell growth and inhibitory rate were measured by MTT assay. Apoptosis induction was detected by fluorescence microscopy, DNA ladder formation and flow cytometry. Results : Trichosanthis Radix extract inhibited the growth of human uterine cervical carcinoma cells in a dose-dependent manner. It induced ME-180 cells to undergo apoptosis including fragmented nuclei and nucleosome-sized DNA fragmentation. Flow cytometric analysis showed the increasing rate of apoptotic cells by Trichosanthis Radix extract. Reduction of mitochondrial membrane potential and increase in caspase-3 activity and were found in ME-180 cells treated with Trichosanthis Radix extract. Conclusion : Our data suggest that Trichosanthis Radix extract inhibit the growth and proliferation of ME-180 cells by apoptotic induction and facilitates its activity via caspase-3 activation initiated by depolarization of mitochondria.

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Non-B, Non-T Acute Lymphoblastic Leukemia in a Cat

  • Sumin Cha;Hyunwoo Kim;Hyeona Bae;Minjeong Kang;Rankyung Jung;Minji Kim;DoHyeon Yu
    • Journal of Veterinary Clinics
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    • v.40 no.4
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    • pp.298-302
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    • 2023
  • A 7-year-old neutered male, domestic shorthair cat presented anorexia and lethargy. The complete blood cell count revealed severe non-regenerative anemia, lymphocytic leukocytosis, neutropenia, and thrombocytopenia. On the peripheral blood smear examination, medium to large lymphoblastic cells with moderate amounts of basophilic cytoplasm were observed in up to 70% of peripheral leukocytes. Feline leukemia and immunodeficiency viruses were not detected using a commercial diagnostic kit. While splenomegaly and blunt margins of the caudoventral liver were observed in abdominal radiography, changes in the intra-abdominal lymph nodes were not remarkable. Ultimately, flow cytometric immunophenotyping from the peripheral blood revealed a negative for B-cell markers (CD21-/CD79a-) and T-cell markers (CD3-/CD4-/CD5-/CD8-). Based on the hematological examination and the immunophenotyping assay, the cat was diagnosed with non-B, non-T acute lymphoblastic leukemia. Here, we report a rare case of non-B, non-T acute lymphoblastic leukemia to raise awareness and provide information on clinical symptoms and laboratory test and immunophenotyping analysis results.

Peritoneal lymphomatosis with pleural and peritoneal effusions in a dog: a case report

  • Minjeong Kang;Hyeona Bae;Rankyung Jung;Minji Kim;Sumin Cha;Tae-Sung Hwang;Hee-Chun Lee;Dong-In Jung;Kyu-Woan Cho;DoHyeon Yu
    • Korean Journal of Veterinary Research
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    • v.63 no.3
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    • pp.25.1-25.5
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    • 2023
  • Peritoneal lymphomatosis (PL) is a rare lymphoid neoplasm in dogs. A nine-year-old spayed female Labrador retriever presented with pleural and peritoneal effusions. Diagnostic imaging revealed diffuse nodular to massive lesions in the mesentery, particularly in the caudal abdomen. While the superficial lymph nodes did not show significant changes, enlargement was observed in the intra-abdominal and intra-thoracic lymph nodes. Cytological and flow cytometric analyses of the effusion indicated the presence of large B-cell lymphocytes expressing CD3-/CD5-/CD14-/CD21-/CD34+/CD45+/CD79a+. PL was diagnosed using diagnostic imaging and fluid analysis. This case report highlights the clinical and diagnostic features of canine PL.