• Journal of Microbiology
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• v.38 no.2
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• pp.109-112
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• 2000

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic hematopoietic growth factor and an activator of lmature myeloid cells and recombinant GM-CSF is increasingly under clinical studies for the treatment of various diseases including cancer, infectious diseases and hematopoietic diseases. We constructed a reconbinant mouse GM-CSF expression plasmid with pelB leader sequence and His. Tag under T7 promoter control, and showed that the construct produced a 20 kDa recombinant protein in 8M urea. We also showed that the 20 kDa recombinant protein prepared in 8M urea sitmulated colony formation in vitro, indicating that the recombinant mGM-CSF can be renatured to its native form to show the colony stimulating activity.

• Archives of Pharmacal Research
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• v.23 no.3
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• pp.252-256
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• 2000

A thymic stromal cell line, TFGD, was established from a thymic tumor mass developed spontaneously in p53 knock out mouse, and was found to produce cytokines that could induce bone marrow hematopoietic stem cells (HSCs) to differentiate into macrophages. The cytokines produced by the TFGD line were assessed by immunoassays. High level of macrophage-colony stimulating factor (M-CSF) and interleukin (IL)-6 was detected in the TFGD-culture supernatant, whereas granulocyte/macrophage-colony stimulating factor (GM-CSF), IL-3, IL-4, IL-5, IL-13, or interferon (IFN)-$\gamma$ was undetectable. Blocking experiments showed that anti-M-CSF monoclonal antibody could neutralize the differentiation-inducing activity shown by the TFGD-culture supernatant. Dot blot analysis of the total RNA isolated from the cultured fetal thymic stromal cells showed that M-CSF transcripts were expressed in the normal thymus. These observations, together with the earlier finding that M-CSF plus IL-6 is the optimal combination of cytokines for the induction of macrophage differentiation from HSCs in vitro, may indicate that thymic macrophages could be generated within the thymus by cytokines involving M-CSF.

• Korean Journal of Pharmacognosy
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• v.21 no.2
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• pp.186-192
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• 1990

This study was conducted to further purify and characterize a polysaccharide fraction from whole parts of Duchesnea indica (Andr.) Focke, which was previously reported to show an antitumor activity by us. Mild acid-treatment of the fraction by 5% acetic acid yielded three subtractions (Fr. A,B and C) on a Sephadex G-25. Fr. A was composed of uronic acid, hexose, pentose and phenolic substance, Fr. C mainly phenolic substance, and Fr. B contained hexose. All three fractions exhibited colony stimulating factor activity.

• Korean Journal of Pharmacognosy
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• v.20 no.2
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• pp.110-116
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• 1989

This study was conducted to purify and characterize a polysaccharide fraction from whole parts of Duchesnea indica (Andr.) Focke, which was previously reported to show an antitumor activity by us. The plant was extracted with 5% ethanol for 3 days in the room temperature. The extract was dialyzed by 5% ethanol, and then concentrated by extraction with $excess\;_N-butanol$, followed by gel filtration named Fr. a and b. Each was composed of gallic acid, hexose, pentose, uronic acid and protein, indicating that both fractions were tannic polysaccharide containing protein. Heat treatment of them yielded gallic acid less polysaccharides. They showed a colony stimulating factor activity.

• Journal of Microbiology
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• v.40 no.1
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• pp.77-81
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• 2002

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic hematspoietic growth factor involved in the development of myeloid cells from bone marrow, and an activator of mature myeloid cells functioning in a variety of antimicrobial and inflammatory responses. Recently, recombinant GM-CSF is increasingly under clinical study for treatment of various diseases including cancer, infectious diseases and hematopoietic diseases as well as for an immune response modulator, In this study, we constructed a recombinant human GM-CSF (rhGM-CSF) expression plasmid with a pelB leader sequence and His. Tag under T7 promoter control. The expression construct was shown to produce a recombinant protein of 20 kDa in the 8M urea preparation, indicating the rhGM-CSF may be expressed as an insoluble inclusion form. The 20 kDa recombinant protein in 8M urea was transformed into the water-so1ub1e form by dialysis against PBS buffer (phosphate buffered saline). The soluble rhGM-CSF protein was shown to stimulate colony formation and cell proliferation in vitro, indicating that the rhGM-CSF could be refolded into its native form to show colony stimulating activity.

• Clinical and Experimental Reproductive Medicine
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• v.30 no.1
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• pp.5-14
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• 2003

Objective: Present study was aimed to verify the effect of granulocyte macrophage-colony stimulating factor (GM-CSF) in the preimplantation development of mouse embryos and the involvement of the mitogen activated protein kiase (MAPK) in the GM-CSF signaling. Methods: Two-cell embryos were cultured for 96 h in the presence or absence of GM-CSF (0, 0.4, 2, 10 ng/ml) and PD98059, a MEK inhibitor (10 ${\mu}M$). Morphological development, cell number per blastocyst, and apoptotic nuclei, were eamined. MAPK activity of embryonic immunoprecipitate by MAPK (ERK1/2) antibody was measured by in vitro phosphorylation of myelin basic protein. Results: At post hCG 122 h the embryonic development among the experimental groups was significantly different (p=0.018). The rate of blastocyst development and cell number per embryo were the highest in 2 ng/ml GM-CSF treatment group. The percent of apoptotic cells of the GM-CSF-treated embryos was the lowest among the group. In blastocysts, GM-CSF treatment transiently increased MAPK activity. PD098059 attenuated the effect of GM-CSF on the morphological development, increase in cell number per blastocyst, down regulation of apoptosis, and upregulation of MAPK activity, suggesting that activation of MAPK cascade possibly mediated the embryotropic effect of GM-CSF. Conclusion: This result suggested that GM-CSF potentiated the development of preimplantation mouse embryos by activation of MAPK.

• Journal of Microbiology and Biotechnology
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• v.10 no.3
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• pp.287-292
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• 2000

Recombinant Aspergillus niger was constructed to express and secrete a biologically active murine granulaocyte macrophage-colony stimulating factor (mGM-CSF). A 500 bp fragment encoding the signal peptide and terminator of glyceraldehyde-3-phosphate dehydrogenase (gpd). The hygromycin phosphotrasferase gene (hph) was used as a selection marker for the fungal transformants. An expression vector was introduced into A. niger ATCC 9642, and a Northern blot analysis indicated the presence of a considerable amount of transcripts from the introduced mGM-CSF. The biological activity of recombinant mGM-CSF (rmGM-CSF) isolated from the culture filtrate was confirmend by measuring the proliferationof the GM-CSF dependent FDC-P1 cell line. It appeared that rmGM-CSF was amenable to the proteolytic activity produced by A. niger, since biological actibity was only observed when the transformants were grown in a protease-repressing medium, and the activity of rmGM-CSF dramatically decreased with an increase of age of the culture. The yield of rmGM-CSF, as determined by ELISA. was 640 ng/l of culture filtrate. Accordingly, its specific activity is estimated to be approximately two-and-a-half times higher than that of a commercial preparation from E. coli.

• Biomolecules & Therapeutics
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• v.6 no.1
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• pp.89-94
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• 1998

CJ50001 is a recombinant human granulocyte colony-stimulating facto, (rHuG-CSF) that stimulates the formation of neutrophils from bone marrow stem cells. It was produced in E. colt and purified through refolding and several processes. We produced CS970125(300) using purified C150001 and additives in order to test the stability of CJ50001. When CS970125(300) was stored at 50'S for more than 1 week, high molecular weight proteins were formed and those proteins were detected by non-reducing SDS-PAGE, gel filtration HPLC, and Western blot. Those proteins showed single band at the same position of CJ50001 in reducing SDS-PAGE. These data indicated that those high molecular weight proteins were the multimers of C150001. In biological assays, iu viro and in viro, the multimers did not have biological activity and inhibitory action to that of CJ 50001. The mutimers did not induce toxicity in mice and rats in acute toxicity test. These results suggest that if Cs970125(300) containing CJ50001 is stored at 5$0^{\circ}C$, CJ50001 will be the multimers that do not have biological activity and inhibitory effect to CJ50001 and do not induce acute toxicity.

• Proceedings of the Zoological Society Korea Conference
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• 1997.10b
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• pp.295.2-295
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• 1997

No Abstract, See Full Text

• Biomolecules & Therapeutics
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• v.2 no.3
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• pp.247-255
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• 1994

As a series of safety studies of DA-3030, a new rhO-CSF, its local irritancy was examined in the rabbits after the following treatment; application into the conjunctival sac of the eye(single), subcutaneous injection(single), intramuscular injection(single), and intravenous injection(8-day repeated). In addition, paravenous irritation of DA-3030 was investigated in mice. The results obtained were as follows. 1. In the result of ocular irritation test, 0.03% solution of DA-3030 could be considered as a non-irritating material. 2. The local irritation of DA-3030 by an injection of 0.5mι of its solution subcutaneously or intramuscularly was negligible and not so much different from that of saline. 3. In the vascular irritancy test, macro- and microscopic observations revealed that the irritating activity of DA-3030 in blood vessels was not different from that of saline when they were injected once a day into vein retroauricularis of rabbits for 8 days.4. The paravenous administration of DA-3030 did not induce any abnormal changes at injection sites except mild swelling in 1 mouse at 3 hours after injection which was thought to be due to slow absorption. The above-mentioned results suggest that DA-3030 has no irritating activity when injected through intravenous or subcutaneous route for clinical practice as 0.03% solution.