• Radiation Oncology Journal
• /
• v.13 no.1
• /
• pp.79-85
• /
• 1995

Purpose : To assess the efficacy of recombinant human granulocyte-macrophage colony-stimulating factor(GM-CSF) in the neutropenia by radiotherapy. Materials and Methods : Eleven patients with various solid tumor were treated with a daily subcutaneous dose of GM-CSF(3-7microgram/kg) for 5days during the radiotherapy. Before and during the course of the study all the patients were monitored by the recording of physical examination, the complete blood count with differential and reticulocyte count and liver function test. Eight patients received prior or concurrent chemotherapy. Results : In 10 patients, the neutrophilic nadir was significantly elevated and the lenght of time that Patients had a neutrophil count below $10^3/mm^3$ a threshold known to be critical to acquiring infective complications was shortened following GM-CSF injection. A significant rise (two fold or greater) of neutrophil count was seen in 10 of 11 patients. In most patients, discontinuation of GM-CSF resulted in a prompt return of granulocyte counts toward baseline. However the neutrophil count remained elevated over $10^3/mm^3$ during radiation therapy, and radiotherapy delays were avoided. Other peripheral blood components including monocytes and platelets also increased after GM-CSF treatment. No significant toxicity was encountered with subcutaneous GM-CSF treatment. Conclusion : GM-CSF was well tolerated by subcutaneous route and induced improvement in the neutropenia caused by radiotherapy.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.9 no.1
• /
• pp.12-16
• /
• 2004

Partitioning of human granulocyte-macrophage colony stimulating factor (hGM-CSF) was achieved in the aqueous two-phase systems (ATPSs) using a crude extract of transgenic tobacco cell suspension culture. This study examined the effects of polyethylene glycol (PEG) molecular weight and concentration and the effects of sodium phosphate concentration in different PEG/sodium phosphate systems on the partition coefficient, K. The best ATPS system was 5% PEG 8,000/1.6 M sodium phosphate after 2 h of incubation at room temperature. In this system, hGM-CSF was partitioned in the PEG-rich phase with a yield of 57.99% and K$\_$hGM-CSF/ of 8.12. In another system, 3% PEG 10,000/1.6 M sodium phosphate, hGM-CSF was also partitioned primarily in the top phase with a yield of 45.66% and K$\_$hGM-CSF/ of 7.64 after 2 h of incubation at room temperature.

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.9 no.6
• /
• pp.423-427
• /
• 2004

A simple purification procedure of bioactive human granulocyte macrophage colony stimulating factor (hGM-CSF) secreted in rice cell suspension culture has previously been described. In this study the protein was purified to apparent homogeneity with an overall yield of $80.1\%$ by ammonium sulfate precipitation and a single chromatographic step involving FPLCanion exchange chromatography. The purified hGM-CSF revealed at least five glycosylated forms ranging from $21.5{\~}29$ kDa, and its biological activity was independent of the glycosylation pattern. This is the first purification report of recombinant hGM-CSF to apparent homogeneity from rice cell suspension cultures.

• Proceedings of the Botanical Society of Korea Conference
• /
• 1999.07a
• /
• pp.45-49
• /
• 1999

Plant cell culture is emerging to express bioactive foreign proteins because it has several advantages in that it is safe, economical, genetically stable and eukaryotic expression system comparing with other expression systems. However several limitations such as slow growth rate, low expression level and lack of well established down stream process need to be answered. As a preliminary approach to produce the immunologically interested molecules through the plant cell culture, we tested if granulocyte-macrophage colony stimulating factors (GM-CSFs) from both murine (mGM-CSF) and human (hGM-CSF) are produced as a biologically active form through plant cell culture. The murine and human GM-CSF genes were cloned into the plant expression vector, pBI121, and Ti-plasmid mediated transformation of tobacco leaves was conducted using Agrobacterium tumefaciens harboring both recombinant GM-CSF (rGM-CSF) genes. Cell suspension culture was established from the leaf-derived calli of transgenic tobacco plant. Northern blot analysis indicated the expression of the introduced mGM-CSF gene in both transgenic plant and cell suspension cultures. In addition, the biological activities of both murine and human GM-CSF from plant cell culture were confirmed by measuring the proliferation of the GM-CSF dependent FDC-PI and TF-1 cells, respectively.

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

• Toxicological Research
• /
• v.23 no.4
• /
• pp.383-389
• /
• 2007

Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) regulates proliferation and differentiation of hematopoietic progenitor cells and modulates function of the mature hematopoietic cells. In the previous study, we reported that hGM-CSF could be produced in transgenic rice cell suspension culture, termed rhGM-CSF. In the present study we examined the single and repeated dose toxicity of rice cells-derived hGM-CSF in SD rats. During single dose toxicity study for 7 days, there were no any toxic effects at any dose of from 10 to $1000{\mu}g/kg$. The lethal dose ($LD_{50}$) was not found in this range. Moreover, repeated dose toxicity study of 14-days period and at the doses of 50 and $200{\mu}g/kg$ (i. v.) of rhGM-CSF did not show any changes in food and water intake. There were also no significant changes in both body and organ weights between the control and the test groups. The hematological and blood biochemical parameters were statistically not different in all the groups. These results suggest that rhGM-CSF has no toxicity in SD rats.

• Toxicological Research
• /
• v.24 no.4
• /
• pp.315-320
• /
• 2008

Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) is a glycoprotein and hematopoietic growth factors that regulates the proliferation of myeloid precursor cells and activates mature granulocytes and macrophages. In a previous study, we reported that hGM-CSF could be produced in transgenic rice cell suspension culture, termed rhGM-CSF. In the present study, we examined the repeated dose toxicity of rhGM-CSF in SD rats. The repeated dose toxicity study was performed at each dose of 50 and 200 ${\mu}g/kg$ subcutaneous administration of rhGM-CSF everyday for 28-days period. The results did not show any changes in food and water intake. There were also no significant changes in both body and organ weights between the control and the tested groups. The hematological and blood biochemical parameters were statistically not different in all groups. These results suggest that rhGM-CSF may show no repeated dose toxicity in SD rats under the conditions.

• Animal cells and systems
• /
• v.16 no.3
• /
• pp.207-214
• /
• 2012

Burns are one of the most devastating forms of trauma and wound healing is a complex and multicellular process, which is executed and regulated by signaling networks involving numerous growth factors, cytokines, and chemokines. Recombinant human granulocyte macrophage colony-stimulating factor (rhGM-CSF) was specifically produced from rice cell culture through use of a recombinant technique in our laboratory. The effect of rhGM-CSF on promotion of deep second-degree burn wound healing on the back skin of a hamster model was evaluated through a randomized and double-blind trial. As macroscopic results, hamster skins of the experimental groups showed earlier recovery by new epidermis than the control groups. Immunohistochemical reactions of proliferating cell nuclear antigen and transforming growth factor-b1, which are indicators of cell proliferation, were more active in the experimental group, compared with the control group. On electron microscopy, basal cells in the epidermis of the experimental group showed oval nuclei, prominent nucleoli, numerous mitochondria and abundant free ribosomes. In addition, fibroblasts contained well-developed rough endoplasmic reticulum with dilated cisternae. Bundles of collagen fibrils filled the extracellular spaces. Particularly, ultrastructural features indicating active metabolism for regeneration of injured skin at 15 days after burn injury, including abundant euchromatin, plentiful free ribosomes, and numerous mitochondria, were observed. These findings suggest that use of rhGM-CSF could result in accelerated deep second-degree burn wound healing in animal models.

• Proceedings of the Korean Society of Life Science Conference
• /
• 2002.12a
• /
• pp.48-60
• /
• 2002

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine that stimulates the production of granulocytes, macrophages, and white blood cells. The effects of osmotic pressure on secretion of human GM-CSF into the culture medium were investigated in suspension cultures of transgenic tobacco cells. An increase in osmotic pressure caused by the addition of mannitol decreased the cell size index, with the effect being more pronounced when cells were measured wet rather than dry. Increased osmotic pressure enhanced the secretion of hGM-CSF. At 90 g/L mannitol, the maximum concentration tested, hGM-CSF was present in the culture medium at 980 ug/L. As the concentration of mannitol increased, the total amount of protein secreted also increased, but was disproportionately enriched in GM-CSF NaCl, another osmoticum, had very similar effects on cell growth and hGM-CSF production, but did not cause enrichment for hGM-CSF Additionally, protein-stabilizing polymer was added to culture broth to enhance stability of secreted recombinant protein. Finally, above two method were applied together to maximize the productivity.

• 대한두경부종양학회:학술대회논문집
• /
• 1994.11a
• /
• pp.20-20
• /
• 1994