• Biotechnology and Bioprocess Engineering:BBE
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• 제9권6호
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• pp.423-427
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• 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.

• Journal of Microbiology and Biotechnology
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• 제10권3호
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• pp.321-326
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• 2000

To examine the feasibility of the long-term production of the human granulocyte colony stimulating factor (hG-CSF) using the $_{L}-arabinose$ promoter system of Escherichia coli, flask relay culture and cyclic fed-batch culture were performed. In the flask relay culture, it was found that the pismid was maintained stably up to about 170 generations in an uninduced condition, whereby the cells could also maintain the capability of expressing hG-CSF expression were maintained stably up to at least 100 generations. In contrast, in the cyclid fed-batch culture, segregational plasmid instability was observed within about 4 generations after induction, even though the cell growth and hG-CSF production reached their maximum balues, 78.0 g/l of dry cell weight and 7.0 g/l of hG-CSF, respectively. It would appear that, when compared to the flask relay culture, the high-cell density and high-level expression of hG-CSF in the cyclic fed-batch cultrure led to the segregational plasmid instability; in other words, a severe metabolic burden existe on the cells due to the high-level expression of hG-CSF. Accordingly, based on these long-term cultures, the segregational and structural plasmid instability was observed and a strategy to overcome such problems could be designed.

• Biotechnology and Bioprocess Engineering:BBE
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• 제9권1호
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• pp.12-16
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• 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.

• Biomolecules & Therapeutics
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• 제2권3호
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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.

• Toxicological Research
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• 제13권3호
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• pp.303-306
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• 1997

Antigenic potential of genetically engineered human granulocyte colony-stimulating factor (CJ-50001) was assessed in guinea pigs and mice. In active systemic anaphylaxis (ASA) test, although CJ-50001 at 50 $\mu\textrm{g}$ /head induced anaphylactic responses, CJ-50001 5 $\mu\textrm{g}$ /head alone or 50 $\mu\textrm{g}$ / head with adjuvant did not induce anaphylactic responses. In passive systemic anaphylaxis test (PCA) or passive hemagglutination test (PHA), CJ-50001 did not induce positive responses. It is concluded that, in light of the fact that CJ-50001 was antigenic only in ASA but not in PCA or PHA and also that CJ-50001 is a foreign human recombinant protein to guinea pigs, CJ-50001 may not induce systemic allergic react-ion in its clinical use in human.

• Asian Pacific Journal of Cancer Prevention
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• 제16권1호
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• pp.299-301
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• 2015

Objectives: To assess safety and efficacy of JiSaiXin (Recombinant Human Granulocyte Colony Stimulating Factor Injection manufactured in China, G-CSF) 150ug per day for three days and whether this regimen could reduce the incidence of febrile neutropenia caused by chemotherapy. Method: From July 2014 to December 2014 patients treated by chemotherapy in our hospital were randomly divided into two groups: Group A with prophylactic use of G-CSF (JiSaiXin) 24 hours after chemotherapy for consecutive 3 days; and Group B with G-CSF (JiSaiXin) after neutropenia. Routine blood tests were performed 7 days and 14 days after chemotherapy. Results: A total of 100 patients fulfilled study criteria, and the incidence of severe neutropenia (grade III/IV) and the incidence of febrile neutropenia in Group A were lower than those in Group B. Nine patients were found severe neutropenia (grade III/IV) in Group B, but one in Group A, three febrile neutropenia in Group B, but 0 in Group A. Conclusions: This study suggested that prophylactic use of G-CSF (JiSaiXin) 150ug per day 24 hours after chemotherapy for consecutive 3 days is safe and could be effective for preventing febrile neutropenia in patients with chemotherapy.

• Journal of Korean Neurosurgical Society
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• 제66권5호
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• pp.511-524
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• 2023

Objective : This animal model aimed to compare the rat group that received brain irradiation and did not receive additional treatment (only saline) and the rat group that underwent brain irradiation and received Granulocyte colony stimulating factor (G-CSF) treatment. In addition, the effects of G-CSF on brain functions were examined by magnetic resonance (MR) imaging and histopathologically. Methods : This study used 24 female Wistar albino rats. Drug administration (saline or G-CSF) was started at the beginning of the study and continued for 15 days after whole-brain radiotherapy (WBRT). WBRT was given on day 7 of the start of the study. At the end of 15 days, the behavioral tests, including the three-chamber sociability test, open field test, and passive avoidance learning test, were done. After the behavioral test, the animals performed the MR spectroscopy procedure. At the end of the study, cervical dislocation was applied to all animals. Results : G-CSF treatment positively affected the results of the three-chamber sociability test, open-space test and passive avoidance learning test, cornu Ammonis (CA) 1, CA3, and Purkinje neuron counts, and the brain levels of brain-derived neurotrophic factor and postsynaptic density protein-95. However, G-CSF treatment reduced the glial fibrillary acidic protein immunostaining index and brain levels of malondialdehyde, tumor necrosis factor-alpha, nuclear factor kappa-B, and lactate. In addition, on MR spectroscopy, G-CSF had a reversible effect on brain lactate levels. Conclusion : In this first designed brain irradiation animal model, which evaluated G-CSF effects, we observed that G-CSF had reparative, neuroprotective and anti-neurodegenerative effects and had increased neurotrophic factor expression, neuronal counts, and morphology changes. In addition, G-CSF had a proven lactate-lowering effect in MR spectroscopy and brain materials.

• 생명과학회지
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• 제20권11호
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• pp.1577-1581
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• 2010

조혈촉진 cytokine인 Granulocyte colony stimulating factor (G-CSF)는 골수세포를 자극하여 granulocyte로 증식, 분화시키는 기능을 가지며, 현재 아주 고가의 치료제로 사용되고 있다. 인간 G-CSF (hG-CSF)를 아직 시도되지 않은 누에 유래 세포주인 BM5 세포에서 발현시키고 생산 효율을 높이기 위해 hG-CSF cDNA를 변형하였다. hG-CSF의 cDNA의 endoplasmic reticulum (ER) signal sequence 부분을 누에의 소포체에서 분비되는 단백질인 prophenoloxidase (PPAE), protein disulfide isomerase (PDI)와 bombyxin (BX)에서 유래한 누에특이 ER signal sequence로 대체한 hG-CSF의 cDNA 함유 벡터를 구축하였다. 이들 벡터를 사용하여 형질전환한 BM5 세포의 배양액에 분비된 G-CSF 단백질을 western blot으로 분석하여 발현을 확인하였다. 누에특이 ER signal sequence들로 대체된 hG-CSF cDNA를 포함하는 벡터에 의한 hG-CSF 단백질 생산이 인간 G-CSF cDNA가 든 벡터에 의한 hG-CSF의 생산보다 월등히 효율적이었다. 또한, PPAE-signal sequence를 포함하는 hG-CSF 단백질은 배양배지에서 형질전환 4일 후에 최고에 달하였고, 7 일째까지 비슷한 양이 배지 내에서 검출되었다. 이상의 결과는 인간유래 유전자가 곤충세포 내에서 발현 될 때 인간유래 유전자 보다는 곤충 유전자발현 시스템에 맞게 변형했을 경우 더 효율적인 단백질 발현을 얻을 수 있음을 보여 준다.

• Biomolecules & Therapeutics
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• 제6권1호
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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.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1999년도 제13회 식물생명공학심포지움 New Approaches to Understand Gene Function in Plants and Application to Plant Biotechnology
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• pp.45-49
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• 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.