• Archives of Pharmacal Research
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• 제20권1호
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• pp.34-38
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• 1997

Recombinant human epidermal growth factor (rhEGF), a polypeptide of 53 amino acid residues, is subject to degradation by numerous enzymes, especially proteases, when it is applied on the skin for the treatment of open wound. Amastatin, aprotinin, bestatin, EDTA, EGTA, gabexate, gentamicin, leupeptin, and TPCK were investigated for the possible protease inhibitors, which may use to protect rhEGF from degradation by the enzymes in the skin. Skin homogenates containing protease inhibitors and rhEGF were incubated at $37^{\circ}C$ for 30 minutes. After the reaction was stopped with trifluoroacetic acid, the amount of rhEGF remaining in the sample was determined with an HPLC method. The percentages of rhEGF degraded, at the skin/PBS ratio of 0.25, in the mouse, rat, and human skin homogenate were 85%, 70%, and 46%, respectively. The degree of degradation of rhEGF in the cytosolic fraction was higher than that in the membrane fraction and these enzyme reactions were completed in 30 minutes. Bestatin, EGTA, and TPCK showed significant inhibitory effects on the degradation of rhEGF in the two fractions (p<0.05), while the other protease inhibitors had no significant inhibitory effects or, even resulted in deleterious effects. Therefore, the formulation containing one or several inhibitors among these effective inhibitors would be a promising topical preparation of rhEGF for the treatment of open wound.

• Biomolecules & Therapeutics
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• 제8권2호
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• pp.171-178
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• 2000

To evaluate the pharmacokinetic properties and tissue distribution of a newly developed recombinant human erythropoietin (GC-rhEPO), we analyzed the plasma and tissue levels of erythropoietin by an ELISA after intravenous (IV) and subcutaneous (SC) adminstration to the male rats at the doses of 20, 100, 500 or 2,500 unit/kg. After single IV bolus injection of GC-rhEPO, the plasma concentration was rapidly increased and decreased with two phases with half-lives of 13.4 min and 2.94 hours. AUC was increased dose- dependently but plasma half-lives remained constant regardless of GC-rhEPO doses. Following SC administration, the plasma concentration increased slowly with half-life of 9.2 hours and reached peak at 8 hours. Mean residence time and bioavailability were 18.2 hours and 44%, respectively. After single IV dose of 100 unit/kg, tissue GC-rhEPO level was higher in bone marrow and spleen, while the depletion rate was slower in liver and bone marrow, indicating the higher affinity of GC-rhEPO to bone marrow. Taken together, the experimental results indicate that GC-rhEPO contained the typical pharmacokinetic properties and the tissue distribution patterns inherent to human erythropoietin.

• Biomolecules & Therapeutics
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• 제8권2호
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• pp.184-193
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• 2000

To evaluate GC-rhEPO, human erythropoietin produced by recombinant DNA technique, its general pharmacological properties were investigated in experimental animals administering intravenously and in vitro test system. GC-rhEPO at doses of 70,700 and 7,000 IU/kg body weight had no influence on general behavior, spontaneous motor activity, thiopental-inducted sleeping time, writhing syndrome induced by acetic acid, strychnine-induced convulsions, charchoal meal propulsion in mice, and body temperature, gastric juice secretion, urine and electrolyte excretion in rats. In anesthetized rabbits, GC-rhEPO (70, 700 and 7,000 lU/kg, i.v.) did not alter respiratory rate, blood pressure, heat rate. In in vitro experiments, GC-rhEPO did not affect the contractions of the isolated ileum of guinea pigs and the muscle twitchs of isolated neuromuscular junction of the rats. In addition, GC-rhEPO did not affect the blood coagulation time and ADP-induced platelet aggregation in plasma of rabbits. Taken together, these results indicate that GC-rhEPO does not induce any adverse effects in the experimental animals.

• 한국작물학회지
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• 제46권4호
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• pp.321-324
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• 2001

Tobacco seeds (Nicotiana tabacum L. cv KF109) were primed in the polyethylene glycol 6000(PEG) solution and then stored at 5 and $25^{\circ}C$ under 40, 60 and 80% relative humidity (RH) conditions for six months. The effect of storage temperature and humidity on mean germination time ($T_{50}$), longevity and germination of the primed tobacco seeds were compared. Untreated seeds (control) stored at $5^{\circ}C$ showed high germinability throughout the entire storage period and humidity, and a decline in germinability showed after 6 months at 60% RH and after 3 months at 80% RH when stored at $25^{\circ}C$, Primed seeds retained high germinability until 6 months at 60% RH and 3 months at 80% RH when stored at $5^{\circ}C$ but showed a significant decline in germinability after 3 months at 40% RH, and 1 months at 60% and 80% RH, respectively when stored at $25^{\circ}C$, Primed seeds were completely lost viability when stored at $25^{\circ}C$ under 60% RH for 6 months and under 80% RH for 3 months.

• 한국동물학회지
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• 제37권4호
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• pp.504-513
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• 1994

The present study was carried out 1) to show the ontogenic development of CA-and GnRH-containing nerve fobres in the median eminence, 2) to simultaneously demonstrate the synaptic contact between these two nenre fibres in the rat median eminence at the ultrastructural level using light and electron microscopic doublelabel immunostainlngs. GnRH-and CA-nenre terminals were detectable in the median eminence at embryonic day 19.5. The CA-newe terminals were obsenred in the entire legion of the extern31 lavers, while GnRH-newe terminals only in the lateral portion. At the 14th postnatal daw, both %ropes of nerve terminals showed a very similar distribution to those of adult one. In the median eminence of adult rats, a substantial overlap existed in the distribution of GnRH fibres with CA-containing nerve fibres. This overlap was most intense throughout the external palisade zone. Furthermore, an electron microscopic double label immunostaining showed that there was a close apposition of CA- and GnRH-nenre fobres. These axo-axonic contacts occurred frequently in the internal and palisade zones, i.e. at the level of the fobre preterminals. These morphological results suggest that the CA-mediated GnRH secretion may occur via sxo-axonic interaction in the median eminence.

• Journal of Microbiology and Biotechnology
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• 제2권2호
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• pp.135-140
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• 1992

The partitioning of recombinant human interleukin-2(rhII-2) in PEG 8000-dextran 38800 aqueous two-phase system has been investigated using three different sources of rhIL-2. In the case of pure rhIL-2, the solubility in a PEG-dextran two-phase system was low and most of rhIL-2 was partitioned into the bottom phase. For the recovery of rhIL-2 from insoluble protein aggregates, the inclusion bodies of recombinant E. coli were solubilized by the treatment with sodium dodecyl sulfate (SDS). The addition of SDS significantly enhanced not only the solubility of rhIL-2 but also the partitioning of rhIL-2 to the top phase. When the ratio of SDS to rhIL-2 was 2.0, the partition coefficient(K) and the recovery yield(Y) at the top phase were 4.5 and 88%, respectively, at pH 6.8. In order to reduce the recovery steps further, SDS was directly added to the intact recombinant E. coli cells and then partitioned into the PEG/dextran aqueous two-phase system. The observed partition coefficient ($K{\cong{3.0$) and recovery yield ($Y{\geq}80%$ )of this method were comparable to the rhIL-2 recovery from insoluble protein aggregates. The results obtained in this work indicate that PEG-dextran two-phase partitioning might provide a simple way for the recovery and partial purification of recombinant proteins which are produced as inclusion bodies.

• 대한치과보철학회지
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• 제50권1호
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• pp.44-52
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• 2012

연구 목적: 본 실험은 거친 표면으로 유의성 있는 줄기세포반응을 나타냈던 4 가지의 티타늄 표면 위에 rhBMP-2를 코팅했을 때 어떤 유의한 줄기세포반응(세포부착, 증식, 분화)이 나타나는지 비교 분석함으로 rhBMP-2 코팅을 위한 가장 적절한 표면을 평가하기 위해 시행되었다. 연구 재료 및 방법: 대조군인 기계절삭표면(machined surface)과 실험군인 양극산화(anodized), RBM, SLA 표면에 rhBMP-2를 코팅한 후 코팅하지 않은 표면과 같이 8가지 표면 위에 인간줄기세포를 배양하였다. 배양 후 24시간 후 SEM을 통해 줄기세포의 부착을 평가하였고 배양 3, 7, 14일후MTT와 ALP 검사를 통해 줄기세포의 증식과 분화반응을 평가하였다. 그리고 배양 7일후RT-PCR 검사를 통해 Type I collagen, osteocalcin, osteopontin의 유전자 발현의 변화를 평가하였다. 결과: SEM 평가에서 4가지 rhBMP-2 표면이 코팅하지 않은 표면에 비해 세포부착 면적이 넓고 긴밀하며 세포돌기가 더 많이 관찰되었다. 양극산화 rhBMP-2코팅표면에서 가장 두드러지게 관찰되었다. MTT 검사에서 크게 의미 있는 차이는 나타나지 않았다. ALP검사에서 양극산화 rhBMP-2코팅 표면은 대조군과 비교해서 (3, 14일) 또 RBM rhBMP-2 코팅 표면과 비교해서 (14일) 유의성 있는 ALP 활성도의 증가를 나타내었다(P<.05). RT-PCR 검사에서 osteocalcin과 osteopontin의 유전자 발현은 양극산화 rhBMP-2코팅 표면에서 높게 나타났다. 결론: 양극산화 rhBMP-2코팅표면이 줄기세포의 부착과 분화실험에서 대조군표면과 rhBMP-2를 코팅한 기계절삭표면이나 RBM 표면에 비해 유의성 있는 증가를 나타냈다(P<.05).

• Clinical and Experimental Pediatrics
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• 제45권4호
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• pp.439-448
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• 2002

목 적 : 신생아 호중구 감소증이 합병된 신생아 패혈증 환아의 치료에 있어서 부가적으로 rhG-CSF(recombinant human granulocye colony-stimulating factor)를 투여함에 있어서 서로 다른 용량의 rhG-CSF를 투여함으로써 나타나는 신생아 패혈증에 합병된 호중구 감소증의 치료와 환아들의 생존율에 미치는 영향을 평가, 비교하려고 하였다(group I/II형 연구). 방 법: 1995년 10월부터 1996년까지 신생아 호중구 감소증이 합병된 신생아 패혈증 환아는 모두 10명으로 이들에게는 $10{\mu}g/kg$을 피하주사 하였고(rhG-CSF $10{\mu}g/kg$ 투여군), 1996년 10월부터 1997년 9 월까지는 신생아 호중구 감소증이 합병된 신생아 패혈증 환아는 모두 12명으로 이들에게는 rhG-CSF를 $5{\mu}g/kg$ 피하주사 하였다(rhG-CSF $5{\mu}g/kg$ 투여군). 각 군의 호중구 증가 정도와 임상적 결과를 서로 비교하였다. 결 과 : RhG-CSF $10{\mu}g/kg$ 투여군은 조발형 신생아 패혈증 1명과 지발형 신생아 패혈증 9명으로 이루어졌고, 모두에게 호중구 감소증이 합병되었다. rhG-CSF $5{\mu}g/kg$ 투여군은 조발형 신생아 패혈증 1명과 지발형 신생아 패혈증 11명이 대상이 되었고, 이들 모두 호중구 감소증이 합병되었다. 두 군간에 출생체중, 재태주령, 항생제 사용, 신생아 패혈증 시기에 기계적 환기요법 투여, 승압제로 dopamine 투여 또는 다른 지지적 요법의 투여에 있어서 차이가 없었다. rhG-CSF 투여 전의 순 호중구 수(ANC)는 rhG-CSF $10{\mu}g/kg$ 투여군이 $1,065{\pm}89$($mean{\pm}SEM$), $5{\mu}g/kg$ 투여군이 $1,053{\pm}131$로 차이가 없었다. 투여 후의 ANC의 증가는 rhG-CSF $10{\mu}g/kg$ 투여군과 $5{\mu}g/kg$ 투여군에서 각각 투여 후 24시간에 7배, 6배, 투여 후 48시간에 10배, 6배, 투여 후 72시간에 8배, 4배, 투여 후 120시간에 8배, 4배로 투여 전에 비하여 두 군 모두에서 각 시간에 의미 있는 증가를 보였다(repeated measure ANOVA와 Kruskall-Wallis test, within subjects effect). 그러나 두 군 간의 차이는 투여 후 48시간에 ANC 최고치에서만 의미 있는 차이를 보였다(student t-test와 Wilcoxon rank sum test). 단핵구 수도 이 기간 동안 의미 있게 증가하였으나 정상범위를 넘지는 않았다. rhG-CSF $10{\mu}g/kg$ 투여군에서 1명의 환아가 자의 퇴원하였고, 1명의 환아가 사망하여 신생아 패혈증에서 회복하여 문제없이 퇴원한 생존율은 자의 퇴원 환아를 제외한 9명 중 8명으로 88.9％였고, rhG-CSF $5{\mu}g/kg$ 투여 군은 12명 중 10명이 생존하여 생존율은 83.3%였다. 두 군 모두에서 특별한 독성이나 부작용은 관찰되지 않았다. 결 론 : RhG-CSF의 투여는 호중구 감소증이 합병된 극심한 신생아 패혈증 환아에서 호중구의 증가를 일으켰다. 두가지 투여 용량에 따르는 효과는 거의 동일하였으며, 단지 투여 후 48시간에 ANC 최고치에서만 의미 있는 차이가 있었다. 두 군의 생존율은 80%이상이었다. 이와 같은 호중구 감소증이 합병된 신생아 패혈증에서 rhG-CSF의 투여 효과는 골수 억압이나 호중구 소모에 의하여 호중구 감소증이 합병된 신생아 패혈증에서 시기적으로 적절히 투여하면 효과적인 치료를 이룰 수 있다는 것을 시사한다. 향후 rhG-CSF의 효능과 부작용에 대하여 무작위 대조실험이 필요시 된다.

• 미생물학회지
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• 제49권2호
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• pp.131-136
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• 2013

본 연구에서는 헤어리베치로부터 뿌리혹 형성능과 질소 고정능이 우수한 Rhizobium 균주를 분리하고자 하였다. 전국의 일곱 군데 헤어리베치 재배 지역에서 수집 후 뿌리혹으로부터 균주를 분리하였고, 헤어리베치에 분리 균주를 재접종하여 뿌리혹 형성능과 질소 고정능을 확인하였다. 그 결과, 52개의 Rhizobium 속 균주를 분리하였고, 이 중 0.5% NaCl 농도 이상에서 자라는 Rhizobium은 16균주이었다. 뿌리혹 형성능은 16개 균주 중 Rhizobium sp. RH1, RH3, RH81, RH82, RH84, RH93의 균주를 접종한 시험구에서 우수하였다. 또한, 질소 고정능을 확인한 결과 Rhizobium sp. RH84가 가장 높은 acetylene reduction activity를 보였다. 이러한 결과는 향후 염의 분포가 불균일한 간척지에 적용 시에도 헤어리베치의 생육을 촉진해 녹비 작물로의 활용 가능성을 높여주는 결과로 판단된다.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제42권2호
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• pp.90-98
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• 2016

Objectives: The aim of this study was to compare the osteogenic effects of demineralized dentin matrix (DDM) combined with recombinant human bone morphogenetic protein-2 (rhBMP-2) in rabbit calvarial defects with DDM and anorganic bovine bone (ABB) combined with rhBMP-2. Materials and Methods: Four round defects with 8-mm diameters were created in each rabbit calvaria. Each defect was treated with one of the following: 1) DDM, 2) ABB/rhBMP-2, or 3) DDM/rhBMP-2. The rhBMP-2 was combined with DDM and ABB according to a stepwise dry and dip lyophilizing protocol. Histological and microcomputed tomography (${\mu}CT$) analyses were performed to measure the amount of bone formation and bone volume after 2- and 8-week healing intervals. Results: Upon histological observation at two weeks, the DDM and ABB/rhBMP-2 groups showed osteoconductive bone formation, while the DDM/rhBMP-2 group showed osteoconductive and osteoinductive bone formation. New bone formation was higher in DDM/rhBMP-2, DDM and ABB decreasing order. The amounts of bone formation were very similar at two weeks; however, at eight weeks, the DDM/rhBMP-2 group showed a twofold greater amount of bone formation compared to the DDM and ABB/rhBMP-2 groups. The ${\mu}CT$ analysis showed markedly increased bone volume in the DDM/rhBMP-2 group at eight weeks compared with that of the DDM group. Notably, there was a slight decrease in bone volume in the ABB/rhBMP-2 group at eight weeks. There were no significant differences among the DDM, ABB/rhBMP-2, and DDM/rhBMP-2 groups at two or eight weeks. Conclusion: Within the limitations of this study, DDM appears to be a suitable carrier for rhBMP-2 in orthotopic sites.