• Toxicological Research
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• v.21 no.3
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• pp.209-218
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• 2005

We investigated effects of recombinant human erythropoietin (rHuEPO) on profiles of mRNA transcripts in 6 male cynomolgus (M. fascicularis) monkey's spleen for 4 weeks. Six monkeys, composed of control and treatment group (Control : M1, M2, M3: Treatment : M4, M5, M6) were intravenously administered 3 times per week without or with a dose of rHuEPO 2730 IU/0.1 ml/kg. After 4 weeks rHuEPO treatment, spleen was removed for RNA isolation. Splenic gene expression was assessed using Affymetrix U133A 2.0 arrays containing 18,400 transcripts and variants, including 14,500 well-characterized human genes. Gene expression pattern was very different between individuals even in same treatment. In rHuEPO treated groups showed number of genes were up- or down-regulated (M4: 79: M5: 48; M6: 73 genes). Six genes (epidermal growth factor receptor, calgranulin A, estrogen receptor binding site associated antigen, matrix metalloproteinase 19, zinc finger and BTB domain containing 16, progestin and adipoQ receptor) were commonly expressed in rHuEPO treated group. The different individual response could be major considering factor in monkey experiment. Further study is needed to clarify the different individual response to rHuEPO in molecular level. This study will be valuable in the fundamental understanding and validation of molecular toxicology for bio-generic drugs including rHuEPO in cynomolgus monkey.

• Korean Journal of Veterinary Research
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• v.39 no.4
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• pp.797-805
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• 1999

T sergenti cDNA library were constructed to get a more broad information about the structural, functional or antigenic properties of the proteins, and analyzes for their partial cDNA sequences and expression sequences tags(ESTg). The mRNA were purified from T sergenti isolates to identify the information of antigen gene, then first and second strand cDNA was synthesized. EcoR I adaptor ligation and Xho I enzyme restriction were used to the synthesized cDNA, and ligated into a Uni-ZAP XR vector. T sergenti cDNA library was constructed with packaging and amplification in vitro. Antibody screening was performed with constructed T sergenti cDNA library using antisera against T sergenti. Among those clones, eight phagemids were rescued from the recombinant in vivo excision with f1 helper phage. Using the analysis of endonuclease restriction and PCR, the recombinant cDNA were proved having a 0.5-3.0kb of inserts. The eight of partial cDNA clones' sequences were obtained and examined for their homology using BLASTN and BLASTX. The eight of sequenced clones were classified into three groups according to the basis of database searches. A total 3,045bp of partial cDNA sequence were determined from six clones. The putatively identified clones contain a cytochrome c gene, a heat shock protein gene, a cyclophilin gene, and a ribosomal protein gene. The unidentified clones have a homology to ATP-binding protein(mtrA) gene of S argillaceus, DNA-binding protein(DBP) gene of Pseudorabies virus 85kDa merozoite protein gene of B bovis, mRNA spm1 protein of T annulata and glycine-rich RNA-binding protein mRNA of O sativa etc.

• 대한생식의학회:학술대회논문집
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• 2004.11a
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• pp.68-69
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• 2004

• IMMUNE NETWORK
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• v.5 no.3
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• pp.157-162
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• 2005

Background: 1-8D gene is a member of human 1-8 interferon inducible gene family and was shown to be overexpressed in fresh colon cancer tissues. Three peptides 1-6, 3-5 and 3-7 derived from human 1-8D gene were shown to have immunogenicity against colon cancer. Methods: To study tumor immunotherapy, of three peptides we established an active immunization model using HHD mice. $D^{b-/-}{\times}{\beta}2$ microglobulin $({\beta}2m)$ null mice transgenic for a chimeric HLA-$A2.1/D^{b-}\;{\beta}2m$ single chain (HHD mice) were challenged with B16/HHD/1-8D tumor cells and were immunized with irradiated peptide-loaded RMA- S/HHD/B7.1 transfectants. In therapy model tumor growth was retarded in HHD mice that were injected with 3-5 peptide-loaded RMA-S/HHD/B7.1. In survival test vaccination with 1-8D-derived peptide protects HHD mice from tumor progression after tumor challenge. Results: These studies show that peptide 3-5 derived from 1-8D gene can be the most effective candidate for the vaccine of immunotherapy against colon cancer and highlight 1-8D gene as putative colon carcinoma associated antigens. Conclusion: We demonstrated that RMA-S/HHD/ B7.1 loaded with 1-8D peptides, especially 3-5, immunization generates potent antitumor immunity against tumor cells in HHD mice and designed active immunization as proper immunotherapeutic protocols.

• Korean Journal Plant Pathology
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• v.13 no.4
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• pp.248-254
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• 1997

Expression vector (pGEX-Tu) for the coat protein (CP) gene of turnip mosaic virus Ca strain (TuMV-Ca) was constructed by incorporation of TuMV CP gene into pGEX-KG vector which had ${\beta}$-galactosidase gene and IPTG (isopropylthio-${\beta}$-D-galactoside) induction site. The results of ELISA and western hybridization indicated that optimal condition of the expression were when IPTG and western hybridization indicated that optimal condition of the expression were when IPTG induction was carried out on YTA medium with ampicillin in 2 hours after the E. coli seed inoculation ($A_{595}$=0.1/ml). TuMV CP gene was expressed with GST (Glutathion S-Transferase) gene fusion system, and the size of fusion protein was estimated to be 59kDa, for TuMV CP was 33 kDa and GST was 26 kDa.

• The Journal of Korean Medicine
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• v.31 no.4
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• pp.79-100
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• 2010

Objective: Production of ROS from glucose toxicity results in injury of pancreatic $\beta$-cells in diabetes models. This study was undertaken to examine the influence of Lespedeza Cuneata extract (LCE) on cytoprotective effects on glucose toxicity, insulin secretion and gene expression in RIN-m5F cells. Methods: First, we measured LCE's antioxidant activity by DPPH free radical-scavenging activity and SOD activity. After the various concentrations of LCE were added to the RIN-m5F cells, we measured cell viability with glucose stimulation by MTT assay and glucose-stimulated insulin secretion. We analyzed gene expression with Agilent whole mouse genome 44K oligo DNA microarray and searched for related pathways in KEGG (Kyoto Encyclopedia of Genes and Genomes). Lastly we measured INS-1, INS-2, INS-R, IRS-1, IRS-2, IRS-3, GLP-1R, and GLP-2R mRNA expression by real time RT-PCR. Results: Free radical-scavenging activity, SOD activity and insulin secretion increased dependent on LCE concentration, but LCE did not show considerable cytoprotective effect on RIN-m5F cells. More than twice expressed gene was 6362 in Oligo DNA chip. In KEGG, the most related pathway was the metabolic pathway. In the insulin signaling pathway, up expressed genes were Irs1, Mapk8, Akt1, and Lipe and down expressed genes were Rhoq, Fbp2, Prkar2b, Gck, and Prkag1. In real time RT-PCR, IRS-2, and IRS-3 expression increased significantly compared to the control group on LCE $12{\mu}g/m{\ell}$ concentration and GCK expression decreased significantly compared to the control group. Conclusions: These results show that LCE encourages insulin secretion and insulin metabolism by complicated gene mechanisms. Further mechanism study and clinical study seem to be necessary about Lespedeza Cuneata.

• Korean Journal of Animal Reproduction
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• v.27 no.4
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• pp.317-324
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• 2003

The present study were performed to analysis the hematocrit and the red blood cells content into the blood plasma of the transgenic pigs harboring recombinent human erythropoietin gene (rhEPO). Mouse whey acidic protein (mWAP) linked to rhEPO gene was microinjected into pronuclei of porcine one-cell zygotes. After delivered of offspring, PCR analyses identified one mWAP-rhEPO transgenic founder offspring(F$_{0}$). The first generation of transgenic pig (F$_{0}$) harboring mWAP-hEPO appeared to be a male, and the second generation (F$_1$) pigs were made by natural mating of F$_{0}$ with domestic swine, and male and female transgenic pigs (F$_1$) were identified by PCR. The blood samples from transgenic and normal pigs were collected for 50 days during lactation and were counted the red blood cell (RBC) numbers and Hematocrit (HCT) content into the blood. The transgenic pigs expressing rhEPO in their blood gave rise to higher RBC numbers and HCT contents than control animals. rhEPO was secreted both in the blood and milk of genetically engineered pigs harboring rhEPO gene. Therefore, this study provides a model regarding the production of transgenic pig carrying hEPO transgene for biomedical research.earch.

• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.68-72
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• 2002

While the biosynthetic gene cluster encoding the pigmented antibiotic actinorhodin is present in the two closely related bacterial species, Streptomyces lividans and Streptomyces coelicolor, it normally is expressed only in S. coelicolor---generating the deep blue colonies responsible for the S. coelicolor name. However, multiple copies of the afsR2 gene, which activates actinorhodin synthesis, result in the ability of S. lividansto also synthesize large amounts of actinorhodin. Here we report that the phenotypic property that historicially distinguishes these two Streptomycesspecies is determined conditionally by the carbon source used for culture. Whereas growth on glucose repressed actinorhodin production in S. lividans, culture on solid media containing glycerol as the sole carbon source dramatically increased the expression of afsR2 mRNA---leading to extensive actinorhodin synthesis by S. lividansand obliterating its phenotypic distinction from S. coelicolor. afsR2 transcription under these conditions was developmentally regulated, rising sharply at the time of aerial mycelium formation and coinciding temporally with the onset of actinorhodin production. Our results, which identify media-dependent parallel pathways that regulate actinorhodin synthesis in S. lividans, demonstrate carbon source control of actinorhodin production through the regulation of afsR2 mRNA synthesis. The nucleotide sequences of afsR2 revealed two putative important domains; the domain containing direct repeats in the middle and the domain homologous to sigma factor sequence in the C-terminal end. In this work, we constructed various sized afsR2-derivatives and compared the actinorhodin stimulating effects in S. lividans TK21. The experimental data indicate that the domain homologous to sigma factor sequence in the C-terminal end of afsR2 plays a critical role as an antibiotic stimulating function. In addition, we also observed that the single copy integration of afsR2 regulatory gene into S. lividans TK21 chromosome significantly activates antibiotic overproduction.

• Journal of Veterinary Clinics
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• v.17 no.2
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• pp.464-469
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• 2000

An epizootic of calf diarrhea occurred in a Korean native cattle farm located in Chonbuk province. The calves that had either bloody or watery diarrhea were 1 to 30 days old. Some of these animals died during the acute course of the disease. Five calves with predominant clinical signs were examined in more detail. Hematological and serum chemical findings were suggestive of dehydration and nutritional insufficiency. Fecal material from the calve was cultured on/in brilliant green agar (BGA), xylose-lysine deoxycholate (XLD) medium, MacConkey agar, eosin methylene blue (EMB) agar and triple sugar iorn (TSI) A bacteria was isolated. which was subsequently identificed as belonging to Salmonella spp. To differentiate Salmoenlla serotype, rfbs gene of S. dublin was amli- find (720 bp) by multiplex (PCR). The rfbS gene sequences of S, dublin ficld isolate(SDC-1) was com- pared with that off S. dublin(S-37) S, dublin(Ahn et al, 1996), S enteritidis(Ahn et al 1996)and S. typhi (Generbak accession No M29682). The identities of nucleotide sequences were 100%. 99.6%, 99.6%, 97.5% respectively.

• Clinical and Experimental Reproductive Medicine
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• v.24 no.1
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• pp.57-65
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• 1997

Cytogenetic observations of loss of the distal portion of the Y chromosome long arm were found to be associated with disrupted spermatogenesis. The existence of a gene involved in the regulation of spermatogenesis, the azoospermia factor (AZF), was postulated. In this study, we screened the AZF region including DAZ and DAZH genes and observed the expression pattern of DAZ and DAZH transcript in infertile men with azoospermia and oligospermia by using a sequence-tagged site (STS)-based PCR method. PCR primers were synthesized for 11 STSs that span Yq interval 6, SRY, DAZ, and DAZH, functional DAZ homologue on chromosome 3. Microdeletions were detected in 4/32 (12.5%) azoospermic men and 1/11 (9%) severe oligospermic men. Only 2 of 5 patients had microdeletions of Yq that contained the DAZ gene, whereas the other 3 patients had deletions extending from intervals 5L-6F proximal to the DAZ gene on Yq. Testis biopsies of the azoospermic patients revealed a variety from Sertoli cell-only syndrome to testicular maturation arrest. Of 4 men with clinical data available, average testis size was R: 13.8 cc, L: 13.8 cc, serum T was $4.0{\pm}1.25$ ng/ml, LH was $3.63{\pm}1.90$ mIU/ml, and FSH was $8.85{\pm}5.13$ mIU/ml. These values did not differ significantly from the remainder of the patients tested. We could not observed the DAZ transcript in 2 patients, who have no mature spermatozoa. In 11.6% of patients microdeletions of the AZF could be detected. These deletions in the AZF region seem to be involved causing spermatogenic failure. But the frequency of microdeletions proximal to DAZ suggests that DAZ is not the only gene associated with spermatogenic failure.