• Reproductive and Developmental Biology
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• v.30 no.3
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• pp.181-187
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• 2006

In recent years the number of patients waiting for organ transplantation has greatly outpaced the supply of human organs available, which leads to a renewed interest in pig-to-human xenotransplantation as an alternative. However, one of the biggest barriers in the xenotransplantation is presence of porcine endogenous retroviruses(PERV) that can infect human cells. In this study, to present a possible solution for this problem we tried to inhibit expression of PERVs using shRNAs(short hairpin RNA) at the level of RNA synthesis and virus release. The shRNA targeting the sequence of PERV A, B type was cloned into pSIREN-RetroQ vector under the control of polymerase-III U6-RNA gene promoter. Quantitative real-time PCR was performed to detect my alterations in mRNA production of PERV A, B targeted by the shRNA in each done. Depending on the target sequence of the shRNA, the transcription of PERV was decreased to as much as 4% and the number of progeny viruses was reduced to less than 1/200,000. Transgenic pigs producing such shRNAs may result in a highly reduced PERV expression in cells and organs, which is a prerequisite for safe xenotransplantations.

• Reproductive and Developmental Biology
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• v.30 no.3
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• pp.157-162
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• 2006

Endogenous 84 amino acid parathyroid hormone(PTH) is synthesized as a pre-pro hormone by the chief cells of the parathyroid glands. Physiological actions of PTH include regulation of bone metabolism, renal tubular reabsorption of calcium and phosphate, and intestinal calcium absorption. In addition, PTH stimulates new bone formation by extraordinary stimulation of osteoblastic activity and decreasing calcium excretion by the kidney. In this study, we constructed and tested retrovirus vectors designed to express the human parathyroid hormone(hPTH) gene under the control of the tetracycline-inducible promoters. To increase the hPTH gene expression at turn-on state, woodchuck hepatitis virus posttranscriptional regulatory element(WPRE) sequence was also introduced into retrovirus vector at downstream region of either the hPTH gene or the sequence encoding reverse tetracycline-controlled transactivator(rtTA). Transformed primary culture cells(porcine fetal fibroblast, PFF, chicken embryonic fibroblast, CEF) were cultured in the medium supplemented with or without doxycycline(tetracycline derivative) for 48 hours, and induction efficiency was measured by comparing the hPTH gene expression level using two step RT-PCR and ELISA Higher hPTH expression($3{\tims}10^4\;pg/ml,\;5.3{\times}10^4\;pg/ml$) and tighter expression control(up to 8 fold) were observed from the vector in which the WPRE sequence was placed at downstream of the hPTH gene. The resulting tetracycline inducible vector system may be helpful in solving serious physiological disturbance problems which have been a major obstacle in successful production of transgenic animals.

• Journal of Veterinary Clinics
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• v.24 no.2
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• pp.94-98
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• 2007

This study was to investigate the effects of ascorbic acid and alpha-tocopherol on the attenuation of renal ischemia-reperfusion (IR) injury in pigs. Ten pigs were subjected to 60 minutes of warm unilateral renal ischemia followed by removal of contralateral kidney and then divided into two groups. Treatment group was performed ascorbic acid and alpha-tocopherol pretreatment 2 days before operation and ascorbic acid with heparin-saline solution irrigation-aspiration. Otherwise, control group used only irrigation-aspiration of heparin-saline solution. Blood samples were collected from these pigs for measurement of serum blood urea nitrogen (BUN) and creatinine values, antioxidant superoxide dismutase (SOD) at pre, day 1, day 3, day 7 and day 14. The kidneys were taken for histopathologic evaluation after euthanasia on postoperative day 14. The levels of BUN were significantly increased in the control group on day 1, day 3 and day 7 (P<0.05). And the level of creatinine was significantly increased in the control group on day 3 (p<0.05). Activity of antioxidant enzymes in plasma revealed significant difference (p<0.05) between control and treatment group at day 14. In histopathologic findings, treatment group was showed less damage than that of control group on the basis of renal tubular damage. It was concluded that ascorbic acid and alpha-tocopherol attenuated renal I/R injury in the pigs.

• Journal of Veterinary Clinics
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• v.24 no.2
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• pp.104-108
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• 2007

The purpose of the study is to evaluate the clinical antagonistic effect of atipamezole(0.25 mg/kg, IM) in cats anesthetized with tiletamine-zolazepam ($Zoletil^{(R)}$, 10 mg/kg, IM) and medetomidine (0.05 mg/kg, IM). Twelve healthy 1 year old Korean mixed breed cats were used for this study. They were 4 males and 8 females. These cats were randomly assigned to two groups. One was control group ($Zoletil^{(R)}$ + medetomidine, ZM), and the other was treatment group ($Zoletil^{(R)}$ + medetomidine and antagonism by atipamezole, ZMA). All cats were examined 15 minutes before, 5, 25, 65 and 105 minutes after administration of tiletamine-zolazepam and medetomidine. Atipamezole was injected intramuscularly 20 minutes after ZM administation. Recovery time, heart rate, respiratory rate, total plasma protein and blood glucose were significantly different between ZM group and ZMA group (P<0.05). However, rectal temperature was not significantly different between ZM group and ZMA group. Two groups were able to induce sternal recumbency within 2 minutes and lateral recumbency within 4 minutes after the anesthetics injection. Mean sternal position time ($mean{\pm}SD$) was $174.0{\pm}44.6\;and\;116.2{\pm}27.3$ minutes, and mean standing position time was $210.8{\pm}45.6\;and\;154.2{\pm}21.1$ minutes in ZM and ZMA group, respectively. In these two groups, adverse effects during recovery time from anesthesia were not seen. As a result, the ZMA group had a faster recovery than the ZM group. Thus it was concluded that atipamezole could exert a useful reversal effect in cats anesthetized with medetomidine-tiletamine/zolazepam combination.

• Korean Journal of Poultry Science
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• v.30 no.4
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• pp.245-251
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• 2003

Three experiments were conducted to investigate the effect of feeding yeast accumulated transgenic ferritin(FRT, Saccharomyces cerevisiae) as a probiotic on the performance, iron contents in the liver, spleen, bone and yolk of laying hens and broiler chicks. Effects of feeding FRT were compared with that of feeding wild-type yeast(W0) and yeast grown on 20 mM ferric citrate-added medium (W20). In Expt 1, to investigate the effect of feeding yeast (control, W0 FRT) on performance and iron content of organs of broiler chicks which were fed basal diet supplemented with 75mg/kg iron(Fe75) or not (Fe0), three hundred sixty one-day-old male broiler chicks were fed a corn-sov based diet for five weeks. Weight gain, feed intake and feed conversion were measured weekly. In Expt 2, fifteen 33-week-old ISA Brown laying hens were placed in individual cages and were fed control, W0 and FRT diets for Four weeks. In Expt 3, twenty four 45-week-old ISA Brown laying hens were placed in individual cages and were fed a basal diet for a week. Then, experimental diets (control, W0, W20, FRT) were fed for three weeks. Iron contents in the liver, heart, spleen and tibia were determined at the end of all experiments. Iron content in yolk was measured weekly (expt 2, 3). The level of yeast added and iron concentration of FRT were $1{\times}10^8$cfu/kg diet and 500 mg/kg cell (DM) respectively in Expt 3, yeast was supplemented at $2{\times}10^{10}$cfu/kg diet and the iron content of FRT was 1000mg/kg cell (DM). In Expt 1. birds fed Fe75 showed significantly higher weight gain compared with Fe0 (P<0.05). However, weight gain and feed intake of birds fed FRT was significantly lower than control (P<0.05). In Expt 2, the iron content of the liver was decreased in the FRT treatment (P<0.05). In Expt 3, iron concentration of the liver and spleen tended to be increased by feeding FRt. However, the iron content of the tibia tended to be decreased in the FRT treatment. These results suggest that feeding FRT as a probiotic cannot improve performance and iron content in organs of broiler chicks and laying hens.

• Journal of Animal Science and Technology
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• v.46 no.4
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• pp.571-584
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• 2004

It is known widely that granulosa cell apoptosis leads follicular atresia and macrophages exert their effects directly and/or indirectly from the initiation to the completion of follicular atresia by phagocytosis of apoptotic bodies and secretion of various cytokines. However, the site of initiation, propagation routes and the elimination methods of apoptotic bodies, and the time and methods of penetration of macrophages into the follicles are not known completely. Using pig(Yorkshire-breed) ovary, immunohistochemical studies with TUNEL for apoptotic bodies and pig macrophage monoclonal antibody 4E9 for macrophages, and light and transmission electron microscopic observations were performed. In the pig, follicular atresia began with the granulosa cell apoptosis, and the apoptosis of theca intema cells occured at the same time. The apoptosis of granulosa cells initiated randomly within the granulosa cell layer and propagated rapidly into the whole layer. Ultrastructura1ly, apoptotic granulosa cells showed characteristic pyknotic and deformed nucleus and intracytoplasmic vesicles. Apoptotic bodies were eliminated by intact granulosa cells and macrophages. Intact granulosa cells ingested apoptotic bodies transiently, soon after they fell into the apoptosis. Finally, apoptotic bodies and degenerated oocyte were phagocytosed by macrophages. Macrophages entered the ovarian follicle at the time of initiation of granulosa cell apoptosis, and migrated with the progression of apoptosis. By elimination of theca cells, macrophages contributed the completion of follicular atresia These results will provide valuable informations on the study of the interrelation between macrophage and ovarian follicular atresia.

• Journal of Plant Biotechnology
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• v.41 no.3
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• pp.146-158
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• 2014

Camelina sativa that belongs to Brassicaceae family is an emerging oilseed crop. Camelina seeds contain approximately 40% storage oils per seed dry weight, which are useful for human and animal diets and industrial applications. Microsomal delta-12 fatty acid desaturase2 (FAD2) catalyzes the conversion of oleic acid to linoleic acid. The polymorphisms of FAD2 genes are correlated with the levels of oleic acids in seed oils. Microsomal delta-12 fatty acid desaturase2 (FAD2) catalyzes the conversion of oleic acid to linoleic acid. The polymorphisms of FAD2 genes are correlated with the levels of oleic acids in seed oils. In this study, three CsFAD2 genes (CsFAD2-1, CsFAD2-2 and CsFAD2-3.1) were isolated from developing seeds of Camelina sativa (L.) cv. CAME. The nucleotide and deduced amino acid sequences of three CsFAD2 genes were compared with those from dicotyledon and monocotyledon plants including Camelina cultivars Sunesone and SRS933. Three histidine motifs (HECGH, HRRHH, and HVAHH) required for FAD activity and a hydrophobic valine or isoleucine residue, which is a SNP (single nucleotide polymorphism) marker related with enzyme activity are well conserved in three CsFAD2s. The expressions of CsFAD2-1 and CsFAD2-3.1 were ubiquitously detected in various Camelina organs, whereas the CsFAD2-2 transcripts were predominantly detected in flowers and developing seeds. The contents of oleic acids decreased, whereas the amounts of linoleic acid increased in dry seeds of transgenic fad2-2 lines expressing each CsFAD2 gene compared with fad2-2 mutant, indicating that three CsFAD2 genes are functionally active. The isolated CsFAD2 genes might be applicable in metabolic engineering of storage oils with high oleic acids in oilseed crops.

• Reproductive and Developmental Biology
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• v.31 no.3
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• pp.161-167
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• 2007

In this study, we constructed and tested retrovirus vectors designed to express the human thrombopoietin gene under the control of the tetracycline-inducible promoters. To increase the hTPO gene expression at him-on state, WPRE sequence was also introduced into retrovirus vector at downstream region of either the hTPO gene or the sequence encoding reverse tetracycline-controlled transactivator (rtTA). Primary culture cells (PFF, porcine fetal fibroblast; CEF, chicken embryonic fibroblast) infected with the recombinant retrovirus were cultured in the medium supplemented with or without doxycycline for 48hr, and induction efficiency was measured by comparing the hTPO gene expression level using RT-PCR, western blot and ELISA. Higher hPTO expression and tighter expression control were observed from the vector in which the WPRE sequence was placed at downstream of the hTPO (in CEF) or rtTA(in PFF) gene. This resulting tetracycline inducible vector system may be helpful in solving serious physiological disturbance problems which have been a major obstacle in successful production of transgenic animals.

• Journal of The Korean Society of Grassland and Forage Science
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• v.24 no.4
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• pp.341-346
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• 2004

To determine lethal temperature of birdsfoot trefoil(BFT) and italian ryegrass(IRG) at heat-stressed conditions, seedlings grown in a small pots fur 4 weeks were subjected to different temperature regimes of heat treatment. No apparent damage was observed BFT and IRG were treated at 45, 50 or $60^{\circ}C$ for 1 h. And also heat treatments at 60, 65 and $70^{\circ}C$ for 1 h, both of them were withered and showed damage symptom on their leaves but it was not lethal conditions for the whole plants. By contrast, most of plants were prominently withered within one day after heat treatment at $80^{\circ}C/60min$. When BFT was exposed to $80^{\circ}C/60$ min, they were died within 6 days but there was found that new shoots were regenerated from the plants that had been treated at $80^{\circ}C$ within 55 min. IRC was also died within 2 days that exposed to $80^{\circ}C/20$ min but there was found that new shoots were regenerated from the plants that had been treated at $80^{\circ}C$ within 15 min. These results indicate that heat killing temperatures of BFT and IRG plants are $80^{\circ}C/60$ min an $80^{\circ}C/20$ min respectively. Simple viability assay system established in this study will be useful for selection and characterization of heat-tolerant transgenic BFT and IRG plants.

• Journal of Veterinary Clinics
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• v.26 no.6
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• pp.568-573
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• 2009

The purpose of this study is to evaluate the analgesic effects on the oral cavity in dogs which are treated with xylazine and electroacupuncture (EA). Furthermore, this study aims to find out its effects on glucose (GLU), serum alanine aminotransferase (ALT), and blood urea nitrogen (BUN) variation values, vital signs (rectal temperature, heart rate, respiratory rate) and pain responses to the noxious stimuli. Eight healthy dogs were randomly assigned to receive either xylazine or EA. Xylazine group dogs with weight of 3.6${\pm}$1.0 kg received 1.5 mg/kg of xylazine intramuscularly. EA group dogs with weight of 3.9${\pm}$1.0 kg received 1 volt (10-15 hz) for 5 minutes, and then 1-9 Volts (25-30 hz) for 60 minutes totally. The acupoints used were LI-3 (San Jian), LI-4 (He Gu) and ST-7 (Xia Guan). All dogs were examined before and 10, 25, 40, 55 and 120 minutes after administration of xylazine or EA. The mean rectal temperatures of the EA group were significantly higher than those of xylazine group after 25, 40 and 55 minutes (p < 0.05). The mean heart rates of the EA group were significantly higher than those of xylazine group after 10, 25, 40 and 55 minutes (p < 0.05). The mean respiratory rates of the EA group were significantly higher than those of xylazine group after 55 and 120 minutes (p < 0.05). The mean GLU concentration of the EA group were significantly lower than those of xylazine group after 55 and 120 minutes (p < 0.05). The sum of mean pain scores (SMPS) of the EA group were significantly higher than those of xylazine group after 10, 25 40 and 55 minutes (p < 0.05). In this study, the pain control of the EA group was shown to be better than that of the xylazine group. Also, there do not appear to be any negative physiologic effects associated with acupuncture-induced surgical analgesia. So, it was considered that these acupoints of EA analgesia might be useful for minor oral surgery in weak patients.