• Genomics & Informatics
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• v.3 no.1
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• pp.24-29
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• 2005

TRPV2 is a non-specific cation channel expressed in sensory neurons, and activated by noxious heat. Particularly, TRPV2 has six transmembrane domains and three ankyrin repeats. TRPV2 has been cloned from various species such as human, rat, and mouse. Oocytes of Xenopus laevis - an African clawed frog ­have been widely used for decades in characterization of various receptors and ion channels. The functional property of rat TRPV2 was also identified by this oocyte expression system. However, no TRPV2 orthologue of Xenopus laevis has been reported so far. Hence, we have focused to clone a TRPV2 orthologue of Xenopus laevis with the aid of bioinformatic tools. Because the genome sequence of Xenopus laevis is not available until now, a genome sequence of Xenopus tropicalis - a close relative species of Xenopus laevis - was used. After a number of bioinformatic searches in silico, a predicted full-length sequence of TRPV2 orthologue of Xenopus tropicalis was found. Based on this predicted sequence, various approaches such as RT-PCR and 5' -RACE technique were applied to clone a full length of Xenopus laevis TRV2. Consequently, a full-length Xenopus laevis TRPV2 was cloned from heart cDNA.

• Journal of Environmental Science International
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• v.18 no.11
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• pp.1247-1259
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• 2009

We investigated the toxic effects of carbaryl on early embryo development in the African clawed frog, Xenopus laevis. To test the toxic effects, frog embryo teratogenesis assays using Xenopus were performed. Embryos were exposed to various concentrations of carbaryl ($5{\sim}320\;{\mu}M$). $LC_{100}$ for carbaryl was $320\;{\mu}M$, and the $LC_{50}$ determined by probit analysis was the concentration of $235.68\;{\mu}M$. Exposure to $160\;{\mu}M$ of carbaryl resulted in 10 different types of severe external malformations. Histological examination revealed dysplasia of the eyes, heart, guts, somatic muscle, dorsal, liver, blood vessel and swelling of the pronephric ducts. Malformation of neural tissue and brain was not severe even in the high dose of carbaryl. Benzidine blood stain showed distinct inhibition of inducing erythrocytes in embryos and animal cap explants. Electron micrographs of embryo revealed retinal detachment, loose photoreceptor lamella and the degeneration of sarcomeres in the carbaryl-treated group. The mitochondrial degeneration was also observed in the test group.

• Journal of Environmental Science International
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• v.20 no.10
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• pp.1221-1232
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• 2011

We investigated the toxic effects of difenoconazole on the development in the African clawed frog, Xenopus laevis. To test the toxic effects, frog embryo teratogenesis assays using Xenopus were performed. Embryos were exposed to various concentrations of difenoconazole (0-30 ${\mu}M$). $LC_{100}$ for difenoconazole was 30 ${\mu}M$, and the $LC_{50}$ determined by probit analysis was 27.19 ${\mu}M$. Exposure to difenoconazole concentrations ${\geq}$5 ${\mu}M$ resulted in 10 different types of severe external malformation. Histological examinations revealed dysplasia of the eye, heart, liver, somatic muscle, and swelling of the pronephric ducts. The tissue-specific toxic effects were investigated with an animal cap assay. Blood cells were normally induced at a high frequency by mSCF and activin A. However, the induction of blood cells was strongly inhibited by the addition of difenoconazole. Electron micrographs of tested embryos showed the degeneration of somatic muscle and the shrinkage of microvilli on pronephric duct. The gene expression of cultivated animal cap explants was investigated by reverse transcriptase-polymerase chain reaction (RT-PCR). It revealed that the expression of the blood-specific marker(${\beta}$-globin II) and muscle-specific marker (XMA) were more strongly inhibited than the neural-specific marker(XEn2) by the addition of difenoconazole.

• Journal of Environmental Science International
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• v.19 no.8
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• pp.1001-1012
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• 2010

We investigated the toxic effects of tebuconazole on development in the African clawed frog, Xenopus laevis. To test the toxic effects, frog embryo teratogenesis assays using Xenopus were performed. Embryos were exposed to various concentrations of tebuconazole($0-100\;{\mu}M$). $LC_{100}$ for tebuconazole was $100\;{\mu}M$, and the $LC_{50}$ determined by probit analysis was $82.35\;{\mu}M$. The exposure to tebuconazole concentrations ${\geq}40\;{\mu}M$ resulted in 11 different types of severe external malformations including gut dysplasia. Histological examinations revealed various dysplasia in the eye, heart, liver, intestine, somatic muscle, and in the pronephric ducts. The tissue-specific toxic effects were investigated with an animal cap assay. Blood cells are generally induced at a high frequency by the combination of mSCF and activin A, however, the induction of blood cells was strongly inhibited by the addition of tebuconazole. Electron micrographs of tested embryos showed many of multivesicular bodies and dysplasia of photo-receptive cell, however, the somatic muscle degeneration was not severe. The gene expression of cultivated animal cap explants was investigated by reverse transcriptase-polymerase chain reaction and revealed that expression of the blood-specific marker, $\beta$ globin II and muscle-specific marker, muscle actin were more strongly inhibited than the neural-specific marker, XEn2.

• Environmental Analysis Health and Toxicology
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• v.16 no.1
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• pp.29-34
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• 2001

The estrogenic potency of di -2-ethylhexyl phthalate (DEHP) using reverse transcriptase-PCR respouse of liver vitellogenin mRNA in male African clawed frog (Xenopus laevis) was studied. Male frogs were injected with DEHP at dose of 300$\mu\textrm{g}$/kg and 300 mg/kg body weight through the dorsal lymph sac. After 4 days, using suitable pair of RT-PCR primers, vitellogenin mRNA induction in the liver was measured and DEHP showed vitellogenin mRNA induction in only the group treated with 300 mg/kg. Any significant histological abnormalities by the exposure of DEHP was not shown in both testis and liver.

• Journal of Pharmaceutical Investigation
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• v.25 no.4
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• pp.299-305
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• 1995

Cloning the gene encoding a dipeptide transporter is necessary for understanding the absorption mechanism of peptides and peptide-like drugs in the gastrointestinal tract. Functional expression of a dipeptide transporter after microinjection into Xenopus laevis oocytes was performed using the mRNA purified from human intestinal HT-29 cells. Fifty nanoliters of purified mRNA (1 mg/mL) were microinjected into healthy oocytes followed by incubation for 4 days in order to express a dipeptide transporter. Functional expression was determined by a uptake assay using 10 Ci/mL $[^3H]-glycylsarcosine$, a dipeptide substate of the transporter. Seasonal variability and batch-to-batch variability were greater in summer. The usage of beveled micropipettes improves viability of oocytes at 4 days after microinjection. Expression of a dipeptide transporter in oocytes after microinjection of mRNA obtained from HT-29 cells was significantly larger than those after microinjection of water or mRNA obtained from the rabbit intestine.

• Journal of Korean Ophthalmic Optics Society
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• v.12 no.3
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• pp.151-158
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• 2007

In this study we investigated the embryo toxicity of three kinds of plant extracts during early development of African clawed frog, Xenopus laevis through FETAX assay (Frog Embryo Teratogenesis Aassay with Xenopus). The plants used in this study were the materials of the Korean herbal medicines, Polygala tenuifolia, Lycium chinensis and Comus officinalis. The test embryos exposed to 1, 10 and $100{\mu}g/ml$ of each plant extract and control embryos were incubated for 96h at $24{\pm}0.5^{\circ}C$. The focus of this study is to elucidate the malformation due to toxicity of plant extracts, especially, to elucidate plant inducing optic malformation. As a result, the growth inhibition of embryos, optic malformation, axial distortion, cephalic and abdominal edema, dysplasia of digestive track and hyper-pigmentation were occurred in all of extracts, and these malformations were increased to the increase of extract concentration. The rate of optic malformation was highest in $100{\mu}g/ml$ of Lycium chinensistreated group and 27% of tested 150 individuals showed optic hernia. The histological results showed enlarged ventriculum in brain, dysplasia of vitreous chamber in eye and unclear retinal layers.

• Korean Journal of Ecology and Environment
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• v.36 no.1 s.102
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• pp.83-94
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• 2003

Concentrated releases of zinc into water usually results from discharges associated with industrial purpose. The released zinc into soil is corroded and released into water. In aquatic environment, exess zinc is toxic to the organisms and causes the growth inhibition and malformation of them as a heavy metal. In this study, excess zinc toxicity was tested by FETAX (frog embryo teratogenetic assay with Xenopus)as in vivo system. Xenopus embryos at st.9 were exposed to $100{\sim}900\;{\mu}M$ of zinc for 7 days and 81% of individuals were survived in 100 ${\mu}M$, and 25% were survived in 1000M of zinc solution. In external malformations, swelled belly and intestinal dysplasia were common, and all of tested individuals showed these malformations in 200 ${\mu}M$ or higher concentration of zinc. In 400 ${\mu}M$ or higher concentration, all of tested tadpoles showed faded heart. Also, hypo-pigmentation, lens hernia and loose digestive track were very frequently found in 100 ${\mu}M$ of zinc. The histological study with paraffin section of zinc treated tadpoles showed following abnormalities; regeneration of photoreceptor on retina, reduced vitreous chamber in eye, reduction of red blood cells in heart, abnormal liver, swelling of pronephric cell, muscle dysplasia and palatal papilloma. These abnormalities may be caused by the degeneration of mitochondria, inhibition of cell adhesion, and the formation of leghemoglobin by zinc due to the substitution of $Ca^{2+}$ by $Zn^{2+}$. The body length was reduced due to the excess zinc. From a statistical result, body lengths of 300 ${\mu}M$ or higher concentrative g개ups was significantly reduced comparing that of control group. Recently, many spontaneous malformations and reduction of amphibians are reported, From the results of present study, excess zinc mi호t be a factor of amphibian reduction, and the control of zinc discharges is very important.

• Toxicological Research
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• v.11 no.1
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• pp.37-42
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• 1995

Effect of cadmium on the early amphibian development was analyzed through FETAX (Frog Embryo Teratogenesis Assay: Xenopus). Embryos manifested concentration-dependent mortality and realformations; shortage of anterior-posterior axis gut realformation, ocular anomalies, bent notochord, misshapen dorsal fin, and derreal blisters. The treatment with 1.5ppm cadmium solution caused 100% mortality and concentration of lppm did not kill the embryos that caused 100% anomaly. The teratogenic index (TI = LC50 /EC50) was 2.8 indicating that $CdCl_2$ is teratogenic for Xenopus laevis. Embryos that were pulsetreated with at early to late blastula stage (St. 3-9) and mid to late blastula stage (St. 6-10) showed relatively strong resistance to cadmium, but the embryos treated at gastrula stage (St. 10-13) showed high mortality. And the embryos treated at tailbud stage (after St. 25) showed highest mortality of any other early stages. Effects of temperature were studied through pulse- treatment during gastrula stage at $20^{\circ}C$ and $30^{\circ}C$. The embryos treated with 7.5ppm at $30^{\circ}C$ and 15ppm at $20^{\circ}C$ caused 100% mortality respectively, indicating that higher temperature had more severe toxic effect. One of the most peculiar effect of cadmium at gastrulation was distortion of the tail. The probable cause of toxic effect of Cd was discussed.

• Journal of Pharmaceutical Investigation
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• v.23 no.3
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• pp.31-40
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• 1993

It is essential to clone the peptide transporter in order to obtain better understanding of its molecular structure, regulation, and substrate specificity. Characteristics of an endogenous peptide transporter in oocytes were studied along with expression of an exogenous proton/peptide cotransporter from rabbit intestine. And further efforts toward cloning the transporter were performed. The presence of an endogenous peptide transporter was detected in Xenopus laevis oocytes by measuring the uptake of $0.25\;{\mu}M\;(10\;{\mu}Ci/ml)\;[^3H]-glycylsarcosine$ (Gly-Sar) at pH 5.5 with or without inhibitors. Uptake of Gly-Sar in oocytes was significantly inhibited by 25 mM Ala-Ala, Gly-Gly, and Gly-Sar (p<0.05), but not by 2.5 mM of Glu-Glu, Ala-Ala, Gly-Gly, Gly-Sar and 25 mM glycine and sarcosine. This result suggests that a selective transporter is involved in the endogenous uptake of dipeptides. Collagenase treatment of oocytes used to strip oocytes from ovarian follicles did not affect the Gly-Sar uptake. Changing pH from 5.5 to 7.5 did not affect the Gly-Sar uptake significantly, suggesting no dependence of the endogenous transporter on a transmembrane proton gradient. An exogenous $H^+/peptide$ cotransporter was expressed after microinjection of polyadenylated messenger ribonucleic acid $[poly\;(A)^+-mRNA]$ obtained from rabbit small intestine. The Gly-Sar uptake in mRNA-injected oocytes was 9 times higher than that in water-injected oocytes. Thus, frog oocytes can be utilized for expression cloning of the genes encoding intestinal $H^+/peptide$ cotransporters. Using the technique size fractionation of mRNA was sucessfully obtained.