Effect of the environmental quality and food chain on trace element concentrations in Heron and Egret chicks at Pyeongtaek colony, Korea.

Kim, Jungsoo,Oh, Jong-Min Chapman Hall ; Kluwer Academic Publishers 2014 Ecotoxicology Vol.23 No.7

<P>Trace element concentrations in the diet can affect the levels in birds. Heron and egret chicks have been recommended as useful biological indicators for monitoring trace element contamination in the aquatic ecosystem. Iron, zinc, manganese, copper, lead and cadmium concentrations were measured in the livers and stomach contents of Grey Heron (Ardea cinerea), Intermediate Egret (Egretta intermedia), Little Egret (Egretta garzetta) and Black-crowned Night Heron (Nycticorax nycticorax) chicks from Pyeongtaek colony, Korea in 2008. Iron, zinc, manganese, copper and cadmium concentrations in livers differed among four heron and egret species. Stomach content concentrations of iron, zinc, copper, lead and cadmium concentrations differed among the four species. Essential elements such as iron, zinc, manganese and copper concentrations were within the range of other heron and egret studies and these levels may be regulated by a normal homeostatic mechanism. Two of 11 (18.2%) Grey Heron and two of 10 (20.0%) Black-crowned Night Heron chicks exceeded the background lead level (<6 ?g/g dw) for birds and were at a level considered lead exposed (6-30 ?g/g dw). Cadmium concentrations did not exceed the background levels for wild birds. In livers of combined heron and egret chicks, lead, but not cadmium concentrations were associated with concentrations in the stomach contents. These results suggest that lead concentrations in chicks can reflect qualities of feeding area such as streams, rivers and paddy fields and chicks represent a bioindicator to evaluate lead contamination in aquatic ecosystems.</P>

Generation of CMAHKO/GTKO/shTNFRI-Fc/HO-1 quadruple gene modified pigs

Kim, G. A.,Lee, E. M.,Jin, J. X.,Lee, S.,Taweechaipaisankul, A.,Hwang, J. I.,Alam, Z.,Ahn, C.,Lee, B. C. Chapman & Hall 2017 Transgenic research Vol.26 No.4

<P>As an alternative source of organs for transplantation into humans, attention has been directed to pigs due to their similarities in biological features and organ size. However, severe immune rejection has prevented successful xenotransplantation using pig organs and tissues. To overcome immune rejection, recently developed genetic engineering systems such as TALEN coupled with somatic cell nuclear transfer (SCNT) to make embryos could be used to produce pigs compatible with xenotransplantation. We used the TALEN system to target the non-Gal antigen cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) gene in pigs that is naturally deleted in humans. Gal-deleted cells expressing both soluble human tumor necrosis factor receptor I IgG(1)-Fc (shTNFRI-Fc) and human hemagglutinin -tagged-human heme oxygenase-1 (hHO-1) were transfected with a TALEN target for CMAH. Cells lacking CMAH were negatively selected using N-glyconeuraminic acid (Neu5Gc)/magnetic beads and the level of Neu5Gc expression of isolated cells were analyzed by FACS and DNA sequencing. Cloned embryos using 3 different genetically modified cell clones were respectively transferred into 3 recipients, with 55.6% (5/9) becoming pregnant and three cloned pigs were produced. Successful genetic disruption of the CMAH gene was confirmed by sequencing, showing lack of expression of CMAH in tail-derived fibroblasts of the cloned piglets. Besides decreased expression of Neu5Gc in piglets produced by SCNT, antibody-mediated complement-dependent cytotoxicity assays and natural antibody binding for examining immuno-reactivity of the quadruple gene modified pigs derived from endothelial cells and fibroblasts were reduced significantly compared to those of wild type animals. We conclude that by combining the TALEN system and transgenic cells, targeting of multiple genes could be useful for generating organs for xenotransplantation. We produced miniature pigs with quadruple modified genes CMAHKO/GTKO/shTNFRI-Fc/hHO-1 that will be suitable for xenotransplantation by overcoming hyperacute, acute and anti-inflammatory rejection.</P>

Differences between immunodeficient mice generated by classical gene targeting and CRISPR/Cas9-mediated gene knockout

Lee, Jae Hoon,Park, Jong-Hyung,Nam, Tae-Wook,Seo, Sun-Min,Kim, Jun-Young,Lee, Han-Kyul,Han, Jong Hyun,Park, Song Yi,Choi, Yang-Kyu,Lee, Han-Woong Chapman & Hall 2018 Transgenic research Vol.27 No.3

<P>Immunodeficient mice are widely used for pre-clinical studies to understand various human diseases. Here, we report the generation of four immunodeficient mouse models using CRISPR/Cas9 system without inserting any foreign gene sequences such as Neo<SUP>R</SUP> cassettes and their characterization. By eliminating any possible effects of adding a Neo<SUP>R</SUP> cassette, our mouse models may allow us to better elucidate the in vivo functions of each gene. Our FVB-<I>Rag2</I><SUP>−/−</SUP>, B6-<I>Rag2</I><SUP>−/−</SUP>, and BALB/c-<I>Prkdc</I><SUP>−/−</SUP> mice showed phenotypes similar to those of the earlier immunodeficient mouse models, including a lack of mature B cells and T cells and an increase in the number of CD45<SUP>+</SUP>DX-5<SUP>+</SUP> natural killer cells. However, B6-<I>Il2rg</I><SUP>−/−</SUP> mice had a unique phenotype, with a lack of mature B cells, increased number of T cells, and decreased number of natural killer cells. Additionally, serum immunoglobulin levels in all four immunodeficient mouse models were significantly reduced when compared to those in wild-type mice with the exception of IgM in B6-<I>Il2r</I>g<SUP>−/−</SUP> mice. These results indicate that our immunodeficient mouse models are a robust tool for in vivo studies of the immune system and will provide new insights into the variation in phenotypic outcomes resulting from different gene-targeting methodologies.</P><P><B>Electronic supplementary material</B></P><P>The online version of this article (10.1007/s11248-018-0069-y) contains supplementary material, which is available to authorized users.</P>

Generation by somatic cell nuclear transfer of GGTA1 knockout pigs expressing soluble human TNFRI-Fc and human HO-1

Kim, Geon A,Lee, Eun Mi,Cho, Bumrae,Alam, Zahid,Kim, Su Jin,Lee, Sanghoon,Oh, Hyun Ju,Hwang, Jong Ik,Ahn, Curie,Lee, Byeong Chun Chapman & Hall 2019 Transgenic research Vol.28 No.1

Evaluation and selection of reference genes for ecotoxicogenomic study of the green alga Closterium ehrenbergii using quantitative real-time PCR.

Lee, Min-Ah,Guo, Ruoyu,Ebenezer, Vinitha,Ki, Jang-Seu Chapman Hall ; Kluwer Academic Publishers 2015 Ecotoxicology Vol.24 No.4

<P>The green alga Closterium ehrenbergii occurs in fresh water environments and has been suggested as a model for ecotoxicological assessment. Quantitative real-time PCR (qRT-PCR), with its high sensitivity and specificity, is a preferred method for reliable quantification of gene expression levels. qRT-PCR requires reference genes to normalize the transcription level of the target gene, and selection of appropriate references is crucial. Here, we evaluated nine housekeeping genes, that is, 18S rRNA, ACT, TUA, TUB, eIF, H4, UBQ, rps4, and GAPDH, using 34 RNA samples of C. ehrenbergii cultured in various environments (e.g. exposure to heat shock, UV, metals, and non-metallic chemicals). Each housekeeping gene tested displayed different ranges of C T values for each experimental condition. The gene stability was determined using the descriptive statistic software geNorm, which showed that ACT, H4, and TUA were the most suitable reference genes for all the conditions tested. In addition, at least three genes were required for proper normalization. With these references, we assessed the expression level of the heat shock protein 70 (HSP70) gene in C. ehrenbergii cells exposed to thermal and toxic contaminant stress and found that it was significantly up-regulated by these stressors. This study provides potential reference genes for gene expression studies on C. ehrenbergii with qRT-PCR.</P>

Trace element contamination in nestling black-tailed gulls (Larus crassirostris) in Korea.

Kim, Jungsoo,Oh, Jong-Min Chapman Hall ; Kluwer Academic Publishers 2015 Ecotoxicology Vol.24 No.4

<P>At Hongdo Island, Gyeongsangnam-do, Korea, a breeding site of black-tailed gull (Larus crassirostris), we collected nestlings from two locations: a 'reference' site (n?=?10) with no known source of lead contamination and 'lighthouse' site (n?=?10) with suspected lead contamination from leaded paint. Iron concentrations in the kidney and bone, manganese in the muscle, copper in the bone, lead in the muscle and bone, and cadmium in the liver, muscle, and bone at the reference site were significantly higher than at the lighthouse. Manganese concentrations in the liver and kidney, and lead in the kidney were significantly greater at the lighthouse than at the reference site. Iron, zinc, manganese, copper, lead and cadmium concentrations had tissue-specific accumulation at both sites. Lead concentrations in 10?% of livers and in 80?% of kidneys at the lighthouse, and in 20?% of livers from the reference were within a range considered toxic (>6.00?μg/g?dw in the liver and kidney). Lead concentrations in 50?% of black-tailed gull nestlings at the reference and 80?% nestlings at the lighthouse were greater in livers than in bones, which is suggestive of acute lead exposure. For cadmium, all liver and kidney concentrations from two sites were at a level considered background in birds. Cadmium concentrations of every sample were higher in kidneys than in livers, suggestive of chronic cadmium exposure. Lead concentrations in gull nestlings in the present study were relatively higher than other gull species worldwide, but cadmium concentrations were relatively lower.</P>

Differential transcription of heat shock protein 90 (HSP90) in the dinoflagellate Prorocentrum minimum by copper and endocrine-disrupting chemicals.

Guo, Ruoyu,Ki, Jang-Seu Chapman Hall ; Kluwer Academic Publishers 2012 Ecotoxicology Vol.21 No.5

<P>The dinoflagellate algae survive variations in water temperature as well as sudden exposures to toxic substances; heat shock proteins (HSPs) seem to function as part of their cell survival strategy. In the present study, we determined the complete open reading frame (ORF) of HSP90 gene in the dinoflagellate Prorocentrum minimum (PmHSP90), and examined the expression levels of the gene after exposure to thermal stressors, copper metal, and endocrine-disrupting chemicals, including bisphenol A (BPA) and polychlorinated biphenyl (PCB). The complete ORF of PmHSP90 was 2,130-bp long, encoding a 709-amino acid-long polypeptide (81.62?kDa), and bearing characteristics of the HSP90 family and conserved domains. Real-time (RT)-PCR analyses revealed different expression patterns after exposure to heat, metals, and chemicals. The expression of PmHSP90 was significantly upregulated by increased thermal stresses, with the highest changes of 2.4-fold and 1.9-fold occurring after 24?h at 25?C and 30?C, respectively. The gene expression dramatically increased (2.1 to 8.9-fold changes) with increasing concentrations of copper (one-way ANOVA, P?<?0.01). Treatment with BPA or PCB, however, did not induce significant changes in PmHSP90 expression. These data suggest that the dinoflagellate HSP90 responds to thermal stressors, but may differentially respond to toxic substances such as metals and endocrine-disrupting chemicals.</P>

Trace element analysis of three tissues from Eurasian otters (Lutra lutra) in South Korea.

Kang, Sukmo,Kang, Jung-Hoon,Kim, Soohee,Lee, Seung Heon,Lee, Seungwoo,Yu, Hee Jeong,Oh, Su-Jun,Park, Jung-Duck,Nam, Ki-Hoan,Han, Sung Yong,Lim, Jong-Deock,Ryu, Doug-Young Chapman Hall ; Kluwer Academic Publishers 2015 Ecotoxicology Vol.24 No.5

<P>Eurasian otters (Lutra lutra) are endangered worldwide, but the specific cause of their decline has not been determined. This study analyzed the concentrations of potentially toxic trace elements, including As, Cd, Pb, Hg, Se, Cu, Mn, and Zn, in the liver, kidney, and lung tissues of Eurasian otters in South Korea. There were high individual variations in the tissue concentrations of all the elements analyzed. The kidneys had the highest concentrations of Cd and Se among the three tissue groups, and the livers had the highest concentrations of Cu, Mn, Zn, and Hg. The Pb and As concentrations in the livers were not significantly different from those in the kidneys, and the lungs had the lowest concentrations of all the elements analyzed. The age-related bioaccumulation of Cd and Hg was evident in the three tissue groups, and of Se in the kidneys. The Pb concentration was higher in the livers of juveniles compared with those of adults and the Zn concentration was higher in the lungs of juveniles. There were no apparent gender differences in the concentrations of the elements analyzed among the tissue groups. The Se concentration correlated with the Hg concentration in the livers and kidneys, and with the Cd concentration in the kidneys. The Hg and Cd levels correlated in the three tissue groups. The Cu and Zn levels also correlated in the livers and kidneys. In general, the element concentrations were within the ranges reported by previous studies of this species from European countries, except for Cd and Hg, the levels of which were mostly lower than those reported previously. These findings may provide baseline information to facilitate the conservation of the Eurasian otter. To the best of our knowledge, this is the first available study of trace element concentrations in the tissues of Eurasian otters from South Korea or Asian countries.</P>

A correction method for streak artifacts in gradient-echo EPI using spin-echo EPI reference data.

Chung, Jun-Young,Han, Yeji,Cho, Zang-Hee,Park, Hyunwook Chapman Hall 2012 Magma Vol.25 No.3

<P>To analyze the streak artifacts in a gradient-echo echo planar imaging (GE-EPI) sequence and to propose a correction method for the Nyquist ghost artifacts that does not cause streak artifacts in the GE-EPI imaging.</P>

Determination of mRNA expression of DMRT93B, vitellogenin, and cuticle 12 in Daphnia magna and their biomarker potential for endocrine disruption.

Kim, Jungkon,Kim, Younghee,Lee, Sangwoo,Kwak, Kyunghee,Chung, Wook-Jin,Choi, Kyungho Chapman Hall ; Kluwer Academic Publishers 2011 Ecotoxicology Vol.20 No.8

<P>We explored the use of molecular genetic biomarkers for endocrine disruption in Daphnia magna after the exposure to fenoxycarb (FOC), a model juvenile hormone analog. For this purpose, the mRNA expression patterns of DMRT93B (DMRT, sex determination), cuticle 12 (CUT, molting), and vitellogenin (VTG, embryo development) were determined in D. magna. Furthermore, these results were compared with developmental abnormality and reproduction performance. The fold changes of CUT and VTG mRNA expression showed significant dose-response relationship with FOC exposure. Relative mRNA expressions of DMRT and CUT showed notable changes at as low as 1 ng/l FOC. After chronic exposure FOC significantly delayed the first day of reproduction and decreased the number of young and growth rate even at 10 ng/l FOC. A concentration-dependant trend in reproduction effect was also observed. Developmental abnormality such as poorly developed second antennae and curved or unextended shell spines were observed. These results suggest that the three mRNAs, i.e., DMRT, CUT, and VTG can be used as biomarkers of endocrine disrupting effects in D. magna.</P>