• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.114-114
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• 2003

Epidermal growth factor (EGF) induces well-documented mitogenic and differentiating effects on murine and bovine preimplantation embryos. However, the effects of EGF on apoptosis and implantation-related gene expression in bovine embryos developing in vitro have not been evaluated. The objective of this study was to determine the effects of exogenous EGF in the presence and absence of BSA on the preimplantation development of bovine embryos. In addition, we measured cell number, apoptosis, and expression of apoptosis and implantation-related genes of the blastocysts that developed in these culture conditions. In vitro produced bovine embryos were randomly cultured in the same medium containing 0 or 10 ng/ml EGF in the presence and absence of 0.8% BSA. More 2-cell embryos developed into blastocysts at day 7 when BSA was present than when BSA was absent. The addition of 10 ng/$m\ell$ EGF into the medium did not significantly increase the developmental rate and the cell numbers per blastocyst. However, addition of EGF in the presence of 0.8% BSA significantly reduced the degree of apoptosis in the blastocysts (P<0.01). To investigate whether EGF modulates mRNA expression of apoptosis-related genes, mRNA was prepared from single blastocysts and each preparation was subjected to RT-PCR for Bcl-2 and Bax transcripts. EGF did not alter the relative abundance of Bax gene expression in the presence of BSA, but increase Bcl-2 (P<0.01) The relative abundance of Interferon tau expression was increased by EGF treatment in the presence of BSA. These results suggest that EGF and BSA synergistically enhance Bcl-2 and interferone tau gene expression, which may result in a net increase in viability in bovine embryos.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.99-99
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• 2003

Epidermal growth factor (EGF) induces well-documented mitegenic and differentiating effects on murine and bovine preimplantation embryos. However, the effects of EGF on apoptosis and implantation-related gene expression in bovine embryos developing in vitro have not been evaluated. The objective of this study was to determine the effects of exogenous EGF in the presence and absence of BSA on the preimplantation development of bovine embryos. In addition, we measured cell number, apoptosis, and expression of apoptosis and implantation-related genes of the blastocysts that developed in these culture conditions. In vitro produced bovine embryos were randomly cultured in the same medium containing 0 or 10 ng/$m\ell$ EGF in the presence and absence of 0.8% BSA. More 2-cell embryos developed into blastocysts at day 7 when BSA was present than when BSA was absent. The addition of 10 ng/$m\ell$ EGF into the medium did not significantly increase the developmental rate and the cell numbers per blastocyst. However, addition of EGF in the presence of 0.8% BSA significantly reduced the degree of apoptosis in the blastocysts (P< 0.01). To investigate whether EGF modulates mRNA expression of apoptosis-related genes, mRNA was prepared from single blastocysts and each preparation was subjected to RT-PCR for Bcl-2 and Bax transcripts. EGF did not alter the relative abundance of Bax gene expression in the presence of BSA, but increase Bcl-2 (P < 0.01). The relative abundance of Interferon tau expression was increased by EGF treatment in the presence of BSA. These results suggest that EGF and BSA synergistically enhance Bcl-2 and interferone tau gene expression, which may result in a net increase in viability in bovine embryos.

• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.341-348
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• 2011

Sloan-Kettering virus gene product of a cellular protooncogene c-Ski is an unique nuclear pro-oncoprotein and belongs to the Ski/Sno proto-oncogene family. Ski plays multiple roles in a variety of cell types, it can induce both oncogenic transformation and terminal muscle differentiation when expressed at high levels. The aim of the present study was to locate Ski protein in rat ovaries in order to predict the possible involvement of Ski in follicular development and atresia. First, expression of c-Ski mRNA in the ovaries of adult female rats was confirmed by RT-PCR. Then, ovaries obtained on the day of estrus were subjected to immunohistochemical analysis for Ski and proliferating cell nuclear antigen (PCNA) in combination with terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL). Ski was expressed in granulosa cells that were positive for TUNEL, but negative for PCNA, regardless of the shape and size of follicles. Expression of Ski in TUNEL-positive granulosa cells, but not in PCNA-positive granulosa cells, was also verified in immature hypophysectomized rats having a single generation of developing and atretic follicles by treatment with equine chorionic gonadotropin (eCG). These results indicate that Ski is profoundly expressed in the granulosa cells of atretic follicles, but not in growing follicles, and suggest that Ski plays a role in apoptosis of granulosa cells during follicular atresia.

• Molecules and Cells
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• v.28 no.1
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• pp.43-48
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• 2009

By screening C. elegans mutants for severe defects in germline proliferation, we isolated a new loss-of-function allele of cdc-25.1, bn115. bn115 and another previously identified loss-of-function allele nr2036 do not exhibit noticeable cell division defects in the somatic tissues but have reduced numbers of germ cells and are sterile, indicating that cdc-25.1 functions predominantly in the germ line during postembryonic development, and that cdc-25.1 activity is probably not required in somatic lineages during larval development. We analyzed cell division of germ cells and somatic tissues in bn115 homozygotes with germline-specific anti-PGL-1 immunofluorescence and GFP transgenes that express in intestinal cells, in distal tip cells, and in gonadal sheath cells, respectively. We also analyzed the expression pattern of cdc-25.1 with conventional and quantitative RT-PCR. In the presence of three other family members of cdc-25.1 in C. elegans, defects are observed only in the germ line but not in the somatic tissues in cdc-25.1 single mutants, and cdc-25.1 is expressed predominantly, if not exclusively, in the germ line during postembryonic stages. Our findings indicate that the function of cdc-25.1 is unique in the germ line but likely redundant with other members in the soma.

• The Korean Journal of Nuclear Medicine
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• v.37 no.3
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• pp.178-189
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• 2003

Purpose: Cellular uptakes of $^{99m}Tc-sestamibi(MIBI)\;and\;\;^{99m}Tc-tetrofosmin$ into cancer cell lines expressing multidrug resistance(MDR) were investigated and compared. The effects of verapamil and cyclosporin A, well-known multidrug resistant reversing agents, on cellular uptakes of both tracers were also compared. Materials and Methods: Doxorubicin-resistant HCT15/CL02 human colorectal cell and doxorubicin-resistant K562(Adr) and vincristine-resistant K562(Vcr) human leukemic cells were studied. RT-PCR analysis was used for the detection of mdr1 mRNA expression. MDR-reversal effects with verapamil and cyclosporine A were evaluated at different drug concentrations after incubation with MIBI and tetrofosmin for 1, 15, 30, 45 and 60 min, using single-cell suspensions at $1{\times}10^6cells/ml$ incubated at $37^{\circ}C$. Radioactivity in supernatants and pellets were measured with gamma well counter. Results: The cellular uptakes of MIBI and tetrofosmin in K562(Adr) and K562(Vcr) were lower than those of parental K562 cell. In HCT15/CL02 cells and K562(Adr) cells, there were no significant difference in cellular uptakes of both tracers, but cellular uptake of MIBI was higher than that of tetrofosmin in K562(Vcr) cells. Coincubation with verapamil resulted in a increase In cellular uptakes of MIBI and tetrofosmin. Verapamil increased cellular uptakes of MIBI and tetrofosmin by HCT15/CL02 cell by 11.9- and 6.8-fold, by K562(Adr) cell by 14.3- and 8-fold and by K562(Vcr) cell by 7- and 5.7-fold in maximum, respectively. Cyciosporin A increased cellular uptakes of MIBI and tetrofosmin by HCT15/CL02 cell by 10- and 2.4-fold, by K562(Adr) cell by 44- and 13-fold and by K562(Vcr) cell by 18.8- and 11.8-fold in maximum, respectively Conclusion: Taking together, MIBI and tetrofosmin are considered as suitable radiopharmaceuticals for defecting multidrug resistance. However, MIBI seems to be a better tracer than tetrofosmin for evaluating MDR reversal effect of the modulators. Since cellular uptakes of both tracers might differ in different cell types, further experiments regarding differences in cellular uptakes between cell types should be explored.

• Development and Reproduction
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• v.12 no.1
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• pp.77-86
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• 2008

This study was carried out to investigate the effect of leukemia inhibitory factor (LIF) on the derivation of mouse ES cells from isolated blastomeres. Two-cell stage mouse embryos were obtained from superovulated BDF1 female mice. Collected embryos were cultured to blastocyst stage in culture medium supplemented with 0, 1,000, 2,500 or 5,000 U/mL of LIF. Cultured blastocysts were examined by counting the number of cells in the inner cell mass (ICM) and trophectoderm (TE) using differential staining method. When 2-cell embryos were cultured with 2,500 U/ml of LIF, the cell numbers of ICM significantly increased in comparing with those of the control($21.0{\pm}4.0$ vs. $15.9{\pm}5.0$, P<0.01) and 1,000 U/mL of LIF-containing group ($21.0{\pm}4.0$ vs. $16.6{\pm}4.9$, P<0.05). We used an ES cell establishment medium with 20% Knockout Serum Replacement and 0.01 mg/mL ACTH instead of fetal bovine serum. Establishing efficacy of ES cell lines were the highest in 2,500 U/mL of LIF-containing group as 36.7% (11/30). This culture medium was applied to the culture of isolated blastomeres and to derivate ES cell lines. Three ES cell lines (21.4%) from isolated blastomeres of 2-cell stage embryos were established. In further experiments, we could establish one ES cell line (4.0%) from single blastomere of 4-cell stage embryo. The subcultured ES cells and their embryoid bodies were characterized by analyzing gene expression for undifferentiation and differentiation marker gene using immunocytochemistry and RT-PCR. In conclusion, LIF supplementation in culture medium could increase the cell number in ICM of blastocysts and support derivation of ES cell lines from isolated blastomeres.

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.1
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• pp.7-12
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• 2010

The full-length cDNA of the porcine peroxisomal ${\Delta}^3$,${\Delta}^2$-enoyl-CoA isomerase (PECI) gene encodes a monofunctional peroxisomal ${\Delta}^3$,${\Delta}^2$-enoyl-CoA isomerase. Cloning and sequencing of the porcine PECI cDNA revealed the presence of an 1185-base pair open reading frame predicted to encode a 394-amino acid protein by the 5'rapid amplification of cDNA ends (5'RACE) and EST sequences. The porcine PECI gene was expressed in seven tissues (heart, liver, spleen, lung, kidney, skeletal muscle, fat) which was revealed by reverse transcriptase-polymerase chain reaction (RT-PCR). The porcine PECI was mapped to SSC71/2 p11-13 using the somatic cell hybrid panel (SCHP) and the radiation hybrid panel (RH) (LOD score 12.84). The data showed that PECI was closely linked to marker S0383. A C/T single nucleotide polymorphism in PECI exon 10 (3'UTR) was detected as a PvuII PCR-RFLP. Association analysis in our experimental pig population showed that different genotypes of PECI gene were significantly associated with the Average Backfat thickness (ABF) (p<0.05) and Buttock backfat thickness (p<0.01).

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.7
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• pp.931-937
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• 2006

The full-length cDNA of the porcine SMPX gene was obtained by the rapid amplification of cDNA ends (RACE). The nucleotide sequences and the predicted protein sequences share high sequence identity with both human and mouse. The promoter of SMPX was sequenced and then analyzed to find the promoter binding sites. The reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that SMPX has a high level of expression in heart and skeletal muscle, a very low expression in lung and spleen and no expression in liver, kidney, fat and brain. Moreover, SMPX has a differential expression level in skeletal muscle, the expression in 65-day embryos being higher than other stages. The porcine SMPX was mapped to SSCXp24 by using a somatic cell hybrid panel (SCHP) and was found closely linked to SW1903 using the radiation hybrid panel IMpRH. An A/G single nucleotide polymorphism (PCR-RFLP) in the 3'-untranslated region (3'-UTR) was detected in eight breeds. The analysis of allele frequency distribution showed that introduced pig breeds (Duroc and Large White) have a higher frequency of allele A while in the Chinese indigenous pig breeds (Qingping pig, Lantang pig, YushanBlack pig, Large Black-White pig, Small Meishan) have a higher frequencies of allele G. The association analysis using an experimental population (188 pigs), which included two cross-bred groups and three pure-blood groups, suggested that the SNP genotype was associated with intramuscular fat content.

• Tuberculosis and Respiratory Diseases
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• v.45 no.5
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• pp.984-991
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• 1998

Background: The 3p deletions has been shown to be the most frequent alteration in lung cancers, strongly suggesting the presence of at least one tumor suppressor gene in this chromosomal region. However, no solid candidate for the tumor suppressor gene(s) on 3p has as yet been identified. Recent attention has focused on a candidate 3p14.2 tumor suppressor gene, FHIT, which is located in a region that is homozygously deleted in multiple tumor cell lines and disrupted by the hereditary renal cell carcinoma t(3;8) chromosomal translocation breakpoint FHIT also spans FRA3B, the most common fragile sites in the human genome. In the present study, we have analyzed expression of the FHIT gene in lung cancer cell lines. Methods: RNA from 21 lung cancer cell lines (16 NSCLC, 5 SCLC) were extracted using standard procedures. Random-primed. first strand cDNAs were synthesized from total RNA and PCR amplication of coding exons 5 to 9 was performed. The RT-PCR products were electrophoresed in 1.5% ethidium bromide-stained agarose gels. Results: 12 of 21(57%) lung cancer cell lines exhibited absent or aberrant FHIT expression [7 of 16(44%) of non-small cell lung cancer and 5 of 5(100%) of small cell lung cancer cell lines]. Conclusion: The result shows that abnormal transcription of the FHIT gene is common in human lung cancer cell lines, especially in small cell lung cancer.

• Journal of Animal Reproduction and Biotechnology
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• v.38 no.3
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• pp.177-187
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• 2023

Background: The grading of Hanwoo (Korean native cattle) is based on four economic traits, and efforts have been continuously made to improve the genetic traits associated with these traits. There is a technology to predict the expected grade based on the 4 economic genetic SNP characteristics of Korean cattle calves using single nucleotide polymorphism (SNP) technology. Selection of highly proliferative, self-renewing, and differentiating satellite cells from cattle is a key technology in the cultured meat industry. Methods: We selected the Hanwoo with high and low-scored of genomic estimated breeding value (GEBV) by using the Hanwoo 50K SNP bead chip. We then isolated the bovine satellite cells from the chuck mass. We then conducted comparative analyses of cell proliferation, immunocytochemistry, qRT-PCR at short- and long-term culture. We also analyzed the differentiation capability at short term culture. Results: Our result showed that the proliferation was significantly high at High scored GEBV (Hs-GEBV) compared to Low scored GEBV (Ls-GEBV) at short- and long-term culture. The expression levels of Pax3 were significantly higher in Hs-GEBV bovine satellite cells at long-term culture. However, there were no significant differences in the expression levels of Pax7 between Hs- and Ls-GEBV bovine satellite cells at short- and long- term culture. The expression levels of MyoG and MyHC were significantly high at Ls-GEBV bovine satellite cells. Conclusions: Our results indicated that selection of bovine satellite cells by Hanwoo 50K SNP bead chip could be effective selection methods for massive producing of satellite cells.