Seunghyung Lee

Effect of cholesterol-loaded-cyclodextrin on sperm viability and acrosome reaction in boar semen cryopreservation

This study was undertaken to examine the effect of cholesterol-loaded-cyclodextrin (CLC) on boar sperm viability and spermatozoa cryosurvival during boar semen cryopreservation, and methyl-β-cyclodextrin (MBCD) was treated for comparing with CLC. Boar semen treated with CLC and MBCD before freezing process to monitor the effect on survival and capacitation status by flow cytometry with appropriate fluorescent probes. Sperm viability was higher in 1.5mg CLC-treated sperm (76.9±1.01%, P<0.05) than un-treated and MBCD-treated sperm before cryopreservation (58.7±1.31% and 60.3±0.31%, respectively). For CTC patterns, F-pattern was higher in CLC treated sperm than MBCD-treated sperm, for B-pattern was higher in CLC-treated sperm than fresh sperm (P<0.05). For AR pattern (an acrosome-reacted sperm) was lower in CLC-treated sperm than MBCD-treated sperm (P<0.05). Moreover, we examined in vitro development of porcine oocytes after in vitro fertilization using CLC-treated frozen-thawed semen, in which CLC treatment prior to freezing and thawing increased the development of oocytes to blastocyst stage in vitro. In conclusion, CLC could protect the viability of spermatozoa from cryodamage prior to cryopreservation in boar semen.

Change of uterine histroph proteins during follicular and luteal phase in pigs

The aim of this study was to examine protein expression patterns of uterine histroph (UH) during the follicular phase (FP) and luteal phase (LP) in pigs. Forty-nine common proteins were identified from FP and LP samples; five were significantly down-regulated (>1.5-fold), while 15 were significantly up-regulated (>1.5-fold) in LPUH compared with FPUH (P<0.05). The 20 differentially-expressed proteins are involved in cell proliferation, cell responses, translation, transport, and metabolism and their molecular functions include nucleic acid binding, oxygen activity, enzymatic activity, growth activity, iron binding, and redox binding. Protein expression of vascular endothelial growth factor D (VEGFD), coatomer subunit gamma-2 (G2COP), collagen alpha 4 chain (COL4), cysteine rich protein 2 (CRP2), myoglobin (MYG), and galactoside 3-L-fucosyltransferase 4 (FUT4) was analyzed by Western blotting. These proteins were significantly higher in LPUH compared to FPUH (P<0.05). These data expand our understanding of changes in the intrauterine environment during the pre-implantation period in pigs.

Effect of Antioxidants on Arsenite Exposed Porcine Embryonic Development

This study was to investigate the role of antioxidants on development in arsenite exposed porcine embryos. Oocytes were collected from porcine ovary, and then matured for 44 h. Maturated oocytes were incubated with sperm for 6 h, and fertilized oocytes with sperm (embryos) cultured for 48 h. After, embryos were culture with arsenite and/or antioxidants (melatonin, silymarin, curcumin and vitamin) for 120 h. Formation of pre-morulae, morulae and blastosysts rate was measured using microscope. In results, 10, 100 and 100 nM arsenite significantly decreased morulae and blastocysts formation compared to control in pigs (P<0.05). 10 μM silymarin and 100 μM vitamin E increased blastocyst formation compared to 10 nM arsenite exposed embryos, but there were no significantly among the treatment, and 1 nM melatonin and 5 μM curcumin did not influence blastocysts formation in 10 nM arsenite exposed embryos. In summary, arsenite decreased embryo development, 10 μM silymarin, 100 μM vitamin E, 1 nM melatonin and 5 μM curcumin had no positive effect to blastocyst formation in arsenite exposed porcine embryos. Therefore, we suggest that little arsenite may have negative effect to embryo development, and silymarin, vitamin E, melatonin and curcumin could not rescue embryo development from damage by arsenite in pigs.

Chondrogenic Differentiation of Porcine Skin-Derived Stem Cells with Different Characteristics of Spontaneous Adipocyte Formation

The purpose of this study is to confirm whether spontaneous adipocyte generation during chondrogenic induction culture affects the chondrogenic differentiation of porcine skin-derived stem cells (pSSCs). For this purpose, chondrogenic differentiation characteristics and specific marker gene expression were analyzed using cell lines showing different characteristics of spontaneous adipocyte formation. Of the four different lines of pSSCs, the pSSCs-IV line showed higher Oil red O (ORO) and glycosaminoglycan (GAG) extraction levels. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the levels of adipogenic markers peroxisome proliferator-activated receptor gamma 2 (PPARγ2) and adipocyte Protein 2 (aP2) mRNAs were significantly higher in pSSCs-IV than those of the other pSSC lines (P<0.05). Among three chondrogenic markers, collagen type II (Col II) and sex determining region Y-box (Sox9) mRNAs were strongly expressed in pSSCs-IV (P<0.05), but not in aggrecan (Agg), which was significantly higher in pSSCs-II (P<0.05). These results demonstrate that the spontaneous adipocyte generation during chondrogenic differentiation has a positive effect on the chondrogenesis of pSSCs. More research is needed on the correlation between adipocyte generation and cartilage formation.

Effect of uterine histotroph on embryo development in pigs

The aim of this study was to investigate the effect of uterine histotroph on embryo development and the expression of cysteine-rich protein 2 (CRP2), coatomer subunit gamma-2 (G2COP), myoglobin (MYG), vascular endothelial growth factor D (VEGFD), collagen alpha 4 chain (COL4) and galactoside 3-L-fucosyltransferase 4 (FUT4) proteins in porcine embryo during pre-implantation. Uterine histotroph (UH) was collected from uterine horn on corpus albican phase, and embryos were cultured in porcine zygote medium with UH for 168 hours. Cleavage and blastocyst formation of embryo were detected at 168 hours after in vitro fertilization. And CRP2, G2COP, MYG, VEGFD, COL4 and FUT4 proteins were observed using confocal laser microscope. In results, embryo cleavage rate was not significantly changed by UH, but blastocyst rate was significantly (P<0.05) decreased in UH-treated embryos. Moreover, CRP2, G2COP, MYG, VEGFD, COL4 and FUT4 proteins were expressed in blastomere. CRP2 in embryo was significantly overexpressed (P<0.05), but not G2COP, MYG, VEGFD, COL4 and FUT4 proteins. In summary, UH on corpus albican phase was increased CRP2 protein in embryo, and inhibited blastocyst formation in preimplantation porcine embryos, suggesting that CRP2 may play an interrupter on embryo development in pigs.