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To assess the effects of slaughter weight and sex on APGS (Animal Products Grading Service) quality and APGS yield grade of Korean Hanwoo (n = 20,881) cattle, data were collected from cow, bull, and steer carcasses during a 1-yr period. Factors used to determine quality grade (marbling, meat color, fat color, texture, and overall maturity score) and yield grade (cold carcass weight, adjusted fat thickness, and longissimus muscle area) by the Korean grading system were recorded. Both yield and quality grades were improved (P 0.01). Longissimus muscle area, adjusted fat thickness, and marbling score increased (P < 0.01) with carcass weight. Bull carcasses showed higher yield but lower quality than those of cows or steers (P < 0.01). The quality grade of steer carcasses was higher (P < 0.01) than that of cow carcasses due to higher marbling scores, lower maturity scores, and heavier carcass weights. Hanwoo carcasses with larger longissimus muscle areas in relation to their carcass weight had lower APGS quality grades. The APGS quality grades were different between yield grade A and B carcasses (P < 0.01), but quality grade was not improved by increased fat thickness beyond the point of yield grade B. Adjusted fat thickness and marbling score showed significant (P < 0.01) differences among all yield grade classes, and this resulted in increased quality grade as yield grade decreased. Adjusted fat thickness showed the strongest correlation (r = -0.63) with yield grade, whereas marbling score had the strongest correlation (r = 0.81) with quality grade. Results showed a negative effect of castration on yield but a positive effect on quality. Also, data showed that Hanwoo carcasses with heavier weights had higher quality grades than those of lighter weight.

bryos reconstructed by using donor cells: iPS cell vs. AID transfected cell* Tae Suk Kim1, Sang Ki Baek2, Song Yi Moon2, Sang Jin Jin2, Yeoung-Gyu Ko3, Sung Woo Kim3, Hae-Geum Park3, Hwan-Hoo Seong3 and Joon Hee Lee2 1Institute of Agriculture & Life Science, College of Agriculture and Life Sciences, Gyeongsang National University, Jinju, 660-701, Republic of Korea, Jin ju, Republic of Korea 2Animal Development & Biotechnology Group, Department of Animal Bioscience, College of Agriculture & Life Sciences, Gyeongsang National University, Jin ju, Republic of Korea 3National Institute of Animal Science, Rural Development Administration, Namwon, Republic of Korea

This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3-L1 cells and HFD adipose tissue.

The effective management of endangered animal genetic resources is one of the most important concerns of modern breeding. Evaluation of genetic diversity and relationship of local breeds is an important factor towards the identification of unique and valuable genetic resources. This study aimed to analyze the genetic diversity and population structure of six Korean native chicken breeds (n = 300), which were compared with three imported breeds in Korea (n = 150). For the analysis of genetic diversity, 30 microsatellite markers from FAO/ISAG recommended diversity panel or previously reported microsatellite markers were used. The number of alleles ranged from 2 to 15 per locus, with a mean of 8.13. The average observed heterozygosity within native breeds varied between 0.46 and 0.59. The overall heterozygote deficiency (F-IT) in native chicken was 0.234 +/- 0.025. Over 30.7% of F-IT was contributed by within-population deficiency (F-IS). Bayesian clustering analysis, using the STRUCTURE software suggested 9 clusters. This study may provide the background for future studies to identify the genetic uniqueness of the Korean native chicken breeds.

DNA methyltransferase1o (Dnmt1o), which is specific to oocyte and preimplantation embryo, plays a role in maintaining DNA methylation in mammalian cells. Here, we investigated the methylation status of CpGs sites in the Dnmt1o 5'-flanking region in germ cells at different stages of oogenesis or spermatogenesis. The methylation levels of the CpG sites at the 5'-flanking regions were hypermethylated in growing oocytes of all follicular stages, while the oocytes in meiotic metaphase II (MII) were demethylated. The methylation pattern within the CpGs sites in the 5'-flanking region, however, was dramatically changed during spermatogenesis. We observed that there was significant non-CpG methylation both in MII oocytes and spermatocytes. Although a low methylation level in non-CpG sites was observed in primary and secondary oocytes, the CpA site of position 25 and CpT site of position 29 within the no-CpG region in the 5'-flanking region of Dnmt1o was highly methylated in MII oocytes. During spermatogenesis, the low degree of methylation at CpG sites in spermatocytes increased to a higher degree in sperm, while the high ratio of methylation in non-CpG sites in spermatocytes decreased. Together, germ cells showed inverted methylation patterns between CpG and non-CpG sites in the Dnmt1o 5'-upstream region, and the methylation pattern during oogenesis did not drastically change, remaining generally hypomethylated at the MII stage.

Several biomarkers are routinely used clinically for predicting preterm labor; however, these factors are either nonspecific or detected too late. Here, we performed protein profiles in preterm- and term-derived human umbilical cord by using 2DE. Approximately 200 different proteins were identified between preterm- and term-delivered umbilical cords. Among them, 48 proteins were identified. A comparison of preterm proteome to that of term proteome revealed potential candidates for biomarkers, such as hypoxia-inducible proteins, phosphorylated heat-shock protein 27 (HSP27), transgelin, vimentin, and transferrin that are specific to preterm umbilical cords. Especially, HSP27 in preterm-derived umbilical cords shows a significant increase in the mono- and tetra-phosphorylation. The real importance of all of HSP27 phosphorylation as well as hypoxia-inducible factor 1alpha, and glyceraldehyde 3-phosphate dehydrogenase require further validation in vitro and in vivo; nevertheless, we believe that they could represent promising diagnostic targets for detection of sudden early delivery. In conclusion, the results of the current study may provide important insights into the molecular mechanisms underlying umbilical cord development.

Background Obesity is a health hazard that is associated with a number of diseases and metabolic abnormalities, such as type-2 diabetes, hypertension, dyslipidemia, and coronary heart disease. In the current study, we investigated the effects of Citrus aurantium flavonoids (CAF) on the inhibition of adipogenesis and adipocyte differentiation in 3T3-L1 cells. Methods During adipocyte differentiation, 3T3-L1 cells were treated with 0, 10, and 50 μg/ml CAF, and then the mRNA and protein expression of adipogenesis-related genes was assayed. We examined the effect of CAF on level of phosphorylated Akt in 3T3-L1 cells treated with CAF at various concentrations during adipocyte differentiation. Results The insulin-induced expression of C/EBPβ and PPARγ mRNA and protein were significantly down-regulated in a dose-dependent manner following CAF treatment. CAF also dramatically decreased the expression of C/EBPα, which is essential for the acquisition of insulin sensitivity by adipocytes. Moreover, the expression of the aP2 and FAS genes, which are involved in lipid metabolism, decreased dramatically upon treatment with CAF. Interestingly, CAF diminished the insulin-stimulated serine phosphorylation of Akt (Ser473) and GSK3β (Ser9), which may reduce glucose uptake in response to insulin and lipid accumulation. Furthermore, CAF not only inhibited triglyceride accumulation during adipogenesis but also contributed to the lipolysis of adipocytes. Conclusions In the present study, we demonstrate that CAF suppressed adipogenesis in 3T3-L1 adipocytes. Our results indicated that CAF down-regulates the expression of C/EBPβ and subsequently inhibits the activation of PPARγ and C/EBPα. The anti-adipogenic activity of CAF was mediated by the inhibition of Akt activation and GSK3β phosphorylation, which induced the down-regulation of lipid accumulation and lipid metabolizing genes, ultimately inhibiting adipocyte differentiation.

In this paper, we report on the effects of a diet containing cloned-cattle meat on the reproductive parameters in pregnant rabbits. The artificially inseminated rabbits (gestation day 0) were fed a diet containing 5% or 10% of normal or cloned-cattle meat during the gestation period. Rabbits fed commercial pellet (no additional supplementations) were used as the control. Supplementation of cloned-cattle meat diets did not have any toxicologically significant effects on reproductive performance in dams (body weight, clinical signs, organ weight, and cesarean section analysis). And it also did not affect on fetal development (body and placental weight, and external, visceral and skeletal findings) compared to the controls. The only difference was a food consumption in the first week of gestation for all meat-based diet groups (p < 0.05, 0.01, and 0.001, respectively). Our results collectively suggest that there are no obvious differences in reproductive parameters in pregnant rabbits fed cloned-cattle meat.

We identified a 3.4-kb 5′-flanking region of the rPL-I gene and examined its promoter activity using rat trophoblast Rcho-1 cells. A regulatory element between base pairs (bp) −2,487 and −2,310 in the 5′-flanking region was essential for maximum promoter activity of the rPL-I gene. This regulatory element was further characterized between bp −2,443 to −2,415 and −2,374 to −2,345. Electrophoretic mobility shift analysis showed that the interaction of nuclear extract proteins from differentiated Rcho-1 cells was inhibited by competition with a GATA-like sequence in the promoter, but not by a mutated GATA sequence. Moreover, the promoter activity of 2487 eLuc containing two novel GATA sites was significantly elevated by co-transfection of a GATA-2 expression vector in proliferating Rcho-1 cells. Our results demonstrate that GATA-2 is involved in multiple promoter regions to activate the specific expression of the rPL-I gene in placental tissue.

The objective of this study was to investigate hormonal and TGF-beta(1) characterizations of delayed parturition in the SCNT recipients (Korean native beef cattle: Hanwoo). The SCNT blastocysts produced by Hanwoo fetal fibroblast cells were transferred into the synchronized Hanwoo recipients. The artificially inseminated Hanwoo recipients (AI-R) were used as control. All AI-R were labored by natural delivery. The SCNT recipients (SCNT-R) with no signs of delivery were operated by Caesarean section. The blood and placentomes were collected during parturition. The weight of placentomes in SCNT-R (n=12, 301+/-41.22 g) was significantly higher than that of AI-R (n=10, 204.8+/-24.89 g) (p

Methylation of DNA is involved in tissue-specific gene control, and establishment of DNA methylation pattern in the genome is thought to be essential for embryonic development. Three isoforms of Dnmt1 (DNA methyltransferase 1) transcripts, Dnmt1s, Dnmt1o, and Dnmt1p, are produced by alternative usage of multiple first exons. Dnmt1s is expressed in somatic cells. Dnmt1p is found only in pachytene spermatocytes, whereas Dnmt1o is specific to oocytes and preimplantation embryos. Here we determined that there is a tissue-dependent differentially methylated region (T-DMR) in the 5' region of Dnmt1o but not in that of the Dnmt1s/1p. The methylation status of the Dnmt1o T-DMR was distinctively different in the oocyte from that in the sperm and adult somatic tissues and changed at each stage from fertilization to blastocyst stage, suggesting that active methylation and demethylation occur during preimplantation development. The T-DMR was highly methylated in somatic cells and embryonic stem cells. Analysis using Dnmt-deficient embryonic stem cell lines revealed that Dnmt1, Dnmt3a, and Dnmt3b are each partially responsible for maintenance of methylation of Dnmt1o T-DMR. In particular, there are compensatory and cooperative roles between Dnmt3a and Dnmt3b. Thus, the regulatory region of Dnmt1o, but not of Dnmt1s/1p, appeared to be a target of DNA methylation. The present study also suggested that the DNA methylation status of the gene region dynamically changes during embryogenesis independently of the change in the bulk DNA methylation status.

The first cell differentiation event in mammalian embryogenesis segregates inner cell mass lineage from the trophectoderm at the blastocyst stage. Oct-4, a member of the POU family of transcription factors, is necessary for the pluripotency of the inner cell mass lineage. Embryonic stem (ES) cells, which contribute to all of embryonic lineages, express the Oct-4 gene. Trophoblast stem (TS) cells, which have the ability to differentiate into trophoblast lineage in vitro, never contribute to embryonic proper tissues in chimeras and differentiate only into trophoblastic cells in the placenta. Expression of the Oct-4 gene was undetectable and severely repressed in trophoblastic lineage, including the stem cells. We found that the culture of TS cells with 5-aza-2'-deoxycytidine or trichostatin A caused the activation of the Oct-4 gene. Analysis of the DNA methylation status of mouse Oct-4 gene upstream region revealed that Oct-4 enhancer/promoter region was hypomethylated in ES cells but hypermethylated in TS cells. Furthermore, in vitro methylation suppressed Oct-4 enhancer/promoter activity in reporter assay. In the placenta of Dnmt1(n/n) mutant mice, most of the CpGs in the enhancer/promoter region were unmethylated, and Oct-4 gene expression was aberrantly detected. Chromatin immunoprecipitation assay revealed that Oct-4 enhancer/promoter region was hyperacetylated in ES cells compared with TS cells, thus demonstrating that DNA methylation status is closely linked to the chromatin structure of the Oct-4 gene. Here we propose that the epigenetic mechanism, consisting of DNA methylation and chromatin remodeling, underlies the developmental stage- and cell type-specific mechanism of Oct-4 gene expression.

This study examined the anti-obesity effect and mechanism of action of blueberry peel extracts (BPE) in 3T3-L1 cells and high-fat diet (HFD)-induced obese rats. The levels of lipid accumulation were measured, along with the changes in the expression of genes and proteins associated with adipocyte differentiation in 3T3-L1 cells. Evidenced by Oil-red O staining and triglyceride assay, BPE dose-dependently inhibited lipid accumulation at concentrations of 0, 50, and 200 µg/ml. BPE decreased the expression of the key adipocyte differentiation regulator C/EBPβ, as well as the C/EBPα and PPARγ genes, during the differentiation of preadipocytes into adipocytes. Moreover, BPE down-regulated adipocyte-specific genes such as aP2 and FAS compared with control adipocytes. The specific mechanism mediating the effects of BP revealed that insulin-stimulated phosphorylation of Akt was strongly decreased, and its downstream substrate, phospho-GSK3β, was downregulated by BPE treatment in 3T3-L1 cells. Together, these data indicated that BP exerted anti-adipogenic activity by inhibiting the expression of PPARγ and C/EBPβ and the Akt signaling pathway in 3T3-L1 adipocytes. Next, we investigated whether BP extracts attenuated HFD-induced obesity in rats. Oral administration of BPE reduced HFD-induced body weight gain significantly without affecting food intake. The epididymal or perirenal adipose tissue weights were lower in rats on an HFD plus BPE compared with the tissue weights of HFD-induced obese rats. Total cholesterol and triglyceride levels in the rats fed BPE were modestly reduced, and the HDL-cholesterol level was significantly increased in HFD plus BP-fed rats compared with those of HFD-fed rats. Taken together, these results demonstrated an inhibitory effect of BP on adipogenesis through the down-regulation of C/EBPβ, C/EBPα, and PPARγ and the reduction of the phospho-Akt adipogenic factor in 3T3-L1 cells. Moreover, BPE reduced body weight gain and inhibited fat accumulation in an HFD-induced animal model of obesity.

Background Somatic cell cloning by nuclear transfer (SCNT) in pig is clearly of great benefit for basic research and biomedical applications. Even though cloned offspring have been successfully produced in pig, SCNT is struggling with the low efficiency. Results In the present study, we investigated differentially expressed proteins of the extraembryonic tissue from pig SCNT fetus compared to control (normal) fetus. We obtained the extraembryonic tissue from embryos at day 35 of pregnancy and examined the protein expression profiles using two-dimensional electrophoresis (2-D) and Western blotting. The extraembryonic tissue of fetus in control pregnancy was compared to the extraembryonic tissue of SCNT fetus, which showed an abnormally small size and shape as well as exhibited abnormal placental morphology compared to control fetus. A proteomic analysis showed that the expression of 33 proteins was significantly increased or decreased in the extraembryonic tissue of SCNT fetus compared to control fetus. The differentially expressed proteins in the extraembryonic tissue of SCNT fetus included ATP or lipid binding proteins, antioxidant proteins, translation elongation factors, and transcription factors. Western blotting analysis indicated that antioxidant enzymes and anti-apoptotic proteins were down-regulated; however, the expression levels of apoptotic proteins, Bax and Hsp27, were increased in the extraembryonic tissue of SCNT fetus. Moreover, immunohistochemical analysis also showed that the expression of the catalase or GPX genes was decreased in the extraembryonic tissue with SCNT fetus compared to those with control fetus. In addition, we observed a significant decrease in DNA methytransferase1 (Dnmt1) expression in SCNT extraembryonic tissue, and the expression levels of Dnmt3a and Dnmt3b were abnormally higher in SCNT fetus compared to control fetus. Moreover, a marked increase in the frequency of TUNEL-positive cells was observed in the extraembryonic tissue in SCNT fetus. Conclusion These results demonstrated that pig SCNT fetus showed abnormal protein expression in the extraembryonic tissue, and extensive apoptosis occurred in the extraembryonic tissue of the SCNT fetus due to an increase in apoptotic protein expression or a decrease in antioxidant protein expression.